Activated STAT5 proteins induce activation of the PI 3-kinase/Akt and Ras/MAPK pathways via the Gab2 scaffolding adapter

PubMed Central

2005-01-01

The active forms of STAT5A (signal transducer and activator of transcription 5A) and STAT5B are able to relieve the cytokine dependence of haematopoietic cells and to induce leukaemia in mice. We have demonstrated previously that activation of the PI3K (phosphoinositide 3-kinase) signalling cascade plays a major role in cell growth and survival induced by these proteins. Interaction between STAT5 and p85, the regulatory subunit of the PI3K, has been suggested to be required for this activation. We show in the present study that the scaffolding protein Gab2 [Grb2 (growth-factor-receptor-bound protein 2)-associated binder-2] is an essential component of this interaction. Gab2 is persistently tyrosine-phosphorylated in Ba/F3 cells expressing caSTAT5 (constitutively activated STAT5), independent of JAK2 (Janus kinase 2) activation where it interacts with STAT5, p85 and Grb2, but not with Shp2 [SH2 (Src homology 2)-domain-containing tyrosine phosphatase] proteins. Interaction of STAT5 with Gab2 was also observed in Ba/F3 cells stimulated with interleukin-3 or expressing the oncogenic fusion protein Tel–JAK2. The MAPKs (mitogen-activated protein kinases) ERK1 (extracellular-signal-regulated kinase 1) and ERK2 were constitutively activated in the caSTAT5-expressing cells and were found to be required for caSTAT5-induced cell proliferation. Overexpression of Gab2-3YF, a mutant of Gab2 incapable of binding PI3K, inhibited the proliferation and survival of caSTAT5-expressing cells as well as ERK1/2 and Akt/protein kinase B phosphorylation. Taken together, our results indicate that Gab2 is required for caSTAT5-induced cell proliferation by regulating both the PI3K/Akt and the Ras/MAPK pathways. PMID:15833084

Constitutive IDO expression in human cancer is sustained by an autocrine signaling loop involving IL-6, STAT3 and the AHR

PubMed Central

Sahm, Felix; Rauschenbach, Katharina J.; Trump, Saskia; Winter, Marcus; Ott, Martina; Ochs, Katharina; Lutz, Christian; Liu, Xiangdong; Anastasov, Natasa; Lehmann, Irina; Höfer, Thomas; von Deimling, Andreas; Wick, Wolfgang; Platten, Michael

2014-01-01

Indoleamine-2,3-dioxygenase (IDO) inhibitors have entered clinical trials based on their ability to restore anti-tumor immunity in preclinical studies. However, the mechanisms leading to constitutive expression of IDO in human tumors are largely unknown. Here we analyzed the pathways mediating constitutive IDO expression in human cancer. IDO-positive tumor cells and tissues showed basal phosphorylation and acetylation of STAT3 as evidenced by western blotting and immunoprecipitation. Inhibition of IL-6 or STAT3 using siRNA and/or pharmacological inhibitors reduced IDO mRNA and protein expression as well as kynurenine formation. In turn, IDO enzymatic activity activated the AHR as shown by the induction of AHR target genes. IDO-mediated AHR activation induced IL-6 expression, while inhibition or knockdown of the AHR reduced IL-6 expression. IDO activity thus sustains its own expression via an autocrine AHR–IL-6–STAT3 signaling loop. Inhibition of the AHR–IL-6–STAT3 signaling loop restored T-cell proliferation in mixed leukocyte reactions performed in the presence of IDO-expressing human cancer cells. Identification of the IDO-AHR-IL-6-STAT3 signaling loop maintaining IDO expression in human cancers reveals novel therapeutic targets for the inhibition of this core pathway promoting immunosuppression of human cancers. The relevance of the IDO-AHR-IL-6-STAT3 transcriptional circuit is underscored by the finding that high expression of its members IDO, STAT3 and the AHR target gene CYP1B1 is associated with reduced relapse-free survival in lung cancer patients. PMID:24657910

5-hydroxy-2-methyl-1,4-naphthoquinone, a vitamin K3 analogue, suppresses STAT3 activation pathway through induction of protein tyrosine phosphatase, SHP-1: potential role in chemosensitization.

PubMed

Sandur, Santosh K; Pandey, Manoj K; Sung, Bokyung; Aggarwal, Bharat B

2010-01-01

The activation of signal transducers and activators of transcription 3 (STAT3) has been linked with carcinogenesis through survival, proliferation, and angiogenesis of tumor cells. Agents that can suppress STAT3 activation have potential not only for prevention but also for treatment of cancer. In the present report, we investigated whether 5-hydroxy-2-methyl-1,4-naphthoquinone (plumbagin), an analogue of vitamin K, and isolated from chitrak (Plumbago zeylanica), an Ayurvedic medicinal plant, can modulate the STAT3 pathway. We found that plumbagin inhibited both constitutive and interleukin 6-inducible STAT3 phosphorylation in multiple myeloma (MM) cells and this correlated with the inhibition of c-Src, Janus-activated kinase (JAK)1, and JAK2 activation. Vanadate, however, reversed the plumbagin-induced downregulation of STAT3 activation, suggesting the involvement of a protein tyrosine phosphatase. Indeed, we found that plumbagin induced the expression of the protein tyrosine phosphatase, SHP-1, and silencing of the SHP-1 abolished the effect of plumbagin. This agent also downregulated the expression of STAT3-regulated cyclin D1, Bcl-xL, and vascular endothelial growth factor; activated caspase-3; induced poly (ADP ribose) polymerase cleavage; and increased the sub-G(1) population of MM cells. Consistent with these results, overexpression of constitutive active STAT3 significantly reduced the plumbagin-induced apoptosis. When compared with AG490, a rationally designed STAT3/JAK2 inhibitor, plumbagin was found more potent in suppressing the proliferation of cells. Plumbagin also significantly potentiated the apoptotic effects of thalidomide and bortezomib in MM cells. Overall, these results suggest that the plumbagin inhibits STAT3 activation pathway through the induction of SHP-1 and this may mediate the sensitization of STAT3 overexpressing cancers to chemotherapeutic agents.

Arctigenin enhances chemosensitivity of cancer cells to cisplatin through inhibition of the STAT3 signaling pathway.

PubMed

Yao, Xiangyang; Zhu, Fenfen; Zhao, Zhihui; Liu, Chang; Luo, Lan; Yin, Zhimin

2011-10-01

Arctigenin is a dibenzylbutyrolactone lignan isolated from Bardanae fructus, Arctium lappa L, Saussureamedusa, Torreya nucifera, and Ipomea cairica. It has been reported to exhibit anti-inflammatory activities, which is mainly mediated through its inhibitory effect on nuclear transcription factor-kappaB (NF-κB). But the role of arctigenin in JAK-STAT3 signaling pathways is still unclear. In present study, we investigated the effect of arctigenin on signal transducer and activator of transcription 3 (STAT3) pathway and evaluated whether suppression of STAT3 activity by arctigenin could sensitize cancer cells to a chemotherapeutic drug cisplatin. Our results show that arctigenin significantly suppressed both constitutively activated and IL-6-induced STAT3 phosphorylation and subsequent nuclear translocation in cancer cells. Inhibition of STAT3 tyrosine phosphorylation was found to be achieved through suppression of Src, JAK1, and JAK2, while suppression of STAT3 serine phosphorylation was mediated by inhibition of ERK activation. Pervanadate reversed the arctigenin-induced downregulation of STAT3 activation, suggesting the involvement of a protein tyrosine phosphatase. Indeed, arctigenin can obviously induce the expression of the PTP SHP-2. Furthermore, the constitutive activation level of STAT3 was found to be correlated to the resistance of cancer cells to cisplatin-induced apoptosis. Arctigenin dramatically promoted cisplatin-induced cell death in cancer cells, indicating that arctigenin enhanced the sensitivity of cancer cells to cisplatin mainly via STAT3 suppression. These observations suggest a novel anticancer function of arctigenin and a potential therapeutic strategy of using arctigenin in combination with chemotherapeutic agents for cancer treatment. Copyright © 2011 Wiley-Liss, Inc.

The critical role that STAT3 plays in glioma-initiating cells: STAT3 addiction in glioma

PubMed Central

Ganguly, Debolina; Fan, Meiyun; Yang, Chuan He; Zbytek, Blazej; Finkelstein, David; Roussel, Martine F.; Pfeffer, Lawrence M.

2018-01-01

Glioma-Initiating Cells (GICs) are thought to be responsible for tumor initiation, progression and recurrence in glioblastoma (GBM). In previous studies, we reported the constitutive phosphorylation of the STAT3 transcription factor in GICs derived from GBM patient-derived xenografts, and that STAT3 played a critical role in GBM tumorigenesis. In this study, we show that CRISPR/Cas9-mediated deletion of STAT3 in an established GBM cell line markedly inhibited tumorigenesis by intracranial injection but had little effect on cell proliferation in vitro. Tumorigenesis was rescued by the enforced expression of wild-type STAT3 in cells lacking STAT3. In contrast, GICs were highly addicted to STAT3 and upon STAT3 deletion GICs were non-viable. Moreover, we found that STAT3 was constitutively activated in GICs by phosphorylation on both tyrosine (Y705) and serine (S727) residues. Therefore, to study STAT3 function in GICs we established an inducible system to knockdown STAT3 expression (iSTAT3-KD). Using this approach, we demonstrated that Y705-STAT3 phosphorylation was critical and indispensable for GIC-induced tumor formation. Both phosphorylation sites in STAT3 promoted GIC proliferation in vitro. We further showed that S727-STAT3 phosphorylation was Y705-dependent. Targeted microarray and RNA sequencing revealed that STAT3 activated cell-cycle regulator genes, and downregulated genes involved in the interferon response, the hypoxia response, the TGFβ pathway, and remodeling of the extracellular matrix. Since STAT3 is an important oncogenic driver of GBM, the identification of these STAT3 regulated pathways in GICs will inform the development of better targeted therapies against STAT3 in GBM and other cancers. PMID:29774125

STAT1 is Constitutively Activated in the T/C28a2 Immortalized Juvenile Human Chondrocyte Line and Stimulated by IL-6 Plus Soluble IL-6R.

PubMed

Meszaros, Evan C; Malemud, Charles J

2015-04-01

T/C28a2 immortalized juvenile human chondrocytes were employed to determine the extent to which activation of Signal Transducers and Activators of Transcription-1 (STAT1) occurred in response to recombinant human interleukin-6 (rhIL-6) or rhIL-6 in combination with the soluble IL-6 receptor (sIL-6R). Two forms of STAT1, STAT1A and STAT1B, were identified on SDS-PAGE and western blotting with anti-STAT1 antibody. Western blotting revealed that STAT1 was constitutively phosphorylated (p-STAT1). Although incubation of T/C28a2 chondrocytes with rhIL-6 (50 ng/ml) increased p-STAT1A by Δ=22.3% after 30 min, this percent difference failed to reach significance by Chi-square analysis. Similarly, no effect of rhIL-6 (Δ=+10.7%) on p-STAT1B was seen at 30 min. In contrast, although the combination of rhIL-6 plus sIL-6R had no effect on p-STAT1A, rhIL-6 plus sIL-6R increased p-STAT1B by Δ=73.3% (p<0.0001) after 30 min compared to the control group and by Δ=56.7% (p<0.0001) compared to rhIL-6 alone. Janex-1, a Janus kinase-3-specific inhibitor (100 μM) partially reduced the effect of rhIL-6 on p-STAT1B by Δ=27.7% (p<0.05). The results of this study showed that STAT1A/STAT1B was constitutively activated in T/C28a2 chondrocytes. Although rhIL-6 increased p-STAT1B to a small extent, the combination of rhIL-6 plus sIL-6R was far more effective in stimulating STAT1B phosphorylation compared to controls or rhIL-6 alone. These data support the likelihood that although JAK3-mediated activation of STAT1 in T/C28a2 chondrocytes may involve the IL-6/IL-6R/gp130 pathway, these results indicated that STAT1 activation in response to IL-6 preferentially involved IL-6 trans -signaling via sIL-6R.

Role of STAT3 in Cancer Metastasis and Translational Advances

PubMed Central

Patil, Prachi; Gude, Rajiv P.

2013-01-01

Signal transducer and activator of transcription 3 (STAT3) is a latent cytoplasmic transcription factor, originally discovered as a transducer of signal from cell surface receptors to the nucleus. It is activated by tyrosine phosphorylation at position 705 leading to its dimerization, nuclear translocation, DNA binding, and activation of gene transcription. Under normal physiological conditions, STAT3 activation is tightly regulated. However, compelling evidence suggests that STAT3 is constitutively activated in many cancers and plays a pivotal role in tumor growth and metastasis. It regulates cellular proliferation, invasion, migration, and angiogenesis that are critical for cancer metastasis. In this paper, we first describe the mechanism of STAT3 regulation followed by how STAT3 is involved in cancer metastasis, then we summarize the various small molecule inhibitors that inhibit STAT3 signaling. PMID:24199193

The Synthetic α-Bromo-2′,3,4,4′-Tetramethoxychalcone (α-Br-TMC) Inhibits the JAK/STAT Signaling Pathway

PubMed Central

Brueggemann, Susanne; Besl, Elisabeth; Al-Rifai, Nafisah; Petkes, Hermina; Amslinger, Sabine; Rascle, Anne

2014-01-01

Signal transducer and activator of transcription STAT5 and its upstream activating kinase JAK2 are essential mediators of cytokine signaling. Their activity is normally tightly regulated and transient. However, constitutive activation of STAT5 is found in numerous cancers and a driving force for malignant transformation. We describe here the identification of the synthetic chalcone α-Br-2′,3,4,4′-tetramethoxychalcone (α-Br-TMC) as a novel JAK/STAT inhibitor. Using the non-transformed IL-3-dependent B cell line Ba/F3 and its oncogenic derivative Ba/F3-1*6 expressing constitutively activated STAT5, we show that α-Br-TMC targets the JAK/STAT pathway at multiple levels, inhibiting both JAK2 and STAT5 phosphorylation. Moreover, α-Br-TMC alters the mobility of STAT5A/B proteins in SDS-PAGE, indicating a change in their post-translational modification state. These alterations correlate with a decreased association of STAT5 and RNA polymerase II with STAT5 target genes in chromatin immunoprecipitation assays. Interestingly, expression of STAT5 target genes such as Cis and c-Myc was differentially regulated by α-Br-TMC in normal and cancer cells. While both genes were inhibited in IL-3-stimulated Ba/F3 cells, expression of the oncogene c-Myc was down-regulated and that of the tumor suppressor gene Cis was up-regulated in transformed Ba/F3-1*6 cells. The synthetic chalcone α-Br-TMC might therefore represent a promising novel anticancer agent for therapeutic intervention in STAT5-associated malignancies. PMID:24595334

Decursin chemosensitizes human multiple myeloma cells through inhibition of STAT3 signaling pathway.

PubMed

Kim, Hyun Jung; Kim, Sung-Moo; Park, Kyung-Ran; Jang, Hyeung-Jin; Na, Young-Soon; Ahn, Kyoo Seok; Kim, Sung-Hoon; Ahn, Kwang Seok

2011-02-01

Recent reports have indicated that decursin can induce apoptosis, suppress tumor growth, and inhibit angiogenesis. In this experiment, we investigated how decursin could potentiate the cytotoxic effects of bortezomib in human multiple myeloma cells. We found that decursin inhibited cell viability in U266, MM.1S and ARH77 cells, but not in peripheral blood mononuclear cells (PBMC). Decursin-induced apoptosis through the activation of caspase-8, -9, and -3 in U266 cells. This correlated with the down-regulating of cyclin D1, bcl-2, bcl-xL, survivin, and the vascular endothelial growth factor (VEGF), which are all regulated by the activation of signal transducers and the activator of transcription 3 (STAT3). Indeed, decursin inhibited constitutive STAT3 activation through inhibition of the activation of Janus-activated kinase 2 (JAK2) in U266 cells. In addition, decursin inhibited interleukin-6-inducible STAT3 activation in a time-dependent manner in MM.1S cells. Interestingly, decursin significantly potentiated the apoptotic effects of bortezomib in U266 cells. These effects of decursin were correlated with the suppression of constitutive STAT3 activation in U266 cells. Overall, these results suggest that decursin is a novel blocker of STAT3 activation and it may be a potential candidate for overcoming chemo-resistance through suppression of this signaling. Copyright © 2010 Elsevier Ireland Ltd. All rights reserved.

Epigenetic silencing of the NR4A3 tumor suppressor, by aberrant JAK/STAT signaling, predicts prognosis in gastric cancer

NASA Astrophysics Data System (ADS)

Yeh, Chung-Min; Chang, Liang-Yu; Lin, Shu-Hui; Chou, Jian-Liang; Hsieh, Hsiao-Yen; Zeng, Li-Han; Chuang, Sheng-Yu; Wang, Hsiao-Wen; Dittner, Claudia; Lin, Cheng-Yu; Lin, Jora M. J.; Huang, Yao-Ting; Ng, Enders K. W.; Cheng, Alfred S. L.; Wu, Shu-Fen; Lin, Jiayuh; Yeh, Kun-Tu; Chan, Michael W. Y.

2016-08-01

While aberrant JAK/STAT signaling is crucial to the development of gastric cancer (GC), its effects on epigenetic alterations of its transcriptional targets remains unclear. In this study, by expression microarrays coupled with bioinformatic analyses, we identified a putative STAT3 target gene, NR4A3 that was downregulated in MKN28 GC daughter cells overexpressing a constitutively activated STAT3 mutant (S16), as compared to an empty vector control (C9). Bisulphite pyrosequencing and demethylation treatment showed that NR4A3 was epigenetically silenced by promoter DNA methylation in S16 and other GC cell lines including AGS cells, showing constitutive activation of STAT3. Subsequent experiments revealed that NR4A3 promoter binding by STAT3 might repress its transcription. Long-term depletion of STAT3 derepressed NR4A3 expression, by promoter demethylation, in AGS GC cells. NR4A3 re-expression in GC cell lines sensitized the cells to cisplatin, and inhibited tumor growth in vitro and in vivo, in an animal model. Clinically, GC patients with high NR4A3 methylation, or lower NR4A3 protein expression, had significantly shorter overall survival. Intriguingly, STAT3 activation significantly associated only with NR4A3 methylation in low-stage patient samples. Taken together, aberrant JAK/STAT3 signaling epigenetically silences a potential tumor suppressor, NR4A3, in gastric cancer, plausibly representing a reliable biomarker for gastric cancer prognosis.

A Single Amino Acid Substitution in the v-Eyk Intracellular Domain Results in Activation of Stat3 and Enhances Cellular Transformation

PubMed Central

Besser, Daniel; Bromberg, Jacqueline F.; Darnell, James E.; Hanafusa, Hidesaburo

1999-01-01

The receptor tyrosine kinase Eyk, a member of the Axl/Tyro3 subfamily, activates the STAT pathway and transforms cells when constitutively activated. Here, we compared the potentials of the intracellular domains of Eyk molecules derived from c-Eyk and v-Eyk to transform rat 3Y1 fibroblasts. The v-Eyk molecule induced higher numbers of transformants in soft agar and stronger activation of Stat3; levels of Stat1 activation by the two Eyk molecules were similar. A mutation in the sequence Y933VPL, present in c-Eyk, to the v-Eyk sequence Y933VPQ led to increased activation of Stat3 and increased transformation efficiency. However, altering another sequence, Y862VNT, present in both Eyk molecules to F862VNT markedly decreased transformation without impairing Stat3 activation. These results indicate that activation of Stat3 enhances transformation efficiency and cooperates with another pathway to induce transformation. PMID:9891073

STAT3/NF-κB-Regulated Lentiviral TK/GCV Suicide Gene Therapy for Cisplatin-Resistant Triple-Negative Breast Cancer

PubMed Central

Kuo, Wei-Ying; Hwu, Luen; Wu, Chun-Yi; Lee, Jhih-Shian; Chang, Chi-Wei; Liu, Ren-Shyan

2017-01-01

Triple-negative breast cancer (TNBC) represents approximately 20% of all breast cancers and appears resistance to conventional cytotoxic chemotherapy, demonstrating a particularly poor prognosis and a significantly worse clinical outcome than other types of cancer. Suicide gene therapy has been used for the in vivo treatment of various solid tumors in recent clinical trials. In tumor microenvironment, STAT3/NF-κB pathways are constitutively activated in stromal cells as well as in cancer stem cells (CSCs). In this study, we have cloned a novel STAT3/NF-κB-based reporter system to drive the expression of herpes simplex virus thymidine kinase (HSV-TK) against breast cancer. Lentiviral vector expressing HSV-TK under the regulation of STAT3/NF-κB fused response element was developed. In this setting, we exploited the constitutive STAT3/NF-κB activation in tumors to achieve higher transgene expression than that driven by a constitutively active CMV promotor in vivo. An orthotropic MDA-MB-231 triple negative breast cancer mouse model was used for evaluating the feasibility of STAT3-NF-κB-TK/GCV suicide gene therapy system. The basal promoter activity of Lenti-CMV-TK and Lenti-STAT3-NF-κB-TK in MDA-MB-231 cells was compared by 3H-FEAU uptake assay. The Lenti-CMV-TK showed ~5 fold higher 3H-FEAU uptake then Lenti -STAT3-NF-κB-TK. In clonogenic assay, cells expressing Lenti-CMV-TK were 2-fold more sensitive to GCV than Lenti-STAT3-NF-κB-TK transduced cells. In vitro effect of STAT3-NF-κB-induced transgene expression was determined by 10ng/mL TNF-α induction and confirmed by western blot analysis and DsRedm fluorescent microscopy. In vivo evaluation of therapeutic effect by bioluminescence and [18F]FHBG microPET imaging indicated that Lenti-STAT3-NF-κB-TK showed more tumor growth retardation than Lenti-CMV-TK when GCV (20 mg/kg) was administered. The invasiveness and expression of cancer stem cell markers were both decreased after STAT3/NF-κB-regulated HSV-TK/GCV therapy. Moreover, STAT3/NF-κB signaling targeting could further sensitize tumor cells to cisplatin. This study successfully established a theranositic approach to treat triple-negative breast cancer via STAT3-NF-κB responsive element-driven suicide gene therapy. This platform may also be an alternative strategy to handle with drug-resistant cancer cells. PMID:28255357

A novel anti-cancer agent Icaritin suppresses hepatocellular carcinoma initiation and malignant growth through the IL-6/Jak2/Stat3 pathway.

PubMed

Zhao, Hong; Guo, Yuming; Li, Shu; Han, Ruiqin; Ying, Jianming; Zhu, Hai; Wang, Yuanyuan; Yin, Li; Han, Yuqing; Sun, Lingzhi; Wang, Zhaoyi; Lin, Qingcong; Bi, Xinyu; Jiao, Yuchen; Jia, Hongying; Zhao, Jianjun; Huang, Zhen; Li, Zhiyu; Zhou, Jianguo; Song, Wei; Meng, Kun; Cai, Jianqiang

2015-10-13

Tumor-initiating cell (TIC) is a subpopulation of cells in tumors that are responsible for tumor initiation and progression. Recent studies indicate that hepatocellular carcinoma-initiating cells (HCICs) confer the high malignancy, recurrence and multi-drug resistance in hepatocellular carcinoma (HCC). In this study, we found that Icaritin, a prenylflavonoid derivative from Epimedium Genus, inhibited malignant growth of HCICs. Icaritin decreased the proportion of EpCAM-positive (a HCICs marker) cells, suppressed tumorsphere formation in vitro and tumor formation in vivo. We also found that Icaritin reduced expression of Interleukin-6 Receptors (IL-6Rs), attenuated both constitutive and IL-6-induced phosphorylation of Janus-activated kinases 2 (Jak2) and Signal transducer and activator of transcription 3 (Stat3), and inhibited Stat3 downstream genes, such as Bmi-1 and Oct4. The inhibitory activity of Icaritin in HCICs was augmented by siRNA-mediated silencing of Stat3 but attenuated by constitutive activation of Stat3.Taken together, our results indicate that Icaritin is able to inhibit malignant growth of HCICs and suggest that Icaritin may be developed into a novel therapeutic agent for effective treatment of HCC.

Mitochondrial translocation of signal transducer and activator of transcription 5 (STAT5) in leukemic T cells and cytokine-stimulated cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chueh, Fu-Yu; Leong, King-Fu; Yu, Chao-Lan, E-mail: chaolan.yu@rosalindfranklin.edu

2010-11-26

Research highlights: {yields} STAT5 interacts with a mitochondrial protein PDC-E2 in a leukemic T cell line LSTRA. {yields} Tyrosine-phosphorylated STAT5, but not STAT3, is present in LSTRA mitochondria. {yields} Cytokines induce mitochondrial translocation of STAT5, but not STAT1 or STAT3. {yields} Cytokine-induced mitochondrial translocation of tyrosine-phosphorylated STAT5 is transient. {yields} Mitochondrial STAT5 binds to a putative STAT5 site in the mitochondrial DNA in vitro. -- Abstract: Signal transducers and activators of transcription (STATs) were first identified as key signaling molecules in response to cytokines. Constitutive STAT activation also has been widely implicated in oncogenesis. We analyzed STAT5-associated proteins in amore » leukemic T cell line LSTRA, which exhibits constitutive tyrosine phosphorylation and activation of STAT5. A cellular protein was found to specifically interact with STAT5 in LSTRA cells by co-immunoprecipitation. Sequencing analysis and subsequent immunoblotting confirmed the identity of this STAT5-associated protein as the E2 component of mitochondrial pyruvate dehydrogenase complex (PDC-E2). Consistent with this interaction, both subcellular fractionation and immunofluorescence microscopy revealed mitochondrial localization of STAT5 in LSTRA cells. Mitochondrial localization of tyrosine-phosphorylated STAT5 also occurred in cytokine-stimulated cells. A time course experiment further demonstrated the transient kinetics of STAT5 mitochondrial translocation after cytokine stimulation. In contrast, cytokine-induced STAT1 and STAT3 activation did not result in their translocation into mitochondria. Furthermore, we showed that mitochondrial STAT5 bound to the D-loop regulatory region of mitochondrial DNA in vitro. It suggests a potential role of STAT5 in regulating the mitochondrial genome. Proliferative metabolism toward aerobic glycolysis is well known in cancer cells as the Warburg effect and is also observed in cytokine-stimulated cells. Our novel findings of cytokine-induced STAT5 translocation into mitochondria and its link to oncogenesis provide important insights into the underlying mechanisms of this characteristic metabolic shift.« less

STATs MEDIATE FIBROBLAST GROWTH FACTOR INDUCED VASCULAR ENDOTHELIAL MORPHOGENESIS

PubMed Central

Yang, Xinhai; Qiao, Dianhua; Meyer, Kristy; Friedl, Andreas

2009-01-01

The fibroblast growth factors (FGFs) play diverse roles in development, wound healing and angiogenesis. The intracellular signal transduction pathways which mediate these pleiotropic activities remain incompletely understood. We show here that the proangiogenic factors FGF2 and FGF8b can activate signal transducers and activators of transcription (STATs) in mouse microvascular endothelial cells. Both FGF2 and FGF8b activate STAT5 and to a lesser extent STAT1, but not STAT3. The FGF2-dependent activation of endothelial STAT5 was confirmed in vivo with the matrigel plug angiogenesis assay. In tissue samples of human gliomas, a tumor type where FGF-induced angiogenesis is important, STAT5 is detected in tumor vessel endothelial cell nuclei, consistent with STAT5 activation. By forced expression of constitutively active or dominant-negative mutant STAT5A in mouse brain endothelial cells, we further show that STAT5 activation is both necessary and sufficient for FGF-induced cell migration, invasion and tube formation, which are key events in vascular endothelial morphogenesis and angiogenesis. In contrast, STAT5 is not required for brain endothelial cell mitogenesis. The cytoplasmic tyrosine kinases Src and Janus kinase 2 (Jak2) both appear to be involved in the activation of STAT5, as their inhibition reduces FGF2 and FGF8b induced STAT5 phosphorylation and endothelial cell tube formation. Constitutively active STAT5A partially restores tube formation in the presence of Src or Jak2 inhibitors. These observations demonstrate that FGFs utilize distinct signaling pathways to induce angiogenic phenotypes. Together, our findings implicate the FGF-Jak2/Src-STAT5 cascade as a critical angiogenic FGF signaling pathway. PMID:19176400

A novel inhibitor of STAT3 homodimerization selectively suppresses STAT3 activity and malignant transformation

PubMed Central

Zhang, Xiaolei; Sun, Ying; Pireddu, Roberta; Yang, Hua; Urlam, Murali K.; Lawrence, Harshani R.; Guida, Wayne C.; Lawrence, Nicholas J.; Sebti, Saïd M.

2014-01-01

STAT3-STAT3 dimerization, which involves reciprocal binding of the STAT3-SH2 domain to phosphorylated tyrosine-705 (Y-705), is required for STAT3 nuclear translocation, DNA binding and transcriptional regulation of downstream target genes. Here we describe a small molecule S3I-1757 capable of disrupting STAT3-STAT3 dimerization, activation and malignant transforming activity. Fluorescence polarization assays and molecular modeling suggest that S3I-1757 interacts with the Y-705 binding site in the SH2 domain and displaces fluorescein-labelled GpYLPQTV phosphotyrosine peptide from binding to STAT3. We generated HA-tagged STAT3 and FLAG-tagged STAT3 and showed using co-immunoprecipitation and co-localization studies that S3I-1757 inhibits STAT3 dimerization and STAT3-EGF receptor binding in intact cells. Treatment of human cancer cells with S3I-1757 (but not a closely related analogue, S3I-1756, that does not inhibit STAT3 dimerization), inhibits selectively the phosphorylation of STAT3 over AKT1 and ERK1/2 (MAPK3/1), nuclear accumulation of P-Y705-STAT3, STAT3-DNA binding and transcriptional activation and suppresses the expression levels of STAT3 target genes such as Bcl-xL (BCL2L1), survivin (BIRC5), cyclin D1 (CCND1) and MMP9. Furthermore, S3I-1757 but not S3I-1756 inhibits anchorage-dependent and -independent growth, migration and invasion of human cancer cells which depend on STAT3. Finally, STAT3-C, a genetically engineered mutant of STAT3 that forms a constitutively dimerized STAT3, rescues cells from the effects of S3I-1757 inhibition. Thus, we have developed S3I-1757 as a STAT3-STAT3 dimerization inhibitor capable of blocking hyper activated STAT3 and suppressing malignant transformation in human cancer cells that depend on STAT3. PMID:23322008

Gastrointestinal Disease in Simian Immunodeficiency Virus-Infected Rhesus Macaques Is Characterized by Proinflammatory Dysregulation of the Interleukin-6-Janus Kinase/Signal Transducer and Activator of Transcription3 Pathway

PubMed Central

Mohan, Mahesh; Aye, Pyone P.; Borda, Juan T.; Alvarez, Xavier; Lackner, Andrew A.

2007-01-01

Gastrointestinal disease and inflammation are common sequelae of human and simian immunodeficiency virus (SIV) infection. Nevertheless, the molecular mechanisms that lead to gastrointestinal dysfunction remain unclear. We investigated regulation of the interleukin (IL)-6-JAK-STAT3 pathway in jejunum and colon, collected at necropsy, from 10 SIV-infected macaques with diarrhea (group 1), 10 non-SIV-infected macaques with diarrhea (group 2), and 7 control uninfected macaques (group 3). All group 1 and 2 macaques had chronic diarrhea, wasting, and colitis, but group 1 animals had more frequent and severe lesions in the jejunum. A significant increase in IL-6 and SOCS-3 gene expression along with constitutive STAT3 activation was observed in the colon of all group 1 and 2 macaques and in the jejunum of only group 1 macaques compared to controls. Further, in colon, histopathology severity scores correlated significantly with IL-6 (groups 1 and 2) and SOCS-3 (group 2) gene expression. In jejunum, a similar correlation was observed only in group 1 animals. Phosphorylated STAT3 (p-STAT3) was localized to lymphocytes (CD3+) and macrophages (CD68+), with fewer CD3+ lymphocytes expressing p-STAT3 in group 1 macaques. Despite high SOCS-3 expression, STAT3 remained constitutively active, providing a possible explanation for persistent intestinal inflammation and immune activation that may favor viral replication and disease progression. PMID:18055558

Withaferin A inhibits JAK/STAT3 signaling and induces apoptosis of human renal carcinoma Caki cells.

PubMed

Um, Hee Jung; Min, Kyoung-Jin; Kim, Dong Eun; Kwon, Taeg Kyu

2012-10-12

Withaferin A, the active component of Withania somnifera, causes cytotoxicity in a variety of tumor cell lines. In this study, we show that withaferin A inhibits constitutive and IL-6-induced phosphorylation of STAT3 (on Tyr705), but not IFN-γ-induced STAT1 phosphorylation. Withaferin A-induced down-regulation of STAT3 activation is associated with a reduction in Janus-activated kinase 2 (JAK2) activity. Withaferin A also down-regulates the expression of STAT3 regulated genes such as Bcl-xL, Bcl-2, cyclin D1 and survivin. The apoptotic effect of withaferin A in Caki human renal cancer cells was investigated. Withaferin A induced dose-dependent apoptotic cell death in Caki cells, as measured by FACS analysis and PARP cleavage. Furthermore, overexpression of STAT3 attenuated withaferin A-induced apoptosis. Taken together, the present study provides strong evidence that down-regulation of the STAT3 signaling pathway mediates withaferin A-induced apoptosis. Copyright © 2012 Elsevier Inc. All rights reserved.

Silibinin and STAT3: A natural way of targeting transcription factors for cancer therapy.

PubMed

Bosch-Barrera, Joaquim; Menendez, Javier A

2015-06-01

Signal transducer and activator of transcription 3 (STAT3) is constitutively activated in many different types of cancer and plays a pivotal role in tumor growth and metastasis. Retrospective studies have established that STAT3 expression or phospho-STAT3 (pSTAT3 or activated STAT3) are poor prognostic markers for breast, colon, prostate and non-small cell lung cancer. Silibinin or silybin is a natural polyphenolic flavonoid which is present in seed extracts of milk thistle (Silybum marianum). Silibinin has been shown to inhibit multiple cancer cell signaling pathways in preclinical models, demonstrating promising anticancer effects in vitro and in vivo. This review summarizes evidence suggesting that silibinin can inhibit pSTAT3 in preclinical cancer models. We also discuss current strategies to overcome the limitations of oral administration of silibinin to cancer patients to translate the bench results to the bed side. Finally, we review the ongoing clinical trials exploring the role of silibinin in cancer. Copyright © 2015 Elsevier Ltd. All rights reserved.

A novel inhibitor of STAT3 homodimerization selectively suppresses STAT3 activity and malignant transformation.

PubMed

Zhang, Xiaolei; Sun, Ying; Pireddu, Roberta; Yang, Hua; Urlam, Murali K; Lawrence, Harshani R; Guida, Wayne C; Lawrence, Nicholas J; Sebti, Saïd M

2013-03-15

STAT3-STAT3 dimerization, which involves reciprocal binding of the STAT3-SH2 domain to phosphorylated tyrosine-705 (Y-705), is required for STAT3 nuclear translocation, DNA binding, and transcriptional regulation of downstream target genes. Here, we describe a small molecule S3I-1757 capable of disrupting STAT3-STAT3 dimerization, activation, and malignant transforming activity. Fluorescence polarization assay and molecular modeling suggest that S3I-1757 interacts with the phospho-Y-705-binding site in the SH2 domain and displaces fluorescein-labeled GpYLPQTV phosphotyrosine peptide from binding to STAT3. We generated hemagglutinin (HA)-tagged STAT3 and FLAG-tagged STAT3 and showed using coimmunoprecipitation and colocalization studies that S3I-1757 inhibits STAT3 dimerization and STAT3-EGF receptor (EGFR) binding in intact cells. Treatment of human cancer cells with S3I-1757 (but not a closely related analog, S3I-1756, which does not inhibit STAT3 dimerization), inhibits selectively the phosphorylation of STAT3 over AKT1 and ERK1/2 (MAPK3/1), nuclear accumulation of P-Y705-STAT3, STAT3-DNA binding, and transcriptional activation and suppresses the expression levels of STAT3 target genes, such as Bcl-xL (BCL2L1), survivin (BIRC5), cyclin D1 (CCND1), and matrix metalloproteinase (MMP)-9. Furthermore, S3I-1757, but not S3I-1756, inhibits anchorage-dependent and -independent growth, migration, and invasion of human cancer cells, which depend on STAT3. Finally, STAT3-C, a genetically engineered mutant of STAT3 that forms a constitutively dimerized STAT3, rescues cells from the effects of S3I-1757 inhibition. Thus, we have developed S3I-1757 as a STAT3-STAT3 dimerization inhibitor capable of blocking hyperactivated STAT3 and suppressing malignant transformation in human cancer cells that depend on STAT3.

Targeting constitutively-activated STAT3 in hypoxic ovarian cancer, using a novel STAT3 inhibitor

PubMed Central

McCann, Georgia A.; Naidu, Shan; Rath, Kellie S.; Bid, Hemant K.; Tierney, Brent J.; Suarez, Adrian; Varadharaj, Saradhadevi; Zhang, Jianying; Hideg, Kálmán; Houghton, Peter; Kuppusamy, Periannan; Cohn, David E.; Selvendiran, Karuppaiyah

2014-01-01

Tumor hypoxia, a feature of many solid tumors including ovarian cancer, is associated with resistance to therapies. We previously demonstrated that hypoxic exposure results in increased expression of phosphorylated signal transducer and activator of transcription 3 (pSTAT3). We hypothesized the activation of STAT3 could lead to chemotherapeutic resistance in ovarian cancer cells in hypoxic conditions. In this study, we demonstrate the level of pSTAT3 Tyr705 is increased in the hypoxic regions of human epithelial ovarian cancer (EOC) specimens, as determined by HIF-1α and CD-31 staining. In vitro mutagenesis studies proved that pSTAT3 Tyr705 is necessary for cell survival and proliferation under hypoxic conditions. In addition, we show that S1PR1, a regulator of STAT3 transcription via the JAK/STAT pathway, is highly expressed in hypoxic ovarian cancer cells (HOCCs). Knock down of S1PR1 in HOCCs reduced pSTAT3 Tyr705 levels and was associated with decreased cell survival. Treatment of HOCCs with the STAT3 inhibitor HO-3867 resulted in a rapid and dramatic decrease in pSTAT3 Tyr705 levels as a result of ubiquitin proteasome degradation. STAT3-target proteins Bcl-xL, cyclin D2 and VEGF showed similar decreases in HO-3867 treated cells. Taken together, these findings suggest that activation of STAT3 Tyr705 promotes cell survival and proliferation in HOCCs, and that S1PR1 is involved in the initiation of STAT3 activation. Targeting hypoxia-mediated STAT3 activation represents a therapeutic option for ovarian cancer and other solid tumors. PMID:25594014

Preclinical characterization of signal transducer and activator of transcription 3 small molecule inhibitors for primary and metastatic brain cancer therapy.

PubMed

Assi, Hikmat H; Paran, Chris; VanderVeen, Nathan; Savakus, Jonathan; Doherty, Robert; Petruzzella, Emanuele; Hoeschele, James D; Appelman, Henry; Raptis, Leda; Mikkelsen, Tom; Lowenstein, Pedro R; Castro, Maria G

2014-06-01

Signal transducer and activator of transcription 3 (STAT3) has been implicated as a hub for multiple oncogenic pathways. The constitutive activation of STAT3 is present in several cancers, including gliomas (GBMs), and is associated with poor therapeutic responses. Phosphorylation of STAT3 triggers its dimerization and nuclear transport, where it promotes the transcription of genes that stimulate tumor growth. In light of this role, inhibitors of the STAT3 pathway are attractive therapeutic targets for cancer. To this end, we evaluated the STAT3-inhibitory activities of three compounds (CPA-7 [trichloronitritodiammineplatinum(IV)], WP1066 [(S,E)-3-(6-bromopyridin-2-yl)-2-cyano-N-(1-phenylethyl)acrylamide, C17H14BrN3O], and ML116 [4-benzyl-1-{thieno[2,3-d]pyrimidin-4-yl}piperidine, C18H19N3S]) in cultured rodent and human glioma cells, including GBM cancer stem cells. Our results demonstrate a potent induction of growth arrest in GBM cells after drug treatment with a concomitant induction of cell death. Although these compounds were effective at inhibiting STAT3 phosphorylation, they also displayed variable dose-dependent inhibition of STAT1, STAT5, and nuclear factor κ light-chain enhancer of activated B cells. The therapeutic efficacy of these compounds was further evaluated in peripheral and intracranial mouse tumor models. Whereas CPA-7 elicited regression of peripheral tumors, both melanoma and GBM, its efficacy was not evident when the tumors were implanted within the brain. Our data suggest poor permeability of this compound to tumors located within the central nervous system. WP1066 and ML116 exhibited poor in vivo efficacy. In summary, CPA-7 constitutes a powerful anticancer agent in models of peripheral solid cancers. Our data strongly support further development of CPA-7-derived compounds with increased permeability to enhance their efficacy in primary and metastatic brain tumors.

Novel Multiplexed Assay for Identifying SH2 Domain Antagonists of STAT Family Proteins

PubMed Central

Takakuma, Kazuyuki; Ogo, Naohisa; Uehara, Yutaka; Takahashi, Susumu; Miyoshi, Nao; Asai, Akira

2013-01-01

Some of the signal transducer and activator of transcription (STAT) family members are constitutively activated in a wide variety of human tumors. The activity of STAT depends on their Src homology 2 (SH2) domain-mediated binding to sequences containing phosphorylated tyrosine. Thus, antagonizing this binding is a feasible approach to inhibiting STAT activation. We have developed a novel multiplexed assay for STAT3- and STAT5b-SH2 binding, based on amplified luminescent proximity homogeneous assay (Alpha) technology. AlphaLISA and AlphaScreen beads were combined in a single-well assay, which allowed the binding of STAT3- and STAT5b-SH2 to phosphotyrosine peptides to be simultaneously monitored. Biotin-labeled recombinant human STAT proteins were obtained as N- and C-terminal deletion mutants. The spacer length of the DIG-labeled peptide, the reaction time, and the concentration of sodium chloride were optimized to establish a HTS system with Z’ values of greater than 0.6 for both STAT3- and STAT5b-SH2 binding. We performed a HTS campaign for chemical libraries using this multiplexed assay and identified hit compounds. A 2-chloro-1,4-naphthalenedione derivative, Compound 1, preferentially inhibited STAT3-SH2 binding in vitro, and the nuclear translocation of STAT3 in HeLa cells. Initial structure activity relationship (SAR) studies using the multiplexed assay showed the 3-substituent effect on both the activity and selectivity of STAT3 and STAT5b inhibition. Therefore, this multiplexed assay is useful for not only searching for potential lead compounds but also obtaining SAR data for developing new STAT3/STAT5b inhibitors. PMID:23977103

Novel multiplexed assay for identifying SH2 domain antagonists of STAT family proteins.

PubMed

Takakuma, Kazuyuki; Ogo, Naohisa; Uehara, Yutaka; Takahashi, Susumu; Miyoshi, Nao; Asai, Akira

2013-01-01

Some of the signal transducer and activator of transcription (STAT) family members are constitutively activated in a wide variety of human tumors. The activity of STAT depends on their Src homology 2 (SH2) domain-mediated binding to sequences containing phosphorylated tyrosine. Thus, antagonizing this binding is a feasible approach to inhibiting STAT activation. We have developed a novel multiplexed assay for STAT3- and STAT5b-SH2 binding, based on amplified luminescent proximity homogeneous assay (Alpha) technology. AlphaLISA and AlphaScreen beads were combined in a single-well assay, which allowed the binding of STAT3- and STAT5b-SH2 to phosphotyrosine peptides to be simultaneously monitored. Biotin-labeled recombinant human STAT proteins were obtained as N- and C-terminal deletion mutants. The spacer length of the DIG-labeled peptide, the reaction time, and the concentration of sodium chloride were optimized to establish a HTS system with Z' values of greater than 0.6 for both STAT3- and STAT5b-SH2 binding. We performed a HTS campaign for chemical libraries using this multiplexed assay and identified hit compounds. A 2-chloro-1,4-naphthalenedione derivative, Compound 1, preferentially inhibited STAT3-SH2 binding in vitro, and the nuclear translocation of STAT3 in HeLa cells. Initial structure activity relationship (SAR) studies using the multiplexed assay showed the 3-substituent effect on both the activity and selectivity of STAT3 and STAT5b inhibition. Therefore, this multiplexed assay is useful for not only searching for potential lead compounds but also obtaining SAR data for developing new STAT3/STAT5b inhibitors.

The inhibitory effects of capillarisin on cell proliferation and invasion of prostate carcinoma cells.

PubMed

Tsui, Ke-Hung; Chang, Ying-Ling; Yang, Pei-Shan; Hou, Chen-Pang; Lin, Yu-Hsiang; Lin, Bing-Wei; Feng, Tsui-Hsia; Juang, Horng-Heng

2018-04-01

Capillarisin (Cap), an active component of Artemisia capillaris root extracts, is characterized by its anti-inflammatory, anti-oxidant and anti-cancer properties. Nevertheless, the functions of Cap in prostate cancer have not been fully explored. We evaluated the potential actions of Cap on the cell proliferation, migration and invasion of prostate carcinoma cells. Cell proliferation and cell cycle distribution were measured by water-soluble tetrazolium-1 and flow cytometry assays. The expression of cyclins, p21, p27, survivin, matrix metallopeptidase (MMP2 and MMP9) were assessed by immunoblotting assays. Effects of Cap on invasion and migration were determined by wound closure and matrigel transmigration assays. The constitutive and interlukin-6 (IL-6)-inducible STAT3 activation of prostate carcinoma cells were determined by immunoblotting and reporter assays. Capillarisin inhibited androgen-independent DU145 and androgen-dependent LNCaP cell growth through the induction of cell cycle arrest at the G0/G1 phase by upregulating p21 and p27 while downregulating expression of cyclin D1, cyclin A and cyclin B. Cap decreased protein expression of survivin, MMP-2, and MMP-9 and therefore blocked the migration and invasion of DU145 cells. Cap suppressed constitutive and IL-6-inducible STAT3 activation in DU145 and LNCaP cells. Our data indicate that Cap blocked cell growth by modulation of p21, p27 and cyclins. The inhibitory effects of Cap on survivin, MMP-2, MMP-9 and STAT3 activation may account for the suppression of invasion in prostate carcinoma cells. Our data suggest that Cap might be a therapeutic agent in treating advanced prostate cancer with constitutive STAT3 or IL-6-inducible STAT3 activation. © 2017 John Wiley & Sons Ltd.

JAK/STAT3 pathway inhibition blocks skeletal muscle wasting downstream of IL-6 and in experimental cancer cachexia.

PubMed

Bonetto, Andrea; Aydogdu, Tufan; Jin, Xiaoling; Zhang, Zongxiu; Zhan, Rui; Puzis, Leopold; Koniaris, Leonidas G; Zimmers, Teresa A

2012-08-01

Cachexia, the metabolic dysregulation leading to sustained loss of muscle and adipose tissue, is a devastating complication of cancer and other chronic diseases. Interleukin-6 and related cytokines are associated with muscle wasting in clinical and experimental cachexia, although the mechanisms by which they might induce muscle wasting are unknown. One pathway activated strongly by IL-6 family ligands is the JAK/STAT3 pathway, the function of which has not been evaluated in regulation of skeletal muscle mass. Recently, we showed that skeletal muscle STAT3 phosphorylation, nuclear localization, and target gene expression are activated in C26 cancer cachexia, a model with high IL-6 family ligands. Here, we report that STAT3 activation is a common feature of muscle wasting, activated in muscle by IL-6 in vivo and in vitro and by different types of cancer and sterile sepsis. Moreover, STAT3 activation proved both necessary and sufficient for muscle wasting. In C(2)C(12) myotubes and in mouse muscle, mutant constitutively activated STAT3-induced muscle fiber atrophy and exacerbated wasting in cachexia. Conversely, inhibiting STAT3 pharmacologically with JAK or STAT3 inhibitors or genetically with dominant negative STAT3 and short hairpin STAT3 reduced muscle atrophy downstream of IL-6 or cancer. These results indicate that STAT3 is a primary mediator of muscle wasting in cancer cachexia and other conditions of high IL-6 family signaling. Thus STAT3 could represent a novel therapeutic target for the preservation of skeletal muscle in cachexia.

JAK/STAT3 pathway inhibition blocks skeletal muscle wasting downstream of IL-6 and in experimental cancer cachexia

PubMed Central

Bonetto, Andrea; Aydogdu, Tufan; Jin, Xiaoling; Zhang, Zongxiu; Zhan, Rui; Puzis, Leopold; Koniaris, Leonidas G.

2012-01-01

Cachexia, the metabolic dysregulation leading to sustained loss of muscle and adipose tissue, is a devastating complication of cancer and other chronic diseases. Interleukin-6 and related cytokines are associated with muscle wasting in clinical and experimental cachexia, although the mechanisms by which they might induce muscle wasting are unknown. One pathway activated strongly by IL-6 family ligands is the JAK/STAT3 pathway, the function of which has not been evaluated in regulation of skeletal muscle mass. Recently, we showed that skeletal muscle STAT3 phosphorylation, nuclear localization, and target gene expression are activated in C26 cancer cachexia, a model with high IL-6 family ligands. Here, we report that STAT3 activation is a common feature of muscle wasting, activated in muscle by IL-6 in vivo and in vitro and by different types of cancer and sterile sepsis. Moreover, STAT3 activation proved both necessary and sufficient for muscle wasting. In C2C12 myotubes and in mouse muscle, mutant constitutively activated STAT3-induced muscle fiber atrophy and exacerbated wasting in cachexia. Conversely, inhibiting STAT3 pharmacologically with JAK or STAT3 inhibitors or genetically with dominant negative STAT3 and short hairpin STAT3 reduced muscle atrophy downstream of IL-6 or cancer. These results indicate that STAT3 is a primary mediator of muscle wasting in cancer cachexia and other conditions of high IL-6 family signaling. Thus STAT3 could represent a novel therapeutic target for the preservation of skeletal muscle in cachexia. PMID:22669242

Signal transducers and activators of transcription (STATs): Novel targets of chemopreventive and chemotherapeutic drugs.

PubMed

Klampfer, Lidija

2006-03-01

A family of latent cytoplasmic transcription factors, signal transducers and activators of transcription (STATs), mediates the responsiveness of cells to several cytokines and growth factors. Although mutations of STATs have not been described in human tumors, the activity of several members of the family, such as STAT1, STAT3 and STAT5, is deregulated in a variety of human tumors. STAT3 and STAT5 acquire oncogenic potential through constitutive phosphorylation on tyrosine, and their activity has been shown to be required to sustain a transformed phenotype. Disruption of STAT3 and STAT5 signaling in transformed cells therefore represents an excellent opportunity for targeted cancer therapy. In contrast to STAT3 and STAT5, STAT1 negatively regulates cell proliferation and angiogenesis and thereby inhibits tumor formation. Consistent with its tumor suppressive properties, STAT1 and its downstream targets have been shown to be reduced in a variety of human tumors and STAT1 deficient mice are highly susceptible to tumor formation. In recent years we have gained mechanistic understanding of the pathways whereby STATs convey signals from the cytoplasm to the nucleus. In addition, several endogenous regulators of the JAK/STAT pathway have been described - and their mechanism of action revealed - that profoundly affect signaling by STATs. Both should greatly facilitate the design of drugs with potential to modulate STAT signaling and to restore the homeostasis in tissues where STATs have gone awry.

Activation of the STAT3 Signaling Pathway Is Associated With Poor Survival in Diffuse Large B-Cell Lymphoma Treated With R-CHOP

PubMed Central

Huang, Xin; Meng, Bin; Iqbal, Javeed; Ding, B. Belinda; Perry, Anamarija M.; Cao, Wenfeng; Smith, Lynette M.; Bi, Chengfeng; Jiang, Chunsun; Greiner, Timothy C.; Weisenburger, Dennis D.; Rimsza, Lisa; Rosenwald, Andreas; Ott, German; Delabie, Jan; Campo, Elias; Braziel, Rita M.; Gascoyne, Randy D.; Cook, James R.; Tubbs, Raymond R.; Jaffe, Elaine S.; Armitage, James O.; Vose, Julie M.; Staudt, Louis M.; McKeithan, Timothy W.; Chan, Wing C.; Ye, B. Hilda; Fu, Kai

2013-01-01

Purpose We previously reported that constitutive STAT3 activation is a prominent feature of the activated B-cell subtype of diffuse large B-cell lymphomas (ABC-DLBCL). In this study, we investigated whether STAT3 activation can risk stratify patients with DLBCL. Patients and Methods By an immunohistochemical method, we investigated phosphotyrosine STAT3 (PY-STAT3) expression from 185 patients with DLBCL treated with R-CHOP (rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone). Cell line-based siRNA experiments were also performed to generate an 11-gene, PY-STAT3 activation signature, which was used to study a previously published cohort of 222 patients with DLBCL. The STAT3 activation status determined by these two methods and by STAT3 mRNA levels were then correlated with survival. Results PY-STAT3 was detected in 37% of DLBCL and enriched in ABC-DLBCL cases (P = .03). PY-STAT3 positivity significantly correlated with poor overall survival (OS; P = .01) and event-free survival (EFS; P = .006). Similar observations were made for high levels of STAT3 mRNA. In multivariable analysis, PY-STAT3 status (P = .02), International Prognostic Index (P = .02), and BCL2 expression (P = .046) were independent prognosticators of OS in this cohort. Among the cell-of-origin subgroups, PY-STAT3 was associated with poor EFS among non–germinal center B-cell DLBCL cases only (P = .027). Similarly, the 11-gene STAT3 activation signature correlated with poor survival in the entire DLBCL cohort (OS, P < .001; EFS, P < .001) as well as the ABC-DLBCL subgroup (OS, P = .029; EFS, P = .025). Conclusion STAT3 activation correlated with poor survival in patients with DLBCL treated with R-CHOP, especially those with tumors of the ABC-DLBCL subtype. PMID:24220563

Activation of Stat3 Transcription Factor by Herpesvirus Saimiri STP-A Oncoprotein

PubMed Central

Chung, Young-Hwa; Cho, Nam-hyuk; Garcia, Maria Ines; Lee, Sun-Hwa; Feng, Pinghui; Jung, Jae U.

2004-01-01

The saimiri transforming protein (STP) oncogene of Herpesvirus saimiri subgroup A strain 11 (STP-A11) is not required for viral replication but is required for lymphoid cell immortalization in culture and lymphoma induction in primates. We previously showed that STP-A11 interacts with cellular Src kinase through its SH2 binding motif and that this interaction elicits Src signal transduction. Here we demonstrate that STP-A11 interacts with signal transducer and activator of transcription 3 (Stat3) independently of Src association and that the amino-terminal short proline-rich motif of STP-A11 and the central linker region of Stat3 are necessary for their interaction. STP-A11 formed a triple complex with Src kinase and Stat3 where Src kinase phosphorylated Stat3, resulting in the nuclear localization and transcriptional activation of Stat3. Consequently, the constitutively active Stat3 induced by STP-A11 elicited cellular signal transduction, which ultimately induced cell survival and proliferation upon serum deprivation. Furthermore, this activity was strongly correlated with the induction of Fos, cyclin D1, and Bcl-XL expression. These results demonstrate that STP-A11 independently targets two important cellular signaling molecules, Src and Stat3, and that these proteins cooperate efficiently to induce STP-A11-mediated transformation. PMID:15163742

Curcumin interacts directly with the Cysteine 259 residue of STAT3 and induces apoptosis in H-Ras transformed human mammary epithelial cells.

PubMed

Hahn, Young-Il; Kim, Su-Jung; Choi, Bu-Young; Cho, Kyung-Cho; Bandu, Raju; Kim, Kwang Pyo; Kim, Do-Hee; Kim, Wonki; Park, Joon Sung; Han, Byung Woo; Lee, Jeewoo; Na, Hye-Kyung; Cha, Young-Nam; Surh, Young-Joon

2018-04-23

Signal transducer and activator of transcription 3 (STAT3) is a transcription factor that is latent but constitutively activated in many types of cancers. It is well known that STAT3 plays a key role in inflammation-associated tumorigenesis. Curcumin is an anti-inflammatory natural compound isolated from the turmeric (Curcuma longa L., Zingiberaceae) that has been extensively used in a traditional medicine over the centuries. In the present study, we have found that curcumin inhibits STAT3 signaling that is persistently overactivated in H-Ras transformed breast epithelial cells (H-Ras MCF10A). Specific cysteine residues present in STAT3 appear to be critical for the activity as well as conformation of this transcription factor. We identified the cysteine residue 259 of STAT3 as a putative site for curcumin binding. Site-directed mutation of this cysteine residue abolished curcumin-induced inactivation of STAT3 and apoptosis in H-Ras MCF10A cells. The α,β-unsaturated carbonyl moiety of curcumin appears to be essential in its binding to STAT3 in H-Ras MCF10A cells. Tetrahydrocurcumin that lacks such electrophilic moiety failed to interact with STAT3 and to induce apoptosis in the same cell line. Taken together, our findings suggest that curcumin can abrogate aberrant activation of STAT3 through direct interaction, thereby inhibiting STAT3-mediated mammary carcinogenesis.

Berberine suppresses tumorigenicity and growth of nasopharyngeal carcinoma cells by inhibiting STAT3 activation induced by tumor associated fibroblasts.

PubMed

Tsang, Chi Man; Cheung, Yuk Chun; Lui, Vivian Wai-Yan; Yip, Yim Ling; Zhang, Guitao; Lin, Victor Weitao; Cheung, Kenneth Chat-Pan; Feng, Yibin; Tsao, Sai Wah

2013-12-31

Cortidis rhizoma (Huanglian) and its major therapeutic component, berberine, have drawn extensive attention in recent years for their anti-cancer properties. Growth inhibitory effects of berberine on multiple types of human cancer cells have been reported. Berberine inhibits invasion, induces cell cycle arrest and apoptosis in human cancer cells. The anti-inflammatory property of berberine, involving inhibition of Signal Transducer and Activator of Transcription 3 (STAT3) activation, has also been documented. In this study, we have examined the effects of berberine on tumorigenicity and growth of nasopharyngeal carcinoma (NPC) cells and their relationship to STAT3 signaling using both in vivo and in vitro models. Berberine effectively inhibited the tumorigenicity and growth of an EBV-positive NPC cell line (C666-1) in athymic nude mice. Inhibition of tumorigenic growth of NPC cells in vivo was correlated with effective inhibition of STAT3 activation in NPC cells inside the tumor xenografts grown in nude mice. In vitro, berberine inhibited both constitutive and IL-6-induced STAT3 activation in NPC cells. Inhibition of STAT3 activation by berberine induced growth inhibition and apoptotic response in NPC cells. Tumor-associated fibroblasts were found to secret IL-6 and the conditioned medium harvested from the fibroblasts also induced STAT3 activation in NPC cells. Furthermore, STAT3 activation by conditioned medium of tumor-associated fibroblasts could be blocked by berberine or antibodies against IL-6 and IL-6R. Our observation that berberine effectively inhibited activation of STAT3 induced by tumor-associated fibroblasts suggests a role of berberine in modulating the effects of tumor stroma on the growth of NPC cells. The effective inhibition of STAT3 activation in NPC cells by berberine supports its potential use in the treatment of NPC.

Stat3 orchestrates interaction between endothelial and tumor cells and inhibition of Stat3 suppresses brain metastasis of breast cancer cells.

PubMed

Lee, Hsueh-Te; Xue, Jianfei; Chou, Ping-Chieh; Zhou, Aidong; Yang, Phillip; Conrad, Charles A; Aldape, Kenneth D; Priebe, Waldemar; Patterson, Cam; Sawaya, Raymond; Xie, Keping; Huang, Suyun

2015-04-30

Brain metastasis is a major cause of morbidity and mortality in patients with breast cancer. Our previous studies indicated that Stat3 plays an important role in brain metastasis. Here, we present evidence that Stat3 functions at the level of the microenvironment of brain metastases. Stat3 controlled constitutive and inducible VEGFR2 expression in tumor-associated brain endothelial cells. Furthermore, inhibition of Stat3 by WP1066 decreased the incidence of brain metastases and increased survival in a preclinical model of breast cancer brain metastasis. WP1066 inhibited Stat3 activation in tumor-associated endothelial cells, reducing their infiltration and angiogenesis. WP1066 also inhibited breast cancer cell invasion. Our results indicate that WP1066 can inhibit tumor angiogenesis and brain metastasis mediated by Stat3 in endothelial and tumor cells.

Activation of Stat1 by mutant fibroblast growth-factor receptor in thanatophoric dysplasia type II dwarfism.

PubMed

Su, W C; Kitagawa, M; Xue, N; Xie, B; Garofalo, S; Cho, J; Deng, C; Horton, W A; Fu, X Y

1997-03-20

The achondroplasia class of chondrodysplasias comprises the most common genetic forms of dwarfism in humans and includes achondroplasia, hypochondroplasia and thanatophoric dysplasia types I and II (TDI and TDII), which are caused by different mutations in a fibroblast growth-factor receptor FGFR3 (ref. 1). The molecular mechanism and the mediators of these FGFR3-related growth abnormalities are not known. Here we show that mutant TDII FGFR3 has a constitutive tyrosine kinase activity which can specifically activate the transcription factor Stat1 (for signal transducer and activator of transcription). Furthermore, expression of TDII FGFR3 induced nuclear translocation of Stat1, expression of the cell-cycle inhibitor p21(WAF1/CIP1), and growth arrest of the cell. Thus, TDII FGFR3 may use Stat1 as a mediator of growth retardation in bone development. Consistent with this, Stat1 activation and increased p21(WAF1/CIP1) expression was found in the cartilage cells from the TDII fetus, but not in those from the normal fetus. Thus, abnormal STAT activation and p21(WAF1/CIP1) expression by the TDII mutant receptor may be responsible for this FGFR3-related bone disease.

OPB-31121, a novel small molecular inhibitor, disrupts the JAK2/STAT3 pathway and exhibits an antitumor activity in gastric cancer cells.

PubMed

Kim, Mi-Jung; Nam, Hyun-Jin; Kim, Hwang-Phill; Han, Sae-Won; Im, Seock-Ah; Kim, Tae-You; Oh, Do-Youn; Bang, Yung-Jue

2013-07-10

We investigated the mechanisms of action and antitumor effects of OPB-31121, a novel STAT3 inhibitor, in gastric cancer cells. OPB-31121 downregulated JAK2 and gp130 expression and inhibited JAK2 phosphorylation which leads to inhibition of STAT3 phosphorylation. OPB-31121 inhibited constitutively activated and IL-6-induced JAK/STAT signaling pathway. OPB-31121 decreased cell proliferation in both gastric cancer cells and in a xenograft model, induced the apoptosis of gastric cancer cells, inhibited the expression of antiapoptotic proteins, and showed synergism with 5-fluorouracil and cisplatin. Taken together, our study suggests that STAT3 inhibition with OPB-31121 can be tested in patients with gastric cancer. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

Suppression of STAT3 NH2 -terminal domain chemosensitizes medulloblastoma cells by activation of protein inhibitor of activated STAT3 via de-repression by microRNA-21.

PubMed

Ray, Sutapa; Coulter, Don W; Gray, Shawn D; Sughroue, Jason A; Roychoudhury, Shrabasti; McIntyre, Erin M; Chaturvedi, Nagendra K; Bhakat, Kishor K; Joshi, Shantaram S; McGuire, Timothy R; Sharp, John G

2018-04-01

Medulloblastoma (MB) is a malignant pediatric brain tumor with poor prognosis. Signal transducers and activators of transcription-3 (STAT3) is constitutively activated in MB where it functions as an oncoprotein, mediating cancer progression and metastasis. Here, we have delineated the functional role of activated STAT3 in MB, by using a cell permeable STAT3-NH 2 terminal domain inhibitor (S3-NTDi) that specifically perturbs the structure/function of STAT3. We have implemented several biochemical experiments using human MB tumor microarray (TMA) and pediatric MB cell lines, derived from high-risk SHH-TP53-mutated and MYC-amplified Non-WNT/SHH tumors. Treatment of MB cells with S3-NTDi leads to growth inhibition, cell cycle arrest, and apoptosis. S3-NTDi downregulated expression of STAT3 target genes, delayed migration of MB cells, attenuated epithelial-mesenchymal transition (EMT) marker expressions and reduced cancer stem-cell associated protein expressions in MB-spheres. To elucidate mechanisms, we showed that S3-NTDi induce expression of pro-apoptotic gene, C/EBP-homologous protein (CHOP), and decrease association of STAT3 to the proximal promoter of CCND1 and BCL2. Of note, S3-NTDi downregulated microRNA-21, which in turn, de-repressed Protein Inhibitor of Activated STAT3 (PIAS3), a negative regulator of STAT3 signaling pathway. Furthermore, combination therapy with S3-NTDi and cisplatin significantly decreased highly aggressive MYC-amplified MB cell growth and induced apoptosis by downregulating STAT3 regulated proliferation and anti-apoptotic gene expression. Together, our results revealed an important role of STAT3 in regulating MB pathogenesis. Disruption of this pathway with S3-NTDi, therefore, may serves as a promising candidate for targeted MB therapy by enhancing chemosensitivity of MB cells and potentially improving outcomes in high-risk patients. © 2017 Wiley Periodicals, Inc.

STAT3 activation in monocytes accelerates liver cancer progression.

PubMed

Wu, Wen-Yong; Li, Jun; Wu, Zheng-Sheng; Zhang, Chang-Le; Meng, Xiang-Ling

2011-12-05

Signal transducer and activator of transcription 3 (STAT3) is an important transcription factor ubiquitously expressed in different cell types. STAT3 plays an essential role in cell survival, proliferation, and differentiation. Aberrantly hyper-activated STAT3 signaling in cancer cells and in the tumor microenvironment has been detected in a wide variety of human cancers and is considered an important factor for cancer initiation, development, and progression. However, the role of STAT3 activation in monocytes in the development of HCC has not been well understood. Immunohistochemical analysis of phosphorylated STAT3 was performed on tissue microarray from HCC patients. Using a co-culture system in vivo, HCC cell growth was determined by the MTT assay. In vivo experiments were conducted with mice given diethylinitrosamine (DEN), which induces HCC was used to investigate the role of STAT3 expression in monocytes on tumor growth. Real-time PCR was used to determine the expression of cell proliferation and cell arrest associated genes in the tumor and nontumor tissue from liver. Phosphorylated STAT3 was found in human hepatocellular carcinoma tissue samples and was expressed in tumor cells and also in monocytes. Phosphorylated STAT3 expression in monocyte was significantly correlated to advanced clinical stage of HCC and a poor prognosis. Using a co-culture system in vivo, monocytes promoted HCC cell growth via the IL-6/STAT3 signaling pathway. The STAT3 inhibitor, NSC 74859, significantly suppressed tumor growth in vivo in mice with diethylinitrosamine (DEN)-induced HCC. In this animal model, blockade of STAT3 with NSC 74859 induced tumor cell apoptosis, while inhibiting both tumor cells and monocytes proliferation. Furthermore, NSC 74859 treatment suppressed cancer associated inflammation in DEN-induce HCC. Our data suggest constitutively activated STAT3 monocytes promote liver tumorigenesis in clinical patients and animal experiments. Thus, STAT3 in tumor infiltrating inflammatory cells may an attractive target for liver cancer therapy.

Constitutive activation of MEK1 in chondrocytes causes Stat1-independent achondroplasia-like dwarfism and rescues the Fgfr3-deficient mouse phenotype

PubMed Central

Murakami, Shunichi; Balmes, Gener; McKinney, Sandra; Zhang, Zhaoping; Givol, David; de Crombrugghe, Benoit

2004-01-01

We generated transgenic mice that express a constitutively active mutant of MEK1 in chondrocytes. These mice showed a dwarf phenotype similar to achondroplasia, the most common human dwarfism, caused by activating mutations in FGFR3. These mice displayed incomplete hypertrophy of chondrocytes in the growth plates and a general delay in endochondral ossification, whereas chondrocyte proliferation was unaffected. Immunohistochemical analysis of the cranial base in transgenic embryos showed reduced staining for collagen type X and persistent expression of Sox9 in chondrocytes. These observations indicate that the MAPK pathway inhibits hypertrophic differentiation of chondrocytes and negatively regulates bone growth without inhibiting chondrocyte proliferation. Expression of a constitutively active mutant of MEK1 in chondrocytes of Fgfr3-deficient mice inhibited skeletal overgrowth, strongly suggesting that regulation of bone growth by FGFR3 is mediated at least in part by the MAPK pathway. Although loss of Stat1 restored the reduced chondrocyte proliferation in mice expressing an achondroplasia mutant of Fgfr3, it did not rescue the reduced hypertrophic zone, the delay in formation of secondary ossification centers, and the achondroplasia-like phenotype. These observations suggest a model in which Fgfr3 signaling inhibits bone growth by inhibiting chondrocyte differentiation through the MAPK pathway and by inhibiting chondrocyte proliferation through Stat1. PMID:14871928

Direct Interaction of Jak1 and v-Abl Is Required for v-Abl-Induced Activation of STATs and Proliferation

PubMed Central

Danial, Nika N.; Losman, Julie A.; Lu, Tianhong; Yip, Natalie; Krishnan, Kartik; Krolewski, John; Goff, Stephen P.; Wang, Jean Y. J.; Rothman, Paul B.

1998-01-01

In Abelson murine leukemia virus (A-MuLV)-transformed cells, members of the Janus kinase (Jak) family of non-receptor tyrosine kinases and the signal transducers and activators of transcription (STAT) family of signaling proteins are constitutively activated. In these cells, the v-Abl oncoprotein and the Jak proteins physically associate. To define the molecular mechanism of constitutive Jak-STAT signaling in these cells, the functional significance of the v-Abl–Jak association was examined. Mapping the Jak1 interaction domain in v-Abl demonstrates that amino acids 858 to 1080 within the carboxyl-terminal region of v-Abl bind Jak1 through a direct interaction. A mutant of v-Abl lacking this region exhibits a significant defect in Jak1 binding in vivo, fails to activate Jak1 and STAT proteins, and does not support either the proliferation or the survival of BAF/3 cells in the absence of cytokine. Cells expressing this v-Abl mutant show extended latency and decreased frequency in generating tumors in nude mice. In addition, inducible expression of a kinase-inactive mutant of Jak1 protein inhibits the ability of v-Abl to activate STATs and to induce cytokine-independent proliferation, indicating that an active Jak1 is required for these v-Abl-induced signaling pathways in vivo. We propose that Jak1 is a mediator of v-Abl-induced STAT activation and v-Abl induced proliferation in BAF/3 cells, and may be important for efficient transformation of immature B cells by the v-abl oncogene. PMID:9774693

STAT3-activated CD36 facilitates fatty acid uptake in chronic lymphocytic leukemia cells

PubMed Central

Rozovski, Uri; Harris, David M.; Li, Ping; Liu, Zhiming; Jain, Preetesh; Ferrajoli, Alessandra; Burger, Jan; Thompson, Phillip; Jain, Nitin; Wierda, William; Keating, Michael J.; Estrov, Zeev

2018-01-01

Although several studies established that unlike normal B cells chronic lymphocytic leukemia (CLL) cells metabolize fatty acids (FA), how CLL cells internalize FA is poorly understood. Because in various cell types CD36 facilitates FA uptake, we wondered whether a similar mechanism is operative CLL. We found that CD36 levels are higher in CLL cells than in normal B cells, and that small interfering RNA, CD36 neutralizing antibodies or sulfosuccinimidyl oleate (SSO) that inhibits CD36 significantly reduced the oxygen consumption of CLL cells incubated with FA. Because CD36 is oeverexpressed and STAT3 is constitutively activated in CLL cells, we wondered whether STAT3 induces CD36 expression. Sequence analysis identified putative STAT3 binding sites in the CD36 gene promoter. Chromatin immunoprecipitation and an electrophoretic mobility shift assay revealed that STAT3 binds to the CD36 gene promoter. A luciferase assay and STAT3-small hairpin RNA, that significantly decreased the levels of CD36 in CLL cells, established that STAT3 activates the transcription of the CD36 gene. Furthermore, SSO induced a dose-dependent apoptosis of CLL cells. Taken together, our data suggest that STAT3 activates CD36 and that CD36 facilitates FA uptake in CLL cells. Whether CD36 inhibition would provide clinical benefits in CLL remains to be determined. PMID:29765537

APC loss in breast cancer leads to doxorubicin resistance via STAT3 activation.

PubMed

VanKlompenberg, Monica K; Leyden, Emily; Arnason, Anne H; Zhang, Jian-Ting; Stefanski, Casey D; Prosperi, Jenifer R

2017-11-28

Resistance to chemotherapy is one of the leading causes of death from breast cancer. We recently established that loss of Adenomatous Polyposis Coli (APC) in the Mouse Mammary Tumor Virus - Polyoma middle T (MMTV-PyMT) transgenic mouse model results in resistance to cisplatin or doxorubicin-induced apoptosis. Herein, we aim to establish the mechanism that is responsible for APC-mediated chemotherapeutic resistance. Our data demonstrate that MMTV-PyMT; Apc Min/+ cells have increased signal transducer and activator of transcription 3 (STAT3) activation. STAT3 can be constitutively activated in breast cancer, maintains the tumor initiating cell (TIC) population, and upregulates multidrug resistance protein 1 (MDR1). The activation of STAT3 in the MMTV-PyMT; Apc Min/+ model is independent of interleukin 6 (IL-6); however, enhanced EGFR expression in the MMTV-PyMT; Apc Min/+ cells may be responsible for the increased STAT3 activation. Inhibiting STAT3 with a small molecule inhibitor A69 in combination with doxorubicin, but not cisplatin, restores drug sensitivity. A69 also decreases doxorubicin enhanced MDR1 gene expression and the TIC population enhanced by loss of APC. In summary, these results have revealed the molecular mechanisms of APC loss in breast cancer that can guide future treatment plans to counteract chemotherapeutic resistance.

Short Stat5-Interacting Peptide Derived from Phospholipase C-β3 Inhibits Hematopoietic Cell Proliferation and Myeloid Differentiation

PubMed Central

Yasudo, Hiroki; Ando, Tomoaki; Xiao, Wenbin; Kawakami, Yuko; Kawakami, Toshiaki

2011-01-01

Constitutive activation of the transcription factor Stat5 in hematopoietic stem/progenitor cells leads to various hematopoietic malignancies including myeloproliferative neoplasm (MPN). Our recent study found that phospholipase C (PLC)-β3 is a novel tumor suppressor involved in MPN, lymphoma and other tumors. Stat5 activity is negatively regulated by the SH2 domain-containing protein phosphatase SHP-1 in a PLC-β3-dependent manner. PLC-β3 can form the multimolecular SPS complex together with SHP-1 and Stat5. The close physical proximity of SHP-1 and Stat5 brought about by interacting with the C-terminal segment of PLC-β3 (PLC-β3-CT) accelerates SHP-1-mediated dephosphorylation of Stat5. Here we identify the minimal sequences within PLC-β3-CT required for its tumor suppressor function. Two of the three Stat5-binding noncontiguous regions, one of which also binds SHP-1, substantially inhibited in vitro proliferation of Ba/F3 cells. Surprisingly, an 11-residue Stat5-binding peptide (residues 988-998) suppressed Stat5 activity in Ba/F3 cells and in vivo proliferation and myeloid differentiation of hematopoietic stem/progenitor cells. Therefore, this study further defines PLC-β3-CT as the Stat5- and SHP-1-binding domain by identifying minimal functional sequences of PLC-β3 for its tumor suppressor function and implies their potential utility in the control of hematopoietic malignancies. PMID:21949826

EBV latent membrane protein 1 activates Akt, NFkappaB, and Stat3 in B cell lymphomas.

PubMed

Shair, Kathy H Y; Bendt, Katherine M; Edwards, Rachel H; Bedford, Elisabeth C; Nielsen, Judith N; Raab-Traub, Nancy

2007-11-01

Latent membrane protein 1 (LMP1) is the major oncoprotein of Epstein-Barr virus (EBV). In transgenic mice, LMP1 promotes increased lymphoma development by 12 mo of age. This study reveals that lymphoma develops in B-1a lymphocytes, a population that is associated with transformation in older mice. The lymphoma cells have deregulated cell cycle markers, and inhibitors of Akt, NFkappaB, and Stat3 block the enhanced viability of LMP1 transgenic lymphocytes and lymphoma cells in vitro. Lymphoma cells are independent of IL4/Stat6 signaling for survival and proliferation, but have constitutively activated Stat3 signaling. These same targets are also deregulated in wild-type B-1a lymphomas that arise spontaneously through age predisposition. These results suggest that Akt, NFkappaB, and Stat3 pathways may serve as effective targets in the treatment of EBV-associated B cell lymphomas.

Berberine suppresses tumorigenicity and growth of nasopharyngeal carcinoma cells by inhibiting STAT3 activation induced by tumor associated fibroblasts

PubMed Central

2013-01-01

Background Cortidis rhizoma (Huanglian) and its major therapeutic component, berberine, have drawn extensive attention in recent years for their anti-cancer properties. Growth inhibitory effects of berberine on multiple types of human cancer cells have been reported. Berberine inhibits invasion, induces cell cycle arrest and apoptosis in human cancer cells. The anti-inflammatory property of berberine, involving inhibition of Signal Transducer and Activator of Transcription 3 (STAT3) activation, has also been documented. Methods In this study, we have examined the effects of berberine on tumorigenicity and growth of nasopharyngeal carcinoma (NPC) cells and their relationship to STAT3 signaling using both in vivo and in vitro models. Results Berberine effectively inhibited the tumorigenicity and growth of an EBV-positive NPC cell line (C666-1) in athymic nude mice. Inhibition of tumorigenic growth of NPC cells in vivo was correlated with effective inhibition of STAT3 activation in NPC cells inside the tumor xenografts grown in nude mice. In vitro, berberine inhibited both constitutive and IL-6-induced STAT3 activation in NPC cells. Inhibition of STAT3 activation by berberine induced growth inhibition and apoptotic response in NPC cells. Tumor-associated fibroblasts were found to secret IL-6 and the conditioned medium harvested from the fibroblasts also induced STAT3 activation in NPC cells. Furthermore, STAT3 activation by conditioned medium of tumor-associated fibroblasts could be blocked by berberine or antibodies against IL-6 and IL-6R. Conclusions Our observation that berberine effectively inhibited activation of STAT3 induced by tumor-associated fibroblasts suggests a role of berberine in modulating the effects of tumor stroma on the growth of NPC cells. The effective inhibition of STAT3 activation in NPC cells by berberine supports its potential use in the treatment of NPC. PMID:24380387

Targeted Blockage of Signal Transducer and Activator of Transcription 5 Signaling Pathway with Decoy Oligodeoxynucleotides Suppresses Leukemic K562 Cell Growth

PubMed Central

Wang, Xiaozhong; Zeng, Jianming; Shi, Mei; Zhao, Shiqiao; Bai, Weijun; Cao, Weixi; Tu, Zhiguang; Huang, Zonggan

2011-01-01

The protein signal transducer and activator of transcription 5 (STAT5) of the JAK/STAT pathway is constitutively activated because of its phosphorylation by tyrosine kinase activity of fusion protein BCR-ABL in chronic myelogenous leukemia (CML) cells. This study investigated the potential therapeutic effect of STAT5 decoy oligodeoxynucleotides (ODN) using leukemia K562 cells as a model. Our results showed that transfection of 21-mer-long STAT5 decoy ODN into K562 cells effectively inhibited cell proliferation and induced cell apoptosis. Further, STAT5 decoy ODN downregulated STAT5 targets bcl-xL, cyclinD1, and c-myc at both mRNA and protein levels in a sequence-specific manner. Collectively, these data demonstrate the therapeutic effect of blocking the STAT5 signal pathway by cis-element decoy for cancer characterized by constitutive STAT5 activation. Thus, our study provides support for STAT5 as a potential target downstream of BCR-ABL for CML treatment and helps establish the concept of targeting STAT5 by decoy ODN as a novel therapy approach for imatinib-resistant CML. PMID:21091189

Heteronemin Is a Novel c-Met/STAT3 Inhibitor Against Advanced Prostate Cancer Cells.

PubMed

Wu, Jian-Ching; Wang, Chiang-Ting; Hung, Han-Chun; Wu, Wen-Jeng; Wu, Deng-Chyang; Chang, Min-Chi; Sung, Ping-Jyun; Chou, Yu-Wei; Wen, Zhi-Hong; Tai, Ming-Hong

2016-12-01

Prostate cancer is one of the most prevalent cancers in men worldwide. Aberrant activation of c-Met/signal transducer and activator of transcription-3 (STAT3) signaling is involved in prostate carcinogenesis, underscoring the demand for developing c-Met/STAT3-targeting drugs. Thus, we first utilized virtual screening strategy to identify STAT3-inhibiting marine compound, heteronemin, and then validated the STAT3-inhibiting function of heteronemin in prostate cancer cells. Human prostate cancer LNCaP, DU145, and PC-3 cell lines were treated with heteronemin for 24 hr, then the cell viability was evaluated by MTT assay. Flow cytometry was performed to analyze the apoptosis in heteronemin-treated cells. Western blot and quantitative real-time PCR were executed to further confirm the c-Met/STAT3 signaling inhibition by heteronemin in DU145 and PC-3 cells. In this study, we employed the virtual screening strategy to identify heteronemin, a spongean sesterterpene, as a potential STAT3 inhibitor from Taiwan marine drugs library. Application of heteronemin potently suppressed the viability and anchorage-independent growth of human prostate cancer cells. Besides, heteronemin induced apoptosis in prostate cancer cells by activation of both intrinsic (caspase-9) and extrinsic (caspase-8) apoptotic pathways. By luciferase assay and expression analysis, it was confirmed that heteronemin inhibited the phosphorylation of c-Met/src/STAT3 signaling axis, STAT3-driven luciferase activities and expression of STAT3-regulated genes including Bcl-xL, Bcl-2, and Cyclin D1. Finally, heteronemin effectively antagonized the hepatocyte growth factor (HGF)-stimulated c-Met/STAT3 activation as well as the proliferation and colonies formation in refractory prostate cancer cells. These findings suggest that heteronemin may constitute a novel c-Met/STAT3-targeting agent for prostate cancer. Prostate 76:1469-1483, 2016. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

STAT proteins: from normal control of cellular events to tumorigenesis.

PubMed

Calò, Valentina; Migliavacca, Manuela; Bazan, Viviana; Macaluso, Marcella; Buscemi, Maria; Gebbia, Nicola; Russo, Antonio

2003-11-01

Signal transducers and activators of transcription (STAT) proteins comprise a family of transcription factors latent in the cytoplasm that participate in normal cellular events, such as differentiation, proliferation, cell survival, apoptosis, and angiogenesis following cytokine, growth factor, and hormone signaling. STATs are activated by tyrosine phosphorylation, which is normally a transient and tightly regulates process. Nevertheless, several constitutively activated STATs have been observed in a wide number of human cancer cell lines and primary tumors, including blood malignancies and solid neoplasias. STATs can be divided into two groups according to their specific functions. One is made up of STAT2, STAT4, and STAT6, which are activated by a small number of cytokines and play a distinct role in the development of T-cells and in IFNgamma signaling. The other group includes STAT1, STAT3, and STAT5, activated in different tissues by means of a series of ligands and involved in IFN signaling, development of the mammary gland, response to GH, and embriogenesis. This latter group of STATS plays an important role in controlling cell-cycle progression and apoptosis and thus contributes to oncogenesis. Although an increased expression of STAT1 has been observed in many human neoplasias, this molecule can be considered a potential tumor suppressor, since it plays an important role in growth arrest and in promoting apoptosis. On the other hand, STAT3 and 5 are considered as oncogenes, since they bring about the activation of cyclin D1, c-Myc, and bcl-xl expression, and are involved in promoting cell-cycle progression, cellular transformation, and in preventing apoptosis.

Negative regulation of STAT3 protein-mediated cellular respiration by SIRT1 protein.

PubMed

Bernier, Michel; Paul, Rajib K; Martin-Montalvo, Alejandro; Scheibye-Knudsen, Morten; Song, Shaoming; He, Hua-Jun; Armour, Sean M; Hubbard, Basil P; Bohr, Vilhelm A; Wang, Lili; Zong, Yaping; Sinclair, David A; de Cabo, Rafael

2011-06-03

In mammals, the transcriptional activity of signal transducer and activator of transcription 3 (STAT3) is regulated by the deacetylase SIRT1. However, whether the newly described nongenomic actions of STAT3 toward mitochondrial oxidative phosphorylation are dependent on SIRT1 is unclear. In this study, Sirt1 gene knock-out murine embryonic fibroblast (MEF) cells were used to delineate the role of SIRT1 in the regulation of STAT3 mitochondrial function. Here, we show that STAT3 mRNA and protein levels and the accumulation of serine-phosphorylated STAT3 in mitochondria were increased significantly in Sirt1-KO cells as compared with wild-type MEFs. Various mitochondrial bioenergetic parameters, such as the oxygen consumption rate in cell cultures, enzyme activities of the electron transport chain complexes in isolated mitochondria, and production of ATP and lactate, indicated that Sirt1-KO cells exhibited higher mitochondrial respiration as compared with wild-type MEFs. Two independent approaches, including ectopic expression of SIRT1 and siRNA-mediated knockdown of STAT3, led to reduction in intracellular ATP levels and increased lactate production in Sirt1-KO cells that were approaching those of wild-type controls. Comparison of profiles of phospho-antibody array data indicated that the deletion of SirT1 was accompanied by constitutive activation of the pro-inflammatory NF-κB pathway, which is key for STAT3 induction and increased cellular respiration in Sirt1-KO cells. Thus, SIRT1 appears to be a functional regulator of NF-κB-dependent STAT3 expression that induces mitochondrial biogenesis. These results have implications for understanding the interplay between STAT3 and SIRT1 in pro-inflammatory conditions.

Negative Regulation of STAT3 Protein-mediated Cellular Respiration by SIRT1 Protein*

PubMed Central

Bernier, Michel; Paul, Rajib K.; Martin-Montalvo, Alejandro; Scheibye-Knudsen, Morten; Song, Shaoming; He, Hua-Jun; Armour, Sean M.; Hubbard, Basil P.; Bohr, Vilhelm A.; Wang, Lili; Zong, Yaping; Sinclair, David A.; de Cabo, Rafael

2011-01-01

In mammals, the transcriptional activity of signal transducer and activator of transcription 3 (STAT3) is regulated by the deacetylase SIRT1. However, whether the newly described nongenomic actions of STAT3 toward mitochondrial oxidative phosphorylation are dependent on SIRT1 is unclear. In this study, Sirt1 gene knock-out murine embryonic fibroblast (MEF) cells were used to delineate the role of SIRT1 in the regulation of STAT3 mitochondrial function. Here, we show that STAT3 mRNA and protein levels and the accumulation of serine-phosphorylated STAT3 in mitochondria were increased significantly in Sirt1-KO cells as compared with wild-type MEFs. Various mitochondrial bioenergetic parameters, such as the oxygen consumption rate in cell cultures, enzyme activities of the electron transport chain complexes in isolated mitochondria, and production of ATP and lactate, indicated that Sirt1-KO cells exhibited higher mitochondrial respiration as compared with wild-type MEFs. Two independent approaches, including ectopic expression of SIRT1 and siRNA-mediated knockdown of STAT3, led to reduction in intracellular ATP levels and increased lactate production in Sirt1-KO cells that were approaching those of wild-type controls. Comparison of profiles of phospho-antibody array data indicated that the deletion of SirT1 was accompanied by constitutive activation of the pro-inflammatory NF-κB pathway, which is key for STAT3 induction and increased cellular respiration in Sirt1-KO cells. Thus, SIRT1 appears to be a functional regulator of NF-κB-dependent STAT3 expression that induces mitochondrial biogenesis. These results have implications for understanding the interplay between STAT3 and SIRT1 in pro-inflammatory conditions. PMID:21467030

Identification of the kinase that activates a nonmetazoan STAT gives insights into the evolution of phosphotyrosine-SH2 domain signaling.

PubMed

Araki, Tsuyoshi; Kawata, Takefumi; Williams, Jeffrey G

2012-07-10

SH2 domains are integral to many animal signaling pathways. By interacting with specific phosphotyrosine residues, they provide regulatable protein-protein interaction domains. Dictyostelium is the only nonmetazoan with functionally characterized SH2 domains, but the cognate tyrosine kinases are unknown. There are no orthologs of the animal tyrosine kinases, but there are very many tyrosine kinase-like kinases (TKLs), a group of kinases which, despite their family name, are classified mainly as serine-threonine kinases. STATs are transcription factors that dimerize via phosphotyrosine-SH2 domain interactions. STATc is activated by phosphorylation on Tyr922 when cells are exposed to the prestalk inducer differentiation inducing factor (DIF-1), a chlorinated hexaphenone. We show that in a null mutant for Pyk2, a tyrosine-specific TKL, exposure to DIF-1 does not activate STATc. Conversely, overexpression of Pyk2 causes constitutive STATc activation. Pyk2 phosphorylates STATc on Tyr922 in vitro and complexes with STATc both in vitro and in vivo. This demonstration that a TKL directly activates a STAT has significant implications for understanding the evolutionary origins of SH2 domain-phosphotyrosine signaling. It also has mechanistic implications. Our previous work suggested that a predicted constitutive STATc tyrosine kinase activity is counterbalanced in vivo by the DIF-1-regulated activity of PTP3, a Tyr922 phosphatase. Here we show that the STATc-Pyk2 complex is formed constitutively by an interaction between the STATc SH2 domain and phosphotyrosine residues on Pyk2 that are generated by autophosphorylation. Also, as predicted, Pyk2 is constitutively active as a STATc kinase. This observation provides further evidence for this highly atypical, possibly ancestral, STAT regulation mechanism.

A single tyrosine of the interleukin-9 (IL-9) receptor is required for STAT activation, antiapoptotic activity, and growth regulation by IL-9.

PubMed

Demoulin, J B; Uyttenhove, C; Van Roost, E; DeLestré, B; Donckers, D; Van Snick, J; Renauld, J C

1996-09-01

Interleukin-9 (IL-9), a T-cell-derived cytokine, interacts with a specific receptor associated with the IL-2 receptor gamma chain. In this report, we analyze the functional domains of the human IL-9 receptor transfected into mouse lymphoid cell lines. Three different functions were examined: growth stimulation in factor-dependent pro-B Ba/F3 cells, protection against dexamethasone-induced apoptosis, and Ly-6A2 induction in BW5147 lymphoma cells. The results indicated that a single tyrosine, at position 116 in the cytoplasmic domain, was required for all three activities. In addition, we observed that human IL-9 reduced the proliferation rate of transfected BW5147 cells, an effect also dependent on the same tyrosine. This amino acid was necessary for IL-9-mediated tyrosine phosphorylation of the receptor and for STAT activation but not for IRS-2/4PS activation or for JAK1 phosphorylation, which depended on a domain closer to the plasma membrane. We also showed that JAK1 was constitutively associated with the IL-9 receptor. Activated STAT complexes induced by IL-9 were found to contain STAT1, STAT3, and STAT5 transcription factors. Moreover, sequence homologies between human IL-9 receptor tyrosine 116 and tyrosines (of other receptors activating STAT3 and STAT5 were observed. Taken together, these data indicate that a single tyrosine of the IL-9 receptor, required for activation of three different STAT proteins, is necessary for distinct activities of this cytokine, including proliferative responses.

Control of glioblastoma tumorigenesis by feed-forward cytokine signaling

PubMed Central

Jahani-Asl, Arezu; Yin, Hang; Soleimani, Vahab D; Haque, Takrima; Luchman, H Artee; Chang, Natasha C; Sincennes, Marie-Claude; Puram, Sidharth V; Scott, Andrew M; Lorimer, Ian A J; Perkins, Theodore J; Ligon, Keith L; Weiss, Samuel; Rudnicki, Michael A; Bonni, Azad

2016-01-01

EGFRvIII-STAT3 signaling is important in glioblastoma pathogenesis. Here, we identified the cytokine receptor OSMR as a direct target gene of the transcription factor STAT3 in mouse astrocytes and human brain tumor stem cells (BTSCs). We found that OSMR functioned as an essential co-receptor for EGFRvIII. OSMR formed a physical complex with EGFRvIII, and depletion of OSMR impaired EGFRvIII-STAT3 signaling. Conversely, pharmacological inhibition of EGFRvIII phosphorylation inhibited the EGFRvIII-OSMR interaction and activation of STAT3. EGFRvIII-OSMR signaling in tumors operated constitutively, whereas EGFR-OSMR signaling in nontumor cells was synergistically activated by the ligands EGF and OSM. Finally, knockdown of OSMR strongly suppressed cell proliferation and tumor growth of mouse glioblastoma cells and human BTSC xenografts in mice, and prolonged the lifespan of those mice. Our findings identify OSMR as a critical regulator of glioblastoma tumor growth that orchestrates a feed-forward signaling mechanism with EGFRvIII and STAT3 to drive tumorigenesis. PMID:27110918

Analysis of the activation status of Akt, NFkappaB, and Stat3 in human diffuse gliomas.

PubMed

Wang, Huamin; Wang, Hua; Zhang, Wei; Huang, Helen J; Liao, Warren S L; Fuller, Gregory N

2004-08-01

Loss of phosphatase and tensin homolog (PTEN) and amplification of the epidermal growth factor receptor (EGFR) gene contribute to the progression of gliomas. As downstream targets of the PTEN and EGFR signaling pathways, Akt, NFkappaB, and signal transducer and activator of transcription-3 (Stat3) have been shown to play important roles in the control of cell proliferation, apoptosis, and oncogenesis. We examined the activation status of Akt, NFkappaB, and Stat3 in 259 diffuse gliomas using tissue microarrays and immunohistochemistry, and evaluated their association with glioma grade. We observed significant positive correlations between the activation status of Akt and NFkappaB and glioma grade. In contrast, only focal immunoreactivity for phospho-Stat3 was observed in < 9% of high-grade gliomas. In addition, we observed a significant correlation between the activation of Akt and NFkappaB. Functional correlation between Akt activation and the activation of NFkappaB was confirmed in U251MG GBM cells in which inhibition of Akt activation either by stable expression of PTEN or by the PI3-kinase inhibitors, wortmannin and LY294002, led to a concomitant decrease in NFkappaB-binding activity. Thus, our results demonstrate that constitutive activation of Akt and NFkappaB, but not Stat3, contributes significantly to the progression of diffuse gliomas, and activation of Akt may lead to NFkappaB activation in high-grade gliomas.

The role of STATs in transcriptional control and their impact on cellular function.

PubMed

Bromberg, J; Darnell, J E

2000-05-15

The STAT proteins (Signal Transducers and Activators of Transcription), were identified in the last decade as transcription factors which were critical in mediating virtually all cytokine driven signaling. These proteins are latent in the cytoplasm and become activated through tyrosine phosphorylation which typically occurs through cytokine receptor associated kinases (JAKs) or growth factor receptor tyrosine kinases. Recently a number of non-receptor tyrosine kinases (for example src and abl) have been found to cause STAT phosphorylation. Phosphorylated STATs form homo- or hetero-dimers, enter the nucleus and working coordinately with other transcriptional co-activators or transcription factors lead to increased transcriptional initiation. In normal cells and in animals, ligand dependent activation of the STATs is a transient process, lasting for several minutes to several hours. In contrast, in many cancerous cell lines and tumors, where growth factor dysregulation is frequently at the heart of cellular transformation, the STAT proteins (in particular Stats 1, 3 and 5) are persistently tyrosine phosphorylated or activated. The importance of STAT activation to growth control in experiments using anti-sense molecules or dominant negative STAT protein encoding constructs performed in cell lines or studies in animals lacking specific STATs strongly indicate that STATs play an important role in controlling cell cycle progression and apoptosis. Stat1 plays an important role in growth arrest, in promoting apoptosis and is implicated as a tumor suppressor; while Stats 3 and 5 are involved in promoting cell cycle progression and cellular transformation and preventing apoptosis. Many questions remain including: (1) a better understanding of how the STAT proteins through association with other factors increase transcription initiation; (2) a more complete definition of the sets of genes which are activated by different STATs and (3) how these sets of activated genes differ as a function of cell type. Finally, in the context of many cancers, where STATs are frequently persistently activated, an understanding of the mechanisms leading to their constitutive activation and defining the potential importance of persistent STAT activation in human tumorigenesis remains. Oncogene (2000).

Activation of the NF-κB pathway by the STAT3 inhibitor JSI-124 in human glioblastoma cells

PubMed Central

McFarland, Braden C.; Gray, G. Kenneth; Nozell, Susan E.; Hong, Suk W.; Benveniste, Etty N.

2013-01-01

Glioblastoma tumors are characterized by their invasiveness and resistance to therapies. The transcription factor STAT3 was recently identified as a master transcriptional regulator in the mesenchymal subtype of GBM, which has generated an increased interest in targeting STAT3. We have evaluated more closely the mechanism of action of one particular STAT3 inhibitor, JSI-124 (cucurbitacin I). In this study, we confirmed that JSI-124 inhibits both constitutive and stimulus-induced JAK2 and STAT3 phosphorylation, and decreases cell proliferation while inducing apoptosis in cultured GBM cells. However, we discovered that prior to the inhibition of STAT3, JSI-124 activates the NF-κB pathway, via NF-κB p65 phosphorylation and nuclear translocation. In addition, JSI-124 treatment induces the expression of IL-6, IL-8 and SOCS3 mRNA, which leads to a corresponding increase in IL-6, IL-8 and SOCS3 protein expression. Moreover, the NF-κB driven SOCS3 expression acts as a negative regulator of STAT3, abrogating any subsequent STAT3 activation and provides a mechanism of STAT3 inhibition following JSI-124 treatment. Chromatin immunoprecipitation analysis confirms that NF-κB p65 in addition to other activating co-factors are found at the promoters of IL-6, IL-8 and SOCS3, following JSI-124 treatment. Using pharmacological inhibition of NF-κB and inducible knockdown of NF-κB p65, we found that JSI-124-induced expression of IL-6, IL-8 and SOCS3 was significantly inhibited, demonstrating an NF-κB dependent mechanism. Our data indicate that although JSI-124 may demonstrate potential anti-tumor effects through inhibition of STAT3, other off-target pro-inflammatory pathways are activated, emphasizing that more careful and thorough pre-clinical investigations must be implemented to prevent potential harmful effects. PMID:23386688

Formononetin-induced oxidative stress abrogates the activation of STAT3/5 signaling axis and suppresses the tumor growth in multiple myeloma preclinical model.

PubMed

Kim, Chulwon; Lee, Seok-Geun; Yang, Woong Mo; Arfuso, Frank; Um, Jae-Young; Kumar, Alan Prem; Bian, Jinsong; Sethi, Gautam; Ahn, Kwang Seok

2018-05-29

Aberrant reactions of signal transducer and transcriptional activator (STAT) are frequently detected in multiple myeloma (MM) cancers and can upregulate the expression of multiple genes related to cell proliferation, survival, metastasis, and angiogenesis. Therefore, agents capable of inhibiting STAT activation can form the basis of novel therapies for MM patients. In the present study, we investigated whether the potential anti-cancer effects of Formononetin (FT), a naturally occurring isoflavone derived from Astragalus membranaceus, Trifolium pratense, Glycyrrhiza glabra, and Pueraria lobata, against MM cell lines and human multiple myeloma xenograft tumors in athymic nu/nu mice model are mediated through the negative regulation of STAT3 and STAT5 pathways. Data from the in vitro studies indicated that FT could significantly inhibit cell viability, and induce apoptosis. Interestingly, FT also suppressed constitutive STAT3 (tyrosine residue 705 and serine residue 727) and STAT5 (tyrosine residue 694/699) activation, which correlated with the suppression of the upstream kinases (JAK1, JAK2, and c-Src) in MM cells, and this effect was found to be mediated via an increased production of reactive oxygen species (ROS) due to GSH/GSSG imbalance. Also, FT abrogated STAT3 and STAT5 DNA binding capacity and nuclear translocation. FT induced cell cycle arrest, downregulated the expression of STAT3-regulated anti-apoptotic, angiogenetic, and proliferative gene products; and this correlated with induction of caspase-3 activation and cleavage of PARP. Intraperitoneal administration of FT significantly suppressed the tumor growth in the multiple myeloma xenograft mouse model without exhibiting any significant adverse effects. Overall, our findings indicate that FT exhibits significant anti-cancer effects in MM that may be primarily mediated through the ROS-regulated inhibition of the STAT3 and STAT5 signaling cascade. Copyright © 2018 Elsevier B.V. All rights reserved.

Transforming properties of the Huntingtin interacting protein 1/ platelet-derived growth factor beta receptor fusion protein.

PubMed

Ross, T S; Gilliland, D G

1999-08-06

We have previously reported that the Huntingtin interacting protein 1 (HIP1) gene is fused to the platelet-derived growth factor beta receptor (PDGFbetaR) gene in a patient with chronic myelomonocytic leukemia. We now show that HIP1/PDGFbetaR oligomerizes, is constitutively tyrosine-phosphorylated, and transforms the murine hematopoietic cell line, Ba/F3, to interleukin-3-independent growth. A kinase-inactive mutant is neither tyrosine-phosphorylated nor able to transform Ba/F3 cells. Oligomerization and kinase activation required the 55-amino acid carboxyl-terminal TALIN homology region but not the leucine zipper domain. Tyrosine phosphorylation of a 130-kDa protein and STAT5 correlates with transformation in cells expressing HIP1/PDGFbetaR and related mutants. A deletion mutant fusion protein that contains only the TALIN homology region of HIP1 fused to PDGFbetaR is incapable of transforming Ba/F3 cells and does not tyrosine-phosphorylate p130 or STAT5, although it is itself constitutively tyrosine-phosphorylated. We have also analyzed cells expressing Tyr --> Phe mutants of HIP1/PDGFbetaR in the known PDGFbetaR SH2 docking sites and report that none of these sites are necessary for STAT5 activation, p130 phosphorylation, or Ba/F3 transformation. The correlation of factor-independent growth of hematopoietic cells with p130 and STAT5 phosphorylation/activation in both the HIP1/PDGFbetaR Tyr --> Phe and deletion mutational variants suggests that both STAT5 and p130 are important for transformation mediated by HIP1/PDGFbetaR.

Ultrasound Targeted Microbubble Destruction-Mediated Delivery of a Transcription Factor Decoy Inhibits STAT3 Signaling and Tumor Growth

PubMed Central

Kopechek, Jonathan A.; Carson, Andrew R.; McTiernan, Charles F.; Chen, Xucai; Hasjim, Bima; Lavery, Linda; Sen, Malabika; Grandis, Jennifer R.; Villanueva, Flordeliza S.

2015-01-01

Signal transducer and activator of transcription 3 (STAT3) is constitutively activated in many cancers where it acts to promote tumor progression. A STAT3-specific transcription factor decoy has been developed to suppress STAT3 downstream signaling, but a delivery strategy is needed to improve clinical translation. Ultrasound-targeted microbubble destruction (UTMD) has been shown to enhance image-guided local delivery of molecular therapeutics to a target site. The objective of this study was to deliver STAT3 decoy to squamous cell carcinoma (SCC) tumors using UTMD to disrupt STAT3 signaling and inhibit tumor growth. Studies performed demonstrated that UTMD treatment with STAT3 decoy-loaded microbubbles inhibited STAT3 signaling in SCC cells in vitro. Studies performed in vivo demonstrated that UTMD treatment with STAT3 decoy-loaded microbubbles induced significant tumor growth inhibition (31-51% reduced tumor volume vs. controls, p < 0.05) in mice bearing SCC tumors. Furthermore, expression of STAT3 downstream target genes (Bcl-xL and cyclin D1) was significantly reduced (34-39%, p < 0.05) in tumors receiving UTMD treatment with STAT3 decoy-loaded microbubbles compared to controls. In addition, the quantity of radiolabeled STAT3 decoy detected in tumors eight hours after treatment was significantly higher with UTMD treatment compared to controls (70-150%, p < 0.05). This study demonstrates that UTMD can increase delivery of a transcription factor decoy to tumors in vivo and that the decoy can inhibit STAT3 signaling and tumor growth. These results suggest that UTMD treatment holds potential for clinical use to increase the concentration of a transcription factor signaling inhibitor in the tumor. PMID:26681983

Disruption of Stat3 reveals a critical role in both the initiation and the promotion stages of epithelial carcinogenesis

PubMed Central

Chan, Keith Syson; Sano, Shigetoshi; Kiguchi, Kaoru; Anders, Joanne; Komazawa, Nobuyasu; Takeda, Junji; DiGiovanni, John

2004-01-01

Constitutive activation of signal transducer and activator of transcription 3 (Stat3) has been found in a wide spectrum of human malignancies. Here, we have assessed the effect of Stat3 deficiency on skin tumor development using the 2-stage chemical carcinogenesis model. The epidermis of Stat3-deficient mice showed a significantly reduced proliferative response following treatment with the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) because of a defect in G1-to-S-phase cell cycle progression. Treatment with the tumor initiator 7,12-dimethylbenz[a]anthracene (DMBA) resulted in a significant increase in the number of keratinocyte stem cells undergoing apoptosis in the bulge region of hair follicles of Stat3-deficient mice compared with nontransgenic littermates. Notably, Stat3-deficient mice were completely resistant to skin tumor development when DMBA was used as the initiator and TPA as the promoter. Abrogation of Stat3 function using a decoy oligonucleotide inhibited the growth of initiated keratinocytes possessing an activated Ha-ras gene, both in vitro and in vivo. In addition, injection of Stat3 decoy into skin tumors inhibited their growth. To our knowledge, these data provide the first evidence that Stat3 is required for de novo epithelial carcinogenesis, through maintaining the survival of DNA-damaged stem cells and through mediating and maintaining the proliferation necessary for clonal expansion of initiated cells during tumor promotion. Collectively, these data suggest that, in addition to its emerging role as a target for cancer therapy, Stat3 may also be a target for cancer prevention strategies. PMID:15343391

Stage-specific disruption of Stat3 demonstrates a direct requirement during both the initiation and promotion stages of mouse skin tumorigenesis.

PubMed

Kataoka, Ken; Kim, Dae Joon; Carbajal, Steve; Clifford, John L; DiGiovanni, John

2008-06-01

Constitutive activation of signal transducer and activator of transcription 3 (Stat3) has been found in a variety of human malignancies and has been suggested to play an important role in carcinogenesis. Recently, our laboratory demonstrated that Stat3 is required for the development of skin tumors via two-stage carcinogenesis using skin-specific loss-of-function transgenic mice. To investigate further the role of Stat3 in each stage of chemical carcinogenesis in mouse skin, i.e. initiation and promotion stages, we generated inducible Stat3-deficient mice (K5.Cre-ER(T2) x Stat3(fl/fl)) that show epidermal-specific disruption of Stat3 following topical treatment with 4-hydroxytamoxifen (TM). The epidermis of inducible Stat3-deficient mice treated with TM showed a significant increase in apoptosis induced by 7,12-dimethylbenz[a]anthracene (DMBA) and reduced proliferation following exposure to 12-O-tetradecanoylphorbol-13-acetate. In two-stage skin carcinogenesis assays, inducible Stat3-deficient mice treated with TM during the promotion stage showed a significant delay of tumor development and a significantly reduced number of tumors compared with control groups. Inducible Stat3-deficient mice treated with TM before initiation with DMBA also showed a significant delay in tumor development and a significantly reduced number of tumors compared with control groups. Finally, treatment of inducible Stat3-deficient mice that had existing skin tumors generated by the two-stage carcinogenesis protocol with TM (by intraperitoneal injection) led to inhibition of tumor growth compared with tumors formed in control groups. Collectively, these results directly demonstrate that Stat3 is required for skin tumor development during both the initiation and promotion stages of skin carcinogenesis in vivo.

6-Shogaol exerts anti-proliferative and pro-apoptotic effects through the modulation of STAT3 and MAPKs signaling pathways.

PubMed

Kim, Sung-Moo; Kim, Chulwon; Bae, Hang; Lee, Jong Hyun; Baek, Seung Ho; Nam, Dongwoo; Chung, Won-Seok; Shim, Bum Sang; Lee, Seok-Geun; Kim, Sung-Hoon; Sethi, Gautam; Ahn, Kwang Seok

2015-10-01

6-shogaol (6SG), one of active ingredients in ginger (Zingiber officinale), is known to exhibit anti-proliferative, anti-metastatic, and pro-apoptotic activities through a mechanism that is not fully elucidated. Because the aberrant activation of STAT3 and MAPKs have been associated with regulation of proliferation, invasion, and metastasis of tumors, we hypothesized that 6SG modulates the activation of STAT3 and MAPKs activation in tumor cells. We found that 6SG strongly inhibited constitutive phosphorylation of STAT3 through inhibition of the activation of upstream JAK2 and c-Src kinases and nuclear translocation of STAT3 on both MDA-MB231 and DU145 cells. Also, 6SG caused the activation of JNK, p38 MAPK, and ERK. Inhibition of ROS generation by N-acetylcysteine (NAC) significantly prevented 6SG-induced apoptosis. 6SG induced apoptosis as characterized by cleavage of PARP, accumulation of cells in subG1 phase, positive Annexin V binding, down-regulation of STAT3-regulated proteins, and activation of caspase-8, -9, -3 in both MDA-MB231 cells. Compared with other analogues of 6SG, such as 6-gingerol (6G), 8-gingerol (8G), and 10-gingerol (10G), 6SG was found to be the most potent blocker of STAT3 activation. We observed that the administration of 6SG alone significantly suppressed the growth of the tumor. As compared to the vehicle control, 6SG also suppressed the expression of STAT3-regulated gene products such as Bcl-2, Bcl-xL, and Survivin in tumor tissues. Overall, these findings suggest that 6SG can interfere with multiple signaling cascades involved in tumorigenesis and can be used as a potential therapeutic candidate for both the prevention and treatment of cancer. © 2014 Wiley Periodicals, Inc.

Increased skin barrier disruption by sodium lauryl sulfate in mice expressing a constitutively active STAT6 in T cells.

PubMed

DaSilva, Sonia C; Sahu, Ravi P; Konger, Raymond L; Perkins, Susan M; Kaplan, Mark H; Travers, Jeffrey B

2012-01-01

Atopic dermatitis (AD) is a pruritic, chronic inflammatory skin disease that affects 10-20% of children and 1-3% of adults worldwide. Recent studies have indicated that the ability of Th2 cytokines, such as interleukin-4 (IL-4) to regulate skin barrier function may be a predisposing factor for AD development. The present studies examined the ability of increased Th2 activity to affect cutaneous barrier function in vivo and epidermal thickening. Mice that express a constitutively active Signal Transducer and Activator of Transcription 6 (STAT6VT) have increased Th2 cells and a predisposition to allergic inflammation were used in these studies, they demonstrate that topical treatment with the irritant sodium lauryl sulfate (SLS) caused increased transepidermal water loss and epidermal thickening in STAT6VT mice over similarly treated wild-type mice. The proliferation marker Ki-67 was increased in the epidermis of STAT6VT compared to the wild-type mice. However, these differences do not appear to be linked to the addition of an irritant as control-treated STAT6VT skin also exhibited elevated Ki-67 levels, suggesting that the increased epidermal thickness in SLS-treated STAT6VT mice is primarily driven by epidermal cell hypertrophy rather than an increase in cellular proliferation. Our results suggest that an environment with increased Th2 cytokines results in abnormal responses to topical irritants.

Increased skin barrier disruption by sodium lauryl sulfate in mice expressing a constitutively active STAT6 in T cells

PubMed Central

DaSilva, Sonia C.; Sahu, Ravi P.; Konger, Raymond L.; Perkins, Susan M.; Kaplan, Mark H.; Travers, Jeffrey B.

2011-01-01

Atopic dermatitis (AD) is a pruritic, chronic inflammatory skin disease that affects 10–20% of children and 1–3% of adults worldwide. Recent studies have indicated that the ability of Th2 cytokines such as interleukin-4 (IL-4) to regulate skin barrier function may be a predisposing factor for AD development. The present studies examined the ability of increased Th2 activity to affect cutaneous barrier function in vivo and epidermal thickening. Mice that express a constitutively active Signal Transducer and Activator of Transcription 6 (STAT6VT) have increased Th2 cells and a predisposition to allergic inflammation were used in these studies; they demonstrate that topical treatment with the irritant sodium lauryl sulfate (SLS) caused increased transepidermal water loss and epidermal thickening in STAT6VT mice over similarly treated wild-type mice. The proliferation marker Ki-67 was increased in the epidermis of STAT6VT compared to wild-type mice. However, these differences do not appear to be linked to the addition of an irritant as control-treated STAT6VT skin also exhibited elevated Ki-67 levels, suggesting that the increased epidermal thickness in SLS-treated STAT6VT mice is primarily driven by epidermal cell hypertrophy rather than an increase in cellular proliferation. Our results suggest that an environment with increased Th2 cytokines results in abnormal responses to topical irritants. PMID:21959772

A Review: Phytochemicals Targeting JAK/STAT Signaling and IDO Expression in Cancer.

PubMed

Arumuggam, Niroshaathevi; Bhowmick, Neil A; Rupasinghe, H P Vasantha

2015-06-01

Cancer remains a major health problem worldwide. Among many other factors, two regulatory defects that are present in most cancer cells are constitutive activation of Janus kinase (JAK)/signal transducer and activator of transcription (STAT) signaling pathway and the induction of indoleamine 2, 3-dioxygenase (IDO), an enzyme that catalyzes tryptophan degradation, through JAK/STAT signaling. Cytokine signaling activates STAT proteins in regulating cell proliferation, differentiation, and survival through modulation of target genes. Many phytochemicals can inhibit both JAK/STAT signaling and IDO expression in antigen-presenting cells by targeting different pathways. Some of the promising phytochemicals that are discussed in this review include resveratrol, cucurbitacin, curcumin, (-)-epigallocatechin gallate, and others. It is now evident that phytochemicals play key roles in inhibition of tumor proliferation and development and provide novel means for therapeutic targeting of cancer. Copyright © 2015 John Wiley & Sons, Ltd.

Concerted suppression of STAT3 and GSK3β is involved in growth inhibition of non-small cell lung cancer by Xanthatin.

PubMed

Tao, Li; Fan, Fangtian; Liu, Yuping; Li, Weidong; Zhang, Lei; Ruan, Junshan; Shen, Cunsi; Sheng, Xiaobo; Zhu, Zhijie; Wang, Aiyun; Chen, Wenxing; Huang, Shile; Lu, Yin

2013-01-01

Xanthatin, a sesquiterpene lactone purified from Xanthium strumarium L., possesses prominent anticancer activity. We found that disruption of GSK3β activity was essential for xanthatin to exert its anticancer properties in non-small cell lung cancer (NSCLC), concurrent with preferable suppression of constitutive activation of STAT3. Interestingly, inactivation of the two signals are two mutually exclusive events in xanthatin-induced cell death. Moreover, we surprisingly found that exposure of xanthatin failed to trigger the presumable side effect of canonical Wnt/β-Catenin followed by GSK3β inactivation. We further observed that the downregulation of STAT3 was required for xanthatin to fine-tune the risk. Thus, the discovery of xanthatin, which has ability to simultaneously orchestrate two independent signaling cascades, may have important implications for screening promising drugs in cancer therapies.

Concerted Suppression of STAT3 and GSK3β Is Involved in Growth Inhibition of Non-Small Cell Lung Cancer by Xanthatin

PubMed Central

Tao, Li; Fan, Fangtian; Liu, Yuping; Li, Weidong; Zhang, Lei; Ruan, Junshan; Shen, Cunsi; Sheng, Xiaobo; Zhu, Zhijie; Wang, Aiyun; Chen, Wenxing; Huang, Shile; Lu, Yin

2013-01-01

Xanthatin, a sesquiterpene lactone purified from Xanthium strumarium L., possesses prominent anticancer activity. We found that disruption of GSK3β activity was essential for xanthatin to exert its anticancer properties in non-small cell lung cancer (NSCLC), concurrent with preferable suppression of constitutive activation of STAT3. Interestingly, inactivation of the two signals are two mutually exclusive events in xanthatin-induced cell death. Moreover, we surprisingly found that exposure of xanthatin failed to trigger the presumable side effect of canonical Wnt/β-Catenin followed by GSK3β inactivation. We further observed that the downregulation of STAT3 was required for xanthatin to fine-tune the risk. Thus, the discovery of xanthatin, which has ability to simultaneously orchestrate two independent signaling cascades, may have important implications for screening promising drugs in cancer therapies. PMID:24312384

Monocytes Induce STAT3 Activation in Human Mesenchymal Stem Cells to Promote Osteoblast Formation

PubMed Central

Nicolaidou, Vicky; Wong, Mei Mei; Redpath, Andia N.; Ersek, Adel; Baban, Dilair F.; Williams, Lynn M.; Cope, Andrew P.; Horwood, Nicole J.

2012-01-01

A major therapeutic challenge is how to replace bone once it is lost. Bone loss is a characteristic of chronic inflammatory and degenerative diseases such as rheumatoid arthritis and osteoporosis. Cells and cytokines of the immune system are known to regulate bone turnover by controlling the differentiation and activity of osteoclasts, the bone resorbing cells. However, less is known about the regulation of osteoblasts (OB), the bone forming cells. This study aimed to investigate whether immune cells also regulate OB differentiation. Using in vitro cell cultures of human bone marrow-derived mesenchymal stem cells (MSC), it was shown that monocytes/macrophages potently induced MSC differentiation into OBs. This was evident by increased alkaline phosphatase (ALP) after 7 days and the formation of mineralised bone nodules at 21 days. This monocyte-induced osteogenic effect was mediated by cell contact with MSCs leading to the production of soluble factor(s) by the monocytes. As a consequence of these interactions we observed a rapid activation of STAT3 in the MSCs. Gene profiling of STAT3 constitutively active (STAT3C) infected MSCs using Illumina whole human genome arrays showed that Runx2 and ALP were up-regulated whilst DKK1 was down-regulated in response to STAT3 signalling. STAT3C also led to the up-regulation of the oncostatin M (OSM) and LIF receptors. In the co-cultures, OSM that was produced by monocytes activated STAT3 in MSCs, and neutralising antibodies to OSM reduced ALP by 50%. These data indicate that OSM, in conjunction with other mediators, can drive MSC differentiation into OB. This study establishes a role for monocyte/macrophages as critical regulators of osteogenic differentiation via OSM production and the induction of STAT3 signalling in MSCs. Inducing the local activation of STAT3 in bone cells may be a valuable tool to increase bone formation in osteoporosis and arthritis, and in localised bone remodelling during fracture repair. PMID:22802946

Inhibition of STAT3 and ErbB2 Suppresses Tumor Growth, Enhances Radiosensitivity, and Induces Mitochondria-Dependent Apoptosis in Glioma Cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gao Ling; Li Fengsheng; Dong Bo

2010-07-15

Purpose: Constitutively activated signal transducer and activator of transcription 3 (STAT3) and ErbB2 are involved in the pathogenesis of many tumors, including astrocytoma. Inactivation of these molecules is reported to result in radiosensitization. The purpose of this study was to investigate whether inhibition of STAT3, ErbB2, or both could enhance radiotherapy in the human glioma model (U251 and U87 cell lines). Methods and Materials: The RNAi plasmids targeting STAT3 or ErbB2 were constructed, and their downregulatory effects on target proteins were examined by immunoblotting. After combination treatment of RNAi with or without irradiation, the cell viability was determined using 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazoliummore » bromide (MTT) and clonogenic assays. The in vivo effect of combined treatment was determined using the U251 xenograft model. The apoptosis caused by the inhibition of STAT3 and ErbB2 was detected, and the mechanism involved in the apoptosis was investigated, including increases in caspase proteins, mitochondrial damage, and the expression of key modulating protein of different apoptosis pathways. Results: Transfection of U251 cells with STAT3 or ErbB2 siRNA plasmids specifically reduced their target gene expressions. Inhibition of STAT3 or ErbB2 greatly decreased glioma cell survival after 2, 4, or 6 Gy irradiation. Inhibition of STAT3 and ErbB2 also enhanced radiation-induced tumor growth inhibition in the U251 xenograft model. Furthermore, the suppression of either STAT3 or ErbB2 could induce U251 cell apoptosis, which was related primarily to the mitochondrial apoptotic pathway. Conclusions: These results indicated that simultaneous inhibition of STAT3 and ErbB2 expression can promote potent antitumor activity and radiosensitizing activity in human glioma.« less

Anti-angiogenic and anti-metastatic activity of JAK inhibitor AZD1480

PubMed Central

Xin, Hong; Herrmann, Andreas; Reckamp, Karen; Zhang, Wang; Pal, Sumanta; Hedvat, Michael; Zhang, Chunyan; Liang, Wei; Scuto, Anna; Weng, Shaobu; Morosini, Deborah; Cao, Zhu A.; Zinda, Michael; Figlin, Robert; Huszar, Dennis; Jove, Richard; Yu, Hua

2011-01-01

STAT3 has important functions in both tumor cells and the tumor microenvironment to facilitate cancer progression. The STAT regulatory kinase JAK has been strongly implicated in promoting oncogenesis of various solid tumors, including through the use of JAK kinase inhibitors such as AZD1480. However, direct evidence that JAK drives STAT3 function and cancer pathogenesis at the level of the tumor microenvironment has yet to be established clearly. In this study, we show that AZD1480 inhibits STAT3 in tumor-associated myeloid cells, reducing their number and inhibiting tumor metastasis. Myeloid cell-mediated angiogenesis was also diminished by AZD1480, with additional direct inhibition of endothelial cell function in vitro and in vivo. AZD1480 blocked lung infiltration of myeloid cells and formation of pulmonary metastases in both mouse syngeneic experimental and spontaneous metastatic models. Furthermore, AZD1480 reduced angiogenesis and metastasis in a human xenograft tumor model. Although the effects of AZD1480 on the tumor microenvironment were important for the observed anti-angiogenic activity, constitutive activation of STAT3 in tumor cells themselves could block these anti-angiogenic effects demonstrating the complexity of the JAK/STAT signaling network in tumor progression. Together, our results indicated that AZD1480 can effectively inhibit tumor angiogenesis and metastasis mediated by STAT3 in stromal cells as well as tumor cells. PMID:21920898

Discovery of novel STAT3 small molecule inhibitors via in silico site-directed fragment-based drug design.

PubMed

Yu, Wenying; Xiao, Hui; Lin, Jiayuh; Li, Chenglong

2013-06-13

Constitutive activation of signal transducer and activator of transcription 3 (STAT3) has been validated as an attractive therapeutic target for cancer therapy. To stop both STAT3 activation and dimerization, a viable strategy is to design inhibitors blocking its SH2 domain phosphotyrosine binding site that is responsible for both actions. A new fragment-based drug design (FBDD) strategy, in silico site-directed FBDD, was applied in this study. A designed novel compound, 5,8-dioxo-6-(pyridin-3-ylamino)-5,8-dihydronaphthalene-1-sulfonamide (LY5), was confirmed to bind to STAT3 SH2 by fluorescence polarization assay. In addition, four out of the five chosen compounds have IC50 values lower than 5 μM for the U2OS cancer cells. 8 (LY5) has an IC50 range in 0.5-1.4 μM in various cancer cell lines. 8 also suppresses tumor growth in an in vivo mouse model. This study has demonstrated the utility of this approach and could be used to other drug targets in general.

The novel curcumin analog FLLL32 decreases STAT3 DNA binding activity and expression, and induces apoptosis in osteosarcoma cell lines

PubMed Central

2011-01-01

Background Curcumin is a naturally occurring phenolic compound shown to have a wide variety of antitumor activities; however, it does not attain sufficient blood levels to do so when ingested. Using structure-based design, a novel compound, FLLL32, was generated from curcumin. FLLL32 possesses superior biochemical properties and more specifically targets STAT3, a transcription factor important in tumor cell survival, proliferation, metastasis, and chemotherapy resistance. In our previous work, we found that several canine and human osteosarcoma (OSA) cell lines, but not normal osteoblasts, exhibit constitutive phosphorylation of STAT3. Compared to curcumin, we hypothesized that FLLL32 would be more efficient at inhibiting STAT3 function in OSA cells and that this would result in enhanced downregulation of STAT3 transcriptional targets and subsequent death of OSA cells. Methods Human and canine OSA cells were treated with vehicle, curcumin, or FLLL32 and the effects on proliferation (CyQUANT®), apoptosis (SensoLyte® Homogeneous AMC Caspase- 3/7 Assay kit, western blotting), STAT3 DNA binding (EMSA), and vascular endothelial growth factor (VEGF), survivin, and matrix metalloproteinase-2 (MMP2) expression (RT-PCR, western blotting) were measured. STAT3 expression was measured by RT-PCR, qRT- PCR, and western blotting. Results Our data showed that FLLL32 decreased STAT3 DNA binding by EMSA. FLLL32 promoted loss of cell proliferation at lower concentrations than curcumin leading to caspase-3- dependent apoptosis, as evidenced by PARP cleavage and increased caspase 3/7 activity; this could be inhibited by treatment with the pan-caspase inhibitor Z-VAD-FMK. Treatment of OSA cells with FLLL32 decreased expression of survivin, VEGF, and MMP2 at both mRNA and protein levels with concurrent decreases in phosphorylated and total STAT3; this loss of total STAT3 occurred, in part, via the ubiquitin-proteasome pathway. Conclusions These data demonstrate that the novel curcumin analog FLLL32 has biologic activity against OSA cell lines through inhibition of STAT3 function and expression. Future work with FLLL32 will define the therapeutic potential of this compound in vivo. PMID:21443800

Regulation of Stat5 by FAK and PAK1 in Oncogenic FLT3 and KIT driven Leukemogenesis

PubMed Central

Chatterjee, Anindya; Ghosh, Joydeep; Ramdas, Baskar; Mali, Raghuveer Singh; Martin, Holly; Kobayashi, Michihiro; Vemula, Sasidhar; Canela, Victor H.; Waskow, Emily R.; Visconte, Valeria; Tiu, Ramon V.; Smith, Catherine C.; Shah, Neil; Bunting, Kevin D.; Boswell, H. Scott; Liu, Yan; Chan, Rebecca J.; Kapur, Reuben

2015-01-01

SUMMARY Oncogenic mutations of FLT3 and KIT receptors are associated with poor survival in patients with acute myeloid leukemia (AML) and myeloproliferative neoplasms (MPN) and currently available drugs are largely ineffective. Although Stat5 has been implicated in regulating several myeloid and lymphoid malignancies, how precisely Stat5 regulates leukemogenesis, including its nuclear translocation to induce gene transcription is poorly understood. In leukemic cells, we show constitutive activation of focal adhesion kinase (FAK), whose inhibition represses leukemogenesis. Downstream of FAK, activation of Rac1 is regulated by RacGEF Tiam1, whose inhibition prolongs the survival of leukemic mice. Inhibition of the Rac1 effector PAK1 prolongs the survival of leukemic mice in part by inhibiting the nuclear translocation of Stat5. These results reveal a leukemic pathway involving FAK/Tiam1/Rac1/PAK1 and demonstrate an essential role for these signaling molecules in regulating the nuclear translocation of Stat5 in leukemogenesis. PMID:25456130

STAT3-mediated SMAD3 activation underlies Oncostatin M-induced Senescence

PubMed Central

Junk, Damian J.; Cipriano, Rocky; Jackson, Mark W.

2017-01-01

ABSTRACT Cytokines in the developing tumor microenvironment (TME) can drive transformation and subsequent progression toward metastasis. Elevated levels of the Interleukin-6 (IL-6) family cytokine Oncostatin M (OSM) in the breast TME correlate with aggressive, metastatic cancers, increased tumor recurrence, and poor patient prognosis. Paradoxically, OSM engages a tumor-suppressive, Signal Transducer and Activator of Transcription 3 (STAT3)-dependent senescence response in normal and non-transformed human mammary epithelial cells (HMEC). Here, we identify a novel link between OSM-activated STAT3 signaling and the Transforming Growth Factor-β (TGF-β) signaling pathway that engages senescence in HMEC. Inhibition of functional TGF-β/SMAD signaling by expressing a dominant-negative TGF-β receptor, treating with a TGF-β receptor inhibitor, or suppressing SMAD3 expression using a SMAD3-shRNA prevented OSM-induced senescence. OSM promoted a protein complex involving activated-STAT3 and SMAD3, induced the nuclear localization of SMAD3, and enhanced SMAD3-mediated transcription responsible for senescence. In contrast, expression of MYC (c-MYC) from a constitutive promoter abrogated senescence and strikingly, cooperated with OSM to promote a transformed phenotype, epithelial-mesenchymal transition (EMT), and invasiveness. Our findings suggest that a novel STAT3/SMAD3-signaling axis is required for OSM-mediated senescence that is coopted during the transformation process to confer aggressive cancer cell properties. Understanding how developing cancer cells bypass OSM/STAT3/SMAD3-mediated senescence may help identify novel targets for future “pro-senescence” therapies aiming to reengage this hidden tumor-suppressive response. PMID:27892764

Functional graphene oxide as a plasmid-based Stat3 siRNA carrier inhibits mouse malignant melanoma growth in vivo

NASA Astrophysics Data System (ADS)

Yin, Di; Li, Yang; Lin, Hang; Guo, Baofeng; Du, Yanwei; Li, Xin; Jia, Huijie; Zhao, Xuejian; Tang, Jun; Zhang, Ling

2013-03-01

Graphene oxide (GO) has attracted intensive interest in the biomedical field in recent years. We investigate whether the use of functional graphene oxide as an efficient delivery system for delivering specific molecular antitumor therapeutics in vivo could achieve a more excellent antitumor effect. Constitutive activation of signal transducer and activator of transcription 3 (Stat3) promotes survival in a wide spectrum of human cancers. In this paper, we study the in vivo behavior of graphene oxide chemically functionalized with polyethylenimine and polyethylene glycol (GO-PEI-PEG) as a plasmid-based Stat3-specific small interfering RNA (siRNA) carrier in mouse malignant melanoma. The in vivo results indicate significant regression in tumor growth and tumor weight after plasmid-based Stat3 siRNA delivered by GO-PEI-PEG treatment. Moreover, there was no significant side effect from GO-PEI-PEG treatment according to histological examination and blood chemistry analysis in mice. Thus, our work is the first success of using GO-PEI-PEG as a promising carrier for plasmid Stat3 siRNA delivery and down-regulation of Stat3 by a polymer-mediated vehicle and suggests the great promise of graphene in biomedical applications such as cancer treatment.

Uterine Deletion of Gp130 or Stat3 Shows Implantation Failure with Increased Estrogenic Responses

PubMed Central

Sun, Xiaofei; Bartos, Amanda; Whitsett, Jeffrey A.

2013-01-01

Leukemia inhibitory factor (LIF), a downstream target of estrogen, is essential for implantation in mice. LIF function is thought to be mediated by its binding to LIF receptor (LIFR) and recruitment of coreceptor GP130 (glycoprotein 130), and this receptor complex then activates signal transducer and activator of transcription (STAT)1/3. However, the importance of LIFR and GP130 acting via STAT3 in implantation remains uncertain, because constitutive inactivation of Lifr, Gp130, or Stat3 shows embryonic lethality in mice. To address this issue, we generated mice with conditional deletion of uterine Gp130 or Stat3 and show that both GP130 and STAT3 are critical for uterine receptivity and implantation. Implantation failure in these deleted mice is associated with higher uterine estrogenic responses prior to the time of implantation. These heightened estrogenic responses are not due to changes in ovarian hormone levels or expression of their nuclear receptors. In the deleted mice, estrogen-responsive gene, Lactoferrin (Ltf), and Mucin 1 protein, were up-regulated in the uterus. In addition, progesterone-responsive genes, Hoxa10 and Indian hedgehog (Ihh), were markedly down-regulated in STAT3-inactivated uteri. These changes in uteri of deleted mice were reflected by the failure of differentiation of the luminal epithelium, which is essential for blastocyst attachment. PMID:23885093

Alantolactone suppresses APOC3 expression and alters lipid homeostasis in L02 liver cells.

PubMed

Yang, Meiting; Zhao, Hanhan; Ai, Huihan; Zhu, Hongbin; Wang, Shuyue; Bao, Yongli; Li, Yuxin

2018-06-05

A high level of APOC3 expression is an independent risk factor for some lipid metabolism-related diseases, such as cardiovascular disease (CVD), nonalcoholic fatty liver disease (NAFLD) and atherosclerosis (AS). This suggests that down-regulating APOC3 expression is a potential way of regulating lipid levels. In this study, we used luciferase reporter screening to identify a natural compound, alantolactone (ALA), that can inhibit the promoter activity of APOC3. ALA decreased APOC3 expression at both mRNA and protein levels. Then we pretreated L02 liver cells with oxLDL to investigate the function of ALA in lipid homeostasis. Intriguingly, ALA attenuated oxLDL-induced foam cell formation by reducing total cholesterol (TC) and triglyceride (TG) contents. Furthermore, these results could be reversed by overexpressing APOC3 protein. ALA inhibited tyrosine phosphorylation (Tyr705pho) of STAT3 to down-regulate APOC3 expression. Intriguingly, overexpression of a wild-type STAT3 or a constitutively active form of STAT3 (STAT3-C) up-regulated APOC3 expression and partly reversed the effect of ALA in oxLDL-induced L02 cells. Overexpression of wild-type STAT3 also increased TC but not TG contents in L02 cells. However, overexpression of STAT3-C significantly increased TC and TG contents, and the effect of ALA was partly attenuated by STAT3-C, although this was not statistically significant. These results suggest that ALA attenuates lipid accumulation through down-regulation of APOC3 expression, at least in part by inhibiting STAT3 signaling. Copyright © 2018 Elsevier B.V. All rights reserved.

STAT3 in Cancer—Friend or Foe?

PubMed Central

Zhang, Hai-Feng; Lai, Raymond

2014-01-01

The roles and significance of STAT3 in cancer biology have been extensively studied for more than a decade. Mounting evidence has shown that constitutive activation of STAT3 is a frequent biochemical aberrancy in cancer cells, and this abnormality directly contributes to tumorigenesis and shapes many malignant phenotypes in cancer cells. Nevertheless, results from more recent experimental and clinicopathologic studies have suggested that STAT3 also can exert tumor suppressor effects under specific conditions. Importantly, some of these studies have demonstrated that STAT3 can function either as an oncoprotein or a tumor suppressor in the same cell type, depending on the specific genetic background or presence/absence of specific coexisting biochemical defects. Thus, in the context of cancer biology, STAT3 can be a friend or foe. In the first half of this review, we will highlight the “evil” features of STAT3 by summarizing its oncogenic functions and mechanisms. The differences between the canonical and non-canonical pathway will be highlighted. In the second half, we will summarize the evidence supporting that STAT3 can function as a tumor suppressor. To explain how STAT3 may mediate its tumor suppressor effects, we will discuss several possible mechanisms, one of which is linked to the role of STAT3β, one of the two STAT3 splicing isoforms. Taken together, it is clear that the roles of STAT3 in cancer are multi-faceted and far more complicated than one appreciated previously. The new knowledge has provided us with new approaches and strategies when we evaluate STAT3 as a prognostic biomarker or therapeutic target. PMID:24995504

Clinicopathological significance and prognostic role of p-STAT3 in patients with hepatocellular carcinoma.

PubMed

Liang, Chaojie; Xu, Yingchen; Ge, Hua; Li, Guangming; Wu, Jixiang

2018-01-01

Constitutive activation of STAT3 through its phosphorylation (p-STAT3) plays a key role in the development and progression of various cancers. However, the relationship between p-STAT3 expression and the clinicopathological features and prognostic value in patients with hepatocellular carcinoma (HCC) remains controversial. We conducted a meta-analysis to evaluate the role of p-STAT3 in HCC. The PubMed, Cochrane Library, Web of Science, EMBASE, Chinese CNKI, and Chinese Wanfang databases were searched using the appropriate terms to find the relevant studies on p-STAT3 and HCC. The relationship between p-STAT3 expression and clinicopathological characteristics and prognostic value was established. Pool odds ratios (ORs) and hazard ratios (HRs) with 95% CIs were calculated using the STATA 14.2 software. The eight articles included in this meta-analysis comprised 752 patients. Expression of p-STAT3 was associated with incidence, age, liver cirrhosis, tumor size, vascular invasion, and TNM stage of HCC, but it was not related to gender, alpha-fetoprotein (AFP), hepatitis B surface antigen (HBsAg), number of tumors, and tumor differentiation. Additionally, the expression of p-STAT3 was related to a poor 3- and 5-year overall survival rate and disease-free survival rate. Expression of p-STAT3 was associated with the incidence, age, liver cirrhosis, tumor size, vascular invasion, and TNM stage. Thus, p-STAT3 can be a reliable prognostic biomarker for HCC. Further high-quality studies with larger numbers of patients are needed.

Characterization of STAT5B phosphorylation correlating with expression of cytokine-inducible SH2-containing protein (CIS).

PubMed

Cooper, John C; Boustead, Jared N; Yu, Chao-Lan

2006-06-01

Cytokine-inducible SH2-containing protein (CIS) is the first identified member of genes encoding for the suppressor of cytokine signaling (SOCS). CIS is also a well-known target gene of signal transducer and activator of transcription 5 (STAT5) pathways, providing normal negative feedback control of signaling by cytokines and growth factors. Three other SOCS genes, SOCS1, SOCS2, and SOCS3, can be silenced by DNA hypermethylation in human cancers, suggesting a potential mechanism for constitutive STAT activation. However, it is not known whether CIS expression is similarly perturbed in tumor cells. We report here the absence of CIS expression in T lymphoma LSTRA that overexpresses the Lck protein tyrosine kinase and exhibits elevated STAT5 activity. Pervanadate-induced CIS expression and STAT5 binding to the CIS promoter in vivo over a short time course implies that mechanisms other than DNA hypermethylation may contribute to defective CIS expression in LSTRA cells. Comparison with cytokine-dependent BaF3 cells stimulated with interleukin-3 (IL-3) further reveals that CIS induction correlates with specific STAT5b post-translational modifications. It exhibits as the slowest migrating form through SDS-polyacrylamide gel electrophoresis (SDS-PAGE) analysis. This distinctly modified STAT5b is the predominant form that binds to the consensus STAT5 sites in the CIS promoter and accumulates in the nucleus. In vitro phosphatase assays and phosphoamino acid analysis suggest the involvement of phosphorylation on residues other than the highly conserved tyrosine and serine sites in this distinct STAT5b mobility shift. All together, our results provide a novel link between incomplete STAT5b phosphorylation and defective SOCS gene expression in cancer cells.

Nuclear localization of pyruvate dehydrogenase complex-E2 (PDC-E2), a mitochondrial enzyme, and its role in signal transducer and activator of transcription 5 (STAT5)-dependent gene transcription.

PubMed

Chueh, Fu-Yu; Leong, King-Fu; Cronk, Robert J; Venkitachalam, Srividya; Pabich, Samantha; Yu, Chao-Lan

2011-07-01

STAT (signal transducer and activator of transcription) proteins play a critical role in cellular response to a wide variety of cytokines and growth factors by regulating specific nuclear genes. STAT-dependent gene transcription can be finely tuned through the association with co-factors in the nucleus. We showed previously that STAT5 (including 5a and 5b) specifically interacts with a mitochondrial enzyme PDC-E2 (E2 subunit of pyruvate dehydrogenase complex) in both leukemic T cells and cytokine-stimulated cells. However, the functional significance of this novel association remains largely unknown. Here we report that PDC-E2 may function as a co-activator in STAT5-dependent nuclear gene expression. Subcellular fractionation analysis revealed that a substantial amount of PDC-E2 was constitutively present in the nucleus of BaF3, an interleukin-3 (IL-3)-dependent cell line. IL-3-induced tyrosine-phosphorylated STAT5 associated with nuclear PDC-E2 in co-immunoprecipitation analysis. These findings were confirmed by confocal immunofluorescence microscopy showing constant nuclear localization of PDC-E2 and its co-localization with STAT5 after IL-3 stimulation. Similar to mitochondrial PDC-E2, nuclear PDC-E2 was lipoylated and associated with PDC-E1. Overexpression of PDC-E2 in BaF3 cells augmented IL-3-induced STAT5 activity as measured by reporter assay with consensus STAT5-binding sites. Consistent with the reporter data, PDC-E2 overexpression in BaF3 cells led to elevated mRNA levels of endogenous SOCS3 (suppressor of cytokine signaling 3) gene, a known STAT5 target. We further identified two functional STAT5-binding sites in the SOCS3 gene promoter important for its IL-3-inducibility. The observation that both cis-acting elements were essential to detect the stimulatory effect by PDC-E2 strongly supports the role of PDC-E2 in up-regulating the transactivating ability of STAT5. All together, our results reveal a novel function of PDC-E2 in the nucleus. It also raises the possibility of nuclear-mitochondrial crosstalk through the interaction between STAT5 and PDC-E2. Copyright © 2011 Elsevier Inc. All rights reserved.

Nuclear localization of pyruvate dehydrogenase complex-E2 (PDC-E2), a mitochondrial enzyme, and its role in signal transducer and activator of transcription 5 (STAT5)-dependent gene transcription

PubMed Central

Chueh, Fu-Yu; Leong, King-Fu; Cronk, Robert J.; Venkitachalam, Srividya; Pabich, Samantha; Yu, Chao-Lan

2011-01-01

STAT (signal transducer and activator of transcription) proteins play a critical role in cellular response to a wide variety of cytokines and growth factors by regulating specific nuclear genes. STAT-dependent gene transcription can be finely tuned through the association with cofactors in the nucleus. We showed previously that STAT5 (including 5a and 5b) specifically interacts with a mitochondrial enzyme PDC-E2 (E2 subunit of pyruvate dehydrogenase complex) in both leukemic T cells and cytokine-stimulated cells. However, the functional significance of this novel association remains largely unknown. Here we report that PDC-E2 may function as a co-activator in STAT5-dependent nuclear gene expression. Subcellular fractionation analysis revealed that a substantial amount of PDC-E2 was constitutively present in the nucleus of BaF3, an interleukin-3 (IL-3)-dependent cell line. IL-3-induced tyrosine-phosphorylated STAT5 associated with nuclear PDC-E2 in co-immunoprecipitation analysis. These findings were confirmed by confocal immunofluorescence microscopy showing constant nuclear localization of PDC-E2 and its co-localization with STAT5 after IL-3 stimulation. Similar to mitochondrial PDC-E2, nuclear PDC-E2 was lipoylated and associated with PDC-E1. Overexpression of PDC-E2 in BaF3 cells augmented IL-3-induced STAT5 activity as measured by reporter assay with consensus STAT5-binding sites. Consistent with the reporter data, PDC-E2 overexpression in BaF3 cells led to elevated mRNA levels of endogenous SOCS3 (suppressor of cytokine signaling 3) gene, a known STAT5 target. We further identified two functional STAT5-binding sites in the SOCS3 gene promoter important for its IL-3-inducibility. The observation that both cis-acting elements were essential to detect the stimulatory effect by PDC-E2 strongly supports the role of PDC-E2 in up-regulating the transactivating ability of STAT5. All together, our results reveal a novel function of PDC-E2 in the nucleus. It also raises the possibility of nuclear-mitochondrial crosstalk through the interaction between STAT5 and PDC-E2. PMID:21397011

Enhanced phosphorylation of STAT1 is dependent on PKR signaling in HLA-B27 expressing U937 monocytic cells

PubMed Central

Ruuska, Marja; Sahlberg, Anna S.; Colbert, Robert A.; Granfors, Kaisa; Penttinen, Markus A.

2011-01-01

Objective To study the phosphorylation of STAT1 in HLA-B27-transfected human monocytic cells and the role of signaling molecules PKR and p38 in STAT1 phosphorylation. Methods U937 human monocytic cell transfectants stably expressing wild type HLA-B27 or mutated HLA-B27 heavy chains (HC) with amino acid substitutions in the B pocket were prepared. Mock transfected cells were prepared using the antibiotic resistance vectors (pSV2neo or RSV5neo) alone. PMA differentiated cells were stimulated with LPS or infected with S. enteritidis. Western blotting and flow cytometry were used to detect the phosphorylation and expression levels of STAT1 protein. Specific inhibitors were added in cell culture to study the role of PKR and p38 on STAT1 phosphorylation. Results STAT1 is constitutively highly phosphorylated on tyrosine 701 residue in HLA-B27 positive monocytic cells when compared to control cells, even prior to stimulation with LPS or bacteria. This phenotype is associated with the expression of HLA-B27 HCs that misfold. In addition, phosphorylation of STAT1 is dependent on PKR. Conclusion Our results show that STAT1 tyrosine 701 is constitutively highly phosphorylated in HLA-B27 expressing monocyte-macrophage cell line. Since phosphorylation of tyrosine 701 on STAT1 is sufficient to induce interferon-dependent genes, constitutive activity of this phosphorylation site may lead to overexpression of interferon-dependent genes, as well as other STAT1-dependent genes, in HLA-B27 monocyte-macrophages. Our results offer a mechanism by which B27 expression alone, without any external trigger, is potentially capable of inducing activation of STAT1, a critical regulator of the inflammatory response. PMID:21968657

MPLW515L Is a Novel Somatic Activating Mutation in Myelofibrosis with Myeloid Metaplasia

PubMed Central

Pikman, Yana; Lee, Benjamin H; Mercher, Thomas; McDowell, Elizabeth; Ebert, Benjamin L; Gozo, Maricel; Cuker, Adam; Wernig, Gerlinde; Moore, Sandra; Galinsky, Ilene; DeAngelo, Daniel J; Clark, Jennifer J; Lee, Stephanie J; Golub, Todd R; Wadleigh, Martha; Gilliland, D. Gary; Levine, Ross L

2006-01-01

Background The JAK2V617F allele has recently been identified in patients with polycythemia vera (PV), essential thrombocytosis (ET), and myelofibrosis with myeloid metaplasia (MF). Subsequent analysis has shown that constitutive activation of the JAK-STAT signal transduction pathway is an important pathogenetic event in these patients, and that enzymatic inhibition of JAK2V617F may be of therapeutic benefit in this context. However, a significant proportion of patients with ET or MF are JAK2V617F-negative. We hypothesized that activation of the JAK-STAT pathway might also occur as a consequence of activating mutations in certain hematopoietic-specific cytokine receptors, including the erythropoietin receptor (EPOR), the thrombopoietin receptor (MPL), or the granulocyte-colony stimulating factor receptor (GCSFR). Methods and Findings DNA sequence analysis of the exons encoding the transmembrane and juxtamembrane domains of EPOR, MPL, and GCSFR, and comparison with germline DNA derived from buccal swabs, identified a somatic activating mutation in the transmembrane domain of MPL (W515L) in 9% (4/45) of JAKV617F-negative MF. Expression of MPLW515L in 32D, UT7, or Ba/F3 cells conferred cytokine-independent growth and thrombopoietin hypersensitivity, and resulted in constitutive phosphorylation of JAK2, STAT3, STAT5, AKT, and ERK. Furthermore, a small molecule JAK kinase inhibitor inhibited MPLW515L-mediated proliferation and JAK-STAT signaling in vitro. In a murine bone marrow transplant assay, expression of MPLW515L, but not wild-type MPL, resulted in a fully penetrant myeloproliferative disorder characterized by marked thrombocytosis (Plt count 1.9–4.0 × 10 12/L), marked splenomegaly due to extramedullary hematopoiesis, and increased reticulin fibrosis. Conclusions Activation of JAK-STAT signaling via MPLW515L is an important pathogenetic event in patients with JAK2V617F-negative MF. The bone marrow transplant model of MPLW515L-mediated myeloproliferative disorders (MPD) exhibits certain features of human MF, including extramedullary hematopoiesis, splenomegaly, and megakaryocytic proliferation. Further analysis of positive and negative regulators of the JAK-STAT pathway is warranted in JAK2V617F-negative MPD. PMID:16834459

MPLW515L is a novel somatic activating mutation in myelofibrosis with myeloid metaplasia.

PubMed

Pikman, Yana; Lee, Benjamin H; Mercher, Thomas; McDowell, Elizabeth; Ebert, Benjamin L; Gozo, Maricel; Cuker, Adam; Wernig, Gerlinde; Moore, Sandra; Galinsky, Ilene; DeAngelo, Daniel J; Clark, Jennifer J; Lee, Stephanie J; Golub, Todd R; Wadleigh, Martha; Gilliland, D Gary; Levine, Ross L

2006-07-01

The JAK2V617F allele has recently been identified in patients with polycythemia vera (PV), essential thrombocytosis (ET), and myelofibrosis with myeloid metaplasia (MF). Subsequent analysis has shown that constitutive activation of the JAK-STAT signal transduction pathway is an important pathogenetic event in these patients, and that enzymatic inhibition of JAK2V617F may be of therapeutic benefit in this context. However, a significant proportion of patients with ET or MF are JAK2V617F-negative. We hypothesized that activation of the JAK-STAT pathway might also occur as a consequence of activating mutations in certain hematopoietic-specific cytokine receptors, including the erythropoietin receptor (EPOR), the thrombopoietin receptor (MPL), or the granulocyte-colony stimulating factor receptor (GCSFR). DNA sequence analysis of the exons encoding the transmembrane and juxtamembrane domains of EPOR, MPL, and GCSFR, and comparison with germline DNA derived from buccal swabs, identified a somatic activating mutation in the transmembrane domain of MPL (W515L) in 9% (4/45) of JAKV617F-negative MF. Expression of MPLW515L in 32D, UT7, or Ba/F3 cells conferred cytokine-independent growth and thrombopoietin hypersensitivity, and resulted in constitutive phosphorylation of JAK2, STAT3, STAT5, AKT, and ERK. Furthermore, a small molecule JAK kinase inhibitor inhibited MPLW515L-mediated proliferation and JAK-STAT signaling in vitro. In a murine bone marrow transplant assay, expression of MPLW515L, but not wild-type MPL, resulted in a fully penetrant myeloproliferative disorder characterized by marked thrombocytosis (Plt count 1.9-4.0 x 10(12)/L), marked splenomegaly due to extramedullary hematopoiesis, and increased reticulin fibrosis. Activation of JAK-STAT signaling via MPLW515L is an important pathogenetic event in patients with JAK2V617F-negative MF. The bone marrow transplant model of MPLW515L-mediated myeloproliferative disorders (MPD) exhibits certain features of human MF, including extramedullary hematopoiesis, splenomegaly, and megakaryocytic proliferation. Further analysis of positive and negative regulators of the JAK-STAT pathway is warranted in JAK2V617F-negative MPD.

6-Shogaol from dried ginger inhibits growth of prostate cancer cells both in vitro and in vivo through inhibition of STAT3 and NF-κB signaling.

PubMed

Saha, Achinto; Blando, Jorge; Silver, Eric; Beltran, Linda; Sessler, Jonathan; DiGiovanni, John

2014-06-01

Despite much recent progress, prostate cancer continues to represent a major cause of cancer-related mortality and morbidity in men. Prostate cancer is the most common nonskin neoplasm and second leading cause of death in men. 6-Shogaol (6-SHO), a potent bioactive compound in ginger (Zingiber officinale Roscoe), has been shown to possess anti-inflammatory and anticancer activity. In the present study, the effect of 6-SHO on the growth of prostate cancer cells was investigated. 6-SHO effectively reduced survival and induced apoptosis of cultured human (LNCaP, DU145, and PC3) and mouse (HMVP2) prostate cancer cells. Mechanistic studies revealed that 6-SHO reduced constitutive and interleukin (IL)-6-induced STAT3 activation and inhibited both constitutive and TNF-α-induced NF-κB activity in these cells. In addition, 6-SHO decreased the level of several STAT3 and NF-κB-regulated target genes at the protein level, including cyclin D1, survivin, and cMyc and modulated mRNA levels of chemokine, cytokine, cell cycle, and apoptosis regulatory genes (IL-7, CCL5, BAX, BCL2, p21, and p27). 6-SHO was more effective than two other compounds found in ginger, 6-gingerol, and 6-paradol at reducing survival of prostate cancer cells and reducing STAT3 and NF-κB signaling. 6-SHO also showed significant tumor growth inhibitory activity in an allograft model using HMVP2 cells. Overall, the current results suggest that 6-SHO may have potential as a chemopreventive and/or therapeutic agent for prostate cancer and that further study of this compound is warranted. ©2014 American Association for Cancer Research.

The spectrum of STAT functions in mammary gland development

PubMed Central

Hughes, Katherine; Watson, Christine J.

2012-01-01

The signal transducer and activator of transcription (STAT) family of transcription factors have a spectrum of functions in mammary gland development. In some cases these roles parallel those of STATs in other organ systems, while in other instances the function of individual STATs in the mammary gland is specific to this tissue. In the immune system, STAT6 is associated with differentiation of T helper cells, while in the mammary gland, it has a fundamental role in the commitment of luminal epithelial cells to the alveolar lineage. STAT5A is required for the production of luminal progenitor cells from mammary stem cells and is essential for the differentiation of milk producing alveolar cells during pregnancy. By contrast, the initiation of regression following weaning heralds a dramatic and specific activation of STAT3, reflecting its pivotal role in the regulation of cell death and tissue remodeling during mammary involution. Although it has been demonstrated that STAT1 is regulated during a mammary developmental cycle, it is not yet determined whether it has a specific, non-redundant function. Thus, the mammary gland constitutes an unusual example of an adult organ in which different STATs are sequentially activated to orchestrate the processes of functional differentiation, cell death and tissue remodeling. PMID:24058764

Regulation of Stat5 by FAK and PAK1 in Oncogenic FLT3- and KIT-Driven Leukemogenesis.

PubMed

Chatterjee, Anindya; Ghosh, Joydeep; Ramdas, Baskar; Mali, Raghuveer Singh; Martin, Holly; Kobayashi, Michihiro; Vemula, Sasidhar; Canela, Victor H; Waskow, Emily R; Visconte, Valeria; Tiu, Ramon V; Smith, Catherine C; Shah, Neil; Bunting, Kevin D; Boswell, H Scott; Liu, Yan; Chan, Rebecca J; Kapur, Reuben

2014-11-20

Oncogenic mutations of FLT3 and KIT receptors are associated with poor survival in patients with acute myeloid leukemia (AML) and myeloproliferative neoplasms (MPNs), and currently available drugs are largely ineffective. Although Stat5 has been implicated in regulating several myeloid and lymphoid malignancies, how precisely Stat5 regulates leukemogenesis, including its nuclear translocation to induce gene transcription, is poorly understood. In leukemic cells, we show constitutive activation of focal adhesion kinase (FAK) whose inhibition represses leukemogenesis. Downstream of FAK, activation of Rac1 is regulated by RacGEF Tiam1, whose inhibition prolongs the survival of leukemic mice. Inhibition of the Rac1 effector PAK1 prolongs the survival of leukemic mice in part by inhibiting the nuclear translocation of Stat5. These results reveal a leukemic pathway involving FAK/Tiam1/Rac1/PAK1 and demonstrate an essential role for these signaling molecules in regulating the nuclear translocation of Stat5 in leukemogenesis. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

Cytokine-Induction of Tumor Necrosis Factor Receptor 2 (TNFR2) is Mediated by STAT3 in Colon Cancer Cells

PubMed Central

Hamilton, Kathryn E.; Simmons, James G.; Ding, Shengli; Van Landeghem, Laurianne; Lund, P. Kay

2011-01-01

The IL-6/STAT3 and TNFα/NFκB pathways are emerging as critical mediators of inflammation-associated colon cancer. TNFR2 expression is increased in inflammatory bowel diseases, the azoxymethane/dextran sodium sulfate (AOM/DSS) model of colitis-associated cancer, and by combined IL-6 and TNFα. The molecular mechanisms that regulate TNFR2 remain undefined. This study used colon cancer cell lines to test the hypothesis that IL-6 and TNFα induce TNFR2 via STAT3 and/or NFκB. Basal and IL-6 + TNFα-induced TNFR2 were decreased by pharmacological STAT3 inhibition. NFκB inhibition had little effect on IL-6 + TNFα-induced TNFR2, but did inhibit induction of endogenous IL-6 and TNFR2 in cells treated with TNFα alone. Chromatin immunoprecipitation (ChIP) revealed cooperative effects of IL-6 + TNFα to induce STAT3 binding to a -1578 STAT response element in the TNFR2 promoter, but no effect on NFκB binding to consensus sites. Constitutively active STAT3 was sufficient to induce TNFR2 expression. Over-expression of SOCS3, a cytokine-inducible STAT3 inhibitor, which reduces tumorigenesis in preclinical models of colitis-associated cancer, decreased cytokine-induced TNFR2 expression and STAT3 binding to the -1578 STAT response element. SOCS3 over-expression also decreased proliferation of colon cancer cells and dramatically decreased anchorage-independent growth of colon cancer cells, even cells over-expressing TNFR2. Collectively, these studies demonstrate that IL-6 and TNFα-induced TNFR2 expression in colon cancer cells is mediated primarily by STAT3, and provide evidence that TNFR2 may contribute to the tumor-promoting roles of STAT3. PMID:21994466

Multifaceted Intervention by the Hsp90 Inhibitor Ganetespib (STA-9090) in Cancer Cells with Activated JAK/STAT Signaling

PubMed Central

Proia, David A.; Foley, Kevin P.; Korbut, Tim; Sang, Jim; Smith, Don; Bates, Richard C.; Liu, Yuan; Rosenberg, Alex F.; Zhou, Dan; Koya, Keizo; Barsoum, James; Blackman, Ronald K.

2011-01-01

There is accumulating evidence that dysregulated JAK signaling occurs in a wide variety of cancer types. In particular, mutations in JAK2 can result in the constitutive activation of STAT transcription factors and lead to oncogenic growth. JAK kinases are established Hsp90 client proteins and here we show that the novel small molecule Hsp90 inhibitor ganetespib (formerly STA-9090) exhibits potent in vitro and in vivo activity in a range of solid and hematological tumor cells that are dependent on JAK2 activity for growth and survival. Of note, ganetespib treatment results in sustained depletion of JAK2, including the constitutively active JAK2V617F mutant, with subsequent loss of STAT activity and reduced STAT-target gene expression. In contrast, treatment with the pan-JAK inhibitor P6 results in only transient effects on these processes. Further differentiating these modes of intervention, RNA and protein expression studies show that ganetespib additionally modulates cell cycle regulatory proteins, while P6 does not. The concomitant impact of ganetespib on both cell growth and cell division signaling translates to potent antitumor efficacy in mouse models of xenografts and disseminated JAK/STAT-driven leukemia. Overall, our findings support Hsp90 inhibition as a novel therapeutic approach for combating diseases dependent on JAK/STAT signaling, with the multimodal action of ganetespib demonstrating advantages over JAK-specific inhibitors. PMID:21533169

Controlling nuclear JAKs and STATs for specific gene activation by IFNγ.

PubMed

Noon-Song, Ezra N; Ahmed, Chulbul M; Dabelic, Rea; Canton, Johnathan; Johnson, Howard M

2011-07-08

We previously showed that gamma interferon (IFNγ) and its receptor subunit, IFNGR1, interacted with the promoter region of IFNγ-activated genes along with transcription factor STAT1α. Recent studies have suggested that activated Janus kinases pJAK2 and pJAK1 also played a role in gene activation by phosphorylation of histone H3 on tyrosine 41. This study addresses the question of the role of activated JAKs in specific gene activation by IFNγ. We carried out chromatin immunoprecipitation (ChIP) followed by PCR in IFNγ treated WISH cells and showed association of pJAK1, pJAK2, IFNGR1, and STAT1 on the same DNA sequence of the IRF-1 gene promoter. The β-actin gene, which is not activated by IFNγ, did not show this association. The movement of activated JAK to the nucleus and the IRF-1 promoter was confirmed by the combination of nuclear fractionation, confocal microscopy and DNA precipitation analysis using the biotinylated GAS promoter. Activated JAKs in the nucleus was associated with phosphorylated tyrosine 41 on histone H3 in the region of the GAS promoter. Unphosphorylated JAK2 was found to be constitutively present in the nucleus and was capable of undergoing activation in IFNγ treated cells, most likely via nuclear IFNGR1. Association of pJAK2 and IFNGR1 with histone H3 in IFNγ treated cells was demonstrated by histone H3 immunoprecipitation. Unphosphorylated STAT1 protein was associated with histone H3 of untreated cells. IFNγ treatment resulted in its disassociation and then re-association as pSTAT1. The results suggest a novel role for activated JAKs in epigenetic events for specific gene activation. Copyright © 2011 Elsevier Inc. All rights reserved.

IL-4/Stat6 activities correlate with apoptosis and metastasis in colon cancer cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li Benhui; Yang Xianzi; Department of Medical Oncology, Taihe Hospital, Yunyang Medical College, Shiyan, Hubei 442000

2008-05-02

IL-4-induced Stat6 signaling is active in a variety of cell types and plays a role in cell proliferation/growth and resistance to apoptosis. Using EMSA, we identified differential IL-4/Stat6 activities in colorectal cancer cell lines, HT-29 being active Stat6{sup high} phenotype and Caco-2 being defective Stat6{sup null} phenotype, respectively. Active Stat6{sup high} HT-29 cells exhibited resistance to apoptosis by flowcytometry and aggressive metastasis by Transwell assay compared with defective Stat6{sup null} Caco-2 cells. Comparing one another using RT-PCR, Stat6{sup high} HT-29 cells expressed more mRNA of anti-apoptotic and pro-metastatic genes Survivin, MDM2, and TMPRSS4, while Stat6{sup null} Caco-2 cells expressed moremore » mRNA of pro-apoptotic and anti-metastatic genes BAX, CAV1, and P53, respectively. This is the first study describing correlations of IL-4/Stat6 activities with apoptosis and metastasis in colon cancer. These findings, together with the observation of constitutive Stat6 activation in many human malignancies, suggest that Stat6 activities could be a biomarker for cancer cell's invasive/metastatic capability.« less

FLT3-ITD confers resistance to the PI3K/Akt pathway inhibitors by protecting the mTOR/4EBP1/Mcl-1 pathway through STAT5 activation in acute myeloid leukemia

PubMed Central

Nogami, Ayako; Oshikawa, Gaku; Okada, Keigo; Fukutake, Shusaku; Umezawa, Yoshihiro; Nagao, Toshikage; Kurosu, Tetsuya; Miura, Osamu

2015-01-01

FLT3-ITD and FLT3-TKD are the most frequent tyrosine kinase mutations in acute myeloid leukemia (AML), with the former associated with poor prognosis. Here, we show that the PI3K inhibitor GDC-0941 or the Akt inhibitor MK-2206 induced apoptosis through the mitochondria-mediated intrinsic pathway more efficiently in hematopoietic 32D cells driven by FLT3-TKD (32D/TKD) than FLT3-ITD (32D/ITD), which robustly activated STAT5. The resistance to GDC-0941 and MK-2206 was gained by expression of the constitutively activated STAT5 mutant STAT5A1*6 in 32D/TKD cells, while it was abrogated by the STAT5 inhibitor pimozide in 32D/ITD cells or FLT3-ITD-expressing human leukemic MV4–11 cells. GDC-0941 or MK-2206 induced dephosphorylation of 4EBP1 more conspicuously in 32D/TKD than in 32D/ITD, which was prevented or augmented by STAT5A1*6 or pimozide, respectively, and correlated with downregulation of the eIF4E/eIF4G complex formation and Mcl-1 expression. Furthermore, exogenous expression of Mcl-1 endowed resistance to GDC-0941 and MK-2206 on 32D/TKD cells. Finally, it was confirmed in primary AML cells with FLT3-ITD that pimozide enhanced 4EBP1 dephosphorylation and Mcl-1 downregulation to augment cytotoxicity of GDC-0941. These data suggest that the robust STAT5 activation by FLT3-ITD protects cells treated with the PI3K/Akt pathway inhibitors from apoptosis by maintaining Mcl-1 expression through the mTORC1/4EBP1/eIF4E pathway. PMID:25826077

FLT3-ITD confers resistance to the PI3K/Akt pathway inhibitors by protecting the mTOR/4EBP1/Mcl-1 pathway through STAT5 activation in acute myeloid leukemia.

PubMed

Nogami, Ayako; Oshikawa, Gaku; Okada, Keigo; Fukutake, Shusaku; Umezawa, Yoshihiro; Nagao, Toshikage; Kurosu, Tetsuya; Miura, Osamu

2015-04-20

FLT3-ITD and FLT3-TKD are the most frequent tyrosine kinase mutations in acute myeloid leukemia (AML), with the former associated with poor prognosis. Here, we show that the PI3K inhibitor GDC-0941 or the Akt inhibitor MK-2206 induced apoptosis through the mitochondria-mediated intrinsic pathway more efficiently in hematopoietic 32D cells driven by FLT3-TKD (32D/TKD) than FLT3-ITD (32D/ITD), which robustly activated STAT5. The resistance to GDC-0941 and MK-2206 was gained by expression of the constitutively activated STAT5 mutant STAT5A1*6 in 32D/TKD cells, while it was abrogated by the STAT5 inhibitor pimozide in 32D/ITD cells or FLT3-ITD-expressing human leukemic MV4-11 cells. GDC-0941 or MK-2206 induced dephosphorylation of 4EBP1 more conspicuously in 32D/TKD than in 32D/ITD, which was prevented or augmented by STAT5A1*6 or pimozide, respectively, and correlated with downregulation of the eIF4E/eIF4G complex formation and Mcl-1 expression. Furthermore, exogenous expression of Mcl-1 endowed resistance to GDC-0941 and MK-2206 on 32D/TKD cells. Finally, it was confirmed in primary AML cells with FLT3-ITD that pimozide enhanced 4EBP1 dephosphorylation and Mcl-1 downregulation to augment cytotoxicity of GDC-0941. These data suggest that the robust STAT5 activation by FLT3-ITD protects cells treated with the PI3K/Akt pathway inhibitors from apoptosis by maintaining Mcl-1 expression through the mTORC1/4EBP1/eIF4E pathway.

Tumor-associated macrophages promote neuroblastoma via STAT3 phosphorylation and up-regulation of c-MYC

PubMed Central

Hadjidaniel, Michael D.; Muthugounder, Sakunthala; Hung, Long T.; Sheard, Michael A.; Shirinbak, Soheila; Chan, Randall Y.; Nakata, Rie; Borriello, Lucia; Malvar, Jemily; Kennedy, Rebekah J.; Iwakura, Hiroshi; Akamizu, Takashi; Sposto, Richard; Shimada, Hiroyuki; DeClerck, Yves A.; Asgharzadeh, Shahab

2017-01-01

Tumor-associated macrophages (TAMs) are strongly associated with poor survival in neuroblastomas that lack MYCN amplification. To study TAM action in neuroblastomas, we used a novel murine model of spontaneous neuroblastoma lacking MYCN amplification, and observed recruitment and polarization of TAMs, which in turn enhanced neuroblastoma proliferation and growth. In both murine and human neuroblastoma cells, we found that TAMs increased STAT3 activation in neuroblastoma cells and transcriptionally up-regulated the MYC oncogene. Analysis of human neuroblastoma tumor specimens revealed that MYC up-regulation correlates with markers of TAM infiltration. In an IL6ko neuroblastoma model, the absence of IL-6 protein had no effect on tumor development and prevented neither STAT3 activation nor MYC up-regulation. In contrast, inhibition of JAK-STAT activation using AZD1480 or the clinically admissible inhibitor ruxolitinib significantly reduced TAM-mediated growth of neuroblastomas implanted subcutaneously in NOD scid gamma mice. Our results point to a unique mechanism in which TAMs promote tumor cells that lack amplification of an oncogene common to the malignancy by up-regulating transcriptional expression of a distinct oncogene from the same gene family, and underscore the role of IL-6-independent activation of STAT3 in this mechanism. Amplification of MYCN or constitutive up-regulation of MYC protein is observed in approximately half of high-risk tumors; our findings indicate a novel role of TAMs as inducers of MYC expression in neuroblastomas lacking independent oncogene activation. PMID:29207662

The RhoU/Wrch1 Rho GTPase gene is a common transcriptional target of both the gp130/STAT3 and Wnt-1 pathways

PubMed Central

SCHIAVONE, Davide; DEWILDE, Sarah; VALLANIA, Francesco; TURKSON, James; CUNTO, Ferdinando DI; POLI, Valeria

2010-01-01

STAT3 (signal transducer and activator of transcription 3) is a transcription factor activated by cytokines, growth factors and oncogenes, whose activity is required for cell survival/proliferation of a wide variety of primary tumours and tumour cell lines. Prominent among its multiple effects on tumour cells is the stimulation of cell migration and metastasis, whose functional mechanisms are however not completely characterized. RhoU/Wrch1 (Wnt-responsive Cdc42 homologue) is an atypical Rho GTPase thought to be constitutively bound to GTP. RhoU was first identified as a Wnt-1-inducible mRNA and subsequently shown to act on the actin cytoskeleton by stimulating filopodia formation and stress fibre dissolution. It was in addition recently shown to localize to focal adhesions and to Src-induced podosomes and enhance cell migration. RhoU overexpression in mammary epithelial cells stimulates quiescent cells to re-enter the cell cycle and morphologically phenocopies Wnt-1-dependent transformation. In the present study we show that Wnt-1-mediated RhoU induction occurs at the transcriptional level. Moreover, we demonstrate that RhoU can also be induced by gp130 cytokines via STAT3, and we identify two functional STAT3-binding sites on the mouse RhoU promoter. RhoU induction by Wnt-1 is independent of β-catenin, but does not involve STAT3. Rather, it is mediated by the Wnt/planar cell polarity pathway through the activation of JNK (c-Jun N-terminal kinase). Both the so-called non-canonical Wnt pathway and STAT3 are therefore able to induce RhoU, which in turn may be involved in mediating their effects on cell migration. PMID:19397496

Protein inhibitor of activated STAT3 inhibits adipogenic gene expression

DOE Office of Scientific and Technical Information (OSTI.GOV)

Deng Jianbei; Hua Kunjie; Caveney, Erica J.

2006-01-20

Protein inhibitor of activated STAT3 (PIAS3), a cytokine-induced repressor of signal transducer and activator of transcription 3 (STAT3) and a modulator of a broad array of nuclear proteins, is expressed in white adipose tissue, but its role in adipogenesis is not known. Here, we determined that PIAS3 was constitutively expressed in 3T3-L1 cells at all stages of adipogenesis. However, it translocated from the nucleus to the cytoplasm 4 days after induction of differentiation by isobutylmethylxanthine, dexamethasone, and insulin (MDI). In ob/ob mice, PIAS3 expression was increased in white adipose tissue depots compared to lean mice and was found in themore » cytoplasm of adipocytes. Overexpression of PIAS3 in differentiating preadipocytes, which localized primarily to the nucleus, inhibited mRNA level gene expression of adipogenic transcription factors C/EBP{alpha} and PPAR{gamma}, as well as their downstream target genes aP2 and adiponectin. PIAS3 also inhibited C/EBP{alpha} promoter activation mediated specifically by insulin, but not dexamethasone or isobutylmethylxanthine. Taken together, these data suggest that PIAS3 may play an inhibitory role in adipogenesis by modulating insulin-activated transcriptional activation events. Increased PIAS3 expression in adipose tissue may play a role in the metabolic disturbances of obesity.« less

Sustained phosphorylation of mutated FGFR3 is a crucial feature of genetic dwarfism and induces apoptosis in the ATDC5 chondrogenic cell line via PLCgamma-activated STAT1.

PubMed

Harada, Daisuke; Yamanaka, Yoshitaka; Ueda, Koso; Nishimura, Riko; Morishima, Tsuneo; Seino, Yoshiki; Tanaka, Hiroyuki

2007-08-01

The most frequent type of rhizomelic dwarfism, achondroplasia (ACH), is caused by mutations in the fibroblast growth factor receptor 3 (FGFR3) gene. Mutations in FGFR3 result in skeletal dysplasias of variable severity, including mild phenotypic effects in hypochondroplasia (HCH), severe phenotypic effects in thanatophoric dysplasia types I (TDI) and II (TDII), and severe but survivable phenotypic effects in severe achondroplasia with developmental delay and acanthosis nigricans (SADDAN). To explore the molecular mechanisms that result in the different phenotypes, we investigated the kinetics of mutated versions of FGFR3. First, we assayed the phosphorylation states of the mutated FGFR3s and found that the level of phosphorylation in TDI-FGFR3 was lower than in ACH-FGFR3, although the other mutants were phosphorylated according to phenotypic severity. Second, we analyzed the duration of the phosphorylation. TDI-FGFR3 was not highly phosphorylated under ligand-free conditions, but the peak phosphorylation levels of TDI-FGFR3 and ACH-FGFR3 were maintained for 30 min after stimulation with FGF-1. Moreover, ligand-dependent phosphorylation of TDI-FGFR3, but not ACH-FGFR3, lasted for more than 8 h after FGF-1 administration. The other mutant proteins showed sustained phosphorylation independent of ligand presence. Third, we investigated the intracellular localization of the mutant proteins. Immunofluorescence analysis showed accumulations of TDII-FGFR3, SADDAN-FGFR3, and a portion of TDI-FGFR3 in the endoplasmic reticulum (ER). Based on these data, we concluded that sustained phosphorylation of FGFR3 causes chondrodysplasia, and the phenotypic severity depends on the proportion of ER-localized mutant FGFR3. In FGFR3 signaling, the transcription factor, signal transducer and activator of transcription 1 (STAT1) inhibit proliferation and induce apoptosis of chondrocytes. Here we reveal that phospholipase C gamma (PLCgamma) mediates FGFR3-induced STAT1 activation. Both PLCgamma and STAT1 were activated by FGFR3 signaling, but a dominant-negative form of PLCgamma (DN-PLCgamma) remarkably reduced STAT1 phosphorylation. Apoptosis assays revealed that the constitutively active forms of FGFR3 (TDII-FGFR3) and STAT1 (STAT1-C) induce apoptosis of chondrogenic ATDC5 cells via caspase activity. DN-PLCgamma reduced the apoptosis of ATDC5 cells expressing TDII-FGFR3, but over-expression of both DN-PLCgamma and STAT1-C induced apoptosis. Therefore, we conclude that a PLCgamma-STAT1 pathway mediates apoptotic signaling by FGFR3.

Intramolecular hydrophobic interactions are critical mediators of STAT5 dimerization

NASA Astrophysics Data System (ADS)

Fahrenkamp, Dirk; Li, Jinyu; Ernst, Sabrina; Schmitz-van de Leur, Hildegard; Chatain, Nicolas; Küster, Andrea; Koschmieder, Steffen; Lüscher, Bernhard; Rossetti, Giulia; Müller-Newen, Gerhard

2016-10-01

STAT5 is an essential transcription factor in hematopoiesis, which is activated through tyrosine phosphorylation in response to cytokine stimulation. Constitutive activation of STAT5 is a hallmark of myeloid and lymphoblastic leukemia. Using homology modeling and molecular dynamics simulations, a model of the STAT5 phosphotyrosine-SH2 domain interface was generated providing first structural information on the activated STAT5 dimer including a sequence, for which no structural information is available for any of the STAT proteins. We identified a novel intramolecular interaction mediated through F706, adjacent to the phosphotyrosine motif, and a unique hydrophobic interface on the surface of the SH2 domain. Analysis of corresponding STAT5 mutants revealed that this interaction is dispensable for Epo receptor-mediated phosphorylation of STAT5 but essential for dimer formation and subsequent nuclear accumulation. Moreover, the herein presented model clarifies molecular mechanisms of recently discovered leukemic STAT5 mutants and will help to guide future drug development.

Intramolecular hydrophobic interactions are critical mediators of STAT5 dimerization

PubMed Central

Fahrenkamp, Dirk; Li, Jinyu; Ernst, Sabrina; Schmitz-Van de Leur, Hildegard; Chatain, Nicolas; Küster, Andrea; Koschmieder, Steffen; Lüscher, Bernhard; Rossetti, Giulia; Müller-Newen, Gerhard

2016-01-01

STAT5 is an essential transcription factor in hematopoiesis, which is activated through tyrosine phosphorylation in response to cytokine stimulation. Constitutive activation of STAT5 is a hallmark of myeloid and lymphoblastic leukemia. Using homology modeling and molecular dynamics simulations, a model of the STAT5 phosphotyrosine-SH2 domain interface was generated providing first structural information on the activated STAT5 dimer including a sequence, for which no structural information is available for any of the STAT proteins. We identified a novel intramolecular interaction mediated through F706, adjacent to the phosphotyrosine motif, and a unique hydrophobic interface on the surface of the SH2 domain. Analysis of corresponding STAT5 mutants revealed that this interaction is dispensable for Epo receptor-mediated phosphorylation of STAT5 but essential for dimer formation and subsequent nuclear accumulation. Moreover, the herein presented model clarifies molecular mechanisms of recently discovered leukemic STAT5 mutants and will help to guide future drug development. PMID:27752093

M-HIFU Inhibits Tumor Growth, Suppresses STAT3 Activity and Enhances Tumor Specific Immunity in a Transplant Tumor Model of Prostate Cancer

PubMed Central

Huang, Xiaoyi; Yuan, Fang; Liang, Meihua; Lo, Hui-Wen; Shinohara, Mari L.; Robertson, Cary; Zhong, Pei

2012-01-01

Objective In this study, we explored the use of mechanical high intensity focused ultrasound (M-HIFU) as a neo-adjuvant therapy prior to surgical resection of the primary tumor. We also investigated the role of signal transducer and activator of transcription 3 (STAT3) in M-HIFU elicited anti-tumor immune response using a transplant tumor model of prostate cancer. Methods RM-9, a mouse prostate cancer cell line with constitutively activated STAT3, was inoculated subcutaneously in C57BL/6J mice. The tumor-bearing mice (with a maximum tumor diameter of 5∼6 mm) were treated by M-HIFU or sham exposure two days before surgical resection of the primary tumor. Following recovery, if no tumor recurrence was observed in 30 days, tumor rechallenge was performed. The growth of the rechallenged tumor, survival rate and anti-tumor immune response of the animal were evaluated. Results No tumor recurrence and distant metastasis were observed in both treatment groups employing M-HIFU + surgery and surgery alone. However, compared to surgery alone, M-HIFU combined with surgery were found to significantly inhibit the growth of rechallenged tumors, down-regulate intra-tumoral STAT3 activities, increase cytotoxic T cells in spleens and tumor draining lymph nodes (TDLNs), and improve the host survival. Furthermore, M-HIFU combined with surgery was found to significantly decrease the level of immunosuppression with concomitantly increased number and activities of dendritic cells, compared to surgery alone. Conclusion Our results demonstrate that M-HIFU can inhibit STAT3 activities, and when combined synergistically with surgery, may provide a novel and promising strategy for the treatment of prostate cancers. PMID:22911830

Resveratrol induces cell cycle arrest and apoptosis in malignant NK cells via JAK2/STAT3 pathway inhibition.

PubMed

Quoc Trung, Ly; Espinoza, J Luis; Takami, Akiyoshi; Nakao, Shinji

2013-01-01

Natural killer (NK) cell malignancies, particularly aggressive NK cell leukaemias and lymphomas, have poor prognoses. Although recent regimens with L-asparaginase substantially improved outcomes, novel therapeutic approaches are still needed to enhance clinical response. Resveratrol, a naturally occurring polyphenol, has been extensively studied for its anti-inflammatory, cardioprotective and anti-cancer activities. In this study, we investigated the potential anti-tumour activities of resveratrol against the NK cell lines KHYG-1, NKL, NK-92 and NK-YS. Resveratrol induced robust G0/G1 cell cycle arrest, significantly suppressed cell proliferation and induced apoptosis in a dose- and time-dependent manner for all four cell lines. In addition, resveratrol suppressed constitutively active STAT3 in all the cell lines and inhibited JAK2 phosphorylation but had no effect on other upstream mediators of STAT3 activation, such as PTEN, TYK2, and JAK1. Resveratrol also induced downregulation of the anti-apoptotic proteins MCL1 and survivin, two downstream effectors of the STAT3 pathway. Finally, resveratrol induced synergistic effect on the apoptotic and antiproliferative activities of L-asparaginase against KHYG-1, NKL and NK-92 cells. These results suggest that resveratrol may have therapeutic potential against NK cell malignancies. Furthermore, our finding that resveratrol is a bonafide JAK2 inhibitor extends its potential benefits to other diseases with dysregulated JAK2 signaling.

Resveratrol Induces Cell Cycle Arrest and Apoptosis in Malignant NK Cells via JAK2/STAT3 Pathway Inhibition

PubMed Central

Quoc Trung, Ly; Espinoza, J. Luis; Takami, Akiyoshi; Nakao, Shinji

2013-01-01

Natural killer (NK) cell malignancies, particularly aggressive NK cell leukaemias and lymphomas, have poor prognoses. Although recent regimens with L-asparaginase substantially improved outcomes, novel therapeutic approaches are still needed to enhance clinical response. Resveratrol, a naturally occurring polyphenol, has been extensively studied for its anti-inflammatory, cardioprotective and anti-cancer activities. In this study, we investigated the potential anti-tumour activities of resveratrol against the NK cell lines KHYG-1, NKL, NK-92 and NK-YS. Resveratrol induced robust G0/G1 cell cycle arrest, significantly suppressed cell proliferation and induced apoptosis in a dose- and time-dependent manner for all four cell lines. In addition, resveratrol suppressed constitutively active STAT3 in all the cell lines and inhibited JAK2 phosphorylation but had no effect on other upstream mediators of STAT3 activation, such as PTEN, TYK2, and JAK1. Resveratrol also induced downregulation of the anti-apoptotic proteins MCL1 and survivin, two downstream effectors of the STAT3 pathway. Finally, resveratrol induced synergistic effect on the apoptotic and antiproliferative activities of L-asparaginase against KHYG-1, NKL and NK-92 cells. These results suggest that resveratrol may have therapeutic potential against NK cell malignancies. Furthermore, our finding that resveratrol is a bonafide JAK2 inhibitor extends its potential benefits to other diseases with dysregulated JAK2 signaling. PMID:23372833

Protein kinase C epsilon, which sensitizes skin to sun's UV radiation-induced cutaneous damage and development of squamous cell carcinomas, associates with Stat3.

PubMed

Aziz, Moammir H; Manoharan, Herbert T; Verma, Ajit K

2007-02-01

Chronic exposure to UV radiation (UVR) is the major etiologic factor in the development of human skin cancers including squamous cell carcinoma (SCC). We have shown that protein kinase C(epsilon) (PKC(epsilon)), a Ca(2+)-independent, phospholipid-dependent serine/threonine kinase, is an endogenous photosensitizer. PKC(epsilon) is among the six isoforms (alpha, delta, epsilon, eta, mu, and zeta) expressed in both mouse and human skin. PKC(epsilon) transgenic mice, which overexpress PKC(epsilon) in the basal epidermal cells and cells of the hair follicle, are highly sensitive to UVR-induced cutaneous damage and development of SCC. We now present that PKC(epsilon)-overexpressing, but not PKC(delta)-overexpressing, transgenic mice, when exposed to a single (4 kJ/m(2)) or repeated (four doses, 2 kJ/m(2)/dose, thrice weekly) UVR, emitted by Kodacel-filtered FS-40 sun lamps, elicit constitutive phosphorylation of signal transducers and activators of transcription 3 (Stat3) at both Tyr705 and Ser727 residues. UVR-induced phosphorylation of Stat3 accompanied increased expression of Stat3-regulated genes (c-myc, cyclin D1, cdc25A, and COX-2). In reciprocal immunoprecipitation/blotting experiments, phosphorylated Stat3 co-immunoprecipitated with PKC(epsilon). As observed in vivo using PKC(epsilon) knockout mice and in vitro in an immunocomplex kinase assay, PKC(epsilon) phosphorylated Stat3 at Ser727 residue. These results indicate for the first time that (a) PKC(epsilon) is a Stat3Ser727 kinase; (b) PKC(epsilon)-mediated phosphorylation of StatSer727 may be essential for transcriptional activity of Stat3; and (c) UVR-induced phosphorylation of Ser727 may be a key component of the mechanism by which PKC(epsilon) imparts sensitivity to UVR-induced development of SCC.

Lemongrass essential oil and citral inhibit Src/Stat3 activity and suppress the proliferation/survival of small-cell lung cancer cells, alone or in combination with chemotherapeutic agents.

PubMed

Maruoka, Takayuki; Kitanaka, Akira; Kubota, Yoshitsugu; Yamaoka, Genji; Kameda, Tomohiro; Imataki, Osamu; Dobashi, Hiroaki; Bandoh, Shuji; Kadowaki, Norimitsu; Tanaka, Terukazu

2018-03-13

Small-cell lung cancer (SCLC) is intractable due to its high propensity for relapse. Novel agents are thus needed for SCLC treatment. Lemongrass essential oil (LG-EO) and its major constituent, citral, have been reported to inhibit the proliferation and survival of several types of cancer cells. However, the precise mechanisms through which LG-EO and citral exert their effects on SCLC cells have not been fully elucidated. SCLC cells express Src and have high levels of Src-tyrosine kinase (Src-TK) activity. In most SCLC cell lines, constitutive phosphorylation of Stat3(Y705), which is essential for its activation, has been detected. Src-TK can phosphorylate Stat3(Y705), and activated Stat3 promotes the expression of the anti-apoptotic factors Bcl-xL and Mcl-1. In the present study, LG-EO and citral prevented Src-TK from phosphorylating Stat3(Y705), resulting in decreased Bcl-xL and Mcl-1 expression, in turn suppressing the proliferation/survival of SCLC cells. To confirm these findings, the wild-type-src gene was transfected into the LU135 SCLC cell line (LU135‑wt-src), in which Src and activated phospho-Stat3(Y705) were overexpressed. The suppression of cell proliferation and the induction of apoptosis by treatment with LG-EO or citral were significantly attenuated in the LU135-wt-src cells compared with the control LU135-mock cells. The signal transducer and activator of transcription 3 (Stat3) signaling pathway is also associated with intrinsic drug resistance. LU135-wt-src cells were significantly resistant to conventional chemotherapeutic agents compared with LU135-mock cells. The combined effects of citral and each conventional chemotherapeutic agent on SCLC cells were also evaluated. The combination treatment exerted additive or more prominent effects on LU135-wt-src, LU165 and MN1112 cells, which are relatively chemoresistant SCLC cells. These findings suggest that either LG-EO or citral, alone or in combination with chemotherapeutic agents, may be a novel therapeutic option for SCLC patients.

Integrated genomic sequencing reveals mutational landscape of T-cell prolymphocytic leukemia

PubMed Central

Kiel, Mark J.; Velusamy, Thirunavukkarasu; Rolland, Delphine; Sahasrabuddhe, Anagh A.; Chung, Fuzon; Bailey, Nathanael G.; Schrader, Alexandra; Li, Bo; Li, Jun Z.; Ozel, Ayse B.; Betz, Bryan L.; Miranda, Roberto N.; Medeiros, L. Jeffrey; Zhao, Lili; Herling, Marco

2014-01-01

The comprehensive genetic alterations underlying the pathogenesis of T-cell prolymphocytic leukemia (T-PLL) are unknown. To address this, we performed whole-genome sequencing (WGS), whole-exome sequencing (WES), high-resolution copy-number analysis, and Sanger resequencing of a large cohort of T-PLL. WGS and WES identified novel mutations in recurrently altered genes not previously implicated in T-PLL including EZH2, FBXW10, and CHEK2. Strikingly, WGS and/or WES showed largely mutually exclusive mutations affecting IL2RG, JAK1, JAK3, or STAT5B in 38 of 50 T-PLL genomes (76.0%). Notably, gain-of-function IL2RG mutations are novel and have not been reported in any form of cancer. Further, high-frequency mutations in STAT5B have not been previously reported in T-PLL. Functionally, IL2RG-JAK1-JAK3-STAT5B mutations led to signal transducer and activator of transcription 5 (STAT5) hyperactivation, transformed Ba/F3 cells resulting in cytokine-independent growth, and/or enhanced colony formation in Jurkat T cells. Importantly, primary T-PLL cells exhibited constitutive activation of STAT5, and targeted pharmacologic inhibition of STAT5 with pimozide induced apoptosis in primary T-PLL cells. These results for the first time provide a portrait of the mutational landscape of T-PLL and implicate deregulation of DNA repair and epigenetic modulators as well as high-frequency mutational activation of the IL2RG-JAK1-JAK3-STAT5B axis in the pathogenesis of T-PLL. These findings offer opportunities for novel targeted therapies in this aggressive leukemia. PMID:24825865

Integrated genomic sequencing reveals mutational landscape of T-cell prolymphocytic leukemia.

PubMed

Kiel, Mark J; Velusamy, Thirunavukkarasu; Rolland, Delphine; Sahasrabuddhe, Anagh A; Chung, Fuzon; Bailey, Nathanael G; Schrader, Alexandra; Li, Bo; Li, Jun Z; Ozel, Ayse B; Betz, Bryan L; Miranda, Roberto N; Medeiros, L Jeffrey; Zhao, Lili; Herling, Marco; Lim, Megan S; Elenitoba-Johnson, Kojo S J

2014-08-28

The comprehensive genetic alterations underlying the pathogenesis of T-cell prolymphocytic leukemia (T-PLL) are unknown. To address this, we performed whole-genome sequencing (WGS), whole-exome sequencing (WES), high-resolution copy-number analysis, and Sanger resequencing of a large cohort of T-PLL. WGS and WES identified novel mutations in recurrently altered genes not previously implicated in T-PLL including EZH2, FBXW10, and CHEK2. Strikingly, WGS and/or WES showed largely mutually exclusive mutations affecting IL2RG, JAK1, JAK3, or STAT5B in 38 of 50 T-PLL genomes (76.0%). Notably, gain-of-function IL2RG mutations are novel and have not been reported in any form of cancer. Further, high-frequency mutations in STAT5B have not been previously reported in T-PLL. Functionally, IL2RG-JAK1-JAK3-STAT5B mutations led to signal transducer and activator of transcription 5 (STAT5) hyperactivation, transformed Ba/F3 cells resulting in cytokine-independent growth, and/or enhanced colony formation in Jurkat T cells. Importantly, primary T-PLL cells exhibited constitutive activation of STAT5, and targeted pharmacologic inhibition of STAT5 with pimozide induced apoptosis in primary T-PLL cells. These results for the first time provide a portrait of the mutational landscape of T-PLL and implicate deregulation of DNA repair and epigenetic modulators as well as high-frequency mutational activation of the IL2RG-JAK1-JAK3-STAT5B axis in the pathogenesis of T-PLL. These findings offer opportunities for novel targeted therapies in this aggressive leukemia. © 2014 by The American Society of Hematology.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Goupille, Olivier; Penglong, Tipparat; Thalassemia Research Center, Mahidol University

The bromodomain and extraterminal (BET) domain family proteins are epigenetic modulators involved in the reading of acetylated lysine residues. The first BET protein inhibitor to be identified, (+)-JQ1, a thienotriazolo-1, 4-diazapine, binds selectively to the acetyl lysine-binding pocket of BET proteins. We evaluated the impact on adipogenesis of this druggable targeting of chromatin epigenetic readers, by investigating the physiological consequences of epigenetic modifications through targeting proteins binding to chromatin. JQ1 significantly inhibited the differentiation of 3T3-L1 preadipocytes into white and brown adipocytes by down-regulating the expression of genes involved in adipogenesis, particularly those encoding the peroxisome proliferator-activated receptor (PPAR-γ), themore » CCAAT/enhancer-binding protein (C/EBPα) and, STAT5A and B. The expression of a constitutively activated STAT5B mutant did not prevent inhibition by JQ1. Thus, the association of BET/STAT5 is required for adipogenesis but STAT5 transcription activity is not the only target of JQ1. Treatment with JQ1 did not lead to the conversion of white adipose tissue into brown adipose tissue (BAT). BET protein inhibition thus interferes with generation of adipose tissue from progenitors, confirming the importance of the connections between epigenetic mechanisms and specific adipogenic transcription factors. - Highlights: • JQ1 prevented the differentiation of 3T3-L1 preadipocytes into white adipocytes. • JQ1 affected clonal cell expansion and abolished lipid accumulation. • JQ1 prevented the differentiation of 3T3-L1 preadipocytes into brown adipocytes. • JQ1 treatment did not lead to the conversion of white adipose tissue into brown adipose tissue. • JQ1 decreased STAT5 expression, but STAT5B{sup ca} expression did not restore adipogenesis.« less

Endosomal accumulation of Toll-like receptor 4 causes constitutive secretion of cytokines and activation of signal transducers and activators of transcription in Niemann-Pick disease type C (NPC) fibroblasts: a potential basis for glial cell activation in the NPC brain.

PubMed

Suzuki, Michitaka; Sugimoto, Yuko; Ohsaki, Yuki; Ueno, Makoto; Kato, Shinsuke; Kitamura, Yukisato; Hosokawa, Hiroshi; Davies, Joanna P; Ioannou, Yiannis A; Vanier, Marie T; Ohno, Kousaku; Ninomiya, Haruaki

2007-02-21

Niemann-Pick disease type C (NPC) is an inherited lipid storage disorder caused by mutations in NPC1 or NPC2 genes. Loss of function of either protein results in the endosomal accumulation of cholesterol and other lipids, progressive neurodegeneration, and robust glial cell activation. Here, we report that cultured human NPC fibroblasts secrete interferon-beta, interleukin-6 (IL-6), and IL-8, and contain increased levels of signal transducers and activators of transcription (STATs). These cells also contained increased levels of Toll-like receptor 4 (TLR4) that accumulated in cholesterol-enriched endosomes/lysosomes, and small interfering RNA knockdown of this receptor reduced cytokine secretion. In the NPC1-/- mouse brain, glial cells expressed TLR4 and IL-6, whereas both glial and neuronal cells expressed STATs. Genetic deletion of TLR4 in NPC1-/- mice reduced IL-6 secretion by cultured fibroblasts but failed to alter STAT levels or glial cell activation in the brain. In contrast, genetic deletion of IL-6 normalized STAT levels and suppressed glial cell activation. These findings indicate that constitutive cytokine secretion leads to activation of STATs in NPC fibroblasts and that this secretion is partly caused by an endosomal accumulation of TLR4. These results also suggest that similar signaling events may underlie glial cell activation in the NPC1-/- mouse brain.

Erythroblast Transformation by the Friend Spleen Focus-Forming Virus Is Associated with a Block in Erythropoietin-Induced STAT1 Phosphorylation and DNA Binding and Correlates with High Expression of the Hematopoietic Phosphatase SHP-1

PubMed Central

Nishigaki, Kazuo; Hanson, Charlotte; Ohashi, Takashi; Spadaccini, Angelo; Ruscetti, Sandra

2006-01-01

Infection of mice with Friend spleen focus-forming virus (SFFV) results in a multistage erythroleukemia. In the first stage, the SFFV envelope glycoprotein interacts with the erythropoietin receptor and a short form of the receptor tyrosine kinase sf-Stk, resulting in constitutive activation of signal transducing molecules and the development of erythropoietin (Epo)-independent erythroid hyperplasia and polycythemia. The second stage results from the outgrowth of a rare virus-infected erythroid cell that expresses nonphysiological levels of the myeloid transcription factor PU.1. These cells exhibit a differentiation block and can be grown as murine erythroleukemia (MEL) cell lines. In this study, we examined SFFV MEL cells to determine whether their transformed phenotype was associated with a block in the activation of any Epo signal-transducing molecules. Our studies indicate that Epo- or SFFV-induced activation of STAT1/3 DNA binding activity is blocked in SFFV MEL cells. The block is at the level of tyrosine phosphorylation of STAT1, although Jak2 phosphorylation is not blocked in these cells. In contrast to Epo, alpha interferon can induce STAT1 phosphorylation and DNA binding in SFFV MEL cells. The SFFV-transformed cells were shown to express elevated levels of the hematopoietic phosphatase SHP-1, and treatment of the cells with a phosphatase inhibitor restored STAT1 tyrosine phosphorylation. MEL cells derived from Friend murine leukemia virus (MuLV) or ME26 MuLV-infected mice, which do not express PU.1, express lower levels of SHP-1 and are not blocked in STAT1/3 DNA-binding activity. Our studies suggest that SFFV-infected erythroid cells become transformed when differentiation signals activated by STAT1/3 are blocked due to high SHP-1 levels induced by inappropriate expression of the PU.1 protein. PMID:16731906

CGI-99 promotes breast cancer metastasis via autocrine interleukin-6 signaling.

PubMed

Lin, C; Liao, W; Jian, Y; Peng, Y; Zhang, X; Ye, L; Cui, Y; Wang, B; Wu, X; Xiong, Z; Wu, S; Li, J; Wang, X; Song, L

2017-06-29

Metastatic relapse remains largely incurable and a major challenge of clinical management in breast cancer, but the underlying mechanisms are poorly understood. Herein, we report that CGI-99 is overexpressed in breast cancer tissues from patients with metastatic recurrence within 5 years. High CGI-99 significantly predicts poorer 5-year metastasis-free patient survival. We find that CGI-99 increases breast cancer stem cell properties, and potentiates efficient tumor lung colonization and outgrowth in vivo. Furthermore, we demonstrate that CGI-99 activates the autocrine interleukin-6 (IL-6)/STAT3 signaling by increasing the accumulation and activity of RNA polymerase II and p300 cofactor at the proximal promoter of IL-6. Importantly, delivery of the IL-6-receptor humanized monoclonal antibody tocilizumab robustly abrogates CGI-99-induced metastasis in vivo. Finally, we find that high levels of CGI-99 are significantly correlated with STAT3 hyperactivation in breast cancer patients. These findings reveal a potential mechanism for constitutive activation of autocrine IL-6/STAT3 signaling and may suggest a novel target for clinical intervention in breast cancer.

Non-genomic STAT5-dependent effects at the endoplasmic reticulum and Golgi apparatus and STAT6-GFP in mitochondria

PubMed Central

Sehgal, Pravin B

2013-01-01

STAT protein species are well-known as transcription factors that regulate nuclear gene expression. Recent novel lines of research suggest new non-genomic functions of STAT5A/B and STAT6. It was discovered in human pulmonary arterial endothelial cells that STAT5A, including STAT5A-GFP, constitutively associated with the Golgi apparatus, and both STAT5A and B with the endoplasmic reticulum. Acute siRNA-mediated knockdown of STAT5A/B led to the rapid development of a dramatic cystic change in the endoplasmic reticulum (ER) characterized by deposition of the ER structural protein reticulon-4 (RTN4; also called Nogo-B) and the ER-resident GTPase atlastin-3 (ATL3) along cyst membranes and cyst-zone boundaries, accompanied by Golgi fragmentation. Functional consequences included reduced anterograde trafficking, an ER stress response (increased GRP78/BiP) and eventual mitochondrial fragmentation. This phenotype was “non-genomic” in that it was elicited in enucleated cytoplasts. In cross-immunopanning assays STAT5A and B species associated with ATL3, and the ER-lumen spacer CLIMP63 (also called cytoskeleton-associated protein 4, CKAP4) but not RTN4. From a disease significance perspective we posit that STAT5, which is known to be affected by estradiol-17β and prolactin, represents the gender-sensitive determinant in the pathogenesis of idiopathic pulmonary hypertension (IPAH), a disease which includes ER/Golgi dysfunctions but with a 2- to 4-fold higher prevalence in postpubertal women. A separate line of recent research produced evidence for the association of STAT6-GFP, but not STAT3-GFP, STAT3-DsRed, or STAT3-Flag, with mitochondria in live-cell, immunofluorescence, and immunoelectron microscopy. An N-terminal truncation of STAT6-GFP (1–459), which lacked the SH2 domain and Tyr-phosphorylation site, constitutively associated with mitochondria. Thus, the emergent new of biology STAT proteins includes non-genomic roles—structurally and functionally—in the three closely related membrane organelles consisting of the endoplasmic reticulum, Golgi apparatus, and mitochondria. PMID:24470974

Apigetrin inhibits gastric cancer progression through inducing apoptosis and regulating ROS-modulated STAT3/JAK2 pathway.

PubMed

Sun, Qian; Lu, Na-Na; Feng, Lei

2018-03-25

Apigetrin (APG), as a flavonoid, has many cellular bioactivities, including regulation of oxidative stress, and induction of apoptosis. However, the means by which APG suppresses human gastric cancer are still little to be understood. In the present study, the anti-cancer effects of APG on human gastric cancer cells were investigated. The results indicated that APG could suppress the proliferation and induce apoptosis in gastric cancer cells. Its role in apoptosis induction was through reducing Bcl-2, and enhancing Bax, Caspase-9/-3 and poly ADP-ribose polymerase (PARP) cleavage. In addition, APG incubation resulted in the generation of intracellular reactive oxygen species (ROS) in cells. Meanwhile, APG suppressed constitutive and interleukin-6 (IL-6)-stimulated signal transducer and activator of transcription 3 (STAT3), Janus kinase 2 gene (JAK2) and Src activation. However, ROS scavenger, N-acety-l-cysteine (NAC), diminished apoptosis induced by APG. And APG-triggered de-phosphorylation of STAT3/JAK2 was rescued by NAC pre-treatment. In vivo, APG administration significantly inhibited the gastric cancer cell xenograft tumorigenesis through inducing apoptosis and inhibiting STAT3/JAK2 pathways. Taken together, the findings above illustrated that APG might be used as a promising candidate against human gastric cancer progression. Copyright © 2018. Published by Elsevier Inc.

A novel MPL point mutation resulting in thrombopoietin-independent activation.

PubMed

Abe, M; Suzuki, K; Inagaki, O; Sassa, S; Shikama, H

2002-08-01

Thrombopoietin (TPO) and its receptor (MPL) are important regulators of megakaryopoiesis. MPL belongs to a cytokine receptor superfamily. To date, all constitutively active MPL mutants have been artificially constructed with amino acid substitutions in the transmembrane domain or extracellular domain of the protein, and they activate signal transduction pathways in Ba/F3 cells that can also be activated by the normal MPL. In this paper, we report a novel spontaneously occurring mutation of MPL, with an amino acid substitution of Trp(508) to Ser(508) in the intracellular domain of MPL, that induces the factor-independent growth of Ba/F3 cells. Examination of intracellular signaling pathways demonstrated that the mutant MPL protein constitutively activates three distinct signaling pathways, SHC-Ras-Raf-MAPK/JNK, JAK-STAT, and PI3K-Akt-Bad.

Stat6 activity-related Th2 cytokine profile and tumor growth advantage of human colorectal cancer cells in vitro and in vivo.

PubMed

Li, Ben Hui; Xu, Shuang Bing; Li, Feng; Zou, Xiao Guang; Saimaiti, Abudukeyoumu; Simayi, Dilixia; Wang, Ying Hong; Zhang, Yan; Yuan, Jia; Zhang, Wen Jie

2012-03-01

Signal transducer and activator of transcription 6 (Stat6) is critical in Th2 polarization of immune cells and active Stat6 activity has been suggested in anti-tumor immunity in animal models. The present study aims at investigating the impact of natural Stat6 activity on tumor microenvironment in human colorectal cancer cells in vitro and in vivo. Using colorectal cancer cell lines HT-29 and Caco-2 whose IL-4/Stat6 activities were known and nude mice as a model, we examined correlative relationships between Stat6 activities and gene expression profiles together with cellular behaviors in vitro and in vivo. HT-29 cells carrying active Stat6 signaling displayed spontaneous expression profiles favoring Th2 cytokines, cell cycle promotion, anti-apoptosis and pro-metastasis with increased mRNA levels of IL-4, IL-13, GATA-3, CDK4, CD44v6 and S100A4 using RT-PCR. In contrast, Caco-2 cells carrying defective Stat6 signaling exhibited spontaneous expression profiles favoring Th1 and Th17 cytokines, cell cycle inhibition, pro-apoptosis and anti-metastasis with elevated mRNA expression of IFNγ, TNFα, IL-12A, IL-17, IL-23, T-bet, CDKN1A, CDKNIB, CDKN2A and NM23-H1. Xenograft tumors of Stat6-active HT-29 cells showed a growth advantage over those of Stat6-defective Caco-2 cells. Furthermore, mice bearing HT-29 tumors expressed increased levels of Th2 cytokines IL-4 and IL-5 in the blood and pro-growth and/or pro-metastasis proteins CDK4 and CD44v6 in the tumor. To the contrary, mice bearing Caco-2 tumors expressed heightened levels of Th1 cytokines IFNγ and TNF in the blood and pro-apoptosis and anti-metastatic proteins p53 and p27(kip1) in the tumor. Colorectal cancer cells carrying active Stat6 signaling may create a microenvironment favoring Th2 cytokines and promoting expression of genes related to pro-growth, pro-metastasis and anti-apoptosis, which leads to a tumor growth advantage in vivo. These findings may imply why Stat6 pathway is constitutively activated in a number of human malignancies. Copyright © 2011 Elsevier Inc. All rights reserved.

Alternative activation of STAT1 and STAT3 in response to interferon-gamma.

PubMed

Qing, Yulan; Stark, George R

2004-10-01

Interferon-gamma (IFNgamma) is a pluripotent cytokine whose major biological effects are mediated through a pathway in which STAT1 is the predominant and essential transcription factor. STAT3 can also be activated weakly by IFNgamma, but the mechanism of activation and function of STAT3 as a part of the interferon response are not known. Here we show that STAT3 activation is much stronger and more prolonged in STAT1-null mouse embryo fibroblasts than in wild-type cells. In response to IFNgamma, SRC-family kinases are required to activate STAT3 (but not STAT1) through tyrosine phosphorylation, whereas the receptor-bound kinases JAK1 and JAK2 are required to activate both STATs. Tyrosine 419 of the IFNgamma receptor subunit 1 (IFNGR1) is required to activate both STATs, suggesting that STAT1 and STAT3 compete with each other for the same receptor phosphotyrosine motif. Activated STAT3 can replace STAT1 in STAT1-null cells to drive the transcription of certain genes, for example, socs-3 and c/ebpdelta, which have gamma-activated sequence motifs in their promoters. Work from Ian Kerr's laboratory reveals that the gp130-linked interleukin-6 receptor, which usually activates STAT3 predominantly, activates STAT1 efficiently when STAT3 is absent. Because STAT1 and STAT3 have opposing biological effects (STAT3 is an oncogene, and STAT1 is a tumor suppressor), the reciprocal activation of these two transcription factors in response to IFNgamma or interleukin-6 suggests that their relative abundance, which may vary substantially in different normal cell types, under different conditions or in tumors is likely to have a major impact on how cells behave in response to different cytokines.

Controlling nuclear JAKs and STATs for specific gene activation by IFN{gamma}

DOE Office of Scientific and Technical Information (OSTI.GOV)

Noon-Song, Ezra N.; Ahmed, Chulbul M.; Dabelic, Rea

2011-07-08

Highlights: {yields} Gamma interferon (IFN{gamma}) and its receptor subunit, IFNGR1, interact with the promoter region of IFN{gamma}-associated genes along with transcription factor STAT1{alpha}. {yields} We show that activated Janus kinases pJAK2 and pJAK1 also associate with IFNGR1 in the nucleus. {yields} The activated Janus kinases are responsible for phosphorylation of tyrosine 41 on histone H3, an important epigenetic event for specific gene activation. -- Abstract: We previously showed that gamma interferon (IFN{gamma}) and its receptor subunit, IFNGR1, interacted with the promoter region of IFN{gamma}-activated genes along with transcription factor STAT1{alpha}. Recent studies have suggested that activated Janus kinases pJAK2 andmore » pJAK1 also played a role in gene activation by phosphorylation of histone H3 on tyrosine 41. This study addresses the question of the role of activated JAKs in specific gene activation by IFN{gamma}. We carried out chromatin immunoprecipitation (ChIP) followed by PCR in IFN{gamma} treated WISH cells and showed association of pJAK1, pJAK2, IFNGR1, and STAT1 on the same DNA sequence of the IRF-1 gene promoter. The {beta}-actin gene, which is not activated by IFN{gamma}, did not show this association. The movement of activated JAK to the nucleus and the IRF-1 promoter was confirmed by the combination of nuclear fractionation, confocal microscopy and DNA precipitation analysis using the biotinylated GAS promoter. Activated JAKs in the nucleus was associated with phosphorylated tyrosine 41 on histone H3 in the region of the GAS promoter. Unphosphorylated JAK2 was found to be constitutively present in the nucleus and was capable of undergoing activation in IFN{gamma} treated cells, most likely via nuclear IFNGR1. Association of pJAK2 and IFNGR1 with histone H3 in IFN{gamma} treated cells was demonstrated by histone H3 immunoprecipitation. Unphosphorylated STAT1 protein was associated with histone H3 of untreated cells. IFN{gamma} treatment resulted in its disassociation and then re-association as pSTAT1. The results suggest a novel role for activated JAKs in epigenetic events for specific gene activation.« less

New Epigenetic Therapeutic Intervention for Metastatic Breast Cancer

DTIC Science & Technology

2016-04-01

also discovered that constitutively activated NF -kB in TNBC sustains prolonged activation of pro-inflammatory cytokines, enabling rapid spread...metastasis) of TNBC tumors. Notably, the functions of both transcription factors Twist and NF -kB in gene activation require lysine acetylation, which signs...proteins including Twist, NF -kB and STAT3. b. Define the molecular basis of the BET BrDs’ selective interactions with effector proteins through structure

A STAT3-decoy oligonucleotide induces cell death in a human colorectal carcinoma cell line by blocking nuclear transfer of STAT3 and STAT3-bound NF-κB

PubMed Central

2011-01-01

Background The transcription factor STAT3 (signal transducer and activator of transcription 3) is frequently activated in tumor cells. Activated STAT3 forms homodimers, or heterodimers with other TFs such as NF-κB, which becomes activated. Cytoplasmic STAT3 dimers are activated by tyrosine phosphorylation; they interact with importins via a nuclear localization signal (NLS) one of which is located within the DNA-binding domain formed by the dimer. In the nucleus, STAT3 regulates target gene expression by binding a consensus sequence within the promoter. STAT3-specific decoy oligonucleotides (STAT3-decoy ODN) that contain this consensus sequence inhibit the transcriptional activity of STAT3, leading to cell death; however, their mechanism of action is unclear. Results The mechanism of action of a STAT3-decoy ODN was analyzed in the colon carcinoma cell line SW 480. These cells' dependence on activated STAT3 was verified by showing that cell death is induced by STAT3-specific siRNAs or Stattic. STAT3-decoy ODN was shown to bind activated STAT3 within the cytoplasm, and to prevent its translocation to the nucleus, as well as that of STAT3-associated NF-κB, but it did not prevent the nuclear transfer of STAT3 with mutations in its DNA-binding domain. The complex formed by STAT3 and the STAT3-decoy ODN did not associate with importin, while STAT3 alone was found to co-immunoprecipitate with importin. Leptomycin B and vanadate both trap STAT3 in the nucleus. They were found here to oppose the cytoplasmic trapping of STAT3 by the STAT3-decoy ODN. Control decoys consisting of either a mutated STAT3-decoy ODN or a NF-κB-specific decoy ODN had no effect on STAT3 nuclear translocation. Finally, blockage of STAT3 nuclear transfer correlated with the induction of SW 480 cell death. Conclusions The inhibition of STAT3 by a STAT3-decoy ODN, leading to cell death, involves the entrapment of activated STAT3 dimers in the cytoplasm. A mechanism is suggested whereby this entrapment is due to STAT3-decoy ODN's inhibition of active STAT3/importin interaction. These observations point to the high potential of STAT3-decoy ODN as a reagent and to STAT3 nucleo-cytoplasmic shuttling in tumor cells as a potential target for effective anti-cancer compounds. PMID:21486470

PLGA nanoparticles codeliver paclitaxel and Stat3 siRNA to overcome cellular resistance in lung cancer cells

PubMed Central

Su, Wen-Pin; Cheng, Fong-Yu; Shieh, Dar-Bin; Yeh, Chen-Sheng; Su, Wu-Chou

2012-01-01

Background: Effective cancer chemotherapy remains an important issue in cancer treatment, and signal transducer and activator of transcription-3 (Stat3) activation leads to cellular resistance of anticancer agents. Polymers are ideal vectors to carry both chemotherapeutics and small interfering ribonucleic acid (siRNA) to enhance antitumor efficacy. In this paper, poly(lactic-co-glycolic acid) (PLGA) nanoparticles loaded with paclitaxel and Stat3 siRNA were successfully synthesized, and their applications in cancer cells were investigated. Methods: Firstly, paclitaxel was enclosed by PLGA nanoparticles through solvent evaporation. They were then coated with cationic polyethylenimine polymer (PLGA-PEI-TAX), enabling it to carry Stat3 siRNA on its surface through electrostatic interactions (PLGA-PEI-TAX-S3SI). The size, zeta potential, deliver efficacy, and release profile of the PLGA nanocomplexes were characterized in vitro. The cellular uptake, intracellular nanoparticle trajectory, and subsequent cellular events were evaluated after treatment with various PLGA nanocomplexes in human lung cancer A549 cells and A549-derived paclitaxel-resistant A549/T12 cell lines with α-tubulin mutation. Results: A549 and A549/T12 cells contain constitutively activated Stat3, and silencing Stat3 by siRNA made both cancer cells more sensitive to paclitaxel. Therefore, PLGA-PEI-TAX-S3SI was synthesized to test its therapeutic role in A549 and A549/T12 cells. Transmission electron microscopy showed the size of PLGA-PEI-TAX-S3SI to be around 250 nm. PLGA-PEI nanoparticles were nontoxic. PLGA-PEI-TAX was taken up by A549 and A549/T12 cells more than free paclitaxel, and they induced more condensed microtubule bundles and had higher cytotoxicity in these cancer cells. Moreover, the yellowish fluorescence observed in the cytoplasm of the cancer cells indicates that the PLGA-PEI nanoparticles were still simultaneously delivering Oregon Green paclitaxel and cyanine-5-labeled Stat3 siRNA 3 hours after treatment. Furthermore, after the cancer cells were incubated with the synthesized PLGA nanocomplexes, PLGA-PEI-TAX-S3SI suppressed Stat3 expression and induced more cellular apoptosis in A549 and A549/T12 cells compared with PLGA-PEI-TAX. Conclusion: The PLGA-PEI-TAX-S3SI complex provides a new therapeutic strategy to control cancer cell growth. PMID:22904633

Stat3-induced S1PR1 expression is critical for persistent Stat3 activation in tumors

PubMed Central

Lee, Heehyoung; Deng, Jiehui; Kujawski, Maciej; Yang, Chunmei; Liu, Yong; Herrmann, Andreas; Kortylewski, Marcin; Horne, David; Somlo, George; Forman, Stephen; Jove, Richard; Yu, Hua

2011-01-01

IL-6/Jak2 signaling is viewed critical for persistent Stat3 activation in cancer. However, IL-6-induced Stat3 activity is transient in normal physiology. Here we identify a mechanism important for persistent Stat3 activation in tumor cells and the tumor microenvironment. We show that sphingosine-1-phosphate receptor 1 (S1PR1), a G-protein-coupled receptor for lysophospholipid sphingosine-1-phosphate (S1P), is elevated in Stat3-positive tumors. Stat3 is a transcription factor for the S1pr1 gene. Enhanced S1pr1 expression activates Stat3 and upregulates Il6 gene expression, thereby accelerating tumor growth and metastasis. Conversely, silencing S1pr1 in tumor cells or immune cells inhibits tumor Stat3 activity, tumor growth and metastasis. S1P/S1PR1-induced Stat3 activation is persistent, in contrast to transient Stat3 activation by IL-6. S1PR1 activates Stat3 in part by upregulating Jak2 tyrosine kinase activity. We demonstrate that Stat3-induced S1pr1 expression, as well as S1P/S1PR1 pathway, is important for persistent Stat3 activation in cancer cells and the tumor microenvironment and for malignant progression. PMID:21102457

Molecular cloning, subcelluar location and expression profile of signal transducer and activator of transcription 2 (STAT2) from turbot, Scophthalmus maximus.

PubMed

Wang, Na; Wang, Xian-Li; Yang, Chang-Geng; Chen, Song-Lin

2013-10-01

Signal transducer and activator of transcription 2 (STAT2) is an important molecule involved in the type I interferon signalling pathway. To date, little STAT2 homologue is available in fish except Atlantic salmon and goldfish. In this paper, STAT2 was firstly cloned and characterized from turbot, a marine flatfish with high economic value. Briefly, turbot STAT2 cDNA is 3206 bp in length encoding a predicted protein of 793 amino acids. The phylogenetic tree shows that turbot STAT2 protein shared the closest relationship with Atlantic salmon. Analysis of subcellular distribution indicates that STAT2 is mainly present in the cytoplasm of TK cells. Stat2 mRNA is constitutively expressed in widespread tissues and induced by several folds in turbot tissues and TK cells after stimulation with Vibrio anguillarum and lymphocystis disease virus (LCDV). Unlike the higher vertebrate STAT2, turbot STAT2 nuclear export signal (NES) exists not in the C-terminal 79 amino acids but in N-terminal 137-312 amino acids (STAT_alpha domain). The nuclear translocation of turbot STAT2 after Poly(I:C) treatment proved its transcription activity in TK cells. All these results suggested that STAT2 may be involved in the immune response in turbot as a transcription factor. Copyright © 2013 Elsevier Ltd. All rights reserved.

Ibrutinib modulates the immunosuppressive CLL microenvironment through STAT3-mediated suppression of regulatory B-cell function and inhibition of the PD-1/PD-L1 pathway.

PubMed

Kondo, K; Shaim, H; Thompson, P A; Burger, J A; Keating, M; Estrov, Z; Harris, D; Kim, E; Ferrajoli, A; Daher, M; Basar, R; Muftuoglu, M; Imahashi, N; Alsuliman, A; Sobieski, C; Gokdemir, E; Wierda, W; Jain, N; Liu, E; Shpall, E J; Rezvani, K

2018-04-01

Ibrutinib, a covalent inhibitor of Bruton Tyrosine Kinase (BTK), is approved for treatment of patients with relapsed/refractory or treatment-naïve chronic lymphocytic leukemia (CLL). Besides directly inhibiting BTK, ibrutinib possesses immunomodulatory properties through targeting multiple signaling pathways. Understanding how this ancillary property of ibrutinib modifies the CLL microenvironment is crucial for further exploration of immune responses in this disease and devising future combination therapies. Here, we investigated the mechanisms underlying the immunomodulatory properties of ibrutinib. In peripheral blood samples collected prospectively from CLL patients treated with ibrutinib monotherapy, we observed selective and durable downregulation of PD-L1 on CLL cells by 3 months post-treatment. Further analysis showed that this effect was mediated through inhibition of the constitutively active signal transducer and activator of transcription 3 (STAT3) in CLL cells. Similar downregulation of PD-1 was observed in CD4+ and CD8+ T cells. We also demonstrated reduced interleukin (IL)-10 production by CLL cells in patients receiving ibrutinib, which was also linked to suppression of STAT3 phosphorylation. Taken together, these findings provide a mechanistic basis for immunomodulation by ibrutinib through inhibition of the STAT3 pathway, critical in inducing and sustaining tumor immune tolerance. The data also merit testing of combination treatments combining ibrutinib with agents capable of augmenting its immunomodulatory effects.

Interleukin-7-induced Stat-5 acts in synergy with Flt-3 signaling to stimulate expansion of hematopoietic progenitor cells.

PubMed

Åhsberg, Josefine; Tsapogas, Panagiotis; Qian, Hong; Zetterblad, Jenny; Zandi, Sasan; Månsson, Robert; Jönsson, Jan-Ingvar; Sigvardsson, Mikael

2010-11-19

The development of lymphoid cells from bone marrow progenitors is dictated by interplay between internal cues such as transcription factors and external signals like the cytokines Flt-3 ligand and Il-7. These proteins are both of large importance for normal lymphoid development; however, it is unclear if they act in direct synergy to expand a transient Il-7R(+)Flt-3(+) population or if the collaboration is created through sequential activities. We report here that Flt-3L and Il-7 synergistically stimulated the expansion of primary Il-7R(+)Flt-3(+) progenitor cells and a hematopoietic progenitor cell line ectopically expressing the receptors. The stimulation resulted in a reduced expression of pro-apoptotic genes and also mediated survival of primary progenitor cells in vitro. However, functional analysis of single cells suggested that the anti-apoptotic effect was additive indicating that the synergy observed mainly depends on stimulation of proliferation. Analysis of downstream signaling events suggested that although Il-7 induced Stat-5 phosphorylation, Flt-3L caused activation of the ERK and AKT signaling pathways. Flt-3L could also drive proliferation in synergy with ectopically expressed constitutively active Stat-5. This synergy could be inhibited with either receptor tyrosine kinase or MAPK inhibitors suggesting that Flt-3L and Il-7 act in synergy by activation of independent signaling pathways to expand early hematopoietic progenitors.

New Epigenetic Therapeutic Intervention for Metastatic Breast Cancer

DTIC Science & Technology

2016-04-01

transcription factor Twist are markedly over-expressed in TNBC but not luminal breast cancer cells. We also discovered that constitutively activated NF -kB in...transcription factors Twist and NF -kB in gene activation require lysine acetylation, which signs to activate the transcriptional machinery in chromatin...including Twist, NF -kB and STAT3. b. Define the molecular basis of the BET BrDs’ selective interactions with effector proteins through structure-guided

JAB1 regulates unphosphorylated STAT3 DNA-binding activity through protein–protein interaction in human colon cancer cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nishimoto, Arata, E-mail: anishimo@yamaguchi-u.ac.jp; Kugimiya, Naruji; Hosoyama, Toru

2013-08-30

Highlights: •JAB1 interacted with unphosphorylated STAT3 in the nucleus. •JAB1 knockdown tended to increase nuclear STAT3 expression. •JAB1 knockdown significantly decreased unphosphorylated STAT3 DNA-binding activity. •JAB1 knockdown significantly decreased MDR1, NANOG, and VEGF expressions. •Nuclear JAB1, but not nuclear STAT3, correlated with STAT3 DNA-binding activity. -- Abstract: Recent studies have revealed that unphosphorylated STAT3 forms a dimer, translocates to the nucleus, binds to the STAT3 binding site, and activates the transcription of STAT3 target genes, thereby playing an important role in oncogenesis in addition to phosphorylated STAT3. Among signaling steps of unphosphorylated STAT3, nuclear translocation and target DNA-binding are themore » critical steps for its activation. Therefore, elucidating the regulatory mechanism of these signaling steps of unphosphorylated STAT3 is a potential step in the discovery of a novel cancer drug. However, the mechanism of unphosphorylated STAT3 binding to the promoter of target genes remains unclear. In this study, we focused on Jun activation domain-binding protein 1 (JAB1) as a candidate protein that regulates unphosphorylated STAT3 DNA-binding activity. Initially, we observed that both unphosphorylated STAT3 and JAB1 existed in the nucleus of human colon cancer cell line COLO205 at the basal state (no cytokine stimulation). On the other hand, phosphorylated STAT3 did not exist in the nucleus of COLO205 cells at the basal state. Immunoprecipitation using nuclear extract of COLO205 cells revealed that JAB1 interacted with unphosphorylated STAT3. To investigate the effect of JAB1 on unphosphorylated STAT3 activity, RNAi studies were performed. Although JAB1 knockdown tended to increase nuclear STAT3 expression, it significantly decreased unphosphorylated STAT3 DNA-binding activity. Subsequently, JAB1 knockdown significantly decreased the expression levels of MDR1, NANOG, and VEGF, which are STAT3 target genes. Furthermore, the expression level of nuclear JAB1, but not nuclear STAT3, correlated with unphosphorylated STAT3 DNA-binding activity between COLO205 and LoVo cells. Taken together, these results suggest that nuclear JAB1 positively regulates unphosphorylated STAT3 DNA-binding activity through protein–protein interaction in human colon cancer cell line COLO205.« less

Metabolism pathways in chronic lymphocytic leukemia

PubMed Central

Rozovski, Uri; Hazan-Halevy, Inbal; Barzilay, Merav; Keating, Michael J.; Estrov, Zeev

2016-01-01

Alterations in CLL cell metabolism have been studied by several investigators. Unlike normal B lymphocytes or other leukemia cells, CLL cells, like adipocytes, store lipids and utilize free fatty acids (FFA) to produce chemical energy. None of the recently identified mutations in CLL directly affects metabolic pathways, suggesting that genetic alterations do not directly contribute to CLL cells’ metabolic reprogramming. Conversely, recent data suggest that activation of STAT3 or downregulation of microRNA-125 levels plays a crucial role in the utilization of FFA to meet CLL cells’ metabolic needs. STAT3, known to be constitutively activated in CLL, increases the levels of lipoprotein lipase that mediates lipoprotein uptake and shifts CLL cells’ metabolism towards utilization of FFA. Herein we review the evidence for altered lipid metabolism, increased mitochondrial activity, and formation of reactive oxygen species in CLL cells, and discuss possible therapeutic strategies to inhibit lipid metabolism pathways in patient with CLL. PMID:26643954

Dysregulation of Suppressor of Cytokine Signaling 3 in Keratinocytes Causes Skin Inflammation Mediated by Interleukin-20 Receptor-Related Cytokines

PubMed Central

Uto-Konomi, Ayako; Miyauchi, Kosuke; Ozaki, Naoko; Motomura, Yasutaka; Suzuki, Yoshie; Yoshimura, Akihiko; Suzuki, Shinobu; Cua, Daniel; Kubo, Masato

2012-01-01

Homeostatic regulation of epidermal keratinocytes is controlled by the local cytokine milieu. However, a role for suppressor of cytokine signaling (SOCS), a negative feedback regulator of cytokine networks, in skin homeostasis remains unclear. Keratinocyte specific deletion of Socs3 (Socs3 cKO) caused severe skin inflammation with hyper-production of IgE, epidermal hyperplasia, and S100A8/9 expression, although Socs1 deletion caused no inflammation. The inflamed skin showed constitutive STAT3 activation and up-regulation of IL-6 and IL-20 receptor (IL-20R) related cytokines, IL-19, IL-20 and IL-24. Disease development was rescued by deletion of the Il6 gene, but not by the deletion of Il23, Il4r, or Rag1 genes. The expression of IL-6 in Socs3 cKO keratinocytes increased expression of IL-20R-related cytokines that further facilitated STAT3 hyperactivation, epidermal hyperplasia and neutrophilia. These results demonstrate that skin homeostasis is strictly regulated by the IL-6-STAT3-SOCS3 axis. Moreover, the SOCS3-mediated negative feedback loop in keratinocytes has a critical mechanistic role in the prevention of skin inflammation caused by hyperactivation of STAT3. PMID:22792286

STAT3 as a potential therapeutic target in ALDH+ and CD44+/CD24+ stem cell-like pancreatic cancer cells.

PubMed

Lin, Li; Jou, David; Wang, Yina; Ma, Haiyan; Liu, Tianshu; Fuchs, James; Li, Pui-Kai; Lü, Jiagao; Li, Chenglong; Lin, Jiayuh

2016-12-01

Persistent activation of signal transducers and activators of transcription 3 (STAT3) is commonly detected in many types of cancer including pancreatic cancer. Whether STAT3 is activated in stem cell-like pancreatic cancer cells and the effect of STAT3 inhibition, is still unknown. Flow cytometry was used to isolate pancreatic cancer stem-like cells which are identified by both aldehyde dehydrogenase (ALDH)-positive (ALDH+) as well as cluster of differentiation (CD) 44-positive/CD24-positive subpopulations (CD44+/CD24+). STAT3 activation and the effects of STAT3 inhibition by STAT3 inhibitors, LLL12, FLLL32, and Stattic in ALDH+ and CD44+/CD24+ cells were examined. Our results showed that ALDH+ and CD44+/CD24+ pancreatic cancer stem-like cells expressed higher levels of phosphorylated STAT3, an active form of STAT3, compared to ALDH-negative (ALDH-) and CD44-negative/CD24-negative (CD44-/CD24-) pancreatic cancer cells, suggesting that STAT3 is activated in pancreatic cancer stem-like cells. Small molecular STAT3 inhibitors inhibited STAT3 phosphorylation, STAT3 downstream target gene expression, cell viability, and tumorsphere formation in ALDH+ and CD44+/CD24+ cells. Our results indicate that STAT3 is a novel therapeutic target in pancreatic cancer stem-like cells and inhibition of activated STAT3 in these cells by STAT3 inhibitors may offer an effective treatment for pancreatic cancer.

Myeloproliferative disease induced by TEL-PDGFRB displays dynamic range sensitivity to Stat5 gene dosage

PubMed Central

Cain, Jennifer A.; Xiang, Zhifu; O'Neal, Julie; Kreisel, Friederike; Colson, AnnaLynn; Luo, Hui; Hennighausen, Lothar

2007-01-01

Expression of the constitutively activated TEL/PDGFβR fusion protein is associated with the t(5;12)(q33;p13) chromosomal translocation found in a subset of patients with chronic myelomonocytic leukemia. TEL/PDGFβR activates multiple signal transduction pathways in cell-culture systems, and expression of the TEL-PDGFRB fusion gene induces myeloproliferative disease (MPD) in mice. We used gene-targeted mice to characterize the contribution of signal transducer and activator of transcription (Stat) and Src family genes to TEL-PDGFRB–mediated transformation in methylcellulose colony and murine bone marrow transduction/transplantation assays. Fetal liver hematopoietic stem and progenitor cells harboring targeted deletion of both Stat5a and Stat5b (Stat5abnull/null) genes were refractory to transformation by TEL-PDGFRB in methylcellulose colony assays. Notably, these cell populations were maintained in Stat5abnull/null fetal livers and succumbed to transformation by c-Myc. Surprisingly, targeted disruption of either Stat5a or Stat5b alone also impaired TEL-PDGFRB–mediated transformation. Survival of TPiGFP→Stat5a−/− and TPiGFP→Stat5a+/− mice was significantly prolonged, demonstrating significant sensitivity of TEL-PDGFRB–induced MPD to the dosage of Stat5a. TEL-PDGFRB–mediated MPD was incompletely penetrant in TPiGFP→Stat5b−/− mice. In contrast, Src family kinases Lyn, Hck, and Fgr and the Stat family member Stat1 were dispensable for TEL-PDGFRB disease. Together, these data demonstrate that Stat5a and Stat5b are dose-limiting mediators of TEL-PDGFRB–induced myeloproliferation. PMID:17218386

Pancreatic adenocarcinoma upregulated factor (PAUF) confers resistance to pancreatic cancer cells against oncolytic parvovirus H-1 infection through IFNA receptor-mediated signaling

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kaowinn, Sirichat; Cho, Il-Rae; Moon, Jeong

2015-04-03

Pancreatic adenocarcinoma upregulated factor (PAUF), a novel oncogene, plays a crucial role in the development of pancreatic cancer, including its metastasis and proliferation. Therefore, PAUF-expressing pancreatic cancer cells could be important targets for oncolytic virus-mediated treatment. Panc-1 cells expressing PAUF (Panc-PAUF) showed relative resistance to parvovirus H-1 infection compared with Panc-1 cells expressing an empty vector (Panc-Vec). Of interest, expression of type I IFN-α receptor (IFNAR) was higher in Panc-PAUF cells than in Panc-Vec cells. Increased expression of IFNAR in turn increased the activation of Stat1 and Tyk2 in Panc-PAUF cells compared with that in Panc-Vec cells. Suppression of Tyk2more » and Stat1, which are important downstream molecules for IFN-α signaling, sensitized pancreatic cancer cells to parvovirus H-1-mediated apoptosis. Further, constitutive suppression of PAUF sensitized Bxpc3 pancreatic cancer cells to parvovirus H-1 infection. Taken together, these results suggested that PAUF conferred resistance to pancreatic cancer cells against oncolytic parvovirus H-1 infection through IFNAR-mediated signaling. - Highlights: • PAUF confers resistance against oncolytic parvovirus H-1 infection. • PAUF enhances the expression of IFNAR in Panc-1 cells. • Increased activation of Tyk2 or Stat1 by PAUF provides resistance to parvovirus H-1-mediated apoptosis. • Constitutive inhibition of PAUF enhances parvovirus H-1-mediated oncolysis of Bxpc3 pancreatic cancer cells.« less

PKCδ activated by c-MET enhances infiltration of human glioblastoma cells through NOTCH2 signaling

PubMed Central

Kang, Seok-Gu; Kim, Rae-Kwon; Cui, Yan-Hong; Lee, Hae-June; Kim, Min-Jung; Lee, Jae-Seong; Kim, In-Gyu; Suh, Yongjoon; Lee, Su-Jae

2016-01-01

Poor prognosis of glioblastoma (GBM) is attributable to the propensity of tumor cells to infiltrate into the brain parenchyma. Protein kinase C (PKC) isozymes are highly expressed or aberrantly activated in GBM. However, how this signaling node translates to GBM cell invasiveness remains unknown. Here, we report that among PKC isoforms, PKCδ is strongly associated with infiltration of GBM cells. Notably, PKCδ enhanced Tyr418 phosphorylation of the non-receptor tyrosine kinase SRC, which in turn activated STAT3 and subsequent NOTCH2 signaling, ultimately leading to GBM cell invasiveness. Furthermore, we showed that PKCδ was aberrantly activated in GBM cells by c-MET, a receptor tyrosine kinase hyperactivated in GBM. In agreement, inhibition either component in the c-MET/PKCδ/SRC/STAT3 signaling axis effectively blocked the NOTCH2 signaling and invasiveness of GBM cells. Taken together, our findings shed a light on the signaling mechanisms behind the constitutive activation of PKCδ signaling in GBM. PMID:26700818

STAT3 Inhibitory Activity of Structurally Simplified Withaferin A Analogues.

PubMed

Tahara, Teruyuki; Streit, Ursula; Pelish, Henry E; Shair, Matthew D

2017-04-07

Signal transducer and activator of transcription 3 (STAT3) is a component of the JAK/STAT pathway. Therapeutic inhibition of STAT3 has been of high interest, as its aberrant activation has been linked to cancer, inflammation, and other human diseases. The withanolide family natural product withaferin A (1) inhibits STAT3 activation. We designed, synthesized, and evaluated simplified withanolide analogues SLW1 (3) and SLW2 (4), and found that SLW1 retained the STAT3 inhibitory activity of withaferin A.

JAK2-V617F-induced MAPK activity is regulated by PI3K and acts synergistically with PI3K on the proliferation of JAK2-V617F-positive cells

PubMed Central

Wolf, Alexandra; Eulenfeld, René; Gäbler, Karoline; Rolvering, Catherine; Haan, Serge; Behrmann, Iris; Denecke, Bernd; Haan, Claude; Schaper, Fred

2013-01-01

The identification of a constitutively active JAK2 mutant, namely JAK2-V617F, was a milestone in the understanding of Philadelphia chromosome-negative myeloproliferative neoplasms. The JAK2-V617F mutation confers cytokine hypersensitivity, constitutive activation of the JAK-STAT pathway, and cytokine-independent growth. In this study we investigated the mechanism of JAK2-V617F-dependent signaling with a special focus on the activation of the MAPK pathway. We observed JAK2-V617F-dependent deregulated activation of the multi-site docking protein Gab1 as indicated by constitutive, PI3K-dependent membrane localization and tyrosine phosphorylation of Gab1. Furthermore, we demonstrate that PI3K signaling regulates MAPK activation in JAK2-V617F-positve cells. This cross-regulation of the MAPK pathway by PI3K affects JAK2-V617F-specific target gene induction, erythroid colony formation, and regulates proliferation of JAK2-V617F-positive patient cells in a synergistically manner. PMID:24069558

Roles of unphosphorylated STATs in signaling.

PubMed

Yang, Jinbo; Stark, George R

2008-04-01

The seven members of the signal transducer and activator of transcription (STAT) family of transcription factors are activated in response to many different cytokines and growth factors by phosphorylation of specific tyrosine residues. The STAT1 and STAT3 genes are specific targets of activated STATs 1 and 3, respectively, resulting in large increases in the levels of these unphosphorylated STATs (U-STATs) in response to the interferons (STAT1) or ligands that active gp130, such as IL-6 (STAT3). U-STATs drive gene expression by novel mechanisms distinct from those used by phosphorylated STAT (P-STAT) dimers. In this review, we discuss the roles of U-STATs in transcription and regulation of gene expression.

The JAK2 Inhibitor, AZD1480, Potently Blocks Stat3 Signaling and Oncogenesis in Solid Tumors

PubMed Central

Hedvat, Michael; Huszar, Dennis; Herrmann, Andreas; Gozgit, Joseph M.; Schroeder, Anne; Sheehy, Adam; Buettner, Ralf; Proia, David; Kowolik, Claudia M.; Xin, Hong; Armstrong, Brian; Bebernitz, Geraldine; Weng, Shaobu; Wang, Lin; Ye, Minwei; McEachern, Kristen; Chen, Huawei; Morosini, Deborah; Bell, Kirsten; Alimzhanov, Marat; Ioannidis, Stephanos; McCoon, Patricia; Cao, Zhu A.; Yu, Hua; Jove, Richard; Zinda, Michael

2009-01-01

Summary Persistent activation of Stat3 is oncogenic and is prevalent in a wide variety of human cancers. Chronic cytokine stimulation is associated with Stat3 activation in some tumors, implicating cytokine receptor-associated Jak family kinases. Using Jak2 inhibitors, we demonstrate a central role of Jaks in modulating basal and cytokine-induced Stat3 activation in human solid tumor cell lines. Inhibition of Jak2 activity is associated with abrogation of Stat3 nuclear translocation and tumorigenesis. The Jak2 inhibitor, AZD1480, suppresses the growth of human solid tumor xenografts harboring persistent Stat3 activity. We demonstrate the essential role of Stat3 downstream of Jaks by inhibition of tumor growth using shRNA targeting Stat3. Our data support a key role of Jak kinase activity in Stat3-dependent tumorigenesis. PMID:19962667

STATs profiling reveals predominantly-activated STAT3 in cholangiocarcinoma genesis and progression.

PubMed

Dokduang, Hasaya; Techasen, Anchalee; Namwat, Nisana; Khuntikeo, Narong; Pairojkul, Chawalit; Murakami, Yoshinori; Loilome, Watcharin; Yongvanit, Puangrat

2014-10-01

We investigated the aberrant expression of the STAT family in humans and liver fluke (Opisthorchis viverrini, Ov)-induced hamster cholangiocarcinoma (CCA) tissues. The expression and phosphorylation of STAT1, STAT2, STAT3, STAT4, STAT5a, STAT5b and STAT6 in human hamster CCA tissues were immunohistochemistry-profiled. Localizations of STAT5 in macrophages and lipopolysaccharide (LPS)-induced macrophage-conditioned media mediated STAT3 activation in CCA cells were demonstrated. The expressions of STAT 1-4 and 6 were detected in the cytoplasm of hyperplastic bile ducts and tumor cells, whereas STAT5a and STAT5b were observed in macrophages and connective tissues surrounding tumor, respectively. The expressions of STAT3 and STAT5b were significantly observed in tumors with a poorer histological differentiation. STAT3 expression was significantly associated with shorter survival of CCA patients and was predominately activated in CCA cell lines. In the CCA-hamsters, STATs expression was gradually increased along the carcinogenesis, especially at 30 days post-infection in which the inflammatory response was markedly observed, showing the correlation between the inflammation and STATs activation. Moreover, LPS-induced macrophage-conditioned media could mediate STAT3 activation in CCA cells. STAT3 is the major STAT, which plays roles in the inflammation that contributes to CCA carcinogenesis and progression and may serve as a marker for a poor prognosis of CCA. © 2014 Japanese Society of Hepato-Biliary-Pancreatic Surgery.

Stat3 phosphorylation is required for embryonic stem cells ground state maintenance in 2i culture media.

PubMed

Wang, Dan; Sang, Hui; Zhang, Kaiyue; Nie, Yan; Zhao, Shuang; Zhang, Yan; He, Ningning; Wang, Yuebing; Xu, Yang; Xie, Xiaoyan; Li, Zongjin; Liu, Na

2017-05-09

Embryonic stem cells (ES cells) can be maintained its undifferentiated state with feeder cells or LIF, which can activate Jak/Stat3 pathway. Recently, it has been reported a new culture condition comprising serum-free medium with ERK and GSK3β inhibitors (2i) could drive ES cells into a state of pluripotency more like inner cell mass (ICM) in mouse blastocysts called ground state. However, although 2i could sustain ES cells self-renewal, LIF is routinely added. The roles of Stat3 activation are still unclear now. Here we investigated whether Jak/Stat3 might also contribute to the induction of ground state pluripotency. We introduced a lentiviral construct with 7-repeat Stat3-binding sequence to drive Renilla luciferase into ES cells, which can be used as a reporter to detect Stat3 activation by noninvasive bioluminescence imaging. Using this ES cells, we investigated the role of Stat3 activation in ground state maintenance. The results showed that Stat3 could be activated by 2i. Stattic, a chemical inhibitor of Stat3 phosphorylation, could effectively inhibit Stat3 activation in ES cells. When Stat3 activation was suppressed, ground state related genes were down regulated, and ES cells could not be maintained the ground state pluripotency even in 2i medium. All of these results indicate Stat3 activation is required in ground state maintenance.

Screening Active Compounds from Garcinia Species Native to China Reveals Novel Compounds Targeting the STAT/JAK Signaling Pathway

PubMed Central

Xu, Linfeng; Lao, Yuanzhi; Zhao, Yanhui; Qin, Jian; Fu, Wenwei; Zhang, Yingjia; Xu, Hongxi

2015-01-01

Natural compounds from medicinal plants are important resources for drug development. In a panel of human tumor cells, we screened a library of the natural products from Garcinia species which have anticancer potential to identify new potential therapeutic leads and discovered that caged xanthones were highly effective at suppressing multiple cancer cell lines. Their anticancer activities mainly depended on apoptosis pathways. For compounds in sensitive cancer line, their mechanisms of mode of action were evaluated. 33-Hydroxyepigambogic acid and 35-hydroxyepigambogic acid exhibited about 1 μM IC50 values against JAK2/JAK3 kinases and less than 1 μM IC50 values against NCI-H1650 cell which autocrined IL-6. Thus these two compounds provided a new antitumor molecular scaffold. Our report describes 33-hydroxyepigambogic acid and 35-hydroxyepigambogic acid that inhibited NCI-H1650 cell growth by suppressing constitutive STAT3 activation via direct inhibition of JAK kinase activity. PMID:26090459

Expression and activation of STAT3 in ischemia-induced retinopathy.

PubMed

Mechoulam, Hadas; Pierce, Eric A

2005-12-01

Signal transducer and activator of transcription protein-3 (STAT3) is a transcription factor that participates in many biological processes, including tumor angiogenesis. The expression and activation of Stat3 in the mouse model of ischemia-induced retinal neovascularization was investigated to evaluate the possible role of STAT3 in retinal vascular disease. Retinal neovascularization was induced in mice pups by exposure to hyperoxia. Gene microarrays were used to identify genes whose expression in the retina is altered at postnatal day (P)12 and P18. The relative levels of Stat3 mRNA were determined by semiquantitative RT-PCR. Stat3 protein levels and the levels of the activated form of Stat3 (pStat3) at P12, P15, P18, and P22 were determined by immunoblot analysis. Stat3 and pStat3 were demonstrated by immunofluorescence in retinal sections at P12, P15, and P18. In a series of microarray experiments, increased Stat3 mRNA levels in the retina were detected at P18. This result was validated by RT-PCR and demonstrated that Stat3 and pStat3 protein levels also increase during the development of neovascularization. Stat3 partially colocalized with blood vessels at the peak of neovascularization. pStat3 colocalized completely with blood vessels in both experimental samples and age-matched controls. pStat3 staining increased notably in the neovascular vessels at P15 and P18 and was more strongly associated with the epiretinal vessels than with inner retinal vessels. It was not detected in larger blood vessels, such as those of the optic nerve. The level of Stat3 expression increased, and pStat3 was observed in association with retinal neovascularization. Activated Stat3 was preferentially localized to neovascular retinal vessels. These data suggest that STAT3 may have a role in proliferative retinopathy.

Gain-of-function STAT1 mutations impair STAT3 activity in patients with chronic mucocutaneous candidiasis (CMC).

PubMed

Zheng, Jie; van de Veerdonk, Frank L; Crossland, Katherine L; Smeekens, Sanne P; Chan, Chun M; Al Shehri, Tariq; Abinun, Mario; Gennery, Andrew R; Mann, Jelena; Lendrem, Dennis W; Netea, Mihai G; Rowan, Andrew D; Lilic, Desa

2015-10-01

Signal transducer and activator of transcription 3 (STAT3) triggered production of Th-17 cytokines mediates protective immunity against fungi. Mutations affecting the STAT3/interleukin 17 (IL-17) pathway cause selective susceptibility to fungal (Candida) infections, a hallmark of chronic mucocutaneous candidiasis (CMC). In patients with autosomal dominant CMC, we and others previously reported defective Th17 responses and underlying gain-of-function (GOF) STAT1 mutations, but how this affects STAT3 function leading to decreased IL-17 is unclear. We also assessed how GOF-STAT1 mutations affect STAT3 activation, DNA binding, gene expression, cytokine production, and epigenetic modifications. We excluded impaired STAT3 phosphorylation, nuclear translocation, and sequestration of STAT3 into STAT1/STAT3 heterodimers and confirm significantly reduced transcription of STAT3-inducible genes (RORC/IL-17/IL-22/IL-10/c-Fos/SOCS3/c-Myc) as likely underlying mechanism. STAT binding to the high affinity sis-inducible element was intact but binding to an endogenous STAT3 DNA target was impaired. Reduced STAT3-dependent gene transcription was reversed by inhibiting STAT1 activation with fludarabine or enhancing histone, but not STAT1 or STAT3 acetylation with histone deacetylase (HDAC) inhibitors trichostatin A or ITF2357. Silencing HDAC1, HDAC2, and HDAC3 indicated a role for HDAC1 and 2. Reduced STAT3-dependent gene transcription underlies low Th-17 responses in GOF-STAT1 CMC, which can be reversed by inhibiting acetylation, offering novel targets for future therapies. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Dysregulation of janus kinases and signal transducers and activators of transcription in cancer

PubMed Central

Costa-Pereira, Ana P; Bonito, Nair A; Seckl, Michael J

2011-01-01

Despite their long recognised pivotal roles in immunological responses, Janus kinases (JAKs) and signal transducers and activators of transcription (STATs) are now seen as important players in cancer development and progression. Indeed, mutations in the JAKs are often found in myeloproliferative disorders (MPDs) and leukaemia, and the constitutive phosphorylation of STATs is a common occurrence in many solid and blood cancer cell lines and primary tumour specimens. More recently, we have also shown that JAKs likely have additional roles in promoting drug resistance in several cancer cell types. JAKs and STATs are thus molecules that may serve as useful targets in the clinic. This review will summarise studies that support this notion. PMID:22016828

Bortezomib inhibits STAT5-dependent degradation of LEF-1, inducing granulocytic differentiation in congenital neutropenia CD34+ cells

PubMed Central

Gupta, Kshama; Kuznetsova, Inna; Klimenkova, Olga; Klimiankou, Maksim; Meyer, Johann; Moore, Malcolm A. S.; Zeidler, Cornelia; Welte, Karl

2014-01-01

The transcription factor lymphoid enhancer–binding factor 1 (LEF-1), which plays a definitive role in granulocyte colony-stimulating factor (G-CSF) receptor-triggered granulopoiesis, is downregulated in granulocytic progenitors of severe congenital neutropenia (CN) patients. However, the exact mechanism of LEF-1 downregulation is unclear. CN patients are responsive to therapeutically high doses of G-CSF and are at increased risk of developing acute myeloid leukemia. The normal expression of LEF-1 in monocytes and lymphocytes, whose differentiation is unaffected in CN, suggests the presence of a granulopoiesis-specific mechanism downstream of G-CSF receptor signaling that leads to LEF-1 downregulation. Signal transducer and activator of transcription 5 (STAT5) is activated by G-CSF and is hyperactivated in acute myeloid leukemia. Here, we investigated the effects of activated STAT5 on LEF-1 expression and functions in hematopoietic progenitor cells. We demonstrated that constitutively active STAT5a (caSTAT5a) inhibited LEF-1–dependent autoregulation of the LEF-1 gene promoter by binding to the LEF-1 protein, recruiting Nemo-like kinase and the E3 ubiquitin-ligase NARF to LEF-1, leading to LEF-1 ubiquitination and a reduction in LEF-1 protein levels. The proteasome inhibitor bortezomib reversed the defective G-CSF–triggered granulocytic differentiation of CD34+ cells from CN patients in vitro, an effect that was accompanied by restoration of LEF-1 protein levels and LEF-1 messenger RNA autoregulation. Taken together, our data define a novel mechanism of LEF-1 downregulation in CN patients via enhanced ubiquitination and degradation of LEF-1 protein by hyperactivated STAT5. PMID:24394665

STAT3 activation is associated with cerebrospinal fluid interleukin-10 (IL-10) in primary central nervous system diffuse large B cell lymphoma.

PubMed

Mizowaki, Takashi; Sasayama, Takashi; Tanaka, Kazuhiro; Mizukawa, Katsu; Takata, Kumi; Nakamizo, Satoshi; Tanaka, Hirotomo; Nagashima, Hiroaki; Nishihara, Masamitsu; Hirose, Takanori; Itoh, Tomoo; Kohmura, Eiji

2015-09-01

Signal transducers and activators of transcription 3 (STAT3) are activated by various cytokines and oncogenes; however, the activity and pathogenesis of STAT3 in diffuse large B cell lymphoma of the central nervous system have not been thoroughly elucidated. We investigated the phosphorylation levels of STAT3 in 40 specimens of primary central nervous system diffuse large B-cell lymphoma (PCNS DLBCL) and analyzed the association between phsopho-STAT3 (pSTAT3) expression and cerebrospinal fluid (CSF) concentration of interleukin-10 (IL-10) or IL-6. Immunohistochemistry and Western blot analysis revealed that most of the specimens in PCNS DLBCL expressed pSTST3 protein, and a strong phosphorylation levels of STAT3 was statistically associated with high CSF IL-10 levels, but not with CSF IL-6 levels. Next, we demonstrated that recombinant IL-10 and CSF containing IL-10 induced the phosphorylation of STAT3 in PCNS DLBCL cells. Furthermore, molecular subtype classified by Hans' algorithm was correlated with pSTAT3 expression levels and CSF IL-10 levels. These results suggest that the STAT3 activity is correlated with CSF IL-10 level, which is a useful marker for STAT3 activity in PCNS DLBCLs.

Activated Rac1 requires gp130 for Stat3 activation, cell proliferation and migration

DOE Office of Scientific and Technical Information (OSTI.GOV)

Arulanandam, Rozanne; Geletu, Mulu; Feracci, Helene

2010-03-10

Rac1 (Rac) is a member of the Rho family of small GTPases which controls cell migration by regulating the organization of actin filaments. Previous results suggested that mutationally activated forms of the Rho GTPases can activate the Signal Transducer and Activator of Transcription-3 (Stat3), but the exact mechanism is a matter of controversy. We recently demonstrated that Stat3 activity of cultured cells increases dramatically following E-cadherin engagement. To better understand this pathway, we now compared Stat3 activity levels in mouse HC11 cells before and after expression of the mutationally activated Rac1 (Rac{sup V12}), at different cell densities. The results revealedmore » for the first time a dramatic increase in protein levels and activity of both the endogenous Rac and Rac{sup V12} with cell density, which was due to inhibition of proteasomal degradation. In addition, Rac{sup V12}-expressing cells had higher Stat3, tyrosine-705 phosphorylation and activity levels at all densities, indicating that Rac{sup V12} is able to activate Stat3. Further examination of the mechanism of Stat3 activation showed that Rac{sup V12} expression caused a surge in mRNA of Interleukin-6 (IL6) family cytokines, known potent Stat3 activators. Knockdown of gp130, the common subunit of this family reduced Stat3 activity, indicating that these cytokines may be responsible for the Stat3 activation by Rac{sup V12}. The upregulation of IL6 family cytokines was required for cell migration and proliferation induced by Rac{sup V12}, as shown by gp130 knockdown experiments, thus demonstrating that the gp130/Stat3 axis represents an essential effector of activated Rac for the regulation of key cellular functions.« less

Multifunctional Effects of a Small-Molecule STAT3 Inhibitor on NASH and Hepatocellular Carcinoma in Mice.

PubMed

Jung, Kwang Hwa; Yoo, Wonbeak; Stevenson, Heather L; Deshpande, Dipti; Shen, Hong; Gagea, Mihai; Yoo, Suk-Young; Wang, Jing; Eckols, T Kris; Bharadwaj, Uddalak; Tweardy, David J; Beretta, Laura

2017-09-15

Purpose: The incidence of hepatocellular carcinoma is increasing in the United States, and liver cancer is the second leading cause of cancer-related mortality worldwide. Nonalcoholic steatohepatitis (NASH) is becoming an important risk for hepatocellular carcinoma, and most patients with hepatocellular carcinoma have underlying liver cirrhosis and compromised liver function, which limit treatment options. Thus, novel therapeutic strategies to prevent or treat hepatocellular carcinoma in the context of NASH and cirrhosis are urgently needed. Experimental Design: Constitutive activation of STAT3 is frequently detected in hepatocellular carcinoma tumors. STAT3 signaling plays a pivotal role in hepatocellular carcinoma survival, growth, angiogenesis, and metastasis. We identified C188-9, a novel small-molecule STAT3 inhibitor using computer-aided rational drug design. In this study, we evaluated the therapeutic potential of C188-9 for hepatocellular carcinoma treatment and prevention. Results: C188-9 showed antitumor activity in vitro in three hepatocellular carcinoma cell lines. In mice with hepatocyte-specific deletion of Pten (Hep Pten - mice), C188-9 treatment blocked hepatocellular carcinoma tumor growth, reduced tumor development, and reduced liver steatosis, inflammation, and bile ductular reactions, resulting in improvement of the pathological lesions of NASH. Remarkably, C188-9 also greatly reduced liver injury in these mice as measured by serum aspartate aminotransferase and alanine transaminase levels. Analysis of gene expression showed that C188-9 treatment of Hep Pten - mice resulted in inhibition of signaling pathways downstream of STAT3, STAT1, TREM-1, and Toll-like receptors. In contrast, C188-9 treatment increased liver specification and differentiation gene pathways. Conclusions: Our results suggest that C188-9 should be evaluated further for the treatment and/or prevention of hepatocellular carcinoma. Clin Cancer Res; 23(18); 5537-46. ©2017 AACR . ©2017 American Association for Cancer Research.

Activation of Stat3 in renal tumors.

PubMed

Guo, Charles; Yang, Guanyu; Khun, Kyle; Kong, Xiantian; Levy, David; Lee, Peng; Melamed, Jonathan

2009-02-28

Signal transducer and activator of transcription 3 (Stat3) plays a vital role in signal transduction pathways that mediate transformation and inhibit apoptosis. Oncogenic Stat3 is persistently activated in several human cancers and transformed cell lines. Previous studies indicate activation of Stat3 in renal cell carcinoma (RCC). However, the detailed characterization of the Stat3 expression pattern in different histologic types of RCC is lacking. We have analyzed the immunoprofile of activated or phosphorylated Stat3 (pStat3) in a tissue microarray of renal tumors of different histologic types, including 42 cases of conventional clear cell type, 24 chromophobe, and 7 papillary, 15 oncocytoma, 7 urothelial carcinoma and 21 normal kidney tissues using an anti-pStat3 antibody (recognizes only activated STAT3). pStat3 nuclear staining was observed in 25 of 42 conventional clear cell RCC (59.5 %), 8 of 24 chromophobe RCC (33.3%), 4 of 7 papillary RCC (57.1%). In the other tumor groups, 4 of 15 oncocytomas (26.7%) and 6 of 7 urothelial carcinomas (85.7%) showed positive nuclear staining. Weak nuclear immunoreactivity for pStat3 was seen in 4 of 21 cases of non-neoplastic kidney tissue (19.0%). The extent of Stat3 activation as determined by nuclear expression of its phosphorylated form is increased in histologic types of renal tumors with greater malignant potential, specifically conventional clear cell RCC, papillary RCC and urothelial carcinoma, only slightly increased in chromophobe RCC, and not increased in oncocytoma. These results suggest a role of Stat3 activation in different types of renal neoplasia, possibly serving as a prognostic marker or therapeutic target.

Activation of Stat3 in renal tumors

PubMed Central

Guo, Charles; Yang, Guanyu; Khun, Kyle; Kong, Xiantian; Levy, David; Lee, Peng; Melamed, Jonathan

2009-01-01

Signal transducer and activator of transcription 3 (Stat3) plays a vital role in signal transduction pathways that mediate transformation and inhibit apoptosis. Oncogenic Stat3 is persistently activated in several human cancers and transformed cell lines. Previous studies indicate activation of Stat3 in renal cell carcinoma (RCC). However, the detailed characterization of the Stat3 expression pattern in different histologic types of RCC is lacking. We have analyzed the immunoprofile of activated or phosphorylated Stat3 (pStat3) in a tissue microarray of renal tumors of different histologic types, including 42 cases of conventional clear cell type, 24 chromophobe, and 7 papillary, 15 oncocytoma, 7 urothelial carcinoma and 21 normal kidney tissues using an anti-pStat3 antibody (recognizes only activated STAT3). pStat3 nuclear staining was observed in 25 of 42 conventional clear cell RCC (59.5 %), 8 of 24 chromophobe RCC (33.3%), 4 of 7 papillary RCC (57.1%). In the other tumor groups, 4 of 15 oncocytomas (26.7%) and 6 of 7 urothelial carcinomas (85.7%) showed positive nuclear staining. Weak nuclear immunoreactivity for pStat3 was seen in 4 of 21 cases of non-neoplastic kidney tissue (19.0%). The extent of Stat3 activation as determined by nuclear expression of its phosphorylated form is increased in histologic types of renal tumors with greater malignant potential, specifically conventional clear cell RCC, papillary RCC and urothelial carcinoma, only slightly increased in chromophobe RCC, and not increased in oncocytoma. These results suggest a role of Stat3 activation in different types of renal neoplasia, possibly serving as a prognostic marker or therapeutic target. PMID:19956438

Tracking STAT nuclear traffic.

PubMed

Reich, Nancy C; Liu, Ling

2006-08-01

Accurate cellular localization is crucial for the effective function of most signalling molecules and nuclear translocation is central to the function of transcription factors. The passage of large molecules between the cytoplasm and nucleus is restricted, and this restriction affords a mechanism to regulate transcription by controlling the access of transcription factors to the nucleus. In this Review, we focus on the signal transducer and activator of transcription (STAT) family of transcription factors. The regulation of the nuclear trafficking of STAT-family members is diverse. Some STAT proteins constitutively shuttle between the nucleus and cytoplasm, whereas others require tyrosine phosphorylation for nuclear localization. In either case, the regulation of nuclear trafficking can provide a target for therapeutic intervention.

Sequential activation of JAKs, STATs and xanthine dehydrogenase/oxidase by hypoxia in lung microvascular endothelial cells.

PubMed

Wang, Guansong; Qian, Pin; Jackson, Fannie R; Qian, Guisheng; Wu, Guangyu

2008-01-01

Xanthine dehydrogenase/oxidase (XDH/XO) is associated with various pathological conditions related to the endothelial injury. However, the molecular mechanism underlying the activation of XDH/XO by hypoxia remains largely unknown. In this report, we determined whether the Janus kinases (JAKs) and signal transducers and activators of transcription (STATs) signaling pathway is involved in hypoxia-induced activation of XDH/XO in primary cultures of lung microvascular endothelial cells (LMVEC). We found that hypoxia significantly increased interleukin 6 (IL6) production in a time-dependent manner in LMVEC. Hypoxia also markedly augmented phosphorylation/activation of JAKs (JAK1, JAK2 and JAK3) and the JAK downstream effectors STATs (STAT3 and STAT5). Hypoxia-induced activation of STAT3 was blocked by IL6 antibodies, the JAK inhibitor AG490 and the suppressor of cytokine signaling 3 (SOCS3), implying that hypoxia-promoted IL6 secretion activates the JAK/STAT pathway in LMVEC. Phosphorylation and DNA-binding activity of STAT3 were also inhibited by the p38 MAPK inhibitor SB203580 and the phosphatidylinositol 3-kinase inhibitor LY294002, suggesting that multiple signaling pathways involved in STAT activation by hypoxia. Importantly, hypoxia promoted XDH/XO activation in LMVEC, which was markedly reversed by inhibiting the JAK-STAT pathway using IL6 antibodies, AG490 and SOCS3. These data demonstrated that JAKs, STATs and XDH/XO were sequentially activated by hypoxia. These data provide the first evidence indicating that the JAK-STAT pathway is involved in hypoxia-mediated XDH/XO activation in LMVEC.

NF-κB-Induced IL-6 Ensures STAT3 Activation and Tumor Aggressiveness in Glioblastoma

PubMed Central

McFarland, Braden C.; Hong, Suk W.; Rajbhandari, Rajani; Twitty, George B.; Gray, G. Kenneth; Yu, Hao; Benveniste, Etty N.; Nozell, Susan E.

2013-01-01

Glioblastoma (GBM) is the most aggressive, neurologically destructive and deadly tumor of the central nervous system (CNS). In GBM, the transcription factors NF-κB and STAT3 are aberrantly activated and associated with tumor cell proliferation, survival, invasion and chemoresistance. In addition, common activators of NF-κB and STAT3, including TNF-α and IL-6, respectively, are abundantly expressed in GBM tumors. Herein, we sought to elucidate the signaling crosstalk that occurs between the NF-κB and STAT3 pathways in GBM tumors. Using cultured GBM cell lines as well as primary human GBM xenografts, we elucidated the signaling crosstalk between the NF-κB and STAT3 pathways utilizing approaches that either a) reduce NF-κB p65 expression, b) inhibit NF-κB activation, c) interfere with IL-6 signaling, or d) inhibit STAT3 activation. Using the clinically relevant human GBM xenograft model, we assessed the efficacy of inhibiting NF-κB and/or STAT3 alone or in combination in mice bearing intracranial xenograft tumors in vivo. We demonstrate that TNF-α-induced activation of NF-κB is sufficient to induce IL-6 expression, activate STAT3, and elevate STAT3 target gene expression in GBM cell lines and human GBM xenografts in vitro. Moreover, the combined inhibition of NF-κB and STAT3 signaling significantly increases survival of mice bearing intracranial tumors. We propose that in GBM, the activation of NF-κB ensures subsequent STAT3 activation through the expression of IL-6. These data verify that pharmacological interventions to effectively inhibit the activity of both NF-κB and STAT3 transcription factors must be used in order to reduce glioma size and aggressiveness. PMID:24244348

NF-κB-induced IL-6 ensures STAT3 activation and tumor aggressiveness in glioblastoma.

PubMed

McFarland, Braden C; Hong, Suk W; Rajbhandari, Rajani; Twitty, George B; Gray, G Kenneth; Yu, Hao; Benveniste, Etty N; Nozell, Susan E

2013-01-01

Glioblastoma (GBM) is the most aggressive, neurologically destructive and deadly tumor of the central nervous system (CNS). In GBM, the transcription factors NF-κB and STAT3 are aberrantly activated and associated with tumor cell proliferation, survival, invasion and chemoresistance. In addition, common activators of NF-κB and STAT3, including TNF-α and IL-6, respectively, are abundantly expressed in GBM tumors. Herein, we sought to elucidate the signaling crosstalk that occurs between the NF-κB and STAT3 pathways in GBM tumors. Using cultured GBM cell lines as well as primary human GBM xenografts, we elucidated the signaling crosstalk between the NF-κB and STAT3 pathways utilizing approaches that either a) reduce NF-κB p65 expression, b) inhibit NF-κB activation, c) interfere with IL-6 signaling, or d) inhibit STAT3 activation. Using the clinically relevant human GBM xenograft model, we assessed the efficacy of inhibiting NF-κB and/or STAT3 alone or in combination in mice bearing intracranial xenograft tumors in vivo. We demonstrate that TNF-α-induced activation of NF-κB is sufficient to induce IL-6 expression, activate STAT3, and elevate STAT3 target gene expression in GBM cell lines and human GBM xenografts in vitro. Moreover, the combined inhibition of NF-κB and STAT3 signaling significantly increases survival of mice bearing intracranial tumors. We propose that in GBM, the activation of NF-κB ensures subsequent STAT3 activation through the expression of IL-6. These data verify that pharmacological interventions to effectively inhibit the activity of both NF-κB and STAT3 transcription factors must be used in order to reduce glioma size and aggressiveness.

STAT4 deficiency reduces the development of atherosclerosis in mice.

PubMed

Taghavie-Moghadam, Parésa L; Gjurich, Breanne N; Jabeen, Rukhsana; Krishnamurthy, Purna; Kaplan, Mark H; Dobrian, Anca D; Nadler, Jerry L; Galkina, Elena V

2015-11-01

Atherosclerosis is a chronic inflammatory process that leads to plaque formation in large and medium sized vessels. T helper 1 (Th1) cells constitute the majority of plaque infiltrating pro-atherogenic T cells and are induced via IFNγ-dependent activation of T-box (Tbet) and/or IL-12-dependent activation of signal transducer and activator of transcription 4 (STAT4). We thus aimed to define a role for STAT4 in atherosclerosis. STAT4-deficiency resulted in a ∼71% reduction (p < 0.001) in plaque burden in Stat4(-/-)Apoe(-/-) vs Apoe(-/-) mice fed chow diet and significantly attenuated atherosclerosis (∼31%, p < 0.01) in western diet fed Stat4(-/-)Apoe(-/-) mice. Surprisingly, reduced atherogenesis in Stat4(-/-)Apoe(-/-) mice was not due to attenuated IFNγ production in vivo by Th1 cells, suggesting an at least partially IFNγ-independent pro-atherogenic role of STAT4. STAT4 is expressed in T cells, but also detected in macrophages (MΦs). Stat4(-/-)Apoe(-/-)in vitro differentiated M1 or M2 MΦs had reduced cytokine production compare to Apoe(-/-) M1 and M2 MΦs that was accompanied by reduced induction of CD69, I-A(b), and CD86 in response to LPS stimulation. Stat4(-/-)Apoe(-/-) MΦs expressed attenuated levels of CCR2 and demonstrated reduced migration toward CCL2 in a transwell assay. Importantly, the percentage of aortic CD11b(+)F4/80(+)Ly6C(hi) MΦs was reduced in Stat4(-/-)Apoe(-/-) vs Apoe(-/-) mice. Thus, this study identifies for the first time a pro-atherogenic role of STAT4 that is at least partially independent of Th1 cell-derived IFNγ, and primarily involving the modulation of MΦ responses. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Epstein-Barr virus-derived EBNA2 regulates STAT3 activation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Muromoto, Ryuta; Ikeda, Osamu; Okabe, Kanako

The Epstein-Barr virus (EBV)-encoded latency protein EBNA2 is a nuclear transcriptional activator that is essential for EBV-induced cellular transformation. Here, we show that EBNA2 interacts with STAT3, a signal transducer for an interleukin-6 family cytokine, and enhances the transcriptional activity of STAT3 by influencing its DNA-binding activity. Furthermore, EBNA2 cooperatively acts on STAT3 activation with LMP1. These data demonstrate that EBNA2 acts as a transcriptional coactivator of STAT3.

Signal transducer and activator of transcription-3 (Stat3) plays a critical role in implantation via progesterone receptor in uterus

PubMed Central

Lee, Jae Hee; Kim, Tae Hoon; Oh, Seo Jin; Yoo, Jung-Yoon; Akira, Shizuo; Ku, Bon Jeong; Lydon, John P.; Jeong, Jae-Wook

2013-01-01

Recent studies have shown that activation of the signal transducer and activator of transcription-3 (Stat3) is required for decidualization, interacting with progesterone receptor (PR) in uterus. Based on previous reports, we hypothesized that crosstalk between STAT3 and PR signaling is required for successful implantation. To identify the interaction between STAT3 and PR isoforms, we performed immunoprecipitation following transient cotransfection and found that STAT3 physically interacted with PR-A, which is known to be important for uterine development and function, but not with PR-B. To further investigate the role of Stat3 in uterine function, Stat3 was conditionally ablated only in the PR-positive cells (PRcre/+ Stat3f/f; Stat3d/d). Our studies revealed that ovarian function and uterine development of Stat3d/d mice were normal. However, Stat3d/d female mice were infertile due to defective embryo implantation. Unlike Stat3f/f mice, Stat3d/d mice exhibited an unclosed uterine lumen. Furthermore, uteri of Stat3d/d mice were unable to undergo a well-characterized hormonally induced decidual reaction. The expression of stromal PR was decreased during decidualization and preimplantation period in Stat3d/d mice, and PR target genes were significantly down-regulated after progesterone induction. Our results suggest that STAT3 and PR crosstalk is required for successful implantation in the mouse uterus.—Lee, J. H., Kim, T. H., Oh, S. J., Yoo, J.-Y., Akira, S., Ku, B. J., Lydon, J. P., Jeong, J.-W. Signal transducer and activator of transcription-3 (Stat3) plays a critical role in implantation via progesterone receptor in uterus. PMID:23531596

Gα73B is a downstream effector of JAK/STAT signalling and a regulator of Rho1 in Drosophila haematopoiesis.

PubMed

Bausek, Nina; Zeidler, Martin P

2014-01-01

JAK/STAT signalling regulates many essential developmental processes including cell proliferation and haematopoiesis, whereas its inappropriate activation is associated with the majority of myeloproliferative neoplasias and numerous cancers. Furthermore, high levels of JAK/STAT pathway signalling have also been associated with enhanced metastatic invasion by cancerous cells. Strikingly, gain-of-function mutations in the single Drosophila JAK homologue, Hopscotch, result in haemocyte neoplasia, inappropriate differentiation and the formation of melanised haemocyte-derived 'tumour' masses; phenotypes that are partly orthologous to human gain-of-function JAK2-associated pathologies. Here we show that Gα73B, a novel JAK/STAT pathway target gene, is necessary for JAK/STAT-mediated tumour formation in flies. In addition, although Gα73B does not affect haemocyte differentiation, it does regulate haemocyte morphology and motility under non-pathological conditions. We show that Gα73B is required for constitutive, but not injury-induced, activation of Rho1 and for the localisation of Rho1 into filopodia upon haemocyte activation. Consistent with these results, we also show that Rho1 interacts genetically with JAK/STAT signalling, and that wild-type levels of Rho1 are necessary for tumour formation. Our findings link JAK/STAT transcriptional outputs, Gα73B activity and Rho1-dependent cytoskeletal rearrangements and cell motility, therefore connecting a pathway associated with cancer with a marker indicative of invasiveness. As such, we suggest a mechanism by which JAK/STAT pathway signalling may promote metastasis.

STATs in NK-Cells: The Good, the Bad, and the Ugly.

PubMed

Gotthardt, Dagmar; Sexl, Veronika

2016-01-01

Natural killer (NK)-cells are major players in the fight against viral infections and transformed cells, but there is increasing evidence attributing a disease-promoting role to NK-cells. Cytokines present in the tumor microenvironment shape NK-cell maturation, function, and effector responses. Many cytokines signal via the Janus kinase (JAK)-signal transducer and activator of transcription (STAT) pathway that is also frequently altered and constitutively active in a broad range of tumor cells. As a consequence, there are currently major efforts to develop therapeutic strategies to target this pathway. Therefore, it is of utmost importance to understand the role and contributions of JAK-STAT molecules in NK-cell biology-only this knowledge will allow us to predict effects of JAK-STAT inhibition for NK-cell functions and to successfully apply precision medicine. We will review the current knowledge on the role of JAK-STAT signaling for NK-cell functions and discuss conditions involved in the switch from NK-cell tumor surveillance to disease promotion.

Altered expression and activation of signal transducers and activators of transcription (STATs) in hepatitis C virus infection: in vivo and in vitro studies.

PubMed

Larrea, E; Aldabe, R; Molano, E; Fernandez-Rodriguez, C M; Ametzazurra, A; Civeira, M P; Prieto, J

2006-08-01

Signal transducers and activators of transcription (STATs) play a critical role in antiviral defence. STAT3 is also important in cell protection against inflammatory damage. STAT proteins are activated by interferons and by hepatoprotective cytokines of the interleukin 6 superfamily, including cardiotrophin 1. We analysed the status of STATs in hepatitis C virus (HCV) infected livers and the relationship between expression and activation of STATs and HCV replication in Huh7 cells transfected with HCV genomic replicon. STAT3alpha expression was reduced in HCV infected livers showing an inverse correlation with serum alanine aminotransferase. In patients with HCV infection, nuclear staining for phosphorylated STAT3 was faint in parenchymal cells (although conspicuous in infiltrating leucocytes), in contrast with strong nuclear staining in hepatocytes from control livers. Expression and activation of STAT1 (a factor activated by both interferon (IFN)-alpha and IFN-gamma) were increased in HCV infected livers, particularly in those with high inflammatory activity. Conversely, phosphorylated STAT2 (a factor selectively activated by IFN-alpha) was undetectable in livers with HCV infection, a finding that was associated with marked downregulation of the two functional subunits of the IFN-alpha receptor. HCV replication in Huh7 cells caused STAT3alpha downregulation and blocked STAT3 phosphorylation by either IFN-alpha or cardiotrophin 1. HCV replication in Huh7 cells also inhibited STAT1 and STAT2 activation by IFN-alpha while there was no impairment of STAT1 phosphorylation by the proinflammatory cytokine IFN-gamma. STAT3 is downregulated in HCV infected livers and in Huh7 cells bearing the full length HCV replicon. HCV replication is associated with impaired Jak-STAT signalling by antiviral and cytoprotective cytokines. These effects may favour viral replication while facilitating the progression of liver disease.

Cross-talk between KLF4 and STAT3 regulates axon regeneration

NASA Astrophysics Data System (ADS)

Qin, Song; Zou, Yuhua; Zhang, Chun-Li

2013-10-01

Cytokine-induced activation of signal transducer and activator of transcription 3 (STAT3) promotes the regrowth of damaged axons in the adult central nervous system (CNS). Here we show that KLF4 physically interacts with STAT3 upon cytokine-induced phosphorylation of tyrosine 705 (Y705) on STAT3. This interaction suppresses STAT3-dependent gene expression by blocking its DNA-binding activity. The deletion of KLF4 in vivo induces axon regeneration of adult retinal ganglion cells (RGCs) via Janus kinase (JAK)-STAT3 signalling. This regeneration can be greatly enhanced by exogenous cytokine treatment, or removal of an endogenous JAK-STAT3 pathway inhibitor called suppressor of cytokine signalling 3 (SOCS3). These findings reveal an unexpected cross-talk between KLF4 and activated STAT3 in the regulation of axon regeneration that might have therapeutic implications in promoting repair of injured adult CNS.

STAT6 is a cargo of exportin 1: Biological relevance in primary mediastinal B-cell lymphoma.

PubMed

Miloudi, Hadjer; Leroy, Karen; Jardin, Fabrice; Sola, Brigitte

2018-06-01

Primary mediastinal B-cell lymphoma (PMBL) is a distinct B-cell lymphoma subtype with unique clinicopathological and molecular features. PMBL cells are characterised by several genetic abnormalities that conduct to the constitutive activation of the Janus kinase 2/signal transducer and activator of transcription 6 (JAK2/STAT6) signalling pathway. Among recurrent genetic changes in PMBL, we previously reported that the XPO1 gene encoding exportin 1 that controls the nuclear export of cargo proteins and RNAs, is mutated (p.E571K) in about 25% of PMBL cases. We therefore hypothesized that STAT6 could be a cargo of XPO1 and that STAT6 cytoplasm/nucleus shuttle could be altered in a subset of PMBL cells. Using immunocytochemistry techniques as well as the proximity ligation assay, we showed that STAT6 bound XPO1 in PBML cell lines and in HEK-293 cells genetically engineered to produce STAT6. Moreover, XPO1-mediated export of STAT6 occurs in cells expressing either a wild-type or the E571K mutated XPO1 protein. Copyright © 2018 Elsevier Inc. All rights reserved.

Tissue-Specific Autoregulation of the stat3 Gene and Its Role in Interleukin-6-Induced Survival Signals in T Cells

PubMed Central

Narimatsu, Masahiro; Maeda, Hisoka; Itoh, Shousaku; Atsumi, Toru; Ohtani, Takuya; Nishida, Keigo; Itoh, Motoyuki; Kamimura, Daisuke; Park, Sung-Joo; Mizuno, Katsunori; Miyazaki, Jun-ichi; Hibi, Masahiko; Ishihara, Katsuhiko; Nakajima, Koichi; Hirano, Toshio

2001-01-01

Signal transducer and activator of transcription 3 (STAT3) mediates signals of various growth factors and cytokines, including interleukin-6 (IL-6). In certain IL-6-responsive cell lines, the stat3 gene is autoregulated by STAT3 through a composite IL-6 response element in its promoter that contains a STAT3-binding element (SBE) and a cyclic AMP-responsive element. To reveal the nature and roles of the stat3 autoregulation in vivo, we generated mice that harbor a mutation in the SBE (stat3mSBE). The intact SBE was crucial for IL-6-induced stat3 gene activation in the spleen, especially in the red pulp region, the kidney, and both mature and immature T lymphocytes. The SBE was not required, however, for IL-6-induced stat3 gene activation in hepatocytes. T lymphocytes from the stat3mSBE/mSBE mice were more susceptible to apoptosis despite the presence of IL-6 than those from wild-type mice. Consistent with this, IL-6-dependent activation of the Pim-1 and junB genes, direct target genes for STAT3, was attenuated in T lymphocytes of the stat3mSBE/mSBE mice. Thus, the tissue-specific autoregulation of the stat3 gene operates in vivo and plays a role in IL-6-induced antiapoptotic signaling in T cells. PMID:11533249

Benzoxathiol derivative BOT-4-one suppresses L540 lymphoma cell survival and proliferation via inhibition of JAK3/STAT3 signaling

PubMed Central

Kim, Byung Hak; Min, Yun Sook; Choi, Jung Sook; Baeg, Gyeong-Hun; Kim, Youngsoo; Shin, Jong Wook; Kim, Tae-Yoon

2011-01-01

Persistently activated JAK/STAT3 signaling pathway plays a pivotal role in various human cancers including major carcinomas and hematologic tumors, and is implicated in cancer cell survival and proliferation. Therefore, inhibition of JAK/STAT3 signaling may be a clinical application in cancer therapy. Here, we report that 2-cyclohexylimino-6-methyl-6,7-dihydro-5H-benzo [1,3]oxathiol-4-one (BOT-4-one), a small molecule inhibitor of JAK/STAT3 signaling, induces apoptosis through inhibition of STAT3 activation. BOT-4-one suppressed cytokine (upd)-induced tyrosine phosphorylation and transcriptional activity of STAT92E, the sole Drosophila STAT homolog. Consequently, BOT-4-one significantly inhibited STAT3 tyrosine phosphorylation and expression of STAT3 downstream target gene SOCS3 in various human cancer cell lines, and its effect was more potent in JAK3-activated Hodgkin's lymphoma cell line than in JAK2-activated breast cancer and prostate cancer cell lines. In addition, BOT-4-one-treated Hodgkin's lymphoma cells showed decreased cell survival and proliferation by inducing apoptosis through down-regulation of STAT3 downstream target anti-apoptotic gene expression. These results suggest that BOT-4-one is a novel small molecule inhibitor of JAK3/STAT3 signaling and may have therapeutic potential in the treatment of human cancers harboring aberrant JAK3/STAT3 signaling, specifically Hodgkin's lymphoma. PMID:21499010

Benzoxathiol derivative BOT-4-one suppresses L540 lymphoma cell survival and proliferation via inhibition of JAK3/STAT3 signaling.

PubMed

Kim, Byung Hak; Min, Yun Sook; Choi, Jung Sook; Baeg, Gyeong Hun; Kim, Young Soo; Shin, Jong Wook; Kim, Tae Yoon; Ye, Sang Kyu

2011-05-31

Persistently activated JAK/STAT3 signaling pathway plays a pivotal role in various human cancers including major carcinomas and hematologic tumors, and is implicated in cancer cell survival and proliferation. Therefore, inhibition of JAK/STAT3 signaling may be a clinical application in cancer therapy. Here, we report that 2-cyclohexylimino-6-methyl-6,7-dihydro-5H-benzo [1,3]oxathiol-4-one (BOT-4-one), a small molecule inhibitor of JAK/STAT3 signaling, induces apoptosis through inhibition of STAT3 activation. BOT-4-one suppressed cytokine (upd)-induced tyrosine phosphorylation and transcriptional activity of STAT92E, the sole Drosophila STAT homolog. Consequently, BOT-4-one significantly inhibited STAT3 tyrosine phosphorylation and expression of STAT3 downstream target gene SOCS3 in various human cancer cell lines, and its effect was more potent in JAK3-activated Hodgkin's lymphoma cell line than in JAK2-activated breast cancer and prostate cancer cell lines. In addition, BOT-4-one-treated Hodgkin's lymphoma cells showed decreased cell survival and proliferation by inducing apoptosis through down-regulation of STAT3 downstream target anti-apoptotic gene expression. These results suggest that BOT-4-one is a novel small molecule inhibitor of JAK3/STAT3 signaling and may have therapeutic potential in the treatment of human cancers harboring aberrant JAK3/STAT3 signaling, specifically Hodgkin's lymphoma.

SIRT1 counteracted the activation of STAT3 and NF-κB to repress the gastric cancer growth.

PubMed

Lu, Juanjuan; Zhang, Liping; Chen, Xiang; Lu, Qiming; Yang, Yuxia; Liu, Jingping; Ma, Xin

2014-01-01

Sirtuin-1 (SIRT1) possesses apparently dual roles in regulation of tumor. Previous reports have documented the crosstalk between SIRT1 with signal transducer and activator of transcription 3 (STAT3) and nuclear factor kappa-B (NF-κB) signaling in leukemia, lymphoma and myeloma. In this study, the purpose was to survey the regulatory effects of SIRT1 on gastric cancer (GC) cells (AGS and MKN-45) and the relationships between SIRT1 and activation of STAT3 and NF-κB in GC cells. We found the SIRT1 activator (resveratrol RSV) contributed to the repression of viability and increase of senescence, which were rescued by SIRT1 inhibitor (nicotinamide NA) and SIRT1 depletion by CCK-8 assay and SA-β-gal assay respectively. Further study found SIRT1 activation (RSV supplement) not only inhibited the activation of STAT3 including STAT3 mRNA level, c-myc mRNA level phosphorylated STAT3 (pSTAT3) proteins and acetylizad STAT3 (acSTAT3) proteins, but also repression of pNF-κB p65 and acNF-κB p65. NA reversed the effects of RSV. In addition, either RSV or NA application could not change the cellular viability and senescence in MKN-45 cells with STAT3 knockdown or NF-κB knockdown. Overall, our findings suggested SIRT1 activation could induced the loss of viability and increases of senescence in GC in vitro. Moreover, our observations revealed SIRT1 displayed growth inhibitory activity in GC cells highly associated with causing repression of activation of STAT3 and NF-κB proteins via deacetylation.

Biologically active leptin-related synthetic peptides activate STAT3 via phosphorylation of ERK1/2 and PI-3K.

PubMed

Lin, Hung-Yun; Yang, Sheng-Huei; Tang, Heng-Yuan; Cheng, Guei-Yun; Davis, Paul J; Grasso, Patricia

2014-07-01

The effects of leptin-related synthetic peptides [d-Leu-4]-OB3 and OB3 on energy balance and glucose homeostasis in ob/ob and db/db mice have been confirmed. The molecular basis of these effects, however, remains unclear. In the present study, we examined the ability of these peptides to activate signal transduction pathways known to be involved in transduction of the leptin signal. In a specific and concentration-dependent manner, [d-Leu-4]-OB3 induced phosphorylation of ERK1/2, PI-3K, Ser-727 STAT3, and Tyr-705 of STAT3. OB3 also induced activation of STAT3 via phosphorylation of ERK1/2, STAT3 Ser-727, STAT3 Tyr-705 and PI-3K p85, but to a lesser degree. Using PD98059 and LY294002, specific inhibitors of MEK and PI-3K, respectively, we were able to identify the signal transduction pathways involved in peptide-induced STAT3 activation. [d-Leu-4]-OB3 induced serine phosphorylation of STAT3 primarily through activation of ERK1/2. Tyrosine phosphorylation of STAT3, however, was induced primarily through activation of PI-3K. Our data suggest that in db/db mice, [d-Leu-4]-OB3 binding to short isoforms of the leptin receptor induces intracellular signaling cascades which do not require OB-Rb activation. These signals may ultimately result in peptide effects on transcriptional and translational events associated with energy balance and glycemic regulation. In summary, we have shown for the first time that, similar to leptin, bioactive leptin-related synthetic peptide analogs activate STAT3 via phosphorylation of serine and tyrosine residues by multiple signal transduction pathways. Copyright © 2014 Elsevier Inc. All rights reserved.

Essential role for Stat5a/b in myeloproliferative neoplasms induced by BCR-ABL1 and JAK2V617F in mice

PubMed Central

Walz, Christoph; Ahmed, Wesam; Lazarides, Katherine; Betancur, Monica; Patel, Nihal; Hennighausen, Lothar; Zaleskas, Virginia M.

2012-01-01

STAT5 proteins are constitutively activated in malignant cells from many patients with leukemia, including the myeloproliferative neoplasms (MPNs) chronic myeloid leukemia (CML) and polycythemia vera (PV), but whether STAT5 is essential for the pathogenesis of these diseases is not known. In the present study, we used mice with a conditional null mutation in the Stat5a/b gene locus to determine the requirement for STAT5 in MPNs induced by BCR-ABL1 and JAK2V617F in retroviral transplantation models of CML and PV. Loss of one Stat5a/b allele resulted in a decrease in BCR-ABL1–induced CML-like MPN and the appearance of B-cell acute lymphoblastic leukemia, whereas complete deletion of Stat5a/b prevented the development of leukemia in primary recipients. However, BCR-ABL1 was expressed and active in Stat5-null leukemic stem cells, and Stat5 deletion did not prevent progression to lymphoid blast crisis or abolish established B-cell acute lymphoblastic leukemia. JAK2V617F failed to induce polycythemia in recipients after deletion of Stat5a/b, although the loss of STAT5 did not prevent the development of myelofibrosis. These results demonstrate that STAT5a/b is essential for the induction of CML-like leukemia by BCR-ABL1 and of polycythemia by JAK2V617F, and validate STAT5a/b and the genes they regulate as targets for therapy in these MPNs. PMID:22234689

Deficiency of activated STAT1 in head and neck cancer cells mediates TAP1-dependent escape from cytotoxic T lymphocytes

PubMed Central

Leibowitz, Michael S.; Filho, Pedro A. Andrade; Ferrone, Soldano; Ferris, Robert L.

2012-01-01

Squamous cell carcinoma of the head and neck (SCCHN) cells can escape recognition by tumor antigen (TA)-specific cytotoxic T lymphocytes (CTL) by downregulation of antigen processing machinery (APM) components, such as the transporter associated with antigen processing (TAP)-1/2 heterodimer. APM component upregulation by interferon gamma (IFN-γ) restores SCCHN cell recognition and susceptibility to lysis by CTL, but the mechanism underlying TAP1/2 downregulation in SCCHN cells is not known. Because IFN-γ activates signal transducer and activator of transcription (STAT)-1, we investigated phosphorylated (p)-STAT1 as a mediator of low basal TAP1/2 expression in SCCHN cells. SCCHN cells were found to express basal total STAT1 but low to undetectable levels of activated STAT1. The association of increased pSTAT1 levels and APM components likely reflects a cause–effect relationship, since STAT1 knockdown significantly reduced both IFN-γ-mediated APM component expression and TA-specific CTL recognition of IFN-γ-treated SCCHN cells. On the other hand, since oncogenic pSTAT3 is overexpressed in SCCHN cells and was found to heterodimerize with pSTAT1, we also tested whether pSTAT3 and pSTAT1:pSTAT3 heterodimers inhibited IFN-γ-induced STAT1 activation and APM component expression. First, STAT3 activation or depletion did not affect basal or IFN-γ-induced expression of pSTAT1 and APM components or recognition of SCCHN cells by TA-specific CTL. Second, pSTAT1:pSTAT3 heterodimers did not interfere with IFN-γ-induced STAT1 binding to the TAP1 promoter or APM protein expression. These findings demonstrate that APM component downregulation is regulated primarily by an IFN-γ-pSTAT1-mediated signaling pathway, independent of oncogenic STAT3 overexpression in SCCHN cells. PMID:21207025

Mesenchymal stem cells protect against obstruction-induced renal fibrosis by decreasing STAT3 activation and STAT3-dependent MMP-9 production

PubMed Central

Matsui, Futoshi; Babitz, Stephen A.; Rhee, Audrey; Hile, Karen L.; Zhang, Hongji

2017-01-01

STAT3 is a transcription factor implicated in renal fibrotic injury, but the role of STAT3 in mesenchymal stem cell (MSC)-induced renoprotection during renal fibrosis remains unknown. We hypothesized that MSCs protect against obstruction-induced renal fibrosis by downregulating STAT3 activation and STAT3-induced matrix metalloproteinase-9 (MMP-9) expression. Male Sprague-Dawley rats underwent renal arterial injection of vehicle or MSCs (1 × 106/rat) immediately before sham operation or induction of unilateral ureteral obstruction (UUO). The kidneys were harvested after 4 wk and analyzed for collagen I and III gene expression, collagen deposition (Masson’s trichrome), fibronectin, α-smooth muscle actin, active STAT3 (p-STAT3), MMP-9, and tissue inhibitor of matrix metalloproteinases 1 (TIMP-1) expression. In a separate arm, the STAT3 inhibitor S3I-201 (10 mg/kg) vs. vehicle was administered to rats intraperitoneally just after induction of UUO and daily for 14 days thereafter. The kidneys were harvested after 2 wk and analyzed for p-STAT3 and MMP-9 expression, and collagen and fibronectin deposition. Renal obstruction induced a significant increase in collagen, fibronectin, α-SMA, p-STAT3, MMP-9, and TIMP-1 expression while exogenously administered MSCs significantly reduced these indicators of obstruction-induced renal fibrosis. STAT3 inhibition with S3I-201 significantly reduced obstruction-induced MMP-9 expression and tubulointerstitial fibrosis. These results demonstrate that MSCs protect against obstruction-induced renal fibrosis, in part, by decreasing STAT3 activation and STAT3-dependent MMP-9 production. PMID:27760767

Differences in antiproliferative effect of STAT3 inhibition in HCC cells with versus without HBV expression

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hong, Yun; Zhou, Lin; Xie, Haiyang

2015-06-05

Chronic infection with hepatitis B virus (HBV) plays an important role in the etiology of hepatocellular carcinoma (HCC). Signal transducer and activator of transcription 3 (STAT3) inactivation could inhibit the tumor growth of HCC. In this study, differential antiproliferative effect of STAT3 inhibition was observed with HBV-related HCC cells being more resistant than non-HBV-related HCC cells. Resistance of HBV-related HCC cells to STAT3 inhibition was positively correlated to the expression of HBV. Enhanced ERK activation after STAT3 blockade was detected in HBV-related HCC cells but not in non-HBV-related HCC cells. Combined ERK and STAT3 inhibition eliminates the discrepancy between themore » two types of HCC cells. Moderate reduced HBV expression was found after STAT3 inhibition. These findings disclose a discrepancy in cellular response to STAT3 inhibition between non-HBV-related and HBV-related HCC cells and underscore the complexity of antiproliferative effect of STAT3 inactivation in HBV-related HCC cells. - Highlights: • HBV endows HCC cells with resistance to STAT3 inactivation on proliferation. • Abnormal ERK activation after STAT3 inhibition in HBV-related HCC cells. • Combined ERK and STAT3 inhibition eliminates the discrepancy. • STAT3 inhibition moderately reduces HBV expression.« less

Withaferin A Inhibits STAT3 and Induces Tumor Cell Death in Neuroblastoma and Multiple Myeloma

PubMed Central

Yco, Lisette P; Mocz, Gabor; Opoku-Ansah, John; Bachmann, André S

2014-01-01

Signal transducer and activator of transcription 3 (STAT3) is an oncogenic transcription factor that has been implicated in many human cancers and has emerged as an ideal target for cancer therapy. Withaferin A (WFA) is a natural product with promising antiproliferative properties through its association with a number of molecular targets including STAT3. However, the effect of WFA in pediatric neuroblastoma (NB) and its interaction with STAT3 have not been reported. In this study, we found that WFA effectively induces dose-dependent cell death in high-risk and drug-resistant NB as well as multiple myeloma (MM) tumor cells, prevented interleukin-6 (IL-6)–mediated and persistently activated STAT3 phosphorylation at Y705, and blocked the transcriptional activity of STAT3. We further provide computational models that show that WFA binds STAT3 near the Y705 phospho-tyrosine residue of the STAT3 Src homology 2 (SH2) domain, suggesting that WFA prevents STAT3 dimer formation similar to BP-1-102, a well-established STAT3 inhibitor. Our findings propose that the antitumor activity of WFA is mediated at least in part through inhibition of STAT3 and provide a rationale for further drug development and clinical use in NB and MM. PMID:25452693

Clustered localization of STAT3 during the cell cycle detected by super-resolution fluorescence microscopy

NASA Astrophysics Data System (ADS)

Gao, Jing; Chen, Junling; Cai, Mingjun; Xu, Haijiao; Jiang, Junguang; Tong, Ti; Wang, Hongda

2017-06-01

Signal transducer and activator of transcription 3 (STAT3) plays a key role in various cellular processes such as cell proliferation, differentiation, apoptosis and immune responses. In particular, STAT3 has emerged as a potential molecular target for cancer therapy. The functional role and standard activation mechanism of STAT3 have been well studied, however, the spatial distribution of STAT3 during the cell cycle is poorly known. Therefore, it is indispensable to study STAT3 spatial arrangement and nuclear-cytoplasimic localization at the different phase of cell cycle in cancer cells. By direct stochastic optical reconstruction microscopy imaging, we find that STAT3 forms various number and size of clusters at the different cell-cycle stage, which could not be clearly observed by conventional fluorescent microscopy. STAT3 clusters get more and larger gradually from G1 to G2 phase, during which time transcription and other related activities goes on consistently. The results suggest that there is an intimate relationship between the clustered characteristic of STAT3 and the cell-cycle behavior. Meanwhile, clustering would facilitate STAT3 rapid response to activating signals due to short distances between molecules. Our data might open a new door to develop an antitumor drug for inhibiting STAT3 signaling pathway by destroying its clusters.

Withaferin A Inhibits STAT3 and Induces Tumor Cell Death in Neuroblastoma and Multiple Myeloma.

PubMed

Yco, Lisette P; Mocz, Gabor; Opoku-Ansah, John; Bachmann, André S

2014-01-01

Signal transducer and activator of transcription 3 (STAT3) is an oncogenic transcription factor that has been implicated in many human cancers and has emerged as an ideal target for cancer therapy. Withaferin A (WFA) is a natural product with promising antiproliferative properties through its association with a number of molecular targets including STAT3. However, the effect of WFA in pediatric neuroblastoma (NB) and its interaction with STAT3 have not been reported. In this study, we found that WFA effectively induces dose-dependent cell death in high-risk and drug-resistant NB as well as multiple myeloma (MM) tumor cells, prevented interleukin-6 (IL-6)-mediated and persistently activated STAT3 phosphorylation at Y705, and blocked the transcriptional activity of STAT3. We further provide computational models that show that WFA binds STAT3 near the Y705 phospho-tyrosine residue of the STAT3 Src homology 2 (SH2) domain, suggesting that WFA prevents STAT3 dimer formation similar to BP-1-102, a well-established STAT3 inhibitor. Our findings propose that the antitumor activity of WFA is mediated at least in part through inhibition of STAT3 and provide a rationale for further drug development and clinical use in NB and MM.

Inhibition of Stat3 activation suppresses caspase-3 and the ubiquitin-proteasome system, leading to preservation of muscle mass in cancer cachexia.

PubMed

Silva, Kleiton Augusto Santos; Dong, Jiangling; Dong, Yanjun; Dong, Yanlan; Schor, Nestor; Tweardy, David J; Zhang, Liping; Mitch, William E

2015-04-24

Cachexia occurs in patients with advanced cancers. Despite the adverse clinical impact of cancer-induced muscle wasting, pathways causing cachexia are controversial, and clinically reliable therapies are not available. A trigger of muscle protein loss is the Jak/Stat pathway, and indeed, we found that conditioned medium from C26 colon carcinoma (C26) or Lewis lung carcinoma cells activates Stat3 (p-Stat3) in C2C12 myotubes. We identified two proteolytic pathways that are activated in muscle by p-Stat3; one is activation of caspase-3, and the other is p-Stat3 to myostatin, MAFbx/Atrogin-1, and MuRF-1 via CAAT/enhancer-binding protein δ (C/EBPδ). Using sequential deletions of the caspase-3 promoter and CHIP assays, we determined that Stat3 activation increases caspase-3 expression in C2C12 cells. Caspase-3 expression and proteolytic activity were stimulated by p-Stat3 in muscles of tumor-bearing mice. In mice with cachexia caused by Lewis lung carcinoma or C26 tumors, knock-out of p-Stat3 in muscle or with a small chemical inhibitor of p-Stat3 suppressed muscle mass losses, improved protein synthesis and degradation in muscle, and increased body weight and grip strength. Activation of p-Stat3 stimulates a pathway from C/EBPδ to myostatin and expression of MAFbx/Atrogin-1 and increases the ubiquitin-proteasome system. Indeed, C/EBPδ KO decreases the expression of MAFbx/Atrogin-1 and myostatin, while increasing muscle mass and grip strength. In conclusion, cancer stimulates p-Stat3 in muscle, activating protein loss by stimulating caspase-3, myostatin, and the ubiquitin-proteasome system. These results could lead to novel strategies for preventing cancer-induced muscle wasting. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Inhibition of Stat3 Activation Suppresses Caspase-3 and the Ubiquitin-Proteasome System, Leading to Preservation of Muscle Mass in Cancer Cachexia*

PubMed Central

Silva, Kleiton Augusto Santos; Dong, Jiangling; Dong, Yanjun; Dong, Yanlan; Schor, Nestor; Tweardy, David J.; Zhang, Liping; Mitch, William E.

2015-01-01

Cachexia occurs in patients with advanced cancers. Despite the adverse clinical impact of cancer-induced muscle wasting, pathways causing cachexia are controversial, and clinically reliable therapies are not available. A trigger of muscle protein loss is the Jak/Stat pathway, and indeed, we found that conditioned medium from C26 colon carcinoma (C26) or Lewis lung carcinoma cells activates Stat3 (p-Stat3) in C2C12 myotubes. We identified two proteolytic pathways that are activated in muscle by p-Stat3; one is activation of caspase-3, and the other is p-Stat3 to myostatin, MAFbx/Atrogin-1, and MuRF-1 via CAAT/enhancer-binding protein δ (C/EBPδ). Using sequential deletions of the caspase-3 promoter and CHIP assays, we determined that Stat3 activation increases caspase-3 expression in C2C12 cells. Caspase-3 expression and proteolytic activity were stimulated by p-Stat3 in muscles of tumor-bearing mice. In mice with cachexia caused by Lewis lung carcinoma or C26 tumors, knock-out of p-Stat3 in muscle or with a small chemical inhibitor of p-Stat3 suppressed muscle mass losses, improved protein synthesis and degradation in muscle, and increased body weight and grip strength. Activation of p-Stat3 stimulates a pathway from C/EBPδ to myostatin and expression of MAFbx/Atrogin-1 and increases the ubiquitin-proteasome system. Indeed, C/EBPδ KO decreases the expression of MAFbx/Atrogin-1 and myostatin, while increasing muscle mass and grip strength. In conclusion, cancer stimulates p-Stat3 in muscle, activating protein loss by stimulating caspase-3, myostatin, and the ubiquitin-proteasome system. These results could lead to novel strategies for preventing cancer-induced muscle wasting. PMID:25787076

Interleukin-6 inhibits hepatic growth hormone signaling via upregulation of Cis and Socs-3.

PubMed

Denson, Lee A; Held, Matthew A; Menon, Ram K; Frank, Stuart J; Parlow, Albert F; Arnold, Dodie L

2003-04-01

Cytokines may cause an acquired growth hormone (GH) resistance in patients with inflammatory diseases. Anabolic effects of GH are mediated through activation of STAT5 transcription factors. We have reported that TNF-alpha suppresses hepatic GH receptor (GHR) gene expression, whereas the cytokine-inducible SH2-containing protein 1 (Cis)/suppressors of cytokine signaling (Socs) genes are upregulated by TNF-alpha and IL-6 and inhibit GH activation of STAT5. However, the relative importance of these mechanisms in inflammatory GH resistance was not known. We hypothesized that IL-6 would prevent GH activation of STAT5 and that this would involve Cis/Socs protein upregulation. GH +/- LPS was administered to TNF receptor 1 (TNFR1) or IL-6 null mice and wild-type (WT) controls. STAT5, STAT3, GHR, Socs 1-3, and Cis phosphorylation and abundance were assessed by using immunoblots, EMSA, and/or real time RT-PCR. TNF-alpha and IL-6 abundance were assessed by using ELISA. GH activated STAT5 in WT and TNFR1 or IL-6 null mice. LPS pretreatment prevented STAT5 activation in WT and TNFR1 null mice; however, STAT5 activation was preserved in IL-6 null mice. GHR abundance did not change with LPS administration. Inhibition of STAT5 activation by LPS was temporally associated with phosphorylation of STAT3 and upregulation of Cis and Socs-3 protein in WT and TNFR1 null mice; STAT3, Cis, and Socs-3 were not induced in IL-6 null mice. IL-6 inhibits hepatic GH signaling by upregulating Cis and Socs-3, which may involve activation of STAT3. Therapies that block IL-6 may enhance GH signaling in inflammatory diseases.

The STAT3 HIES mutation is a gain-of-function mutation that activates genes via AGG-element carrying promoters.

PubMed

Xu, Li; Ji, Jin-Jun; Le, Wangping; Xu, Yan S; Dou, Dandan; Pan, Jieli; Jiao, Yifeng; Zhong, Tianfei; Wu, Dehong; Wang, Yumei; Wen, Chengping; Xie, Guan-Qun; Yao, Feng; Zhao, Heng; Fan, Yong-Sheng; Chin, Y Eugene

2015-10-15

Cytokine or growth factor activated STAT3 undergoes multiple post-translational modifications, dimerization and translocation into nuclei, where it binds to serum-inducible element (SIE, 'TTC(N3)GAA')-bearing promoters to activate transcription. The STAT3 DNA binding domain (DBD, 320-494) mutation in hyper immunoglobulin E syndrome (HIES), called the HIES mutation (R382Q, R382W or V463Δ), which elevates IgE synthesis, inhibits SIE binding activity and sensitizes genes such as TNF-α for expression. However, the mechanism by which the HIES mutation sensitizes STAT3 in gene induction remains elusive. Here, we report that STAT3 binds directly to the AGG-element with the consensus sequence 'AGG(N3)AGG'. Surprisingly, the helical N-terminal region (1-355), rather than the canonical STAT3 DBD, is responsible for AGG-element binding. The HIES mutation markedly enhances STAT3 AGG-element binding and AGG-promoter activation activity. Thus, STAT3 is a dual specificity transcription factor that promotes gene expression not only via SIE- but also AGG-promoter activity. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

Stat3 inhibition activates tumor macrophages and abrogates glioma growth in mice.

PubMed

Zhang, Leying; Alizadeh, Darya; Van Handel, Michelle; Kortylewski, Marcin; Yu, Hua; Badie, Behnam

2009-10-01

As the main effector-cell population of the central nervous system, microglia (MG) are considered to play an important immunoregulatory function in a number of pathological conditions such as inflammation, trauma, degenerative disease, and brain tumors. Recent studies, however, have suggested that the anti-neoplastic function of MG may be suppressed in malignant brain tumors. Considering the proposed suppressive role of signal transducers and activators of transcription 3 (Stat3) in antitumor immunity, we evaluated the role of Stat3 inhibition on MG and macrophage (MP) activation and tumor growth in a murine glioma model. N9 MG cells were exposed to GL261 glioma conditioned medium (GL261-CM) and evaluated for Stat3 activity and cytokine expression. Furthermore, the role of Stat3 inhibition on MG and MP activation was studied both in vitro and in vivo. Finally, the effect of Stat3 inhibition on tumor growth was assessed in intracranial GL261 gliomas. GL261-CM increased Stat3 activity in N9 cells in vitro and resulted in overexpression of IL-10 and IL-6, and downregulation of IL1-beta, a pro-inflammatory cytokine. Inhibition of Stat3 by CPA-7 or siRNA reversed glioma-induced cytokine expression profile in N9 cells. Furthermore, inactivation of Stat3 in intracranial GL261 tumors by siRNA resulted in MG/MP activation and tumor growth inhibition. Glioma-induced MG and MP suppression may be mediated thorough Stat3. Inhibition of Stat3 function in tumor MG/MP may result in their activation and can potentially be used as an adjunct immunotherapy approach for gliomas.

STAT3 inhibition as a therapeutic strategy for leukemia.

PubMed

Kanna, Rubashruti; Choudhary, Gaurav; Ramachandra, Nandini; Steidl, Ulrich; Verma, Amit; Shastri, Aditi

2017-11-22

Leukemia is characterized by selective overgrowth of malignant hematopoietic stem cells (HSC's) that interfere with HSC differentiation. Cytoreductive chemotherapy can kill rapidly dividing cancerous cells but cannot eradicate the malignant HSC pool leading to relapses. Leukemic stem cells have several dysregulated pathways and the Janus kinases (JAKs) and signal transducer and activator of transcription (STAT) pathway are prominent among them. STAT3 is an important transcription factor that regulates cell growth, proliferation, and inhibits apoptosis. High STAT3 expression in leukemia has been associated with an increased risk for relapse and decreased overall survival. Multiple strategies for interfering with STAT3 activity in leukemic cells include inhibition of STAT3 phosphorylation, interfering with STAT3 interactions, preventing nuclear transfer, inhibiting transcription and causing interference in STAT: DNA binding. A better understanding of key interactions and upstream mediators of STAT3 activity will help facilitate the development of effective cancer therapies and may result in durable remissions.

LRIG1 inhibits STAT3-dependent inflammation to maintain corneal homeostasis

PubMed Central

Nakamura, Takahiro; Hamuro, Junji; Takaishi, Mikiro; Simmons, Szandor; Maruyama, Kazuichi; Zaffalon, Andrea; Bentley, Adam J.; Kawasaki, Satoshi; Nagata-Takaoka, Maho; Fullwood, Nigel J.; Itami, Satoshi; Sano, Shigetoshi; Ishii, Masaru; Barrandon, Yann; Kinoshita, Shigeru

2013-01-01

Corneal integrity and transparency are indispensable for good vision. Cornea homeostasis is entirely dependent upon corneal stem cells, which are required for complex wound-healing processes that restore corneal integrity following epithelial damage. Here, we found that leucine-rich repeats and immunoglobulin-like domains 1 (LRIG1) is highly expressed in the human holoclone-type corneal epithelial stem cell population and sporadically expressed in the basal cells of ocular-surface epithelium. In murine models, LRIG1 regulated corneal epithelial cell fate during wound repair. Deletion of Lrig1 resulted in impaired stem cell recruitment following injury and promoted a cell-fate switch from transparent epithelium to keratinized skin-like epidermis, which led to corneal blindness. In addition, we determined that LRIG1 is a negative regulator of the STAT3-dependent inflammatory pathway. Inhibition of STAT3 in corneas of Lrig1–/– mice rescued pathological phenotypes and prevented corneal opacity. Additionally, transgenic mice that expressed a constitutively active form of STAT3 in the corneal epithelium had abnormal features, including corneal plaques and neovascularization similar to that found in Lrig1–/– mice. Bone marrow chimera experiments indicated that LRIG1 also coordinates the function of bone marrow–derived inflammatory cells. Together, our data indicate that LRIG1 orchestrates corneal-tissue transparency and cell fate during repair, and identify LRIG1 as a key regulator of tissue homeostasis. PMID:24316976

Activation of an IL-6:STAT3-dependent Transcriptome in Pediatric-onset Inflammatory Bowel Disease

PubMed Central

Carey, Rebecca; Jurickova, Ingrid; Ballard, Edgar; Bonkowski, Erin; Han, Xiaonan; Xu, Huan; Denson, Lee A.

2008-01-01

Background: While activation of the IL-6-dependent transcription factor signal transducer and activator of transcription 3 (STAT3) has been implicated in the pathogenesis of inflammatory bowel disease (IBD), a direct effect on mucosal gene expression and inflammation has not been shown. We hypothesized that a proinflammatory IL-6:STAT3-dependent biological network would be up regulated in pediatric-onset IBD patients, and would be associated with the severity of mucosal inflammation. Methods: Patients with pediatric-onset IBD were enrolled at diagnosis and during therapy. Serum cytokine analysis was performed using Bioplex. STAT3 phosphorylation (pSTAT3) in peripheral blood leukocytes (PBLs) was assessed by flow cytometry. Immunohistochemistry of colonic mucosa was used to localize pSTAT3 and STAT3 target genes. Microarray analysis was used to determine RNA expression profiles from colon biopsies. Results: Circulating IL-6 was upregulated in active IBD patients at diagnosis and during therapy. STAT3 activation was increased in PB granulocytes, IL-6-stimulated CD3+/CD4+ lymphocytes, and affected colon biopsies of IBD patients. The frequency of pSTAT3+PB granulocytes and colon epithelial and lamina propria cells was highly correlated with the degree of mucosal inflammation. Microarray and Ingenuity Systems bioinformatics analysis identified IL-6:STAT3-dependent biological networks upregulated in IBD patients which control leukocyte recruitment, HLA expression, angiogenesis, and tissue remodeling. Conclusions: A proinflammatory IL6:STAT3 biologic network is upregulated in active pediatric IBD patients at diagnosis and during therapy. Specific targeting of this network may be effective in reducing mucosal inflammation. PMID:18069684

Role of JAK-STAT signaling in the pathogenesis of myeloproliferative disorders.

PubMed

Levine, Ross L; Wernig, Gerlinde

2006-01-01

The identification of JAK2V617F mutations in polycythemia vera (PV), essential thrombocytosis (ET), and myelofibrosis (MF) represents an important advance in our understanding of these myeloproliferative disorders (MPD). Most, if not all, patients with PV and a significant number of patients with ET and MF are JAK2V617F positive, and the mutation likely arises in the hematopoietic stem cell compartment. JAK2V617F is a constitutively active tyrosine kinase that is able to activate JAK-STAT signaling most efficiently when co-expressed with the erythropoietin receptor (EPOR), the thrombopoietin receptor (MPL), or the granulocyte colony-stimulating factor receptor (GCSFR). Data from murine models supports the central role of JAK2V617F in the pathogenesis of MPD, as expression of JAK2V617F in a bone marrow transplantation assay results in polycythemia and myelofibrosis in recipient mice. Activation of JAK-STAT signaling by JAK2V617F in some, but not all MPD patients with ET and MF led to the identification of the constitutively active MPLW515L allele in ET and MF. Small molecule inhibitors of JAK-STAT signaling are currently being developed, which offer potential for molecularly targeted therapy for patients with PV, ET, and MF. Despite these advances, many questions remain regarding the role of a single disease allele in three phenotypically distinct MPD, the potential clinical efficacy of JAK2 inhibitors, and the identity of oncogenic alleles in JAK2V617F/MPLW515-negative MPD.

Identification of STAT1 and STAT3 Specific Inhibitors Using Comparative Virtual Screening and Docking Validation

PubMed Central

Szelag, Malgorzata; Czerwoniec, Anna; Wesoly, Joanna; Bluyssen, Hans A. R.

2015-01-01

Signal transducers and activators of transcription (STATs) facilitate action of cytokines, growth factors and pathogens. STAT activation is mediated by a highly conserved SH2 domain, which interacts with phosphotyrosine motifs for specific STAT-receptor contacts and STAT dimerization. The active dimers induce gene transcription in the nucleus by binding to a specific DNA-response element in the promoter of target genes. Abnormal activation of STAT signaling pathways is implicated in many human diseases, like cancer, inflammation and auto-immunity. Searches for STAT-targeting compounds, exploring the phosphotyrosine (pTyr)-SH2 interaction site, yielded many small molecules for STAT3 but sparsely for other STATs. However, many of these inhibitors seem not STAT3-specific, thereby questioning the present modeling and selection strategies of SH2 domain-based STAT inhibitors. We generated new 3D structure models for all human (h)STATs and developed a comparative in silico docking strategy to obtain further insight into STAT-SH2 cross-binding specificity of a selection of previously identified STAT3 inhibitors. Indeed, by primarily targeting the highly conserved pTyr-SH2 binding pocket the majority of these compounds exhibited similar binding affinity and tendency scores for all STATs. By comparative screening of a natural product library we provided initial proof for the possibility to identify STAT1 as well as STAT3-specific inhibitors, introducing the ‘STAT-comparative binding affinity value’ and ‘ligand binding pose variation’ as selection criteria. In silico screening of a multi-million clean leads (CL) compound library for binding of all STATs, likewise identified potential specific inhibitors for STAT1 and STAT3 after docking validation. Based on comparative virtual screening and docking validation, we developed a novel STAT inhibitor screening tool that allows identification of specific STAT1 and STAT3 inhibitory compounds. This could increase our understanding of the functional role of these STATs in different diseases and benefit the clinical need for more drugable STAT inhibitors with high specificity, potency and excellent bioavailability. PMID:25710482

Targeting colon cancer stem cells using a new curcumin analogue, GO-Y030

PubMed Central

Lin, L; Liu, Y; Li, H; Li, P-K; Fuchs, J; Shibata, H; Iwabuchi, Y; Lin, J

2011-01-01

Background: Persistent activation of signal transducers and activators of transcription 3 (STAT3) is commonly detected in many types of cancer, including colon cancer. To date, whether STAT3 is activated and the effects of STAT3 inhibition by a newly developed curcumin analogue, GO-Y030, in colon cancer stem cells are still unknown. Methods: Flow cytometry was used to isolate colon cancer stem cells, which are characterised by both aldehyde dehydrogenase (ALDH)-positive and CD133-positive subpopulations (ALDH+/CD133+). The levels of STAT3 phosphorylation and the effects of STAT3 inhibition by a newly developed curcumin analogue, GO-Y030, that targets STAT3 in colon cancer stem cells were examined. Results: Our results observed that ALDH+/CD133+ colon cancer cells expressed higher levels of phosphorylated STAT3 than ALDH-negative/CD133-negative colon cancer cells, suggesting that STAT3 is activated in colon cancer stem cells. GO-Y030 and curcumin inhibited STAT3 phosphorylation, cell viability, tumoursphere formation in colon cancer stem cells. GO-Y030 also reduced STAT3 downstream target gene expression and induced apoptosis in colon cancer stem cells. Furthermore, GO-Y030 suppressed tumour growth of cancer stem cells from both SW480 and HCT-116 colon cancer cell lines in the mouse model. Conclusion: Our results indicate that STAT3 is a novel therapeutic target in colon cancer stem cells, and inhibition of activated STAT3 in cancer stem cells by GO-Y030 may offer an effective treatment for colorectal cancer. PMID:21694723

Targeting colon cancer stem cells using a new curcumin analogue, GO-Y030.

PubMed

Lin, L; Liu, Y; Li, H; Li, P-K; Fuchs, J; Shibata, H; Iwabuchi, Y; Lin, J

2011-07-12

Persistent activation of signal transducers and activators of transcription 3 (STAT3) is commonly detected in many types of cancer, including colon cancer. To date, whether STAT3 is activated and the effects of STAT3 inhibition by a newly developed curcumin analogue, GO-Y030, in colon cancer stem cells are still unknown. Flow cytometry was used to isolate colon cancer stem cells, which are characterised by both aldehyde dehydrogenase (ALDH)-positive and CD133-positive subpopulations (ALDH(+)/CD133(+)). The levels of STAT3 phosphorylation and the effects of STAT3 inhibition by a newly developed curcumin analogue, GO-Y030, that targets STAT3 in colon cancer stem cells were examined. Our results observed that ALDH(+)/CD133(+) colon cancer cells expressed higher levels of phosphorylated STAT3 than ALDH-negative/CD133-negative colon cancer cells, suggesting that STAT3 is activated in colon cancer stem cells. GO-Y030 and curcumin inhibited STAT3 phosphorylation, cell viability, tumoursphere formation in colon cancer stem cells. GO-Y030 also reduced STAT3 downstream target gene expression and induced apoptosis in colon cancer stem cells. Furthermore, GO-Y030 suppressed tumour growth of cancer stem cells from both SW480 and HCT-116 colon cancer cell lines in the mouse model. Our results indicate that STAT3 is a novel therapeutic target in colon cancer stem cells, and inhibition of activated STAT3 in cancer stem cells by GO-Y030 may offer an effective treatment for colorectal cancer.

Activation of peroxisome proliferator-activated receptor-β/-δ (PPAR-β/-δ) ameliorates insulin signaling and reduces SOCS3 levels by inhibiting STAT3 in interleukin-6-stimulated adipocytes.

PubMed

Serrano-Marco, Lucía; Rodríguez-Calvo, Ricardo; El Kochairi, Ilhem; Palomer, Xavier; Michalik, Liliane; Wahli, Walter; Vázquez-Carrera, Manuel

2011-07-01

It has been suggested that interleukin (IL)-6 is one of the mediators linking obesity-derived chronic inflammation with insulin resistance through activation of STAT3, with subsequent upregulation of suppressor of cytokine signaling 3 (SOCS3). We evaluated whether peroxisome proliferator-activated receptor (PPAR)-β/-δ prevented activation of the IL-6-STAT3-SOCS3 pathway and insulin resistance in adipocytes. Adipocytes and white adipose tissue from wild-type and PPAR-β/-δ-null mice were used to evaluate the effect of PPAR-β/-δ on the IL-6-STAT3-SOCS3 pathway. First, we observed that the PPAR-β/-δ agonist GW501516 prevented both IL-6-dependent reduction in insulin-stimulated Akt phosphorylation and glucose uptake in adipocytes. In addition, this drug treatment abolished IL-6-induced SOCS3 expression in differentiated 3T3-L1 adipocytes. This effect was associated with the capacity of the drug to prevent IL-6-induced STAT3 phosphorylation on Tyr(705) and Ser(727) residues in vitro and in vivo. Moreover, GW501516 prevented IL-6-dependent induction of extracellular signal-related kinase (ERK)1/2, a serine-threonine-protein kinase involved in serine STAT3 phosphorylation. Furthermore, in white adipose tissue from PPAR-β/-δ-null mice, STAT3 phosphorylation (Tyr(705) and Ser(727)), STAT3 DNA-binding activity, and SOCS3 protein levels were higher than in wild-type mice. Several steps in STAT3 activation require its association with heat shock protein 90 (Hsp90), which was prevented by GW501516 as revealed in immunoprecipitation studies. Consistent with this finding, the STAT3-Hsp90 association was enhanced in white adipose tissue from PPAR-β/-δ-null mice compared with wild-type mice. Collectively, our findings indicate that PPAR-β/-δ activation prevents IL-6-induced STAT3 activation by inhibiting ERK1/2 and preventing the STAT3-Hsp90 association, an effect that may contribute to the prevention of cytokine-induced insulin resistance in adipocytes. © 2011 by the American Diabetes Association.

Expression of the JAK/STAT Signaling Pathway in Bullous Pemphigoid and Dermatitis Herpetiformis

PubMed Central

Wozniacka, A.; Waszczykowska, E.; Zebrowska, A.

2017-01-01

A family of eleven proteins comprises the Janus kinases (JAK) and signal transducers and activators of transcription (STAT) signaling pathway, which enables transduction of signal from cytokine receptor to the nucleus and activation of transcription of target genes. Irregular functioning of the cascade may contribute to pathogenesis of autoimmune diseases; however, there are no reports concerning autoimmune bullous diseases yet to be published. The aim of this study was to evaluate the expression of proteins constituting the JAK/STAT signaling pathway in skin lesions and perilesional area in dermatitis herpetiformis (DH) and bullous pemphigoid (BP), as well as in the control group. Skin biopsies were collected from 21 DH patients, from 20 BP patients, and from 10 healthy volunteers. The localization and expression of selected STAT and JAK proteins were examined by immunohistochemistry and immunoblotting. We found significantly higher expression of JAK/STAT proteins in skin lesions in patients with BP and DH, in comparison to perilesional skin and the control group, which may be related to proinflammatory cytokine network and induction of inflammatory infiltrate in tissues. Our findings suggest that differences in the JAK and STAT expression may be related to distinct cytokines activating them and mediating neutrophilic and/or eosinophilic infiltrate. PMID:29203970

STATs in cancer inflammation and immunity: a leading role for STAT3.

PubMed

Yu, Hua; Pardoll, Drew; Jove, Richard

2009-11-01

Commensurate with their roles in regulating cytokine-dependent inflammation and immunity, signal transducer and activator of transcription (STAT) proteins are central in determining whether immune responses in the tumour microenvironment promote or inhibit cancer. Persistently activated STAT3 and, to some extent, STAT5 increase tumour cell proliferation, survival and invasion while suppressing anti-tumour immunity. The persistent activation of STAT3 also mediates tumour-promoting inflammation. STAT3 has this dual role in tumour inflammation and immunity by promoting pro-oncogenic inflammatory pathways, including nuclear factor-kappaB (NF-kappaB) and interleukin-6 (IL-6)-GP130-Janus kinase (JAK) pathways, and by opposing STAT1- and NF-kappaB-mediated T helper 1 anti-tumour immune responses. Consequently, STAT3 is a promising target to redirect inflammation for cancer therapy.

Mesenchymal stem cells protect against obstruction-induced renal fibrosis by decreasing STAT3 activation and STAT3-dependent MMP-9 production.

PubMed

Matsui, Futoshi; Babitz, Stephen A; Rhee, Audrey; Hile, Karen L; Zhang, Hongji; Meldrum, Kirstan K

2017-01-01

STAT3 is a transcription factor implicated in renal fibrotic injury, but the role of STAT3 in mesenchymal stem cell (MSC)-induced renoprotection during renal fibrosis remains unknown. We hypothesized that MSCs protect against obstruction-induced renal fibrosis by downregulating STAT3 activation and STAT3-induced matrix metalloproteinase-9 (MMP-9) expression. Male Sprague-Dawley rats underwent renal arterial injection of vehicle or MSCs (1 × 10 6 /rat) immediately before sham operation or induction of unilateral ureteral obstruction (UUO). The kidneys were harvested after 4 wk and analyzed for collagen I and III gene expression, collagen deposition (Masson's trichrome), fibronectin, α-smooth muscle actin, active STAT3 (p-STAT3), MMP-9, and tissue inhibitor of matrix metalloproteinases 1 (TIMP-1) expression. In a separate arm, the STAT3 inhibitor S3I-201 (10 mg/kg) vs. vehicle was administered to rats intraperitoneally just after induction of UUO and daily for 14 days thereafter. The kidneys were harvested after 2 wk and analyzed for p-STAT3 and MMP-9 expression, and collagen and fibronectin deposition. Renal obstruction induced a significant increase in collagen, fibronectin, α-SMA, p-STAT3, MMP-9, and TIMP-1 expression while exogenously administered MSCs significantly reduced these indicators of obstruction-induced renal fibrosis. STAT3 inhibition with S3I-201 significantly reduced obstruction-induced MMP-9 expression and tubulointerstitial fibrosis. These results demonstrate that MSCs protect against obstruction-induced renal fibrosis, in part, by decreasing STAT3 activation and STAT3-dependent MMP-9 production. Copyright © 2017 the American Physiological Society.

Proinflammatory Cytokines IL-6 and TNF-α Increased Telomerase Activity through NF-κB/STAT1/STAT3 Activation, and Withaferin A Inhibited the Signaling in Colorectal Cancer Cells.

PubMed

Chung, Seyung S; Wu, Yong; Okobi, Quincy; Adekoya, Debbie; Atefi, Mohammad; Clarke, Orette; Dutta, Pranabananda; Vadgama, Jaydutt V

2017-01-01

There are increasing evidences of proinflammatory cytokine involvement in cancer development. Here, we found that two cytokines, IL-6 and TNF- α , activated colorectal cancer cells to be more invasive and stem-like. Combined treatment of IL-6 and TNF- α phosphorylated transcription factors STAT3 in a synergistic manner. STAT3, STAT1, and NF- κ B physically interacted upon the cytokine stimulation. STAT3 was bound to the promoter region of human telomerase reverse transcriptase (hTERT). IL-6 and TNF- α stimulation further enhanced STAT3 binding affinity. Stem cell marker Oct-4 was upregulated in colorectal cancer cells upon IL-6 and TNF- α stimulation. Withaferin A, an anti-inflammatory steroidal lactone, inhibited the IL-6- and TNF- α -induced cancer cell invasion and decreased colonosphere formation. Notably, withaferin A inhibited STAT3 phosphorylation and abolished the STAT3, STAT1, and NF- κ B interactions. Oct-4 expression was also downregulated by withaferin A inhibition. The binding of STAT3 to the hTERT promoter region and telomerase activity showed reduction with withaferin A treatments. Proinflammatory cytokine-induced cancer cell invasiveness is mediated by a STAT3-regulated mechanism in colorectal cancer cells. Our data suggest that withaferin A could be a promising anticancer agent that effectively inhibits the progression of colorectal cancer.

Interferon-γ biphasically regulates angiotensinogen expression via a JAK-STAT pathway and suppressor of cytokine signaling 1 (SOCS1) in renal proximal tubular cells

PubMed Central

Satou, Ryousuke; Miyata, Kayoko; Gonzalez-Villalobos, Romer A.; Ingelfinger, Julie R.; Navar, L. Gabriel; Kobori, Hiroyuki

2012-01-01

Renal inflammation modulates angiotensinogen (AGT) production in renal proximal tubular cells (RPTCs) via inflammatory cytokines, including interleukin-6, tumor necrosis factor α, and interferon-γ (IFN-γ). Among these, the effects of IFN-γ on AGT regulation in RPTCs are incompletely delineated. This study aimed to elucidate mechanisms by which IFN-γ regulates AGT expression in RPTCs. RPTCs were incubated with or without IFN-γ up to 48 h. AGT expression, STAT1 and STAT3 activities, and SOCS1 expression were evaluated. RNA interference studies against STAT1, SOCS1, and STAT3 were performed to elucidate a signaling cascade. IFN-γ decreased AGT expression at 6 h (0.61±0.05, ratio to control) and 12 h (0.47±0.03). In contrast, longer exposure for 24 and 48 h increased AGT expression (1.76±0.18, EC50=3.4 ng/ml, and 1.45±0.08, respectively). IFN-γ treatment for 6 h strongly induced STAT1 phosphorylation and SOCS1 augmentation, and decreased STAT3 activity. However, STAT1 phosphorylation and SOCS1 augmentation waned at 24 h, while STAT3 activity increased. RNA interference studies revealed that activation of STAT1-SOCS1 axis decreased STAT3 activity. Thus, IFN-γ biphasically regulates AGT expression in RPTCs via STAT3 activity modulated by STAT1-SOCS1 axis, suggesting the STAT1-SOCS1 axis is important in IFN-γ-induced activation of the intrarenal renin-angiotensin system.—Satou, R., Miyata, K., Gonzalez-Villalobos, R. A., Ingelfinger, J. R., Navar, L. G., Kobori, H. Interferon-γ biphasically regulates angiotensinogen expression via a JAK-STAT pathway and suppressor of cytokine signaling 1 (SOCS1) in renal proximal tubular cells. PMID:22302831

Metformin ameliorates IL-6-induced hepatic insulin resistance via induction of orphan nuclear receptor small heterodimer partner (SHP) in mouse models.

PubMed

Kim, Y D; Kim, Y H; Cho, Y M; Kim, D K; Ahn, S W; Lee, J M; Chanda, D; Shong, M; Lee, C H; Choi, H S

2012-05-01

IL-6 is a proinflammatory cytokine associated with the pathogenesis of hepatic diseases. Metformin is an anti-diabetic drug used for the treatment of type 2 diabetes, and orphan nuclear receptor small heterodimer partner (SHP, also known as NR0B2), a transcriptional co-repressor, plays an important role in maintaining metabolic homeostasis. Here, we demonstrate that metformin-mediated activation of AMP-activated protein kinase (AMPK) increases SHP protein production and regulates IL-6-induced hepatic insulin resistance. We investigated metformin-mediated SHP production improved insulin resistance through the regulation of an IL-6-dependent pathway (involving signal transducer and activator of transcription 3 [STAT3] and suppressor of cytokine signalling 3 [SOCS3]) in both Shp knockdown and Shp null mice. IL-6-induced STAT3 transactivation and SOCS3 production were significantly repressed by metformin, adenoviral constitutively active AMPK (Ad-CA-AMPK), and adenoviral SHP (Ad-SHP), but not in Shp knockdown, or with the adenoviral dominant negative form of AMPK (Ad-DN-AMPK). Chromatin immunoprecipitation (ChIP), co-immunoprecipitation (Co-IP) and protein localisation studies showed that SHP inhibits DNA binding of STAT3 on the Socs3 gene promoter via interaction and colocalisation within the nucleus. Upregulation of inflammatory genes and downregulation of hepatic insulin signalling by acute IL-6 treatment were observed in wild-type mice but not in Shp null mice. Finally, chronic IL-6 exposure caused hepatic insulin resistance, leading to impaired insulin tolerance and elevated gluconeogenesis, and these phenomena were aggravated in Shp null mice. Our results demonstrate that SHP upregulation by metformin may prevent hepatic disorders by regulating the IL-6-dependent pathway, and that this pathway can help to ameliorate the pathogenesis of cytokine-mediated metabolic dysfunction.

STAT3 Activation Promotes Oncolytic HSV1 Replication in Glioma Cells

PubMed Central

Okemoto, Kazuo; Wagner, Benjamin; Meisen, Hans; Haseley, Amy; Kaur, Balveen; Chiocca, Ennio Antonio

2013-01-01

Recent studies report that STAT3 signaling is a master regulator of mesenchymal transformation of gliomas and that STAT3 modulated genes are highly expressed in the mesenchymal transcriptome of gliomas. A currently studied experimental treatment for gliomas consists of intratumoral injection of oncolytic viruses (OV), such as oncolytic herpes simplex virus type 1 (oHSV). We have described one particular oHSV (rQNestin34.5) that exhibits potent anti-glioma activity in animal models. Here, we hypothesized that alterations in STAT3 signaling in glioma cells may affect the replicative ability of rQNestin34.5. In fact, human U251 glioma cells engineered to either over-express STAT3 or with genetic down-regulation of STAT3 supported oHSV replication to a significantly higher or lesser degree, respectively, when compared to controls. Administration of pharmacologic agents that increase STAT3 phosphorylation/activation (Valproic Acid) or increase STAT3 levels (Interleukin 6) also significantly enhanced oHSV replication. Instead, administration of inhibitors of STAT3 phosphorylation/activation (LLL12) significantly reduced oHSV replication. STAT3 led to a reduction in interferon signaling in oHSV infected cells and inhibition of interferon signaling abolished the effect of STAT3 on oHSV replication. These data thus indicate that STAT3 signaling in malignant gliomas enhances oHSV replication, likely by inhibiting the interferon response in infected glioma cells, thus suggesting avenues for possible potentiation of oncolytic virotherapy. PMID:23936533

Sustained activation of STAT5 is essential for chromatin remodeling and maintenance of mammary-specific function

DOE Office of Scientific and Technical Information (OSTI.GOV)

Xu, Ren; Nelson, Celeste M.; Muschler, John L.

2009-06-03

Epithelial cells, once dissociated and placed in two-dimensional (2D) cultures, rapidly lose tissue-specific functions. We showed previously that in addition to prolactin, signaling by laminin-111 was necessary to restore functional differentiation of mammary epithelia. Here, we elucidate two additional aspects of laminin-111 action. We show that in 2D cultures, the prolactin receptor is basolaterally localized and physically segregated from its apically placed ligand. Detachment of the cells exposes the receptor to ligation by prolactin leading to signal transducers and activators of transcription protein 5 (STAT5) activation, but only transiently and not sufficiently for induction of milk protein expression. We showmore » that laminin-111 reorganizes mammary cells into polarized acini, allowing both the exposure of the prolactin receptor and sustained activation of STAT5. The use of constitutively active STAT5 constructs showed that the latter is necessary and sufficient for chromatin reorganization and {beta}-casein transcription. These results underscore the crucial role of continuous laminin signaling and polarized tissue architecture in maintenance of transcription factor activation, chromatin organization, and tissue-specific gene expression.« less

Chemical and Hormonal Effects on STAT5b-Dependent Sexual Dimorphism of the Liver Transcriptome

PubMed Central

Oshida, Keiyu; Waxman, David J.; Corton, J. Christopher

2016-01-01

The growth hormone (GH)-activated transcription factor signal transducer and activator of transcription 5b (STAT5b) is a key regulator of sexually dimorphic gene expression in the liver. Suppression of hepatic STAT5b signaling is associated with lipid metabolic dysfunction leading to steatosis and liver cancer. In the companion publication, a STAT5b biomarker gene set was identified and used in a rank-based test to predict both increases and decreases in liver STAT5b activation status/function with high (≥ 97%) accuracy. Here, this computational approach was used to identify chemicals and hormones that activate (masculinize) or suppress (feminize) STAT5b function in a large, annotated mouse liver and primary hepatocyte gene expression compendium. Exposure to dihydrotestosterone and thyroid hormone caused liver masculinization, whereas glucocorticoids, fibroblast growth factor 15, and angiotensin II caused liver feminization. In mouse models of diabetes and obesity, liver feminization was consistently observed and was at least partially reversed by leptin or resveratrol exposure. Chemical-induced feminization of male mouse liver gene expression profiles was a relatively frequent phenomenon: of 156 gene expression biosets from chemically-treated male mice, 29% showed feminization of liver STAT5b function, while <1% showed masculinization. Most (93%) of the biosets that exhibited feminization of male liver were also associated with activation of one or more xenobiotic-responsive receptors, most commonly constitutive activated receptor (CAR) or peroxisome proliferator-activated receptor alpha (PPARα). Feminization was consistently associated with increased expression of peroxisome proliferator-activated receptor gamma (Pparg) but not other lipogenic transcription factors linked to steatosis. GH-activated STAT5b signaling in mouse liver is thus commonly altered by diverse chemicals, and provides a linkage between chemical exposure and dysregulated gene expression associated with adverse effects on the liver. PMID:26959237

GPR30 mediates anorectic estrogen-induced STAT3 signaling in the hypothalamus.

PubMed

Kwon, Obin; Kang, Eun Seok; Kim, Insook; Shin, Sora; Kim, Mijung; Kwon, Somin; Oh, So Ra; Ahn, Young Soo; Kim, Chul Hoon

2014-11-01

Estrogen plays an important role in the control of energy balance in the hypothalamus. Leptin-independent STAT3 activation (i.e., tyrosine(705)-phosphorylation of STAT3, pSTAT3) in the hypothalamus is hypothesized as the primary mechanism of the estrogen-induced anorexic response. However, the type of estrogen receptor that mediates this regulation is unknown. We investigated the role of the G protein-coupled receptor 30 (GPR30) in estradiol (E2)-induced STAT3 activation in the hypothalamus. Regulation of STAT3 activation by E2, G-1, a specific agonist of GPR30 and G-15, a specific antagonist of GPR30 was analyzed in vitro and in vivo. Effect of GPR30 activation on eating behavior was analyzed in vivo. E2 stimulated pSTAT3 in cells expressing GPR30, but not expressing estrogen receptor ERα and ERβ. G-1 induced pSTAT3, and G-15 inhibited E2-induced pSTAT3 in primary cultures of hypothalamic neurons. A cerebroventricular injection of G-1 increased pSTAT3 in the arcuate nucleus of mice, which was associated with a decrease in food intake and body weight gain. These results suggest that GPR30 is the estrogen receptor that mediates the anorectic effect of estrogen through the STAT3 pathway in the hypothalamus. Copyright © 2014 Elsevier Inc. All rights reserved.

Propofol mediates signal transducer and activator of transcription 3 activation and crosstalk with phosphoinositide 3-kinase/AKT.

PubMed

Shravah, Jayant; Wang, Baohua; Pavlovic, Marijana; Kumar, Ujendra; Chen, David Dy; Luo, Honglin; Ansley, David M

2014-01-01

We previously demonstrated that propofol, an intravenous anesthetic with anti-oxidative properties, activated the phosphoinositide 3-kinase (PI3K)/AKT pathway to increase the expression of B cell lymphoma (Bcl)-2 and, therefore the anti-apoptotic potential on cardiomyocytes. Here, we wanted to determine if propofol can also activate the Janus kinase (JAK) 2/signal transducer and activator of transcription (STAT) 3 pathway, another branch of cardioprotective signaling. The cellular response of nuclear factor kappa B (NFκB) and STAT3 was also evaluated. Cardiac H9c2 cells were treated by propofol alone or in combination with pretreatment by inhibitors for JAK2/STAT3 or PI3K/AKT pathway. STAT3 and AKT phosphorylation, and STAT3 translocation were measured by western blotting and immunofluorescence staining, respectively. Propofol treatment significantly increased STAT3 phosphorylation at both tyrosine 705 and serine 727 residues. Sustained early phosphorylation of STAT3 was observed with 25~75 μM propofol at 10 and 30 min. Nuclear translocation of STAT3 was seen at 4 h after treatment with 50 μM propofol. In cultured H9c2 cells, we further demonstrated that propofol-induced STAT3 phosphorylation was reduced by pretreatment with PI3K/AKT pathway inhibitors wortmannin or API-2. Conversely, pretreatment with JAK2/STAT3 pathway inhibitor AG490 or stattic inhibited propofol-induced AKT phosphorylation. In addition, propofol induced NFκB p65 subunit perinuclear translocation. Inhibition or knockdown of STAT3 was associated with increased levels of the NFκB p65 subunit. Our results suggest that propofol induces an adaptive response by dual activation and crosstalk of cytoprotective PI3K/AKT and JAK2/STAT3 pathways. Rationale to apply propofol clinically as a preemptive cardioprotectant during cardiac surgery is supported by our findings.

Potent and Selective Phosphopeptide Mimetic Prodrugs Targeted to the Src Homology 2 (SH2) Domain of Signal Transducer and Activator of Transcription 3

PubMed Central

Mandal, Pijus K.; Gao, Fengqin; Lu, Zhen; Ren, Zhiyong; Ramesh, Rajagopal; Birtwistle, J. Sanderson; Kaluarachchi, Kumaralal K.; Chen, Xiaomin; Bast, Robert C.; Liao, Warren S.; McMurray, John S.

2011-01-01

Signal transducer and activator of transcription 3 (Stat3), a target for anticancer drug design, is activated by recruitment to phosphotyrosine residues on growth factor and cytokine receptors via its SH2 domain. We report here structure-activity relationship studies on phosphopeptide mimics targeted to the SH2 domain of Stat3. Inclusion of a methyl group on the β-position of the pTyr mimic, 4-phosphocinfnamide, enhanced affinity 2–3 fold. Bis-pivaloyloxymethyl prodrugs containing β-methyl cinnamide, dipeptide scaffolds Haic and Nle-cis-3,4-methanoproline, and glutamine surrogates were highly potent, completely inhibiting phosphorylation of Stat3 Tyr705 at 0.5–1 µM in a variety of cancer cell lines. The inhibitors were selective for Stat3 over Stat1, Stat5, Src, and p85 of PI3K, indicating ability to discriminate individual SH2 domains in intact cells. At concentrations that completely inhibited Stat3 phosphorylation, the prodrugs were not cytotoxic to a panel of tumor cells, thereby showing clear distinction between cytotoxicity and effects downstream of activated Stat3. PMID:21486047

STATs in Lung Development: Distinct Early and Late Expression, Growth Modulation and Signaling Dysregulation in Congenital Diaphragmatic Hernia.

PubMed

Piairo, Paulina; Moura, Rute S; Baptista, Maria João; Correia-Pinto, Jorge; Nogueira-Silva, Cristina

2018-01-01

Congenital diaphragmatic hernia (CDH) is a life-threatening developmental anomaly, intrinsically combining severe pulmonary hypoplasia and hypertension. During development, signal transducers and activators of transcription (STAT) are utilized to elicit cell growth, differentiation, and survival. We used the nitrofen-induced CDH rat model. At selected gestational time points, lungs were divided into two experimental groups, i.e., control or CDH. We performed immunohistochemistry and western blotting analysis to investigate the developmental expression profile of the complete family of STATs (STAT1-6), plus specific STATs activation (p-STAT3, p-STAT6) and regulation by SOCS (SOCS3) in normal lungs against those of diseased lungs. The normal fetal lung explants were treated with piceatannol (STAT3 inhibitor) in vitro followed by morphometrical analysis. Molecular profiling of STATs during the lung development revealed distinct early and late expression signatures. Experimental CDH altered the STATs expression, activation, and regulation in the fetal lungs. In particular, STAT3 and STAT6 were persistently over-expressed and early over-activated. Piceatannol treatment dose-dependently stimulated the fetal lung growth. These findings suggest that STATs play an important role during normal fetal lung development and CDH pathogenesis. Moreover, functionally targeting STAT signaling modulates fetal lung growth, which highlights that STAT3 and STAT6 signaling might be promising therapeutic targets in reducing or preventing pulmonary hypoplasia in CDH. © 2018 The Author(s). Published by S. Karger AG, Basel.

Increased expression of activated pSTAT3 and PIM-1 in the pulmonary vasculature of experimental congenital diaphragmatic hernia.

PubMed

Hofmann, Alejandro D; Takahashi, Toshiaki; Duess, Johannes; Gosemann, Jan-Hendrik; Puri, Prem

2015-06-01

Signal transducer and activator of transcription (STAT) protein family (STAT1-6) regulates diverse cellular processes. Recently, the isoform STAT3 has been implicated to play a central role in the pathogenesis of pulmonary hypertension (PH). In human PH activated STAT3 (pSTAT3) was shown to directly trigger expression of the provirus integration site for Moloney murine leukemia virus (Pim-1), which promotes proliferation and resistance to apoptosis in SMCs. We designed this study to investigate the hypothesis that pSTAT3 and Pim-1 pulmonary vascular expression is increased in nitrofen-induced CDH. Pregnant rats were exposed to nitrofen or vehicle on D9.5. Fetuses were sacrificed on D21 and divided into nitrofen (n=16) and control group (n=16). QRT-PCR, western blotting, and confocal-immunofluorescence were performed to determine pulmonary gene and protein expression levels of pSTAT3 and Pim-1. Pulmonary Pim-1 gene expression was significantly increased in the CDH group compared to controls. Western blotting and confocal-microscopy confirmed increased pulmonary protein expression of Pim-1 and increased activation of pSTAT3 in CDH lungs compared to controls. Markedly increased gene and protein expression of Pim-1 and activated pSTAT3 in the pulmonary vasculature of nitrofen-induced CDH lungs suggest that pSTAT3 and Pim-1 are important mediators of PH in nitrofen-induced CDH. Copyright © 2015. Published by Elsevier Inc.

Nuclear localization of activated STAT6 and STAT3 in epidermis of prurigo nodularis.

PubMed

Fukushi, S; Yamasaki, K; Aiba, S

2011-11-01

Prurigo nodularis (PN) is a chronic dermatitis characterized by discrete, raised, and firm papulonodules with intense pruritus. The pathogenesis still remains to be elucidated. To clarify the role of Th1 and Th2 cytokines in the pathogenesis of PN. We examined the cytokine signatures, such as phosphorylation of STAT1, STAT3 and STAT6, HLA-DR and hyaluronan accumulation, to reveal the Th1 and Th2 cytokine influence on the lesional epidermis of PN. We first optimized antigen retrieval methods to detect these signatures with antibodies for phospho-STAT1 (pSTAT1), phospho-STAT3 (pSTAT3), phospho-STAT6 (pSTAT6), HLA-DR and hyaluronic acid binding protein (HABP) on the formalin-fixed paraffin-embedded sections of psoriasis, lichen planus and atopic dermatitis biopsy samples. Activation of STAT1 and STAT6 in epidermis by Th1 and Th2 cytokines was further confirmed in a cultured skin equivalent model treated with interferon-γ or interleukin (IL)-4/IL-13. With the relevant immunostaining methods, we examined the cytokine signatures in 22 cases of PN. The results revealed that (i) the entire epidermis of 19 cases was stained with anti-pSTAT6 antibody, (ii) 21 cases demonstrated nuclear staining with anti-pSTAT3 antibody, (iii) the entire epidermis of 21 cases was stained with HABP, (iv) the epidermis of eight cases showed scattered staining with anti-pSTAT1 antibody, and (v) six cases were positive for HLA-DR membrane expression. These data indicated that Th2 cytokines related to STAT6 activation together with some unknown stimuli that activate STAT3 play a principal role in the pathogenesis of PN. © 2011 The Authors. BJD © 2011 British Association of Dermatologists.

STAT3 and importins are novel mediators of early molecular and cellular responses in experimental duodenal ulceration.

PubMed

Khomenko, Tetyana; Deng, Xiaoming; Ahluwalia, Amrita; Tarnawski, Andrzej; Patel, Khushin N; Sandor, Zsuzsanna; Szabo, Sandor

2014-02-01

Signal transducer and activator of transcription 3 (STAT3) is a transcription factor that directly upregulates VEGF, Ref-1, p21, and anti-apoptotic genes such as Bcl-xL. In this study, we hypothesized that STAT3 signaling is activated and provides a critical protective role that is required for enterocyte survival during the early phases of cysteamine-induced duodenal ulcers. We studied the effect of inhibition of STAT3 activity on cysteamine-induced duodenal ulcers in rats and egr-1 knockout mice using STAT3/DNA binding assay, immunohistochemistry, immunoblot, and quantitative reverse transcriptase PCR analyses. We found that G-quartet oligodeoxynucleotides T40214, a specific inhibitor of STAT3/DNA binding, aggravated cysteamine-induced duodenal ulcers in rats 2.8-fold (p < 0.05). In the pre-ulcerogenic stage, cysteamine induced STAT3 tyrosine phosphorylation, its translocation to nuclei, an increased expression and nuclear translocation of importin α and β in the rat duodenal mucosa. Cysteamine enhanced the binding of STAT3 to its DNA consensus sequences at 6, 12, and 24 h after cysteamine by 1.5-, 1.8-, and 3.5-fold, respectively, and activated the expression of STAT3 target genes such as VEGF, Bcl-xL, Ref-1, and STAT3-induced feedback inhibitor, a suppressor of cytokine signaling 3. We also demonstrated that egr-1 knockout mice, which are more susceptible to cysteamine-induced duodenal ulcers, had lower levels of STAT3 expression, its phosphorylation, expression of importin α or β, and STAT3/DNA binding than wild-type mice in response to cysteamine. Thus, STAT3 represents an important new molecular mechanism in experimental duodenal ulceration.

In silico simulations of STAT1 and STAT3 inhibitors predict SH2 domain cross-binding specificity.

PubMed

Szelag, Malgorzata; Sikorski, Krzysztof; Czerwoniec, Anna; Szatkowska, Katarzyna; Wesoly, Joanna; Bluyssen, Hans A R

2013-11-15

Signal transducers and activators of transcription (STATs) comprise a family of transcription factors that are structurally related and which participate in signaling pathways activated by cytokines, growth factors and pathogens. Activation of STAT proteins is mediated by the highly conserved Src homology 2 (SH2) domain, which interacts with phosphotyrosine motifs for specific contacts between STATs and receptors and for STAT dimerization. By generating new models for human (h)STAT1, hSTAT2 and hSTAT3 we applied comparative in silico docking to determine SH2-binding specificity of the STAT3 inhibitor stattic, and of fludarabine (STAT1 inhibitor). Thus, we provide evidence that by primarily targeting the highly conserved phosphotyrosine (pY+0) SH2 binding pocket stattic is not a specific hSTAT3 inhibitor, but is equally effective towards hSTAT1 and hSTAT2. This was confirmed in Human Micro-vascular Endothelial Cells (HMECs) in vitro, in which stattic inhibited interferon-α-induced phosphorylation of all three STATs. Likewise, fludarabine inhibits both hSTAT1 and hSTAT3 phosphorylation, but not hSTAT2, by competing with the highly conserved pY+0 and pY-X binding sites, which are less well-preserved in hSTAT2. Moreover we observed that in HMECs in vitro fludarabine inhibits cytokine and lipopolysaccharide-induced phosphorylation of hSTAT1 and hSTAT3 but does not affect hSTAT2. Finally, multiple sequence alignment of STAT-SH2 domain sequences confirmed high conservation between hSTAT1 and hSTAT3, but not hSTAT2, with respect to stattic and fludarabine binding sites. Together our data offer a molecular basis that explains STAT cross-binding specificity of stattic and fludarabine, thereby questioning the present selection strategies of SH2 domain-based competitive small inhibitors. © 2013 Elsevier B.V. All rights reserved.

mPGES-1-derived prostaglandin E2 stimulates Stat3 to promote podocyte apoptosis.

PubMed

Yu, Jing; Wu, Yimei; Wang, Lu; Zhang, Wen; Xu, Man; Song, Jiayu; Fu, Yu; Cui, Yiyun; Gong, Wei; Li, Shuzhen; Xia, Weiwei; Huang, Songming; Zhang, Aihua; Jia, Zhanjun

2017-11-01

We previously reported that microsomal prostaglandin E synthase-1 (mPGES-1) contributed to adriamycin (Adr)-induced podocyte apoptosis. However, the molecular mechanism remains unclear. Here we studied the role of mPGES-1/PGE2 cascade in activating Stat3 signaling and the contribution of Stat3 in PGE2- and Adr-induced podocyte apoptosis. In murine podocytes, PGE2 dose- and time-dependently increased the phosphorylation of Stat3 in line with the enhanced cell apoptosis and reduced podocyte protein podocin. In agreement with the increased Stat3 phosphorylation, Stat3-derived cytokines including IL-6, IL-17, MCP-1, and ICAM-1 were significantly upregulated following PGE2 treatment. By application of a specific Stat3 inhibitor S3I-201, PGE2-induced podocyte apoptosis was largely abolished in parallel with a blockade of podocin reduction. Next, we observed that Adr treatment also enhanced p-Stat3 and activated mPGES-1/PGE2 cascade. Blockade of Stat3 by S3I-201 significantly ameliorated Adr-induced cell apoptosis and podocin reduction. More interestingly, silencing mPGES-1 in podocytes by mPGES-1 siRNA blocked Adr-induced increments of Stat-3 phosphorylation, PGE2 production, and Stat3-derived inflammatory cytokines. Taken together, this study suggested that mPGES-1-derived PGE2 could activate Stat3 signaling to promote podocyte apoptosis. Targeting mPGES-1/PGE2/Stat3 signaling might be a potential strategy for the treatment of podocytopathy.

Inhibition of the STAT3 signaling pathway contributes to apigenin-mediated anti-metastatic effect in melanoma

PubMed Central

Cao, Hui-Hui; Chu, Jian-Hong; Kwan, Hiu-Yee; Su, Tao; Yu, Hua; Cheng, Chi-Yan; Fu, Xiu-Qiong; Guo, Hui; Li, Ting; Tse, Anfernee Kai-Wing; Chou, Gui-Xin; Mo, Huan-Biao; Yu, Zhi-Ling

2016-01-01

Signal transducer and activator of transcription 3 (STAT3) signaling is constantly activated in human melanoma, and promotes melanoma metastasis. The dietary flavonoid apigenin is a bioactive compound that possesses low toxicity and exerts anti-metastatic activity in melanoma. However, the anti-metastasis mechanism of apigenin has not been fully elucidated. In the present study, we showed that apigenin suppressed murine melanoma B16F10 cell lung metastasis in mice, and inhibited cell migration and invasion in human and murine melanoma cells. Further study indicated that apigenin effectively suppressed STAT3 phosphorylation, decreased STAT3 nuclear localization and inhibited STAT3 transcriptional activity. Apigenin also down-regulated STAT3 target genes MMP-2, MMP-9, VEGF and Twist1, which are involved in cell migration and invasion. More importantly, overexpression of STAT3 or Twist1 partially reversed apigenin-impaired cell migration and invasion. Our data not only reveal a novel anti-metastasis mechanism of apigenin but also support the notion that STAT3 is an attractive and promising target for melanoma treatment. PMID:26911838

Glutamine Deprivation Causes Hydrogen Peroxide-induced Interleukin-8 Expression via Jak1/Stat3 Activation in Gastric Epithelial AGS Cells

PubMed Central

Lee, Yun Mi; Kim, Mi Jung; Kim, Youngha; Kim, Hyeyoung

2015-01-01

Background: The Janus kinase (Jak)/Signal transducers of activated transcription (Stat) pathway is an upstream signaling pathway for NF-κB activation in Helicobacter pylori-induced interleukin (IL)-8 production in gastric epithelial AGS cells. H. pylori activates NADPH oxidase and produces hydrogen peroxide, which activates Jak1/Stat3 in AGS cells. Therefore, hydrogen peroxide may be critical for IL-8 production via Jak/Stat activation in gastric epithelial cells. Glutamine is depleted during severe injury and stress and contributes to the formation of glutathione (GSH), which is involved in conversion of hydrogen peroxide into water as a cofactor for GSH peroxidase. Methods: We investigated whether glutamine deprivation induces hydrogen peroxide-mediated IL-8 production and whether hydrogen peroxide activates Jak1/Stat3 to induce IL-8 in AGS cells. Cells were cultured in the presence or absence of glutamine or hydrogen peroxide, with or without GSH or a the Jak/Stat specific inhibitor AG490. Results: Glutamine deprivation decreased GSH levels, but increased levels of hydrogen peroxide and IL-8, an effect that was inhibited by treatment with GSH. Hydrogen peroxide induced the activation of Jak1/Stat3 time-dependently. AG490 suppressed hydrogen peroxide- induced activation of Jak1/Stat3 and IL-8 expression in AGS cells, but did not affect levels of reactive oxygen species in AGS cells. Conclusions: In gastric epithelial AGS cells, glutamine deprivation increases hydrogen peroxide levels and IL-8 expression, which may be mediated by Jak1/Stat3 activation. Glutamine supplementation may be beneficial for preventing gastric inflammation by suppressing hydrogen peroxide-mediated Jak1/Stat3 activation and therefore, reducing IL-8 production. Scavenging hydrogen peroxide or targeting Jak1/Stat3 may also prevent oxidant-mediated gastric inflammation. PMID:26473156

Proinflammatory Cytokines IL-6 and TNF-α Increased Telomerase Activity through NF-κB/STAT1/STAT3 Activation, and Withaferin A Inhibited the Signaling in Colorectal Cancer Cells

PubMed Central

Okobi, Quincy; Adekoya, Debbie; Atefi, Mohammad; Clarke, Orette; Dutta, Pranabananda; Vadgama, Jaydutt V.

2017-01-01

There are increasing evidences of proinflammatory cytokine involvement in cancer development. Here, we found that two cytokines, IL-6 and TNF-α, activated colorectal cancer cells to be more invasive and stem-like. Combined treatment of IL-6 and TNF-α phosphorylated transcription factors STAT3 in a synergistic manner. STAT3, STAT1, and NF-κB physically interacted upon the cytokine stimulation. STAT3 was bound to the promoter region of human telomerase reverse transcriptase (hTERT). IL-6 and TNF-α stimulation further enhanced STAT3 binding affinity. Stem cell marker Oct-4 was upregulated in colorectal cancer cells upon IL-6 and TNF-α stimulation. Withaferin A, an anti-inflammatory steroidal lactone, inhibited the IL-6- and TNF-α-induced cancer cell invasion and decreased colonosphere formation. Notably, withaferin A inhibited STAT3 phosphorylation and abolished the STAT3, STAT1, and NF-κB interactions. Oct-4 expression was also downregulated by withaferin A inhibition. The binding of STAT3 to the hTERT promoter region and telomerase activity showed reduction with withaferin A treatments. Proinflammatory cytokine-induced cancer cell invasiveness is mediated by a STAT3-regulated mechanism in colorectal cancer cells. Our data suggest that withaferin A could be a promising anticancer agent that effectively inhibits the progression of colorectal cancer. PMID:28676732

The JAK2 pathway is activated in idiopathic pulmonary fibrosis.

PubMed

Milara, Javier; Hernandez, Gracia; Ballester, Beatriz; Morell, Anselm; Roger, Inés; Montero, P; Escrivá, Juan; Lloris, José M; Molina-Molina, Maria; Morcillo, Esteban; Cortijo, Julio

2018-02-06

Idiopathic pulmonary fibrosis (IPF) is the most rapidly progressive and fatal fibrotic disorder, with no curative therapies. The signal transducer and activator of transcription 3 (STAT3) protein is activated in lung fibroblasts and alveolar type II cells (ATII), thereby contributing to lung fibrosis in IPF. Although activation of Janus kinase 2 (JAK2) has been implicated in proliferative disorders, its role in IPF is unknown. The aim of this study was to analyze JAK2 activation in IPF, and to determine whether JAK2/STAT3 inhibition is a potential therapeutic strategy for this disease. JAK2/p-JAK2 and STAT3/pSTAT3 expression was evaluated using quantitative real time-PCR, western blotting, and immunohistochemistry. Compared to human healthy lung tissue (n = 10) both proteins were upregulated in the lung tissue of IPF patients (n = 12). Stimulating primary ATII and lung fibroblasts with transforming growth factor beta 1 or interleukin (IL)-6/IL-13 activated JAK2 and STAT3, inducing epithelial to mesenchymal and fibroblast to myofibroblast transitions. Dual p-JAK2/p-STAT3 inhibition with JSI-124 or silencing of JAK2 and STAT3 genes suppressed ATII and the fibroblast to myofibroblast transition, with greater effects than the sum of those obtained using JAK2 or STAT3 inhibitors individually. Dual rather than single inhibition was also more effective for inhibiting fibroblast migration, preventing increases in fibroblast senescence and Bcl-2 expression, and ameliorating impaired autophagy. In rats administered JSI-124, a dual inhibitor of p-JAK2/p-STAT3, at a dose of 1 mg/kg/day, bleomycin-induced lung fibrosis was reduced and collagen deposition in the lung was inhibited, as were JAK2 and STAT3 activation and several markers of fibrosis, autophagy, senescence, and anti-apoptosis. JAK2 and STAT3 are activated in IPF, and their dual inhibition may be an attractive strategy for treating this disease.

Targeted inhibition of STATs and IRFs as a potential treatment strategy in cardiovascular disease.

PubMed

Szelag, Malgorzata; Piaszyk-Borychowska, Anna; Plens-Galaska, Martyna; Wesoly, Joanna; Bluyssen, Hans A R

2016-07-26

Key factors contributing to early stages of atherosclerosis and plaque development include the pro-inflammatory cytokines Interferon (IFN)α, IFNγ and Interleukin (IL)-6 and Toll-like receptor 4 (TLR4) stimuli. Together, they trigger activation of Signal Transducer and Activator of Transcription (STAT) and Interferon Regulatory Factor (IRF) families. In particular, STAT1, 2 and 3; IRF1 and 8 have recently been recognized as prominent modulators of inflammation, especially in immune and vascular cells during atherosclerosis. Moreover, inflammation-mediated activation of these STATs and IRFs coordinates a platform for synergistic amplification leading to pro-atherogenic responses.Searches for STAT3-targeting compounds, exploring the pTyr-SH2 interaction area of STAT3, yielded many small molecules including natural products. Only a few inhibitors for other STATs, but none for IRFs, are described. Promising results for several STAT3 inhibitors in recent clinical trials predicts STAT3-inhibiting strategies may find their way to the clinic. However, many of these inhibitors do not seem STAT-specific, display toxicity and are not very potent. This illustrates the need for better models, and screening and validation tools for novel STAT and IRF inhibitors.This review presents a summary of these findings. It postulates STAT1, STAT2 and STAT3 and IRF1 and IRF8 as interesting therapeutic targets and targeted inhibition could be a potential treatment strategy in CVDs. In addition, it proposes a pipeline approach that combines comparative in silico docking of STAT-SH2 and IRF-DBD models with in vitro STAT and IRF activation inhibition validation, as a novel tool to screen multi-million compound libraries and identify specific inhibitors for STATs and IRFs.

Targeted inhibition of STATs and IRFs as a potential treatment strategy in cardiovascular disease

PubMed Central

Szelag, Malgorzata; Piaszyk-Borychowska, Anna; Plens-Galaska, Martyna; Wesoly, Joanna; Bluyssen, Hans A.R.

2016-01-01

Key factors contributing to early stages of atherosclerosis and plaque development include the pro-inflammatory cytokines Interferon (IFN)α, IFNγ and Interleukin (IL)-6 and Toll-like receptor 4 (TLR4) stimuli. Together, they trigger activation of Signal Transducer and Activator of Transcription (STAT) and Interferon Regulatory Factor (IRF) families. In particular, STAT1, 2 and 3; IRF1 and 8 have recently been recognized as prominent modulators of inflammation, especially in immune and vascular cells during atherosclerosis. Moreover, inflammation-mediated activation of these STATs and IRFs coordinates a platform for synergistic amplification leading to pro-atherogenic responses. Searches for STAT3-targeting compounds, exploring the pTyr-SH2 interaction area of STAT3, yielded many small molecules including natural products. Only a few inhibitors for other STATs, but none for IRFs, are described. Promising results for several STAT3 inhibitors in recent clinical trials predicts STAT3-inhibiting strategies may find their way to the clinic. However, many of these inhibitors do not seem STAT-specific, display toxicity and are not very potent. This illustrates the need for better models, and screening and validation tools for novel STAT and IRF inhibitors. This review presents a summary of these findings. It postulates STAT1, STAT2 and STAT3 and IRF1 and IRF8 as interesting therapeutic targets and targeted inhibition could be a potential treatment strategy in CVDs. In addition, it proposes a pipeline approach that combines comparative in silico docking of STAT-SH2 and IRF-DBD models with in vitro STAT and IRF activation inhibition validation, as a novel tool to screen multi-million compound libraries and identify specific inhibitors for STATs and IRFs. PMID:27166190

STATs: An Old Story, Yet Mesmerizing.

PubMed

Abroun, Saeid; Saki, Najmaldin; Ahmadvand, Mohammad; Asghari, Farahnaz; Salari, Fatemeh; Rahim, Fakher

2015-01-01

Signal transducers and activators of transcription (STATs) are cytoplasmic transcription factors that have a key role in cell fate. STATs, a protein family comprised of seven members, are proteins which are latent cytoplasmic transcription factors that convey signals from the cell surface to the nucleus through activation by cytokines and growth factors. The signaling pathways have diverse biological functions that include roles in cell differentiation, proliferation, development, apoptosis, and inflammation which place them at the center of a very active area of research. In this review we explain Janus kinase (JAK)/STAT signaling and focus on STAT3, which is transient from cytoplasm to nucleus after phosphorylation. This procedure controls fundamental biological processes by regulating nuclear genes controlling cell proliferation, survival, and development. In some hematopoietic disorders and cancers, overexpression and activation of STAT3 result in high proliferation, suppression of cell differentiation and inhibition of cell maturation. This article focuses on STAT3 and its role in malignancy, in addition to the role of microRNAs (miRNAs) on STAT3 activation in certain cancers.

Suppression of autophagy augments the radiosensitizing effects of STAT3 inhibition on human glioma cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yuan, Xiaopeng; Du, Jie; Hua, Song

Radiotherapy is an essential component of the standard therapy for newly diagnosed glioblastoma. To increase the radiosensitivity of glioma cells is a feasible solution to improve the therapeutic effects. It has been suggested that inhibition of signal transducer and activator of transcription 3 (STAT3) can radiosensitize glioma cells, probably via the activation of mitochondrial apoptotic pathway. In this study, human malignant glioma cells, U251 and A172, were treated with an STAT3 inhibitor, WP1066, or a short hairpin RNA plasmid targeting STAT3 to suppress the activation of STAT3 signaling. The radiosensitizing effects of STAT3 inhibition were confirmed in glioma cells. Intriguingly,more » combination of ionizing radiation exposure and STAT3 inhibition triggered a pronounced increase of autophagy flux. To explore the role of autophagy, glioma cells were treated with 3-methyladenine or siRNA for autophagy-related gene 5, and it was demonstrated that inhibition of autophagy further strengthened the radiosensitizing effects of STAT3 inhibition. Accordingly, more apoptotic cells were induced by the dual inhibition of autophagy and STAT3 signaling. In conclusion, our data revealed a protective role of autophagy in the radiosensitizing effects of STAT3 inhibition, and inhibition of both autophagy and STAT3 might be a potential therapeutic strategy to increase the radiosensitivity of glioma cells. - Highlights: • Inactivation of STAT3 signaling radiosensitizes malignant glioma cells. • STAT3 inhibition triggers a significant increase of autophagy flux induced by ionizing radiation in glioma cells. • Suppression of autophagy further strengthens the radiosensitizing effects of STAT3 inhibition in glioma cells. • Dual inhibition of autophagy and STAT3 induce massive apoptotic cells upon exposure to ionizing radiation.« less

Activation of Peroxisome Proliferator–Activated Receptor-β/-δ (PPAR-β/-δ) Ameliorates Insulin Signaling and Reduces SOCS3 Levels by Inhibiting STAT3 in Interleukin-6–Stimulated Adipocytes

PubMed Central

Serrano-Marco, Lucía; Rodríguez-Calvo, Ricardo; El Kochairi, Ilhem; Palomer, Xavier; Michalik, Liliane; Wahli, Walter; Vázquez-Carrera, Manuel

2011-01-01

OBJECTIVE It has been suggested that interleukin (IL)-6 is one of the mediators linking obesity-derived chronic inflammation with insulin resistance through activation of STAT3, with subsequent upregulation of suppressor of cytokine signaling 3 (SOCS3). We evaluated whether peroxisome proliferator–activated receptor (PPAR)-β/-δ prevented activation of the IL-6-STAT3-SOCS3 pathway and insulin resistance in adipocytes. RESEARCH DESIGN AND METHODS Adipocytes and white adipose tissue from wild-type and PPAR-β/-δ-null mice were used to evaluate the effect of PPAR-β/-δ on the IL-6-STAT3-SOCS3 pathway. RESULTS First, we observed that the PPAR-β/-δ agonist GW501516 prevented both IL-6–dependent reduction in insulin-stimulated Akt phosphorylation and glucose uptake in adipocytes. In addition, this drug treatment abolished IL-6–induced SOCS3 expression in differentiated 3T3-L1 adipocytes. This effect was associated with the capacity of the drug to prevent IL-6–induced STAT3 phosphorylation on Tyr705 and Ser727 residues in vitro and in vivo. Moreover, GW501516 prevented IL-6–dependent induction of extracellular signal–related kinase (ERK)1/2, a serine-threonine-protein kinase involved in serine STAT3 phosphorylation. Furthermore, in white adipose tissue from PPAR-β/-δ–null mice, STAT3 phosphorylation (Tyr705 and Ser727), STAT3 DNA-binding activity, and SOCS3 protein levels were higher than in wild-type mice. Several steps in STAT3 activation require its association with heat shock protein 90 (Hsp90), which was prevented by GW501516 as revealed in immunoprecipitation studies. Consistent with this finding, the STAT3-Hsp90 association was enhanced in white adipose tissue from PPAR-β/-δ–null mice compared with wild-type mice. CONCLUSIONS Collectively, our findings indicate that PPAR-β/-δ activation prevents IL-6–induced STAT3 activation by inhibiting ERK1/2 and preventing the STAT3-Hsp90 association, an effect that may contribute to the prevention of cytokine-induced insulin resistance in adipocytes. PMID:21617181

Dynamic mathematical modeling of IL13-induced signaling in Hodgkin and primary mediastinal B-cell lymphoma allows prediction of therapeutic targets.

PubMed

Raia, Valentina; Schilling, Marcel; Böhm, Martin; Hahn, Bettina; Kowarsch, Andreas; Raue, Andreas; Sticht, Carsten; Bohl, Sebastian; Saile, Maria; Möller, Peter; Gretz, Norbert; Timmer, Jens; Theis, Fabian; Lehmann, Wolf-Dieter; Lichter, Peter; Klingmüller, Ursula

2011-02-01

Primary mediastinal B-cell lymphoma (PMBL) and classical Hodgkin lymphoma (cHL) share a frequent constitutive activation of JAK (Janus kinase)/STAT signaling pathway. Because of complex, nonlinear relations within the pathway, key dynamic properties remained to be identified to predict possible strategies for intervention. We report the development of dynamic pathway models based on quantitative data collected on signaling components of JAK/STAT pathway in two lymphoma-derived cell lines, MedB-1 and L1236, representative of PMBL and cHL, respectively. We show that the amounts of STAT5 and STAT6 are higher whereas those of SHP1 are lower in the two lymphoma cell lines than in normal B cells. Distinctively, L1236 cells harbor more JAK2 and less SHP1 molecules per cell than MedB-1 or control cells. In both lymphoma cell lines, we observe interleukin-13 (IL13)-induced activation of IL4 receptor α, JAK2, and STAT5, but not of STAT6. Genome-wide, 11 early and 16 sustained genes are upregulated by IL13 in both lymphoma cell lines. Specifically, the known STAT-inducible negative regulators CISH and SOCS3 are upregulated within 2 hours in MedB-1 but not in L1236 cells. On the basis of this detailed quantitative information, we established two mathematical models, MedB-1 and L1236 model, able to describe the respective experimental data. Most of the model parameters are identifiable and therefore the models are predictive. Sensitivity analysis of the model identifies six possible therapeutic targets able to reduce gene expression levels in L1236 cells and three in MedB-1. We experimentally confirm reduction in target gene expression in response to inhibition of STAT5 phosphorylation, thereby validating one of the predicted targets.

Small-molecule targeting of signal transducer and activator of transcription (STAT) 3 to treat non-small cell lung cancer.

PubMed

Lewis, Katherine M; Bharadwaj, Uddalak; Eckols, T Kris; Kolosov, Mikhail; Kasembeli, Moses M; Fridley, Colleen; Siller, Ricardo; Tweardy, David J

2015-11-01

Lung cancer is the leading cause of cancer death in both men and women. Non-small cell lung cancer (NSCLC) has an overall 5-year survival rate of 15%. While aberrant STAT3 activation has previously been observed in NSCLC, the scope of its contribution is uncertain and agents that target STAT3 for treatment are not available clinically. We determined levels of activated STAT3 (STAT3 phosphorylated on Y705, pSTAT3) and the two major isoforms of STAT3 (α and β) in protein extracts of 8 NSCLC cell lines, as well as the effects of targeting STAT3 in vitro and in vivo in NSCLC cells using short hairpin (sh) RNA and two novel small-molecule STAT3 inhibitors, C188-9 and piperlongumine (PL). Levels of pSTAT3, STAT3α, and STATβ were increased in 7 of 8 NSCLC cell lines. Of note, levels of pSTAT3 were tightly correlated with levels of STAT3β, but not STAT3α. Targeting of STAT3 in A549 cells using shRNA decreased tSTAT3 by 75%; this was accompanied by a 47-78% reduction in anchorage-dependent and anchorage-independent growth and a 28-45% reduction in mRNA levels for anti-apoptotic STAT3 gene targets. C188-9 and PL (@30 μM) each reduced pSTAT3 levels in all NSCLC cell lines tested by ≥50%, reduced anti-apoptotic protein mRNA levels by 25-60%, and reduced both anchorage-dependent and anchorage-independent growth of NSCLC cell lines with IC50 values ranging from 3.06 to 52.44 μM and 0.86 to 11.66 μM, respectively. Treatment of nude mice bearing A549 tumor xenografts with C188-9 or PL blocked tumor growth and reduced levels of pSTAT3 and mRNA encoding anti-apoptotic proteins. STAT3 is essential for growth of NSCLC cell lines and tumors and its targeting using C188-9 or PL may be a useful strategy for treatment. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

MMPP Attenuates Non-Small Cell Lung Cancer Growth by Inhibiting the STAT3 DNA-Binding Activity via Direct Binding to the STAT3 DNA-Binding Domain.

PubMed

Son, Dong Ju; Zheng, Jie; Jung, Yu Yeon; Hwang, Chul Ju; Lee, Hee Pom; Woo, Ju Rang; Baek, Song Yi; Ham, Young Wan; Kang, Min Woong; Shong, Minho; Kweon, Gi Ryang; Song, Min Jong; Jung, Jae Kyung; Han, Sang-Bae; Kim, Bo Yeon; Yoon, Do Young; Choi, Bu Young; Hong, Jin Tae

2017-01-01

Rationale: Signal transducer and activator of transcription-3 (STAT3) plays a pivotal role in cancer biology. Many small-molecule inhibitors that target STAT3 have been developed as potential anticancer drugs. While designing small-molecule inhibitors that target the SH2 domain of STAT3 remains the leading focus for drug discovery, there has been a growing interest in targeting the DNA-binding domain (DBD) of the protein. Methods: We demonstrated the potential antitumor activity of a novel, small-molecule (E)-2-methoxy-4-(3-(4-methoxyphenyl)prop-1-en-1-yl)phenol (MMPP) that directly binds to the DBD of STAT3, in patient-derived non-small cell lung cancer (NSCLC) xenograft model as well as in NCI-H460 cell xenograft model in nude mice. Results: MMPP effectively inhibited the phosphorylation of STAT3 and its DNA binding activity in vitro and in vivo . It induced G1-phase cell cycle arrest and apoptosis through the regulation of cell cycle- and apoptosis-regulating genes by directly binding to the hydroxyl residue of threonine 456 in the DBD of STAT3. Furthermore, MMPP showed a similar or better antitumor activity than that of docetaxel or cisplatin. Conclusion: MMPP is suggested to be a potential candidate for further development as an anticancer drug that targets the DBD of STAT3.

Cyclophilins contribute to Stat3 signaling and survival of multiple myeloma cells.

PubMed

Bauer, K; Kretzschmar, A K; Cvijic, H; Blumert, C; Löffler, D; Brocke-Heidrich, K; Schiene-Fischer, C; Fischer, G; Sinz, A; Clevenger, C V; Horn, F

2009-08-06

Signal transducer and activator of transcription 3 (Stat3) is the major mediator of interleukin-6 (IL-6) family cytokines. In addition, Stat3 is known to be involved in the pathophysiology of many malignancies. Here, we show that the cis-trans peptidyl-prolyl isomerase cyclophilin (Cyp) B specifically interacts with Stat3, whereas the highly related CypA does not. CypB knockdown inhibited the IL-6-induced transactivation potential but not the tyrosine phosphorylation of Stat3. Binding of CypB to Stat3 target promoters and alteration of the intranuclear localization of Stat3 on CypB depletion suggested a nuclear function of Stat3/CypB interaction. By contrast, CypA knockdown inhibited Stat3 IL-6-induced tyrosine phosphorylation and nuclear translocation. The Cyp inhibitor cyclosporine A (CsA) caused similar effects. However, Stat1 activation in response to IL-6 or interferon-gamma was not affected by Cyp silencing or CsA treatment. As a result, Cyp knockdown shifted IL-6 signaling to a Stat1-dominated pathway. Furthermore, Cyp depletion or treatment with CsA induced apoptosis in IL-6-dependent multiple myeloma cells, whereas an IL-6-independent line was not affected. Thus, Cyps support the anti-apoptotic action of Stat3. Taken together, CypA and CypB both play pivotal roles, yet at different signaling levels, for Stat3 activation and function. These data also suggest a novel mechanism of CsA action.

Gain-of-function mutations in the gene encoding the tyrosine phosphatase SHP2 induce hydrocephalus in a catalytically dependent manner.

PubMed

Zheng, Hong; Yu, Wen-Mei; Waclaw, Ronald R; Kontaridis, Maria I; Neel, Benjamin G; Qu, Cheng-Kui

2018-03-20

Catalytically activating mutations in Ptpn11 , which encodes the protein tyrosine phosphatase SHP2, cause 50% of Noonan syndrome (NS) cases, whereas inactivating mutations in Ptpn11 are responsible for nearly all cases of the similar, but distinct, developmental disorder Noonan syndrome with multiple lentigines (NSML; formerly called LEOPARD syndrome). However, both types of disease mutations are gain-of-function mutations because they cause SHP2 to constitutively adopt an open conformation. We found that the catalytic activity of SHP2 was required for the pathogenic effects of gain-of-function, disease-associated mutations on the development of hydrocephalus in the mouse. Targeted pan-neuronal knockin of a Ptpn11 allele encoding the active SHP2 E76K mutant resulted in hydrocephalus due to aberrant development of ependymal cells and their cilia. These pathogenic effects of the E76K mutation were suppressed by the additional mutation C459S, which abolished the catalytic activity of SHP2. Moreover, ependymal cells in NSML mice bearing the inactive SHP2 mutant Y279C were also unaffected. Mechanistically, the SHP2 E76K mutant induced developmental defects in ependymal cells by enhancing dephosphorylation and inhibition of the transcription activator STAT3. Whereas STAT3 activity was reduced in Ptpn11 E76K/+ cells, the activities of the kinases ERK and AKT were enhanced, and neural cell-specific Stat3 knockout mice also manifested developmental defects in ependymal cells and cilia. These genetic and biochemical data demonstrate a catalytic-dependent role of SHP2 gain-of-function disease mutants in the pathogenesis of hydrocephalus. Copyright © 2018 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

STAT3, p-STAT3 and HIF-1α are associated with vasculogenic mimicry and impact on survival in gastric adenocarcinoma

PubMed Central

SONG, YAN-YAN; SUN, LI-DAN; LIU, MIN-LI; LIU, ZHONG-LIANG; CHEN, FEI; ZHANG, YING-ZHE; ZHENG, YAN; ZHANG, JIAN-PING

2014-01-01

Vasculogenic mimicry (VM) formation is important for invasion and metastasis of tumor cells in gastric adenocarcinoma (GAC). The present study aimed to investigate the association between signal transducer and activator of transcription-3 (STAT3), phosphor-STAT3 (p-STAT3), hypoxia-inducible factor-1α (HIF-1α) and VM formation in GAC, and discuss their clinical significance and correlation with the prognosis of patients with GAC. The expression levels of STAT3, p-STAT3, HIF-1α and VM were assessed in 60 cases of patients with GAC and 20 cases of patients with gastritis on tissue microarrays by immunohistochemical methods. The expression levels of STAT3, p-STAT3, HIF-1α and VM were higher in patients with GAC (particularly in poorly differentiated GAC) than in those with gastritis (P<0.05). The expression levels of STAT3, p-STAT3 and HIF-1α were higher in VM tissues compared with non-VM tissues (P<0.05). Positive correlations existed between STAT3, p-STAT3, HIF-1α and VM expression (P<0.05). The expression levels of STAT3, p-STAT3 and HIF-1α, VM, status of lymph node metastasis and tumor differentiation degree were associated with the overall survival time of patients with GAC (P<0.05). However, only p-STAT3 and VM expression were identified as the independent risk factors of GAC OS when analyzed with multivariate analysis. p-STAT3 and VM play a significant role in indicating the prognosis of patients with GAC. STAT3 activation may play a positive role in VM formation of GAC by the STAT3-p-STAT3-HIF-1α-VM effect axis. PMID:24959290

SPARC expression induces cell cycle arrest via STAT3 signaling pathway in medulloblastoma cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chetty, Chandramu; Dontula, Ranadheer; Ganji, Purnachandra Nagaraju

2012-01-13

Highlights: Black-Right-Pointing-Pointer Ectopic expression of SPARC impaired cell proliferation in medulloblastoma cells. Black-Right-Pointing-Pointer SPARC expression induces STAT3 mediated cell cycle arrest in medulloblastoma cells. Black-Right-Pointing-Pointer SPARC expression significantly inhibited pre-established tumor growth in nude-mice. -- Abstract: Dynamic cell interaction with ECM components has profound influence in cancer progression. SPARC is a component of the ECM, impairs the proliferation of different cell types and modulates tumor cell aggressive features. We previously reported that SPARC expression significantly impairs medulloblastoma tumor growth in vivo. In this study, we demonstrate that expression of SPARC inhibits medulloblastoma cell proliferation. MTT assay indicated a dose-dependent reductionmore » in tumor cell proliferation in adenoviral mediated expression of SPARC full length cDNA (Ad-DsRed-SP) in D425 and UW228 cells. Flow cytometric analysis showed that Ad-DsRed-SP-infected cells accumulate in the G2/M phase of cell cycle. Further, immunoblot and immunoprecipitation analyses revealed that SPARC induced G2/M cell cycle arrest was mediated through inhibition of the Cyclin-B-regulated signaling pathway involving p21 and Cdc2 expression. Additionally, expression of SPARC decreased STAT3 phosphorylation at Tyr-705; constitutively active STAT3 expression reversed SPARC induced G2/M arrest. Ad-DsRed-SP significantly inhibited the pre-established orthotopic tumor growth and tumor volume in nude-mice. Immunohistochemical analysis of tumor sections from mice treated with Ad-DsRed-SP showed decreased immunoreactivity for pSTAT3 and increased immunoreactivity for p21 compared to tumor section from mice treated with mock and Ad-DsRed. Taken together our studies further reveal that STAT3 plays a key role in SPARC induced G2/M arrest in medulloblastoma cells. These new findings provide a molecular basis for the mechanistic understanding of the effects of SPARC on medulloblastoma tumor cell proliferation.« less

STAT3 and STAT1 mediate IL-11–dependent and inflammation-associated gastric tumorigenesis in gp130 receptor mutant mice

PubMed Central

Ernst, Matthias; Najdovska, Meri; Grail, Dianne; Lundgren-May, Therese; Buchert, Michael; Tye, Hazel; Matthews, Vance B.; Armes, Jane; Bhathal, Prithi S.; Hughes, Norman R.; Marcusson, Eric G.; Karras, James G.; Na, Songqing; Sedgwick, Jonathon D.; Hertzog, Paul J.; Jenkins, Brendan J.

2008-01-01

Deregulated activation of STAT3 is frequently associated with many human hematological and epithelial malignancies, including gastric cancer. While exaggerated STAT3 signaling facilitates an antiapoptotic, proangiogenic, and proproliferative environment for neoplastic cells, the molecular mechanisms leading to STAT3 hyperactivation remain poorly understood. Using the gp130Y757F/Y757F mouse model of gastric cancer, which carries a mutated gp130 cytokine receptor signaling subunit that cannot bind the negative regulator of cytokine signaling SOCS3 and is characterized by hyperactivation of the signaling molecules STAT1 and STAT3, we have provided genetic evidence that IL-11 promotes chronic gastric inflammation and associated tumorigenesis. Expression of IL-11 was increased in gastric tumors in gp130Y757F/Y757F mice, when compared with unaffected gastric tissue in wild-type mice, while gp130Y757F/Y757F mice lacking the IL-11 ligand–binding receptor subunit (IL-11Rα) showed normal gastric STAT3 activation and IL-11 expression and failed to develop gastric tumors. Furthermore, reducing STAT3 activity in gp130Y757F/Y757F mice, either genetically or by therapeutic administration of STAT3 antisense oligonucleotides, normalized gastric IL-11 expression and alleviated gastric tumor burden. Surprisingly, the genetic reduction of STAT1 expression also reduced gastric tumorigenesis in gp130Y757F/Y757F mice and coincided with reduced gastric inflammation and IL-11 expression. Collectively, our data have identified IL-11 as a crucial cytokine promoting chronic gastric inflammation and associated tumorigenesis mediated by excessive activation of STAT3 and STAT1. PMID:18431520

Contribution of JAK2 mutations to T-cell lymphoblastic lymphoma development.

PubMed

Roncero, A M; López-Nieva, P; Cobos-Fernández, M A; Villa-Morales, M; González-Sánchez, L; López-Lorenzo, J L; Llamas, P; Ayuso, C; Rodríguez-Pinilla, S M; Arriba, M C; Piris, M A; Fernández-Navarro, P; Fernández, A F; Fraga, M F; Santos, J; Fernández-Piqueras, J

2016-01-01

The JAK-STAT pathway has a substantial role in lymphoid precursor cell proliferation, survival and differentiation. Nonetheless, the contribution of JAK2 to T-cell lymphoblastic lymphoma (T-LBL) development remains poorly understood. We have identified one activating TEL-JAK2 translocation and four missense mutations accumulated in 2 out of 16 T-LBL samples. Two of them are novel JAK2 mutations and the other two are reported for the first time in T-LBL. Notably, R683G and I682T might have arisen owing to RNA editing. Mutated samples showed different mutated transcripts suggesting sub-clonal heterogeneity. Functional approaches revealed that two JAK2 mutations (H574R and R683G) constitutively activate JAK-STAT signaling in γ2A cells and can drive the proliferation of BaF3-EpoR cytokine-dependent cell line. In addition, aberrant hypermethylation of SOCS3 might contribute to enhance the activation of JAK-STAT signaling. Of utmost interest is that primary T-LBL samples harboring JAK2 mutations exhibited increased expression of LMO2, suggesting a mechanistic link between JAK2 mutations and the expression of LMO2, which was confirmed for the four missense mutations in transfected γ2A cells. We therefore propose that active JAK2 contribute to T-LBL development by two different mechanisms, and that the use of pan-JAK inhibitors in combination with epigenetic drugs should be considered in future treatments.

Contribution of JAK2 mutations to T-cell lymphoblastic lymphoma development

PubMed Central

Roncero, A M; López-Nieva, P; Cobos-Fernández, M A; Villa-Morales, M; González-Sánchez, L; López-Lorenzo, J L; Llamas, P; Ayuso, C; Rodríguez-Pinilla, S M; Arriba, M C; Piris, M A; Fernández-Navarro, P; Fernández, A F; Fraga, M F; Santos, J; Fernández-Piqueras, J

2016-01-01

The JAK-STAT pathway has a substantial role in lymphoid precursor cell proliferation, survival and differentiation. Nonetheless, the contribution of JAK2 to T-cell lymphoblastic lymphoma (T-LBL) development remains poorly understood. We have identified one activating TEL-JAK2 translocation and four missense mutations accumulated in 2 out of 16 T-LBL samples. Two of them are novel JAK2 mutations and the other two are reported for the first time in T-LBL. Notably, R683G and I682T might have arisen owing to RNA editing. Mutated samples showed different mutated transcripts suggesting sub-clonal heterogeneity. Functional approaches revealed that two JAK2 mutations (H574R and R683G) constitutively activate JAK-STAT signaling in γ2A cells and can drive the proliferation of BaF3-EpoR cytokine-dependent cell line. In addition, aberrant hypermethylation of SOCS3 might contribute to enhance the activation of JAK-STAT signaling. Of utmost interest is that primary T-LBL samples harboring JAK2 mutations exhibited increased expression of LMO2, suggesting a mechanistic link between JAK2 mutations and the expression of LMO2, which was confirmed for the four missense mutations in transfected γ2A cells. We therefore propose that active JAK2 contribute to T-LBL development by two different mechanisms, and that the use of pan-JAK inhibitors in combination with epigenetic drugs should be considered in future treatments. PMID:26216197

Novel quinazoline-based sulfonamide derivative (3D) induces apoptosis in colorectal cancer by inhibiting JAK2-STAT3 pathway.

PubMed

Al-Obeed, Omar; Vaali-Mohammed, Mansoor-Ali; Eldehna, Wagdy M; Al-Khayal, Khayal; Mahmood, Amer; Abdel-Aziz, Hatem A; Zubaidi, Ahmed; Alafeefy, Ahmed; Abdulla, Maha; Ahmad, Rehan

2018-01-01

Colorectal cancer (CRC) is a major worldwide health problem owing to its high prevalence and mortality rate. Developments in screening, prevention, biomarker, personalized therapies and chemotherapy have improved detection and treatment. However, despite these advances, many patients with advanced metastatic tumors still succumb to the disease. New anticancer agents are needed for treating advanced stage CRC as most of the deaths occur due to cancer metastasis. A recently developed novel sulfonamide derivative 4-((2-(4-(dimethylamino) phenyl)quinazolin-4-yl)amino)benzenesulfonamide (3D) has shown potent antitumor effect; however, the mechanism underlying the antitumor effect remains unknown. 3D-mediated inhibition on cell viability was evaluated by MTT and real-time cell proliferation was measured by xCelligence RTDP instrument. Western blotting was used to measure pro-apoptotic, anti-apoptotic proteins and JAK2-STAT3 phosphorylation. Flow cytometry was used to measure ROS production and apoptosis. Our study revealed that 3D treatment significantly reduced the viability of human CRC cells HT-29 and SW620. Furthermore, 3D treatment induced the generation of reactive oxygen species (ROS) in human CRC cells. Confirming our observation, N-acetylcysteine significantly inhibited apoptosis. This is further evidenced by the induction of p53 and Bax; release of cytochrome c; activation of caspase-9, caspase-7 and caspase-3; and cleavage of PARP in 3D-treated cells. This compound was found to have a significant effect on the inhibition of antiapoptotic proteins Bcl2 and BclxL. The results further demonstrate that 3D inhibits JAK2-STAT3 pathway by decreasing the constitutive and IL-6-induced phosphorylation of STAT3. 3D also decreases STAT3 target genes such as cyclin D1 and survivin. Furthermore, a combination study of 3D with doxorubicin (Dox) also showed more potent effects than single treatment of Dox in the inhibition of cell viability. Taken together, these findings indicate that 3D induces ROS-mediated apoptosis and inhibits JAK2-STAT3 signaling in CRC.

High mobility group box 1 induces the activation of the Janus kinase 2 and signal transducer and activator of transcription 3 (JAK2/STAT3) signaling pathway in pancreatic acinar cells in rats, while AG490 and rapamycin inhibit their activation.

PubMed

Wang, Guoliang; Zhang, Jingchao; Dui, Danhua; Ren, Haoyuan; Liu, Jin

2016-11-10

The pathogenesis of severe acute pancreatitis (SAP) remains unclear. The Janus kinase and signal transducer and activator of transcription (JAK/STAT) pathway is important for various cytokines and growth factors. This study investigated the effect of the late inflammatory factor high mobility group box 1 (HMGB1) on the activation of JAK2/STAT3 in pancreatic acinar cells and the inhibitory effects of AG490 (a JAK2 inhibitor) and rapamycin (a STAT3 inhibitor) on this pathway. Rat pancreatic acinar cells were randomly divided into the control, HMGB1, AG490, and rapamycin groups. The mRNA levels of JAK2 and STAT3 at 10, 30, 60, and 120 minutes were detected using reverse transcription polymerase chain reaction (RT-PCR). The protein levels of JAK2 and STAT3 at 60 and 120 minutes were observed using Western blotting. Compared with the control group, the HMGB1 group exhibited significantly increased levels of JAK2 mRNA at each time point; STAT3 mRNA at 30, 60, and 120 minutes; and JAK2 and STAT3 proteins at 60 and 120 minutes (p < 0.01). Compared with the HMGB1 group, the AG490 and rapamycin groups both exhibited significantly decreased levels of JAK2 mRNA at each time point (p < 0.05); STAT3 mRNA at 30, 60, and 120 minutes (p < 0.01); and JAK2 and STAT3 proteins at 60 and 120 minutes (p < 0.01). HMGB1 induces the activation of the JAK2/STAT3 signaling pathway in rat pancreatic acinar cells, and this activation can be inhibited by AG490 and rapamycin. The results of this study may provide new insights for the treatment of SAP.

Truncated p110 ERBB2 induces mammary epithelial cell migration, invasion and orthotopic xenograft formation, and is associated with loss of phosphorylated STAT5.

PubMed

Ward, T M; Iorns, E; Liu, X; Hoe, N; Kim, P; Singh, S; Dean, S; Jegg, A-M; Gallas, M; Rodriguez, C; Lippman, M; Landgraf, R; Pegram, M D

2013-05-09

Truncated-ERBB2 isoforms (t-ERBB2s), resulting from receptor proteolysis or alternative translation of the ERBB2 mRNA, exist in a subset of human breast tumors. t-ERBB2s lack the receptor extracellular domain targeted by therapeutic anti-ERBB2 antibodies and antibody-drug conjugates, including trastuzumab, trastuzumab-DM1 and pertuzumab. In clinical studies, expression of t-ERBB2 in breast tumors correlates with metastasis as well as trastuzumab resistance. By using a novel immuno-microarray method, we detect a significant t-ERBB2 fraction in 18 of 31 (58%) of immunohistochemistry (IHC)3+ ERBB2+ human tumor specimens, and further show that t-ERBB2 isoforms are phosphorylated in a subset of IHC3+ samples (10 of 31, 32%). We investigated t-ERBB2 biological activity via engineered expression of full-length and truncated ERBB2 isoforms in human mammary epithelial cells (HMECs), including HMEC and MCF10A cells. Expression of p110 t-ERBB2, but not p95m (m=membrane, also 648CTF) or intracellular ERBB2s, significantly enhanced cell migration and invasion in multiple cell types. In addition, only expression of the p110 isoform led to human breast epithelial cell (HMLE) xenograft formation in vivo. Expression of t-ERBB2s did not result in hyperactivation of the phosphoinositide kinase-3/AKT or mitogen-activated protein kinase signaling pathways in these cells; rather, phosphoproteomic array profiling revealed attenuation of phosphorylated signal transducer and activator of transcription 5 (STAT5) in p110-t-ERBB2-expressing cells compared to controls. Short hairpin-mediated silencing of STAT5 phenocopied p110-t-ERBB2-driven cell migration and invasion, while expression of constitutively active STAT5 reversed these effects. Thus, we provide novel evidence that (1) expression of p110 t-ERBB2 is sufficient for full transformation of HMEC, yielding in vivo xenograft formation, and (2) truncated p110 t-ERBB2 expression is associated with decreased phosphorylation of STAT5.

Truncated p110 ERBB2 induces mammary epithelial cell migration, invasion and orthotopic xenograft formation, and is associated with loss of phosphorylated STAT5

PubMed Central

Ward, T M; Iorns, E; Liu, X; Hoe, N; Kim, P; Singh, S; Dean, S; Jegg, A-M; Gallas, M; Rodriguez, C; Lippman, M; Landgraf, R; Pegram, M D

2013-01-01

Truncated-ERBB2 isoforms (t-ERBB2s), resulting from receptor proteolysis or alternative translation of the ERBB2 mRNA, exist in a subset of human breast tumors. t-ERBB2s lack the receptor extracellular domain targeted by therapeutic anti-ERBB2 antibodies and antibody–drug conjugates, including trastuzumab, trastuzumab-DM1 and pertuzumab. In clinical studies, expression of t-ERBB2 in breast tumors correlates with metastasis as well as trastuzumab resistance. By using a novel immuno-microarray method, we detect a significant t-ERBB2 fraction in 18 of 31 (58%) of immunohistochemistry (IHC)3+ ERBB2+ human tumor specimens, and further show that t-ERBB2 isoforms are phosphorylated in a subset of IHC3+ samples (10 of 31, 32%). We investigated t-ERBB2 biological activity via engineered expression of full-length and truncated ERBB2 isoforms in human mammary epithelial cells (HMECs), including HMEC and MCF10A cells. Expression of p110 t-ERBB2, but not p95m (m=membrane, also 648CTF) or intracellular ERBB2s, significantly enhanced cell migration and invasion in multiple cell types. In addition, only expression of the p110 isoform led to human breast epithelial cell (HMLE) xenograft formation in vivo. Expression of t-ERBB2s did not result in hyperactivation of the phosphoinositide kinase-3/AKT or mitogen-activated protein kinase signaling pathways in these cells; rather, phosphoproteomic array profiling revealed attenuation of phosphorylated signal transducer and activator of transcription 5 (STAT5) in p110-t-ERBB2-expressing cells compared to controls. Short hairpin-mediated silencing of STAT5 phenocopied p110-t-ERBB2-driven cell migration and invasion, while expression of constitutively active STAT5 reversed these effects. Thus, we provide novel evidence that (1) expression of p110 t-ERBB2 is sufficient for full transformation of HMEC, yielding in vivo xenograft formation, and (2) truncated p110 t-ERBB2 expression is associated with decreased phosphorylation of STAT5. PMID:22751112

Temporal regulation of Stat5 activity in determination of cell differentiation program

PubMed Central

Hoshino, Akemi; Fujii, Hodaka

2007-01-01

Although Stat5 is activated by various cytokines, only ethrytopoietin (Epo) and a small number of cytokines induce Stat5-dependent erythroid differentiation. Here, by using a reporter gene system to monitor transcriptional activity of Stat5, we showed that Epo but not interleukin (IL)-3 supports sustained activation of Stat5, which induces globin gene expression. IL-3 or IL-2 stimulation inhibits Epo-induced globin gene expression. The acidic region of the IL-2 receptor β chain was essential for this inhibition. These results underscore the importance of temporal regulation of Stat activity for regulation of cytokine-specific cell differentiation. PMID:17511959

Dihydroartemisinin as a Putative STAT3 Inhibitor, Suppresses the Growth of Head and Neck Squamous Cell Carcinoma by Targeting Jak2/STAT3 Signaling

PubMed Central

Jia, Lifeng; Song, Qi; Zhou, Chenyang; Li, Xiaoming; Pi, Lihong; Ma, Xiuru; Li, Hui; Lu, Xiuying; Shen, Yupeng

2016-01-01

Developing drugs that can effectively block STAT3 activation may serve as one of the most promising strategy for cancer treatment. Currently, there is no putative STAT3 inhibitor that can be safely and effectively used in clinic. In the present study, we investigated the potential of dihydroartemisinin (DHA) as a putative STAT3 inhibitor and its antitumor activities in head and neck squamous cell carcinoma (HNSCC). The inhibitory effects of DHA on STAT3 activation along with its underlying mechanisms were studied in HNSCC cells. The antitumor effects of DHA against HNSCC cells were explored both in vitro and in vivo. An investigation on cooperative effects of DHA with cisplatin in killing HNSCC cells was also implemented. DHA exhibited remarkable and specific inhibitory effects on STAT3 activation via selectively blocking Jak2/STAT3 signaling. Besides, DHA significantly inhibited HNSCC growth both in vitro and in vivo possibly through induction of apoptosis and attenuation of cell migration. DHA also synergized with cisplatin in tumor inhibition in HNSCC cells. Our findings demonstrate that DHA is a putative STAT3 inhibitor that may represent a new and effective drug for cancer treatment and therapeutic sensitization in HNSCC patients. PMID:26784960

STAT3 activation in pressure-overloaded feline myocardium: role for integrins and the tyrosine kinase BMX.

PubMed

Willey, Christopher D; Palanisamy, Arun P; Johnston, Rebecca K; Mani, Santhosh K; Shiraishi, Hirokazu; Tuxworth, William J; Zile, Michael R; Balasubramanian, Sundaravadivel; Kuppuswamy, Dhandapani

2008-06-27

Growth, survival and cytoskeletal rearrangement of cardiomyocytes are critical for cardiac hypertrophy. Signal transducer and activator of transcription-3 (STAT3) activation is an important cardioprotective factor associated with cardiac hypertrophy. Although STAT3 activation has been reported via signaling through Janus Kinase 2 (JAK2) in several cardiac models of hypertrophy, the importance of other nonreceptor tyrosine kinases (NTKs) has not been explored. Utilizing an in vivo feline right ventricular pressure-overload (RVPO) model of hypertrophy, we demonstrate that in 48 h pressure-overload (PO) myocardium, STAT3 becomes phosphorylated and redistributed to detergent-insoluble fractions with no accompanying JAK2 activation. PO also caused increased levels of phosphorylated STAT3 in both cytoplasmic and nuclear fractions. To investigate the role of other NTKs, we used our established in vitro cell culture model of hypertrophy where adult feline cardiomyocytes are embedded three-dimensionally (3D) in type-I collagen and stimulated with an integrin binding peptide containing an Arg-Gly-Asp (RGD) motif that we have previously shown to recapitulate the focal adhesion complex (FAC) formation of 48 h RVPO. RGD stimulation of adult cardiomyocytes in vitro caused both STAT3 redistribution and activation that were accompanied by the activation and redistribution of c-Src and the TEC family kinase, BMX, but not JAK2. However, infection with dominant negative c-Src adenovirus was unable to block RGD-stimulated changes on either STAT3 or BMX. Further analysis in vivo in 48 h PO myocardium showed the presence of both STAT3 and BMX in the detergent-insoluble fraction with their complex formation and phosphorylation. Therefore, these studies indicate a novel mechanism of BMX-mediated STAT3 activation within a PO model of cardiac hypertrophy that might contribute to cardiomyocyte growth and survival.

STAT3 signaling pathway is necessary for cell survival and tumorsphere forming capacity in ALDH{sup +}/CD133{sup +} stem cell-like human colon cancer cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lin, Li, E-mail: lin.796@osu.edu; Division of Cardiology, Department of Internal Medicine, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030; Fuchs, James

2011-12-16

Highlights: Black-Right-Pointing-Pointer The phosphorylated or activated form of STAT3 was expressed in colon cancer stem-like cells. Black-Right-Pointing-Pointer STAT3 inhibitor, FLLL32 inhibits P-STAT3 and STAT3 target genes in colon cancer stem-like cells. Black-Right-Pointing-Pointer Inhibition of STAT3 resulted in decreased cell viability and reduced numbers of tumorspheres. Black-Right-Pointing-Pointer STAT3 is required for survival and tumorsphere forming capacity in colon cancer stem-like cells. Black-Right-Pointing-Pointer Targeting STAT3 in cancer stem-like cells may offer a novel treatment approach for colon cancer. -- Abstract: Persistent activation of Signal Transducers and Activators of Transcription 3 (STAT3) is frequently detected in colon cancer. Increasing evidence suggests the existencemore » of a small population of colon cancer stem or cancer-initiating cells may be responsible for tumor initiation, metastasis, and resistance to chemotherapy and radiation. Whether STAT3 plays a role in colon cancer-initiating cells and the effect of STAT3 inhibition is still unknown. Flow cytometry was used to isolate colon cancer stem-like cells from three independent human colon cancer cell lines characterized by both aldehyde dehydrogenase (ALDH)-positive and CD133-positive subpopulation (ALDH{sup +}/CD133{sup +}). The effects of STAT3 inhibition in colon cancer stem-like cells were examined. The phosphorylated or activated form of STAT3 was expressed in colon cancer stem-like cells and was reduced by a STAT3-selective small molecular inhibitor, FLLL32. FLLL32 also inhibited the expression of potential STAT3 downstream target genes in colon cancer stem-like cells including survivin, Bcl-XL, as well as Notch-1, -3, and -4, which may be involved in stem cell function. Furthermore, FLLL32 inhibited cell viability and tumorsphere formation as well as induced cleaved caspase-3 in colon cancer stem-like cells. FLLL32 is more potent than curcumin as evidenced with lower IC50 in colon cancer stem-like cells. In summary, our results indicate that STAT3 is a novel therapeutic target in colon cancer stem-like cells and inhibition of STAT3 in cancer stem-like cells may offer a potential treatment for colorectal cancer.« less

Signal transducer and activator of transcription 5B (STAT5B) modulates adipocyte differentiation via MOF.

PubMed

Gao, Peng; Zhang, Yuchao; Liu, Yuantao; Chen, Jicui; Zong, Chen; Yu, Cong; Cui, Shang; Gao, Weina; Qin, Dandan; Sun, Wenchuan; Li, Xia; Wang, Xiangdong

2015-12-01

The role and mechanism of signal transducer and activator of transcription 5B (STAT5B) in adipogenesis remain unclear. In this study, our data showed that Males absent on the first (MOF) protein expression was increased during 3 T3-L1 preadipocytes differentiation accompanied with STAT5B expression increasing. Over-expression STAT5B enhanced MOF promoter trans-activation in HeLa cells. Mutagenesis assay and ChIP analysis exhibited that STAT5B was able to bind MOF promoter. Knocking-down STAT5B in 3 T3-L1 preadipocytes led to decreased expression of MOF, but resulted in increased expression of peroxisome proliferator-activated receptor γ (PPARγ), CCAAT/enhancer-binding protein α (C/EBPα) and fatty acid-binding protein 4 (Fabp4), which were important factors or enzymes for adipogenesis. We also found that knocking-down MOF in 3 T3-L1 preadipocytes resulted in increased expression of PPARγ, C/EBPα and Fabp4, which was in the same trend as STAT5B knocking-down. Over-expression MOF resulted in reduced promoter trans-activation activity of C/EBPα. These results suggest that STAT5B and MOF work as negative regulators in adipogenesis, and STAT5B modulates preadipocytes differentiation partially by regulating MOF expression. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

Native and reconstituted HDL activate Stat3 in ventricular cardiomyocytes via ERK1/2: role of sphingosine-1-phosphate.

PubMed

Frias, Miguel A; James, Richard W; Gerber-Wicht, Christine; Lang, Ursula

2009-05-01

High-density lipoprotein (HDL) has been reported to have cardioprotective properties independent from its cholesterol transport activity. The influence of native HDL and reconstituted HDL (rHDL) on Stat3, the transcription factor playing an important role in myocardium adaptation to stress, was analysed in neonatal rat ventricular cardiomyocytes. We have investigated modulating the composition of rHDL as a means of expanding its function and potential cardioprotective effects. Stat3 phosphorylation and activation were determined by western blotting and electrophoretic mobility shift assay (EMSA). In ventricular cardiomyocytes, HDL and the HDL constituent sphingosine-1-phosphate (S1P) induce a concentration- and time-dependent increase in Stat3 activation. They also enhance extracellular signal-regulated kinases (ERK1/2) and p38 mitogen-activated protein kinase (MAPK) phosphorylation. U0126, a specific inhibitor of MEK1/2, the upstream activator of ERK1/2, abolishes HDL- and S1P-induced Stat3 activation, whereas the p38 MAPK blocker SB203580 has no significant effect. Inhibition of the tyrosine kinase family Src (Src) caused a significant reduction of Stat3 activation, whereas inhibition of phosphatidylinositol 3-kinase (PI3K) had no effect. S1P and rHDL containing S1P have a similar strong stimulatory action on Stat3, ERK1/2, and p38 MAPK comparable to native HDL. S1P-free rHDL has a much weaker effect. Experiments with agonists and antagonists of the S1P receptor subtypes indicate that HDL and S1P activate Stat3 mainly through the S1P2 receptor. In ventricular cardiomyocytes, addition of S1P to rHDL enhances its therapeutic potential by improving its capacity to activate Stat3. Activation of Stat3 occurs mainly via the S1P constituent and the lipid receptor S1P2 requiring stimulation of ERK1/2 and Src but not p38 MAPK or PI3K. The study underlines the therapeutic potential of tailoring rHDL to confront particular clinical situations.

Multidimensional Single Cell Based STAT Phosphorylation Profiling Identifies a Novel Biosignature for Evaluation of Systemic Lupus Erythematosus Activity

PubMed Central

Huang, Xinfang; Guo, Yanzhi; Bao, Chunde; Shen, Nan

2011-01-01

Introduction Dysregulated cytokine action on immune cells plays an important role in the initiation and progress of systemic lupus erythematosus (SLE), a complex autoimmune disease. Comprehensively quantifying basal STATs phosphorylation and their signaling response to cytokines should help us to better understand the etiology of SLE. Methods Phospho-specific flow cytometry was used to measure the basal STAT signaling activation in three immune cell types of peripheral-blood mononuclear cells from 20 lupus patients, 9 rheumatoid arthritis (RA) patients and 13 healthy donors (HDs). A panel of 27 cytokines, including inflammatory cytokines, was measured with Bio-Plex™ Human Cytokine Assays. Serum Prolactin levels were measured with an immunoradiometric assay. STAT signaling responses to inflammatory cytokines (interferon α [IFNα], IFNγ, interleukin 2 [IL2], IL6, and IL10) were also monitored. Results We observed the basal activation of STAT3 in SLE T cells and monocytes, and the basal activation of STAT5 in SLE T cells and B cells. The SLE samples clustered into two main groups, which were associated with the SLE Disease Activity Index 2000, their erythrocyte sedimentation rate, and their hydroxychloroquine use. The phosphorylation of STAT5 in B cells was associated with cytokines IL2, granulocyte colony-stimulating factor (G-CSF), and IFNγ, whereas serum prolactin affected STAT5 activation in T cells. The responses of STAT1, STAT3, and STAT5 to IFNα were greatly reduced in SLE T cells, B cells, and monocytes, except for the STAT1 response to IFNα in monocytes. The response of STAT3 to IL6 was reduced in SLE T cells. Conclusions The basal activation of STATs signaling and reduced response to cytokines may be helpful us to identify the activity and severity of SLE. PMID:21799742

STATs: An Old Story, Yet Mesmerizing

PubMed Central

Abroun, Saeid; Saki, Najmaldin; Ahmadvand, Mohammad; Asghari, Farahnaz; Salari, Fatemeh; Rahim, Fakher

2015-01-01

Signal transducers and activators of transcription (STATs) are cytoplasmic transcription factors that have a key role in cell fate. STATs, a protein family comprised of seven members, are proteins which are latent cytoplasmic transcription factors that convey signals from the cell surface to the nucleus through activation by cytokines and growth factors. The signaling pathways have diverse biological functions that include roles in cell differentiation, proliferation, development, apoptosis, and inflammation which place them at the center of a very active area of research. In this review we explain Janus kinase (JAK)/STAT signaling and focus on STAT3, which is transient from cytoplasm to nucleus after phosphorylation. This procedure controls fundamental biological processes by regulating nuclear genes controlling cell proliferation, survival, and development. In some hematopoietic disorders and cancers, overexpression and activation of STAT3 result in high proliferation, suppression of cell differentiation and inhibition of cell maturation. This article focuses on STAT3 and its role in malignancy, in addition to the role of microRNAs (miRNAs) on STAT3 activation in certain cancers. PMID:26464811

Leptin Suppresses the Rewarding Effects of Running via STAT3 Signaling in Dopamine Neurons.

PubMed

Fernandes, Maria Fernanda A; Matthys, Dominique; Hryhorczuk, Cécile; Sharma, Sandeep; Mogra, Shabana; Alquier, Thierry; Fulton, Stephanie

2015-10-06

The adipose hormone leptin potently influences physical activity. Leptin can decrease locomotion and running, yet the mechanisms involved and the influence of leptin on the rewarding effects of running ("runner's high") are unknown. Leptin receptor (LepR) signaling involves activation of signal transducer and activator of transcription-3 (STAT3), including in dopamine neurons of the ventral tegmental area (VTA) that are essential for reward-relevant behavior. We found that mice lacking STAT3 in dopamine neurons exhibit greater voluntary running, an effect reversed by viral-mediated STAT3 restoration. STAT3 deletion increased the rewarding effects of running whereas intra-VTA leptin blocked it in a STAT3-dependent manner. Finally, STAT3 loss-of-function reduced mesolimbic dopamine overflow and function. Findings suggest that leptin influences the motivational effects of running via LepR-STAT3 modulation of dopamine tone. Falling leptin is hypothesized to increase stamina and the rewarding effects of running as an adaptive means to enhance the pursuit and procurement of food. Copyright © 2015 Elsevier Inc. All rights reserved.

Minocycline Suppresses Interleukine-6, Its Receptor System and Signaling Pathways and Impairs Migration, Invasion and Adhesion Capacity of Ovarian Cancer Cells: In Vitro and In Vivo Studies

PubMed Central

Ataie-Kachoie, Parvin; Morris, David L.; Pourgholami, Mohammad H.

2013-01-01

Interleukin (IL)-6 has been shown to be a major contributing factor in growth and progression of ovarian cancer. The cytokine exerts pro-tumorigenic activity through activation of several signaling pathways in particular signal transducer and activator of transcription (STAT3) and extracellular signal-regulated kinase (ERK)1/2. Hence, targeting IL-6 is becoming increasingly attractive as a treatment option in ovarian cancer. Here, we investigated the effects of minocycline on IL-6 and its signaling pathways in ovarian cancer. In vitro, minocycline was found to significantly suppress both constitutive and IL-1β or 4-hydroxyestradiol (4-OH-E2)-stimulated IL-6 expression in human ovarian cancer cells; OVCAR-3, SKOV-3 and CAOV-3. Moreover, minocycline down-regulated two major components of IL-6 receptor system (IL-6Rα and gp130) and blocked the activation of STAT3 and ERK1/2 pathways leading to suppression of the downstream product MCL-1. In female nude mice bearing intraperitoneal OVCAR-3 tumors, acute administration (4 and 24 h) of minocycline (30 mg/kg) led to suppression of IL-6. Even single dose of minocycline was effective at significantly lowering plasma and tumor IL-6 levels. In line with this, tumoral expression of p-STAT3, p-ERK1/2 and MCL-1 were decreased in minocycline-treated mice. Evaluation of the functional implication of minocycline on metastatic activity revealed the capacity of minocycline to inhibit cellular migration, invasion and adhesion associated with down-regulation of matrix metalloproteinases (MMP)-2 and 9. Thus, the data suggest a potential role for minocycline in suppressing IL-6 expression and activity. These effects may prove to be an important attribute to the upcoming clinical trials of minocycline in ovarian cancer. PMID:23593315

STAT3 Oligonucleotide Inhibits Tumor Angiogenesis in Preclinical Models of Squamous Cell Carcinoma

PubMed Central

Klein, Jonah D.; Sano, Daisuke; Sen, Malabika; Myers, Jeffrey N.; Grandis, Jennifer R.; Kim, Seungwon

2014-01-01

Purpose Signal transducer and activator of transcription 3 (STAT3) has shown to play a critical role in head and neck squamous cell carcinoma (HNSCC) and we have recently completed clinical trials of STAT3 decoy oligonucleotide in patients with recurrent or metastatic HNSCC. However, there is limited understanding of the role of STAT3 in modulating other aspects of tumorigenesis such as angiogenesis. In this study, we aimed to examine the effects of STAT3 decoy oligonucleotide on tumor angiogenesis. Experimental Design A STAT3 decoy oligonucleotide and small interfering RNA (siRNA) were used to inhibit STAT3 in endothelial cells in vitro and in vivo. The biochemical effects of STAT3 inhibition were examined in conjunction with the consequences on proliferation, migration, apoptotic staining, and tubule formation. Additionally, we assessed the effects of STAT3 inhibition on tumor angiogenesis using murine xenograft models. Results STAT3 decoy oligonucleotide decreased proliferation, induces apoptosis, decreased migration, and decreased tubule formation of endothelial cells in vitro. The STAT3 decoy oligonucleotide also inhibited tumor angiogenesis in murine tumor xenografts. Lastly, our data suggest that the antiangiogenic effects of STAT3 decoy oligonucleotide were mediatedthrough the inhibition of both STAT3 and STAT1. Conclusions The STAT3 decoy oligonucleotidewas found to be an effective antiangiogenic agent, which is likely to contribute to the overall antitumor effects of this agent in solid tumors.Taken together with the previously demonstrated antitumor activity of this agent, STAT3 decoy oligonucleotide represents a promising single agent approach to targeting both the tumor and vascular compartments in various malignancies. PMID:24404126

A homolog of teleostean signal transducer and activator of transcription 3 (STAT3) from rock bream, Oplegnathus fasciatus: Structural insights, transcriptional modulation, and subcellular localization.

PubMed

Bathige, S D N K; Thulasitha, William Shanthakumar; Umasuthan, Navaneethaiyer; Jayasinghe, J D H E; Wan, Qiang; Nam, Bo-Hye; Lee, Jehee

2017-04-01

Signal transducer and activator of transcription 3 (STAT3) is one of the crucial transcription factors in the Janus kinase (JAK)/STAT signaling pathway, and it was previously considered as acute phase response factor. A number of interleukins (ILs) such as IL-5, IL-6, IL-9, IL-10, IL-12, and IL-22 are known to be involved in activation of STAT3. In addition, various growth factors and pathogenic or oxidative stresses mediate the activation of a wide range of functions via STAT3. In this study, a STAT3 homolog was identified and functionally characterized from rock bream (RbSTAT3), Oplegnathus fasciatus. In silico characterization revealed that the RbSTAT3 amino acid sequence shares highly conserved common domain architectural features including N-terminal domain, coiled coil domain, DNA binding domain, linker domain, and Src homology 2 (SH2) domains. In addition, a fairly conserved transcriptional activation domain (TAD) was located at the C-terminus. Comparison of RbSTAT3 with other counterparts revealed higher identities (>90%) with fish orthologs. The genomic sequence of RbSTAT3 was obtained from a bacterial artificial chromosome (BAC) library, and was identified as a multi-exonic gene (24 exons), as found in other vertebrates. Genomic structural comparison and phylogenetic studies have showed that the evolutionary routes of teleostean and non-teleostean vertebrates were distinct. Quantitative real time PCR (qPCR) analysis revealed that the spatial distribution of RbSTAT3 mRNA expression was ubiquitous and highly detectable in blood, heart, and liver tissues. Transcriptional modulation of RbSTAT3 was examined in blood and liver tissues after challenges with bacteria (Edwardsiella tarda and Streptococcus iniae), rock bream irido virus (RBIV), and immune stimulants (LPS and poly (I:C)). Significant changes in RbSTAT3 transcription were also observed in response to tissue injury. In addition, the transcriptional up-regulation of RbSTAT3 was detected in rock bream heart cells upon recombinant rock bream IL-10 (rRbIL-10) treatment. Subcellular localization and nuclear translocation of rock bream STAT3 following poly (I:C) treatment were also demonstrated. Taken together, the results of the current study provide important evidence for potential roles of rock bream STAT3 in the immune system and wound healing processes. Copyright © 2017 Elsevier B.V. All rights reserved.

Role of Signal Transducer and Activator of Transcription 3 in Neuronal Survival and Regeneration

PubMed Central

Dziennis, Suzan; Alkayed, Nabil J.

2009-01-01

Synopsis Signal Transducers and Activators of Transcription (STATs) comprise a family of transcription factors that mediate a wide variety of biological functions in the central and peripheral nervous systems. Injury to neural tissue induces STAT activation, and STATs are increasingly recognized for their role in neuronal survival. In this review, we discuss the role of STAT3 during neural development and following ischemic and traumatic injury in brain, spinal cord and peripheral nerves. We focus on STAT3 because of the expanding body of literature that investigates protective and regenerative effects of growth factors, hormones and cytokines that use STAT3 to mediate their effect, in part through transcriptional upregulation of neuroprotective and neurotrophic genes. Defining the endogenous molecular mechanisms that lead to neuroprotection by STAT3 after injury might identify novel therapeutic targets against acute neural tissue damage as well as chronic neurodegenerative disorders. PMID:19145989

Modulation of the caveolin-3 localization to caveolae and STAT3 to mitochondria by catecholamine-induced cardiac hypertrophy in H9c2 cardiomyoblasts

PubMed Central

Jeong, Kyuho; Kwon, Hayeong; Min, Chanhee

2009-01-01

We investigated the effect of phenylephrine (PE)- and isoproterenol (ISO)-induced cardiac hypertrophy on subcellular localization and expression of caveolin-3 and STAT3 in H9c2 cardiomyoblast cells. Caveolin-3 localization to plasma membrane was attenuated and localization of caveolin-3 to caveolae in the plasma membrane was 24.3% reduced by the catecholamine-induced hypertrophy. STAT3 and phospho-STAT3 were up-regulated but verapamil and cyclosporin A synergistically decreased the STAT3 and phospho-STAT3 levels in PE- and ISO-induced hypertrophic cells. Both expression and activation of STAT3 were increased in the nucleus by the hypertrophy. Immunofluorescence analysis revealed that the catecholamine-induced hypertrophy promoted nuclear localization of pY705-STAT3. Of interest, phosphorylation of pS727-STAT3 in mitochondria was significantly reduced by catecholamine-induced hypertrophy. In addition, mitochondrial complexes II and III were greatly down-regulated in the hypertrophic cells. Our data suggest that the alterations in nuclear and mitochondrial activation of STAT3 and caveolae localization of caveolin-3 are related to the development of the catecholamine-induced cardiac hypertrophy. PMID:19299911

STAT inhibitors for cancer therapy

PubMed Central

2013-01-01

Signal Transducer and Activator of Transcription (STAT) proteins are a family of cytoplasmic transcription factors consisting of 7 members, STAT1 to STAT6, including STAT5a and STAT5b. STAT proteins are thought to be ideal targets for anti-cancer therapy since cancer cells are more dependent on the STAT activity than their normal counterparts. Inhibitors targeting STAT3 and STAT5 have been developed. These included peptidomimetics, small molecule inhibitors and oligonucleotides. This review summarized advances in preclinical and clinical development of these compounds. PMID:24308725

Arsenic trioxide mediates HAPI microglia inflammatory response and subsequent neuron apoptosis through p38/JNK MAPK/STAT3 pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mao, Jiamin

Arsenic is a widely distributed toxic metalloid all over the world. Inorganic arsenic species are supposed to affect astrocytic functions and to cause neuron apoptosis in CNS. Microglias are the key cell type involved in innate immune responses in CNS, and microglia activation has been linked to inflammation and neurotoxicity. In this study, using ELISA, we showed that Arsenic trioxide up-regulated the expression and secretion of IL-1β in a dose-dependent manner and a time-dependent manner in cultured HAPI microglia cells. The secretion of IL-1β caused the apoptosis of SH-SY5Y. These pro-inflammatory responses were inhibited by the STAT3 blocker, AG490 andmore » P38/JNK MAPK blockers SB202190, SP600125. Further, Arsenic trioxide exposure could induce phosphorylation and activation of STAT3, and the translocation of STAT3 from the cytosol to the nucleus in this HAPI microglia cell line. Thus, the STAT3 signaling pathway can be activated after Arsenic trioxide treatment. However, P38/JNK MAPK blockers SB202190, SP600125 also obviously attenuated STAT3 activation and transnuclear transport induced by Arsenic trioxide. In concert with these results, we highlighted that the secretion of IL-1β and STAT3 activation induced by Arsenic trioxide can be mediated by elevation of P38/JNK MAPK in HAPI microglia cells and then induced the toxicity of neurons. - Highlights: • Arsenic trioxide exposure induced expression of IL-β in HAPI microglia. • Arsenic trioxide exposure induced activation of MAPK pathways in HAPI microglia. • Arsenic trioxide exposure induced activation of STAT3 pathways in HAPI microglia. • The expression of IL-β though P38/JNK MAPK/STAT3 pathways in HAPI microglia.« less

Mitochondrial GRIM-19 as a potential therapeutic target for STAT3-dependent carcinogenesis of gastric cancer

PubMed Central

Zhao, Xiaodong; Bao, Liming; Huang, Daochao; Song, Lihua; Li, Yang

2016-01-01

Aberrant STAT3 activation occurs in most human gastric cancers (GCs) and contributes to the malignant progression of GC, but mechanism(s) underlying aberrant STAT3 remain largely unknown. Here we demonstrated that the gene associated with retinoid interferon-induced mortality 19 (GRIM-19) was severely depressed or lost in GC and chronic atrophic gastritis (CAG) tissues and its loss contributed to GC tumorigenesis partly by activating STAT3 signaling. In primary human GC tissues, GRIM-19 was frequently depressed or lost and this loss correlated with advanced clinical stage, lymph node metastasis, H. pylori infection and poor overall survival of GC patients. In CAG tissues, GRIM-19 was progressively decreased along with its malignant transformation. Functionally, we indentified an oncogenic role of GRIM-19 loss in promoting GC tumorigenesis. Ectopic GRIM-19 expression suppressed GC tumor formation in vitro and in vivo by inducing cell cycle arrest and apoptosis. Moreover, we revealed that GRIM-19 inhibited STAT3 transcriptional activation and its downstream targets by reducing STAT3 nuclear distribution. Conversely, knockdown of GRIM-19 induced aberrant STAT3 activation and accelerated GC cell growth in vitro and in vivo, and this could be partly attenuated by the blockage of STAT3 activation. In addition, we observed subcellular redistributions of GRIM-19 characterized by peri-nuclear aggregates, non-mitochondria cytoplasmic distribution and nuclear invasion, which should be responsible for reduced STAT3 nuclear distribution. Our studies suggest that mitochondrial GRIM-19 could not only serve as an valuable prognostic biomarker for GC development, but also as a potential therapeutic target for STAT3-dependent carcinogenesis of GC. PMID:27167343

[Expressions of VEGF/VEGFRs and activation of STATs in ovarian carcinoma].

PubMed

Chen, Bing-Ya; Ye, Da-Feng; Xie, Xing; Chen, Huai-Zeng; Lü, Wei-Guo

2005-01-01

To study the expressions of VEGF/VEGFRs and activation of STATs in ovarian epithelial carcinoma, and to elucidate direct effect of VEGF on ovarian carcinoma cells. Tissue samples from 42 women with primary ovarian epithelial carcinoma (OVCA), 29 with begnin ovarian tumor (OVBT) and 11 with normal ovarian tissue (NOV) were collected. LSAB immunohistochemical staining was used to determine the expression of VEGF, VEGFR1, VEGFR2 and activated STATS (P-STAT1, P-STAT3, P-STAT5, P-STAT6) proteins. (1) Semi-quantitative scoring showed that VEGF expression in OVCA was significantly higher than that in OVBT and NOV (P < 0.01). Expressions of VEGFR1 and VEGFR2 were significantly elevated in OVCA, including tumor cells and stromal vascular endothelial cells (P < 0.01, compared with OVBT and NOV). There was no difference in VEGFRs expressions between OVBT and NOV. (2) In OVCA, tumor cells and endothelial cells expressed P-STAT3 and P-STAT5 at significantly higher levels than those in OVBT and NOV (P = 0.000). The staining of P-STAT1 and P-STAT6 was weak with no significant differences among OVCA, OVBT and NOV. (3) Expressions of VEGFR1 and VEGFR2 in endothelial cells were significantly correlated with P-STAT5 and P-STAT3, respectively (P = 0.006 and 0.001). In cancer cells, VEGF, VEGFR1 and VEGFR2 were all significantly correlated with P-STAT3 and P-STAT5 (P = 0.000), but not with P-STAT1 or P-STAT6. VEGF affects ovarian carcinoma cells via VEGFRs, and STATs probably participate in intracellular signaling of VEGF.

IL-27 inhibits epithelial-mesenchymal transition and angiogenic factor production in a STAT1-dominant pathway in human non-small cell lung cancer

PubMed Central

2013-01-01

Background Interleukin-27 signaling is mediated by the JAK-STAT pathway via activation of STAT1 and STAT3, which have tumor suppressive and oncogenic activities, respectively. Epithelial–mesenchymal transition (EMT) and angiogenesis are key processes in carcinogenesis. Although IL-27 has been shown to have potent anti-tumor activity in various cancer models, the role of IL-27 in EMT and angiogenesis is poorly understood. In this study, we investigated the role of IL-27 in regulating EMT and angiogenesis through modulation of the STAT pathways in human non-small cell lung carcinoma (NSCLC) cells. Methods STAT activation following IL-27 exposure was measured in human NSCLC cell lines. Expression of epithelial (E-cadherin, γ-catenin) and mesenchymal (N-cadherin, vimentin) markers were assessed by Western blot analysis. Production of pro-angiogenic factors (VEGF, IL-8/CXCL8, CXCL5) were examined by ELISA. Cell motility was examined by an in vitro scratch and transwell migration assays. Selective inhibitors of STAT1 (STAT1 siRNAs) and STAT3 (Stattic) were used to determine whether both STAT1 and STAT3 are required for IL-27 mediated inhibition of EMT and secretion of angiogenic factors. Results Our results demonstrate that IL-27 stimulation in NSCLC resulted in 1) STAT1 and STAT3 activation in a JAK-dependent manner, 2) development of epithelial phenotypes, including a decrease in the expression of a transcriptional repressor for E-cadherin (SNAIL), and mesenchymal marker (vimentin) with a reciprocal increase in the expression of epithelial markers, 3) inhibition of cell migration, and 4) reduced production of pro-angiogenic factors. STAT1 inhibition in IL-27–treated cells reversed the IL-27 effect with resultant increased expression of Snail, vimentin and the pro-angiogenic factors. The inhibition of STAT3 activation had no effect on the development of the epithelial phenotype. Conclusion IL-27 induces mesenchymal to epithelial transition and inhibits the production of pro-angiogenic factors in a STAT1–dominant pathway. These findings highlight the importance of STAT1 in repressing lung carcinogenesis and describe a new anti-tumor mechanism of IL-27. PMID:24274066

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sakaeda, Yoshiichi; Hiroi, Miki; Shimojima, Takahiro

Sulindac, a non-steroidal anti-inflammatory drug, has been shown to exert an anti-tumor effect on several types of cancer. To determine the effect of sulindac on intracellular signaling pathways in host immune cells such as macrophages, we investigated the effect of the drug on interferon gamma (IFN{gamma})-induced expression of signal transducer and activator of transcription 1 (STAT1) and other genes in mouse macrophage-like cell line RAW264.7 cells. Sulindac, but not aspirin or sodium salicylate, inhibited IFN{gamma}-induced expression of the CXC ligand 9 (CXCL9) mRNA, a chemokine for activated T cells, whereas the interferon-induced expression of CXCL10 or IFN regulatory factor-1 wasmore » not affected by sulindac. Luciferase reporter assay demonstrated that sulindac inhibited IFN{gamma}-induced promoter activity of the CXCL9 gene. Surprisingly, sulindac had no inhibitory effect on IFN{gamma}-induced STAT1 activation; however, constitutive nuclear factor {kappa}B activity was suppressed by the drug. These results indicate that sulindac selectively inhibited IFN{gamma}-inducible gene expression without inhibiting STAT1 activation.« less

Defects along the T(H)17 differentiation pathway underlie genetically distinct forms of the hyper IgE syndrome.

PubMed

Al Khatib, Shadi; Keles, Sevgi; Garcia-Lloret, Maria; Karakoc-Aydiner, Elif; Reisli, Ismail; Artac, Hasibe; Camcioglu, Yildiz; Cokugras, Haluk; Somer, Ayper; Kutukculer, Necil; Yilmaz, Mustafa; Ikinciogullari, Aydan; Yegin, Olcay; Yüksek, Mutlu; Genel, Ferah; Kucukosmanoglu, Ercan; Baki, Ali; Bahceciler, Nerin N; Rambhatla, Anupama; Nickerson, Derek W; McGhee, Sean; Barlan, Isil B; Chatila, Talal

2009-08-01

The hyper IgE syndrome (HIES) is characterized by abscesses, eczema, recurrent infections, skeletal and connective tissue abnormalities, elevated serum IgE, and diminished inflammatory responses. It exists as autosomal-dominant and autosomal-recessive forms that manifest common and distinguishing clinical features. A majority of those with autosomal-dominant HIES have heterozygous mutations in signal transducer and activator of transcription (STAT)-3 and impaired T(H)17 differentiation. To elucidate mechanisms underlying different forms of HIES. A cohort of 25 Turkish children diagnosed with HIES were examined for STAT3 mutations by DNA sequencing. Activation of STAT3 by IL-6 and IL-21 and STAT1 by IFN-alpha was assessed by intracellular staining with anti-phospho (p)STAT3 and -pSTAT1 antibodies. T(H)17 and T(H)1 cell differentiation was assessed by measuring the production of IL-17 and IFN-gamma, respectively. Six subjects had STAT3 mutations affecting the DNA binding, Src homology 2, and transactivation domains, including 3 novel ones. Mutation-positive but not mutation-negative subjects with HIES exhibited reduced phosphorylation of STAT3 in response to cytokine stimulation, whereas pSTAT1 activation was unaffected. Both patient groups exhibited impaired T(H)17 responses, but whereas STAT3 mutations abrogated early steps in T(H)17 differentiation, the defects in patients with HIES with normal STAT3 affected more distal steps. In this cohort of Turkish children with HIES, a majority had normal STAT3, implicating other targets in disease pathogenesis. Impaired T(H)17 responses were evident irrespective of the STAT3 mutation status, indicating that different genetic forms of HIES share a common functional outcome.

JAK-STAT signalling and the atrial fibrillation promoting fibrotic substrate

PubMed Central

Chen, Yu; Surinkaew, Sirirat; Naud, Patrice; Qi, Xiao-Yan; Gillis, Marc-Antoine; Shi, Yan-Fen; Tardif, Jean-Claude; Dobrev, Dobromir; Nattel, Stanley

2017-01-01

Aims Left-atrial (LA) fibrosis is an important feature of many atrial fibrillation (AF) substrates. The JAK-STAT system contributes to cardiac remodelling, but its role in AF is unknown. Here we investigated JAK-STAT changes in an AF-model and their potential contributions to LA-fibrosis. Methods and results LA-remodelling was studied in dogs with heart failure (HF) induced by ventricular tachypacing (VTP, 240 bpm), and in mice with left-ventricular (LV) dysfunction due to myocardial infarction (MI). The selective STAT-3 inhibitor S3I-201 was administered to fibroblasts in vitro or mice in vivo (10 mg/kg/d, osmotic mini-pump). HF-dogs developed LA-selective fibrosis and AF-susceptibility at 1-week VTP. The mRNA-expression of platelet-derived growth factor (PDGF, a JAK-STAT activator) isoforms A, C and D, as well as JAK2, increased in LA fibroblasts from 1-week VTP. HF upregulated protein-expression of PDGF-receptor-β and phosphorylated (activated) signal transducer and activator of transcription 3 (STAT3) in LA. PDGF-AB stimulation of LA fibroblasts increased PDGFR-α, STAT3 and phosphorylated-STAT3 expression, as well as collagen-1 and fibronectin-1 protein secretion (by 1.6- to 20-fold), with smaller changes in LV fibroblasts. Phosphorylated-STAT3 and collagen upregulation were suppressed by the JAK2 inhibitor AG-490, PDGF receptor inhibitor AG1296 and STAT3-inhibitor SI3-201. In vivo S3I-201 treatment of MI-mice attenuated LA-fibrosis, LA-dilation and P-wave duration changes versus vehicle-control. Conclusions HF activates the LA JAK-STAT system and enhances PDGF-signalling. JAK-STAT inhibition reduces the profibrotic effects of PDGF stimulation on canine fibroblasts in vitro while attenuating in vivo LA-fibrosis and remodelling in post-MI mice, suggesting that the JAK/STAT pathway contributes to LA-fibrogenesis and might be a potential target for LA-fibrosis prevention. PMID:28158495

Inhibition of interleukin-3- and interferon- α-induced JAK/STAT signaling by the synthetic α-X-2',3,4,4'-tetramethoxychalcones α-Br-TMC and α-CF3-TMC.

PubMed

Jobst, Belinda; Weigl, Julia; Michl, Carina; Vivarelli, Fabio; Pinz, Sophia; Amslinger, Sabine; Rascle, Anne

2016-11-01

The JAK/STAT pathway is an essential mediator of cytokine signaling, often upregulated in human diseases and therefore recognized as a relevant therapeutic target. We previously identified the synthetic chalcone α-bromo-2',3,4,4'-tetramethoxychalcone (α-Br-TMC) as a novel JAK2/STAT5 inhibitor. We also found that treatment with α-Br-TMC resulted in a downward shift of STAT5 proteins in SDS-PAGE, suggesting a post-translational modification that might affect STAT5 function. In the present study, we show that a single cysteine within STAT5 is responsible for the α-Br-TMC-induced protein shift, and that this modification does not alter STAT5 transcriptional activity. We also compared the inhibitory activity of α-Br-TMC to that of another synthetic chalcone, α-trifluoromethyl-2',3,4,4'-tetramethoxychalcone (α-CF3-TMC). We found that, like α-Br-TMC, α-CF3-TMC inhibits JAK2 and STAT5 phosphorylation in response to interleukin-3, however without altering STAT5 mobility in SDS-PAGE. Moreover, we demonstrate that both α-Br-TMC and α-CF3-TMC inhibit interferon-α-induced activation of STAT1 and STAT2, by inhibiting their phosphorylation and the expression of downstream interferon-stimulated genes. Together with the previous finding that α-Br-TMC and α-CF3-TMC inhibit the response to inflammation by inducing Nrf2 and blocking NF-κB activities, our data suggest that synthetic chalcones might be useful as anti-inflammatory, anti-cancer and immunomodulatory agents in the treatment of human diseases.

S100B Attenuates Microglia Activation in Gliomas: Possible Role of STAT3 Pathway

PubMed Central

Zhang, Leying; Liu, Wei; Alizadeh, Darya; Zhao, Dongchang; Farrukh, Omar; Lin, Jeffrey; Badie, Sam A.; Badie, Behnam

2010-01-01

Despite significant infiltration into tumors, the effector function of macrophages (MPs) and microglia (MG) appears to be suppressed in gliomas. Although STAT3 pathway is thought to play a role in this process, the exact mechanism by which gliomas induce STAT3 activation in MPs and MG is not known. Because activation of receptor for advanced glycation end products (RAGE) can induce STAT3, and because gliomas express high levels of S100B, a RAGE ligand, we hypothesized that MP/MG STAT3 activity may be modulated through S100B-RAGE interaction. Exposure of N9 MG and bone marrow-derived monocytes (BMM) to GL261 glioma condition medium (GCM) and low (nM) levels of S100B increased RAGE expression, induced STAT3 and suppressed MG function in vitro. Furthermore, neutralization of S100B in GCM, partially reversed IL-1β suppression in BMM, suggesting that the inhibitory effect of GCM to be in part due to S100B. Finally, blockage of S100B-RAGE interaction inhibited STAT3 activation in N9 MG and in glioma MG/MP in vivo. These findings suggest that the RAGE pathway may play an important role in STAT3 induction in glioma-associated MG/MPs, and that this process may be mediated through S100B. PMID:21264954

S100B attenuates microglia activation in gliomas: possible role of STAT3 pathway.

PubMed

Zhang, Leying; Liu, Wei; Alizadeh, Darya; Zhao, Dongchang; Farrukh, Omar; Lin, Jeffrey; Badie, Sam A; Badie, Behnam

2011-03-01

Despite significant infiltration into tumors, the effector function of macrophages (MPs) and microglia (MG) appears to be suppressed in gliomas. Although STAT3 pathway is thought to play a role in this process, the exact mechanism by which gliomas induce STAT3 activation in MPs and MG is not known. Because activation of receptor for advanced glycation end products (RAGE) can induce STAT3, and because gliomas express high levels of S100B, a RAGE ligand, we hypothesized that MP/MG STAT3 activity may be modulated through S100B-RAGE interaction. Exposure of N9 MG and bone marrow-derived monocytes (BMM) to GL261 glioma condition medium (GCM) and low (nM) levels of S100B increased RAGE expression, induced STAT3 and suppressed MG function in vitro. Furthermore, neutralization of S100B in GCM, partially reversed IL-1β suppression in BMM, suggesting that the inhibitory effect of GCM to be in part due to S100B. Finally, blockage of S100B-RAGE interaction inhibited STAT3 activation in N9 MG and in glioma MG/MP in vivo. These findings suggest that the RAGE pathway may play an important role in STAT3 induction in glioma-associated MG/MPs, and that this process may be mediated through S100B. Copyright © 2010 Wiley-Liss, Inc.

Secoisolariciresinol diglucoside prevents the oxidative stress-induced apoptosis of myocardial cells through activation of the JAK2/STAT3 signaling pathway.

PubMed

Huang, Guiqiong; Huang, Xiaofang; Liu, Min; Hua, Yue; Deng, Bo; Jin, Wen; Yan, Wen; Tan, Zhangbin; Wu, Yifen; Liu, Bin; Zhou, Yingchun

2018-06-01

Myocardial cell apoptosis mediated by oxidative stress has previously been identified as a key process in ischemic heart disease. Secoisolariciresinol diglucoside (SDG), a polyphenolic plant lignan primarily found in flaxseed, has been demonstrated to effectively protect myocardial cells from apoptosis. In the present study, the role of the Janus kinase 2 (JAK2)/signal transducer and activator of transcription 3 (STAT3) was investigated in mediating the protective effect of SDG. Findings of the present study revealed that treatment with H2O2 reduced cell viability and induced apoptosis in H9C2 rat cardiomyocytes. However, SDG was able to reduce the effect of H2O2 in a dose‑dependent manner. H2O2 reduced the expression level of phosphorylated STAT3 and inhibited the levels of B‑cell lymphoma‑extra‑large and induced myeloid leukemia cell differentiation protein, which are the STAT3 target genes. Conversely, SDG rescued phosphorylation of STAT3 and increased the levels of STAT3 target genes. Treatment with SDG alone led to a dose‑dependent increased phosphorylation of JAK2 and STAT3, without activating Src. Furthermore, the anti‑apoptotic effects of SDG were partially abolished by a JAK2/STAT3 inhibitor. In addition, molecular docking revealed that SDG may bind to the protein kinase domain of JAK2, at a binding energy of ‑8.258 kcal/mol. Molecular dynamics simulations revealed that JAK2‑SDG binding was stable. In conclusion, activation of the JAK2/STAT3 signaling pathway contributed to the anti‑apoptotic activity of SDG, which may be a potential JAK2 activator.

The role of the JAK2-STAT3 pathway in pro-inflammatory responses of EMF-stimulated N9 microglial cells

PubMed Central

2010-01-01

Background In several neuropathological conditions, microglia can become overactivated and cause neurotoxicity by initiating neuronal damage in response to pro-inflammatory stimuli. Our previous studies have shown that exposure to electromagnetic fields (EMF) activates cultured microglia to produce tumor necrosis factor (TNF)-α and nitric oxide (NO) through signal transduction involving the activator of transcription STAT3. Here, we investigated the role of STAT3 signaling in EMF-induced microglial activation and pro-inflammatory responses in more detail than the previous study. Methods N9 microglial cells were treated with EMF exposure or a sham treatment, with or without pretreatment with an inhibitor (Pyridone 6, P6) of the Janus family of tyrosine kinases (JAK). The activation state of microglia was assessed via immunoreaction using the microglial marker CD11b. Levels of inducible nitric oxide synthase (iNOS), TNF-α and NO were measured using real-time reverse transcription-polymerase chain reaction (RT-PCR), enzyme-linked immunosorbent assay (ELISA) and the nitrate reductase method. Activation of JAKs and STAT3 proteins was evaluated by western blotting for specific tyrosine phosphorylation. The ability of STAT3 to bind to DNA was detected with an electrophoresis mobility shift assay (EMSA). Results EMF was found to significantly induce phosphorylation of JAK2 and STAT3, and DNA-binding ability of STAT3 in N9 microglia. In addition, EMF dramatically increased the expression of CD11b, TNF-α and iNOS, and the production of NO. P6 strongly suppressed the phosphorylation of JAK2 and STAT3 and diminished STAT3 activity in EMF-stimulated microglia. Interestingly, expression of CD11b as well as gene expression and production of TNF-α and iNOS were suppressed by P6 at 12 h, but not at 3 h, after EMF exposure. Conclusions EMF exposure directly triggers initial activation of microglia and produces a significant pro-inflammatory response. Our findings confirm that the JAK2-STAT3 pathway may not mediate this initial microglial activation but does promote pro-inflammatory responses in EMF-stimulated microglial cells. Thus, the JAK2-STAT3 pathway might be a therapeutic target for reducing pro-inflammatory responses in EMF-activated microglia. PMID:20828402

Signal transducer and activator of transcription 3 is the dominant mediator of the anti-inflammatory effects of IL-10 in human macrophages.

PubMed

Williams, Lynn; Bradley, Laura; Smith, Alexandra; Foxwell, Brian

2004-01-01

The signaling mechanism by which the anti-inflammatory cytokine IL-10 mediates suppression of proinflammatory cytokine synthesis remains largely unknown. Macrophage-specific STAT3-null mice have demonstrated that STAT3 plays a critical role in the suppression of LPS-induced TNF-alpha release, although the mechanism by which STAT3 mediates this inhibition is still not clear. Using an adenoviral system, we have expressed a dominant negative (DN) STAT3 in human macrophages to broaden the investigation to determine the role of STAT3 in IL-10-mediated anti-inflammatory signaling and gene expression. Overexpression of STAT3 DN completely inhibited IL-10-induced suppressor of cytokine signaling 3, tissue inhibitor of MMP-1, TNF receptor expression, and the recently identified IL-10-inducible genes, T cell protein tyrosine phosphatase and signaling lymphocyte activation molecule. STAT3 DN also blocked IL-10-mediated inhibition of MHC class II and COX2 expression. In agreement with the studies in STAT3-null mice, overexpression of the STAT3 DN completely reversed the ability of IL-10 to inhibit LPS-mediated TNF-alpha and IL-6 production. However, real-time PCR analysis showed that STAT3 DN expression did not affect immediate suppression of TNF-alpha mRNA, but did reverse the suppression observed at later time points, suggesting a biphasic regulation of TNF-alpha mRNA levels by IL-10. In conclusion, although STAT3 does appear to be the dominant mediator of the majority of IL-10 functions, there are elements of its anti-inflammatory activity that are STAT3 independent.

Reciprocal activation between STAT3 and miR-181b regulates the proliferation of esophageal cancer stem-like cells via the CYLD pathway.

PubMed

Xu, Dan-Dan; Zhou, Peng-Jun; Wang, Ying; Zhang, Li; Fu, Wu-Yu; Ruan, Bi-Bo; Xu, Hai-Peng; Hu, Chao-Zhi; Tian, Lu; Qin, Jin-Hong; Wang, Sheng; Wang, Xiao; Li, Yi-Cheng; Liu, Qiu-Ying; Ren, Zhe; Zhang, Rong; Wang, Yi-Fei

2016-05-17

Recent studies have suggested that cancer cells contain subpopulations that can initiate tumor growth, self-renew, and maintain tumor cell growth. However, for esophageal cancer cells, the relationship between STAT3, microRNAs and cancer stem cells remains unclear. Serum-free culture was used to enrich esophageal cancer stem-like cells (ECSLC). Flow cytometry determined the proportion of ECSLC. qPCR were performed to examine expression level of stemness factors, mesenchymal markers, ATP-binding cassette (ABC) transporters, STAT3, miR-181b, CYLD. Western blot were performed to analyze the expression of STAT3, p-STAT3 and CYLD (cylindromatosis). BALB/c mice xenograft studies were conducted to evaluate the tumorigenicity of enriched ECSLC. Sphere formation assay and colony formation assays were employed to analyze the relationship between STAT3 and miR-181b. Luciferase assays were used to evaluate activity which CYLD is a target of miR-181b. Sphere formation cells (SFCs) with properties of ECSLC were enriched. Enriched SFCs in serum-free suspension culture exhibited cancer stem-like cell properties and increased single-positive CD44 + CD24-, stemness factor, mesenchymal marker expression ABC transporters and tumorigenicity in vivo compared with the parental cells. Additionally, we found that reciprocal activation between STAT3 and miR-181b regulated SFCs proliferation. Moreover, STAT3 directly activated miR-181b transcription in SFCs and miR-181b then potentiated p-STAT3 activity. Luciferase assays indicated that CYLD was a direct and functional target of miR-181b. The mutual regulation between STAT3 and miR-181b in SFCs was required for proliferation and apoptosis resistance. STAT3 and miR-181b control each other's expression in a positive feedback loop that regulates SFCs via CYLD pathway. These findings maybe is helpful for targeting ECSLC and providing approach for esophageal cancer treatments.

TNF-α-inducing protein of Helicobacter pylori induces epithelial-mesenchymal transition (EMT) in gastric cancer cells through activation of IL-6/STAT3 signaling pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chen, Guodong; Tang, Na; Wang, Chao

Tumor necrosis factor (TNF)-α-inducing protein (Tipα) is a newly identified carcinogenic factor secreted by Helicobacter pylori (H. pylori). Although it has been proved that Tipα is a strong inducer of epithelial-mesenchymal transition (EMT), a crucial process of migration, the exact molecular mechanism is unknown. Current evidence indicates that the oncogenic transcription factor signal transducers and activators of transcription 3 (STAT3) is inappropriately activated in multiple malignancies, including gastric cancer. In this study, we showed that Tipα significantly down-regulated the expression of EMT-related markers E-cadherin as well as up-regulated N-cadherin and vimentin in SGC7901 cells, with typical morphological changes of EMT. Tipα alsomore » promoted proliferation and migration of SGC7901 cells. Furthermore, Tipα activated interleukin-6 (IL-6)/STAT3 signaling pathway in SGC7901 cells. The effects of Tipα treatment observed was abolished when we block IL-6/STAT3 signaling pathway. Altogether, our data demonstrated that Tipα may accelerate tumor aggressiveness in gastric cancer by promoting EMT through activation of IL-6/STAT3 pathway. - Highlights: • Tipα induces EMT and activates IL-6/STAT3 pathway in gastric cancer cells. • IL-6/STAT3 pathway inhibition reverses Tipα-induced proliferation and migration in gastric cancer cells. • Tipα induces EMT in gastric cancer cells via IL-6/STAT3 pathway activation.« less

STAT3 Activation in Pressure-Overloaded Feline Myocardium: Role for Integrins and the Tyrosine Kinase BMX

PubMed Central

Willey, Christopher D.; Palanisamy, Arun P.; Johnston, Rebecca K.; Mani, Santhosh K.; Shiraishi, Hirokazu; Tuxworth, William J.; Zile, Michael R.; Balasubramanian, Sundaravadivel; Kuppuswamy, Dhandapani

2008-01-01

Growth, survival and cytoskeletal rearrangement of cardiomyocytes are critical for cardiac hypertrophy. Signal transducer and activator of transcription-3 (STAT3) activation is an important cardioprotective factor associated with cardiac hypertrophy. Although STAT3 activation has been reported via signaling through Janus Kinase 2 (JAK2) in several cardiac models of hypertrophy, the importance of other nonreceptor tyrosine kinases (NTKs) has not been explored. Utilizing an in vivo feline right ventricular pressure-overload (RVPO) model of hypertrophy, we demonstrate that in 48 h pressure-overload (PO) myocardium, STAT3 becomes phosphorylated and redistributed to detergent-insoluble fractions with no accompanying JAK2 activation. PO also caused increased levels of phosphorylated STAT3 in both cytoplasmic and nuclear fractions. To investigate the role of other NTKs, we used our established in vitro cell culture model of hypertrophy where adult feline cardiomyocytes are embedded three-dimensionally (3D) in type-I collagen and stimulated with an integrin binding peptide containing an Arg-Gly-Asp (RGD) motif that we have previously shown to recapitulate the focal adhesion complex (FAC) formation of 48 h RVPO. RGD stimulation of adult cardiomyocytes in vitro caused both STAT3 redistribution and activation that were accompanied by the activation and redistribution of c-Src and the TEC family kinase, BMX, but not JAK2. However, infection with dominant negative c-Src adenovirus was unable to block RGD-stimulated changes on either STAT3 or BMX. Further analysis in vivo in 48 h PO myocardium showed the presence of both STAT3 and BMX in the detergent-insoluble fraction with their complex formation and phosphorylation. Therefore, these studies indicate a novel mechanism of BMX-mediated STAT3 activation within a PO model of cardiac hypertrophy that might contribute to cardiomyocyte growth and survival. PMID:18612371

STAT3-Activated GM-CSFRα Translocates to the Nucleus and Protects CLL Cells from Apoptosis

PubMed Central

Li, Ping; Harris, David; Liu, Zhiming; Rozovski, Uri; Ferrajoli, Alessandra; Wang, Yongtao; Bueso-Ramos, Carlos; Hazan-Halevy, Inbal; Grgurevic, Srdana; Wierda, William; Burger, Jan; O'Brien, Susan; Faderl, Stefan; Keating, Michael; Estrov, Zeev

2014-01-01

Here it was determined that Chronic Lymphocytic Leukemia (CLL) cells express the α-subunit but not the β-subunit of the granulocyte-macrophage colony-stimulating factor receptor (GM-CSFR/CSF3R). GM-CSFRα was detected on the surface, in the cytosol, and the nucleus of CLL cells via confocal microscopy, cell fractionation, and GM-CSFRα antibody epitope mapping. Because STAT3 is frequently activated in CLL and the GM-CSFRα promoter harbors putative STAT3 consensus binding sites, MM1 cells were transfected with truncated forms of the GM-CSFRα promoter, then stimulated with IL-6 to activate STAT3 to identify STAT3 binding sites. Chromatin immunoprecipitation (ChIP) and an electoromobility shift assay (EMSA) confirmed STAT3 occupancy to those promoter regions in both IL-6 stimulated MM1 and CLL cells. Transfection of MM1 cells with STAT3 siRNA or CLL cells with STAT3 shRNA significantly down-regulated GM-CSFRα mRNA and protein levels. RNA transcripts, involved in regulating cell-survival pathways, and the proteins KAP1 (TRIM28) and ISG15 co-immunoprecipitated with GM-CSFRα. GM-CSFRα-bound KAP1 enhanced the transcriptional activity of STAT3, whereas ISG15 inhibited the NF-κB pathway. Nevertheless, overexpression of GM-CSFRα protected MM1 cells from dexamethasone-induced apoptosis, and GM-CSFRα knockdown induced apoptosis in CLL cells, suggesting that GM-CSFRα provides a ligand-independent survival advantage. PMID:24836891

Ultraviolet B exposure activates Stat3 signaling via phosphorylation at tyrosine705 in skin of SKH1 hairless mouse: a target for the management of skin cancer?

PubMed

Ahsan, Haseeb; Aziz, Moammir Hasan; Ahmad, Nihal

2005-07-22

Understanding the molecular determinants of ultraviolet (UV) response may lead to the development of novel targets; and therefore, better approaches for the management of cancers, which mainly arise due to the exposure of skin to UV (particularly its UVB spectrum). Signal transducer and activator of transcription (Stat) proteins have been shown to activate multiple signaling pathways to contribute to oncogenesis. Here, we studied the regulation of Stat3 during UVB exposure-mediated responses in the skin of SKH-1 hairless mouse, a model regarded to possess relevance to human situations. Our data demonstrated that a single UVB (180 mJ/cm(2)) exposure to the skin of SKH-1 hairless mice resulted in significant upregulation in (i) protein levels of Stat3 and (ii) phosphorylation of Stat3 at tyrosine(705). Further, the activation of Stat3 was found to be associated with a decrease in apoptotic response of UVB and a gradual time-dependent increase in leukocyte infiltration and hyperplasia. In conclusion, we have demonstrated, for the first time, that UVB exposure to skin resulted in an activation of pro-survival protein Stat3. Based on our observation, we suggest that Stat3 could serve as a target for the management of UVB exposure-mediated damages including skin cancer.

A biomarker-based screen of a gene expression compendium ...

EPA Pesticide Factsheets

Computational approaches were developed to identify factors that regulate Nrf2 in a large gene expression compendium of microarray profiles including >2000 comparisons which queried the effects of chemicals, genes, diets, and infectious agents on gene expression in the mouse liver. A gene expression biomarker of 48 genes which accurately predicted Nrf2 activation was used to identify factors which resulted in a gene expression profile with significant correlation to the biomarker. A number of novel insights were made. Chemicals that activated the xenosensor constitutive activated receptor (CAR) consistently activated Nrf2 across hundreds of profiles, possibly downstream of Cyp-induced increases in oxidative stress. Nrf2 activation was also found to be negatively regulated by the growth hormone (GH)- and androgen-regulated transcription factor STAT5b, a transcription factor suppressed by CAR. Nrf2 was activated when STAT5b was suppressed in female mice vs. male mice, after exposure to estrogens, or in genetic mutants in which GH signaling was disrupted. A subset of the mutants that show STAT5b suppression and Nrf2 activation result in increased resistance to environmental stressors and increased longevity. This study describes a novel approach for understanding the network of factors that regulate the Nrf2 pathway and highlights novel interactions between Nrf2, CAR and STAT5b transcription factors. (This abstract does not represent EPA policy.) Computational appr

Critical functions of RhoB in support of glioblastoma tumorigenesis

PubMed Central

Ma, Yufang; Gong, Yuanying; Cheng, Zhixiang; Loganathan, Sudan; Kao, Crystal; Sarkaria, Jann N.; Abel, Ty W.; Wang, Jialiang

2015-01-01

Background RhoB is a member of the Rho small GTPase family that regulates cytoskeletal dynamics and vesicle trafficking. The RhoB homologs, RhoA and RhoC, have been shown to promote cancer progression and metastasis. In contrast, the functions of RhoB in human cancers are context dependent. Although expression of RhoB inversely correlates with disease progression in several epithelial cancers, recent data suggest that RhoB may support malignant phenotypes in certain cancer types. Methods We assessed RhoB protein levels in glioma surgical specimens and patient-derived xenografts. The roles of RhoB in glioblastoma were determined by loss-of-function and gain-of-function assays in vitro and in vivo. The impact on p53 and STAT3 signaling was investigated. Results RhoB expression was similar in tumor specimens compared with normal neural tissues obtained from epilepsy surgery. RhoB was expressed in the vast majority of xenograft tumors and spheroid cultures. Knockdown of RhoB induced cell-cycle arrest and apoptosis and compromised in vivo tumorigenic potential. However, overexpression of wild-type RhoB or a constitutively active mutant (RhoB-V14) did not significantly affect cell growth, which suggests that RhoB is not a rate-limiting oncogenic factor and is consistent with the scarcity of RhoB mutations in human cancer. Knockdown of RhoB reduced basal STAT3 activity and impaired cytokine-induced STAT3 activation. In glioblastoma tumors retaining wild-type p53, depletion of RhoB also activated p53 and induced expression of p21CIP1/WAF1. Conclusions Our data suggest that RhoB belongs to an emerging class of “nononcogene addiction” factors that are essential for maintenance of malignant phenotypes in human cancers. PMID:25216671

The kinase activity of fibroblast growth factor receptor 3 with activation loop mutations affects receptor trafficking and signaling.

PubMed

Lievens, Patricia M-J; Mutinelli, Chiara; Baynes, Darcie; Liboi, Elio

2004-10-08

Amino acid substitutions at the Lys-650 codon within the activation loop kinase domain of fibroblast growth factor receptor 3 (FGFR3) result in graded constitutive phosphorylation of the receptor. Accordingly, the Lys-650 mutants are associated with dwarfisms with graded clinical severity. To assess the importance of the phosphorylation level on FGFR3 maturation along the secretory pathway, hemagglutinin A-tagged derivatives were studied. The highly activated SADDAN (severe achondroplasia with developmental delay and acanthosis nigricans) mutant accumulates in its immature and phosphorylated form in the endoplasmic reticulum (ER), which fails to be degraded. Furthermore, the Janus kinase (Jak)/STAT pathway is activated from the ER by direct recruitment of Jak1. Abolishing the autocatalytic property of the mutated FGFR3 by replacing the critical Tyr-718 reestablishes the receptor full maturation and inhibits signaling. Differently, the low activated hypochondroplasia mutant is present as a mature phosphorylated form on the plasma membrane, although with a delayed transition in the ER, and is completely processed. Signaling does not occur in the presence of brefeldin A; instead, STAT1 is activated when protein secretion is blocked with monensin, suggesting that the hypochondroplasia receptor signals at the exit from the ER. Our results suggest that kinase activity affects FGFR3 trafficking and determines the spatial segregation of signaling pathways. Consequently, the defect in down-regulation of the highly activated receptors results in the increased signaling capacity from the intracellular compartments, and this may determine the severity of the diseases.

An ENU mutagenesis-derived mouse model with a dominant Jak1 mutation resembling phenotypes of systemic autoimmune disease.

PubMed

Sabrautzki, Sibylle; Janas, Eva; Lorenz-Depiereux, Bettina; Calzada-Wack, Julia; Aguilar-Pimentel, Juan A; Rathkolb, Birgit; Adler, Thure; Cohrs, Christian; Hans, Wolfgang; Diener, Susanne; Fuchs, Helmut; Gailus-Durner, Valerie; Busch, Dirk H; Höfler, Heinz; Ollert, Markus; Strom, Tim M; Wolf, Eckhard; Neff, Frauke; Hrabě de Angelis, Martin

2013-08-01

Within the Munich, Germany, N-ethyl-N-nitrosourea mouse mutagenesis program, we isolated a dominant Jak1 mouse model resembling phenotypic characteristics related to autoimmune disease. Chromosomal sequencing revealed a new Jak1 (p.Ser645Pro) point mutation at the conserved serine of the pseudokinase domain, corresponding to a somatic human mutation (p.Ser646Phe) inducing a constitutive activation of the Janus kinase (JAK)/STAT pathway. Morphologically, all Jak1(S645P+/-) mice showed a progressive structural deterioration of ears starting at the age of 4 months, with mononuclear cell infiltration into the dermis. Female mutant mice, in particular, developed severe skin lesions in the neck from 7 months of age. The IHC analysis of these lesions showed an activation of Stat3 downstream to Jak1(S645P) and elevated tissue levels of IL-6. Histopathological analysis of liver revealed a nodular regenerative hyperplasia. In the spleen, the number of Russell bodies was doubled, correlating with significant increased levels of all immunoglobulin isotypes and anti-DNA antibodies in serum. Older mutant mice developed thrombocytopenia and altered microcytic red blood cell counts. Jak1(S645P+/-) mice showed phenotypes related to impaired bone metabolism as increased carboxy-terminal collagen cross-link-1 levels and alkaline phosphatase activities in plasma, hypophosphatemia, and strongly decreased bone morphometric values. Taken together, Jak1(S645P+/-) mice showed an increased activation of the IL-6-JAK-STAT pathway leading to a systemic lupus erythematosus-like phenotype and offering a new valuable tool to study the role of the JAK/STAT pathway in disease development. Copyright © 2013 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

Primary Murine CD4+ T Cells Fail to Acquire the Ability to Produce Effector Cytokines When Active Ras Is Present during Th1/Th2 Differentiation

PubMed Central

Janardhan, Sujit V.; Marks, Reinhard; Gajewski, Thomas F.

2014-01-01

Constitutive Ras signaling has been shown to augment IL-2 production, reverse anergy, and functionally replace many aspects of CD28 co-stimulation in CD4+ T cells. These data raise the possibility that introduction of active Ras into primary T cells might result in improved functionality in pathologic situations of T cell dysfunction, such as cancer or chronic viral infection. To test the biologic effects of active Ras in primary T cells, CD4+ T cells from Coxsackie-Adenovirus Receptor Transgenic mice were transduced with an adenovirus encoding active Ras. As expected, active Ras augmented IL-2 production in naive CD4+ T cells. However, when cells were cultured for 4 days under conditions to promote effector cell differentiation, active Ras inhibited the ability of CD4+ T cells to acquire a Th1 or Th2 effector cytokine profile. This differentiation defect was not due to deficient STAT4 or STAT6 activation by IL-12 or IL-4, respectively, nor was it associated with deficient induction of T-bet and GATA-3 expression. Impaired effector cytokine production in active Ras-transduced cells was associated with deficient demethylation of the IL-4 gene locus. Our results indicate that, despite augmenting acute activation of naïve T cells, constitutive Ras signaling inhibits the ability of CD4+ T cells to properly differentiate into Th1/Th2 effector cytokine-producing cells, in part by interfering with epigenetic modification of effector gene loci. Alternative strategies to potentiate Ras pathway signaling in T cells in a more regulated fashion should be considered as a therapeutic approach to improve immune responses in vivo. PMID:25397617

Roles of Stat3 and ERK in G-CSF signaling.

PubMed

Kamezaki, Kenjirou; Shimoda, Kazuya; Numata, Akihiko; Haro, Takashi; Kakumitsu, Haruko; Yoshie, Masumi; Yamamoto, Masahiro; Takeda, Kiyoshi; Matsuda, Tadashi; Akira, Shizuo; Ogawa, Katsuhiro; Harada, Mine

2005-02-01

G-CSF specifically stimulates the proliferation and differentiation of cells that are committed to the neutrophil-granulocyte lineage. Although Stat3 was thought to be essential for the transduction of G-CSF-induced cell proliferation and differentiation signals, mice deficient for Stat3 in hematopoietic cells show neutrocytosis and infiltration of cells into the digestive tract. The number of progenitor cells in the neutrophil lineage is not changed, and G-CSF-induced proliferation of progenitor cells and prolonged neutrophil survival were observed in Stat3-deficient mice. In hematopoietic cells from Stat3-deficient mice, trace levels of SOCS3, a negative regulator of granulopoiesis, were observed, and SOCS3 expression was not induced by G-CSF stimulation. Stat3-null bone marrow cells displayed a significant activation of extra-cellular regulated kinase 1 (ERK1)/ERK2 under basal conditions, and the activation of ERK was enhanced and sustained by G-CSF stimulation. Furthermore, the augmented proliferation of Stat3-deficient bone marrow cells in response to G-CSF was dramatically decreased by addition of a MEK1 inhibitor. These results indicate that Stat3 functions as a negative regulator of G-CSF signaling by inducing SOCS3 expression and that ERK activation is the major factor responsible for inducing the proliferation of hematopoietic cells in response to G-CSF.

Reciprocal activation of α5-nAChR and STAT3 in nicotine-induced human lung cancer cell proliferation.

PubMed

Zhang, Yao; Jia, Yanfei; Li, Ping; Li, Huanjie; Xiao, Dongjie; Wang, Yunshan; Ma, Xiaoli

2017-07-20

Cigarette smoking is the top environmental risk factor for lung cancer. Nicotine, the addictive component of cigarettes, induces lung cancer cell proliferation, invasion and migration via the activation of nicotinic acetylcholine receptors (nAChRs). Genome-wide association studies (GWAS) show that CHRNA5 gene encoding α5-nAChR is especially relevant to lung cancer. However, the mechanism of this subunit in lung cancer is not clear. In the present study, we demonstrate that the expression of α5-nAChR is correlated with phosphorylated STAT3 (pSTAT3) expression, smoking history and lower survival of non-small cell lung cancer (NSCLC) samples. Nicotine increased the levels of α5-nAChR mRNA and protein in NSCLC cell lines and activated the JAK2/STAT3 signaling cascade. Nicotine-induced activation of JAK2/STAT3 signaling was inhibited by the silencing of α5-nAChR. Characterization of the CHRNA5 promoter revealed four STAT3-response elements. ChIP assays confirmed that the CHRNA5 promoter contains STAT3 binding sites. By silencing STAT3 expression, nicotine-induced upregulation of α5-nAChR was suppressed. Downregulation of α5-nAChR and/or STAT3 expression inhibited nicotine-induced lung cancer cell proliferation. These results suggest that there is a feedback loop between α5-nAChR and STAT3 that contributes to the nicotine-induced tumor cell proliferation, which indicates that α5-nAChR is an important therapeutic target involved in tobacco-associated lung carcinogenesis. Copyright © 2017 Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, and Genetics Society of China. Published by Elsevier Ltd. All rights reserved.

STAT3/IRF1 Pathway Activation Sensitizes Cervical Cancer Cells to Chemotherapeutic Drugs.

PubMed

Walch-Rückheim, Barbara; Pahne-Zeppenfeld, Jennifer; Fischbach, Jil; Wickenhauser, Claudia; Horn, Lars Christian; Tharun, Lars; Büttner, Reinhard; Mallmann, Peter; Stern, Peter; Kim, Yoo-Jin; Bohle, Rainer Maria; Rübe, Christian; Ströder, Russalina; Juhasz-Böss, Ingolf; Solomayer, Erich-Franz; Smola, Sigrun

2016-07-01

Neoadjuvant radio/chemotherapy regimens can markedly improve cervical cancer outcome in a subset of patients, while other patients show poor responses, but may encounter severe adverse effects. Thus, there is a strong need for predictive biomarkers to improve clinical management of cervical cancer patients. STAT3 is considered as a critical antiapoptotic factor in various malignancies. We therefore investigated STAT3 activation during cervical carcinogenesis and its impact on the response of cervical cancer cells to chemotherapeutic drugs. Tyr705-phosphorylated STAT3 increased from low-grade cervical intraepithelial neoplasia (CIN1) to precancerous CIN3 lesions. Notably, pTyr705-STAT3 activation significantly declined from CIN3 to invasive cancer, also when compared in the same clinical biopsy. pTyr705-STAT3 was also low or absent in cultured human cervical cancer cell lines, consistent with the in vivo expression data. Unexpectedly, IL6-type cytokine signaling inducing STAT3 activation rendered cervical cancer cells significantly more susceptible to chemotherapeutic drugs, that is, cisplatin or etoposide. This chemosensitization was STAT3-dependent and we identified IFN regulatory factor-1 (IRF1) as the STAT3-inducible mediator required for cell death enhancement. In line with these data, pTyr705-STAT3 significantly correlated with nuclear IRF1 expression in cervical cancer in vivo Importantly, high IRF1 expression in pretreatment cervical cancer biopsy cells was associated with a significantly better response to neoadjuvant radio/chemotherapy of the patients. In summary, our study has identified a key role of the STAT3/IRF1 pathway for chemosensitization in cervical cancer. Our results suggest that pretherapeutic IRF1 expression should be evaluated as a novel predictive biomarker for neoadjuvant radio/chemotherapy responses. Cancer Res; 76(13); 3872-83. ©2016 AACR. ©2016 American Association for Cancer Research.

The role of signal transducer and activator of transcription 3 in Rift Valley fever virus infection

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pinkham, Chelsea; An, Soyeon; Lundberg, Lindsay

Rift Valley fever (RVF) is a zoonotic disease that can cause severe illness in humans and livestock, triggering spontaneous abortion in almost 100% of pregnant ruminants. In this study, we demonstrate that signal transducer and activator of transcription 3 (STAT3) is phosphorylated on its conserved tyrosine residue (Y705) following RVFV infection. This phosphorylation was dependent on a major virulence factor, the viral nonstructural protein NSs. Loss of STAT3 had little effect on viral replication, but rather resulted in cells being more susceptible to RVFV-induced cell death. Phosphorylated STAT3 translocated to the nucleus, coinciding with inhibition of fos, jun, and nr4a2more » gene expression, and the presence of STAT3 and NSs at the nr4a2 promoter. NSs was found predominantly in the cytoplasm of STAT3 null cells, indicating that STAT3 influences NSs nuclear localization. Collectively, these data demonstrate that STAT3 functions in a pro-survival capacity through modulation of NSs localization. - Highlights: • STAT3 is phosphorylated on tyrosine residue 705 following RVFV infection. • Phosphorylation of STAT3 was dependent on the viral protein NSs. • STAT3 -/- MEFs were more susceptible to RVFV-induced cell death. • Loss of STAT3 led to an increase in pro-apoptotic gene expression. • STAT3 functions in a pro-survival capacity by modulation of NSs localization.« less

Regulation of STATs by polycystin-1 and their role in polycystic kidney disease.

PubMed

Weimbs, Thomas; Olsan, Erin E; Talbot, Jeffrey J

2013-04-01

Autosomal-dominant polycystic kidney disease (ADPKD) is a common genetic disease caused by mutations in the gene coding for polycystin-1 (PC1). PC1 can regulate STAT transcription factors by a novel, dual mechanism. STAT3 and STAT6 are aberrantly activated in renal cysts. Genetic and pharmacological approaches to inhibit STAT3 or STAT6 have led to promising results in ADPKD mouse models. Here, we review current findings that lead to a model of PC1 as a key regulator of STAT signaling in renal tubule cells. We discuss how PC1 may orchestrate appropriate epithelial responses to renal injury, and how this system may lead to aberrant STAT activation in ADPKD thereby causing inappropriate activation of tissue repair programs that culminate in renal cyst growth and fibrosis.

Analysis of the STAT3 interactome using in-situ biotinylation and SILAC.

PubMed

Blumert, Conny; Kalkhof, Stefan; Brocke-Heidrich, Katja; Kohajda, Tibor; von Bergen, Martin; Horn, Friedemann

2013-12-06

Signal transducer and activator of transcription 3 (STAT3) is activated by a variety of cytokines and growth factors. To generate a comprehensive data set of proteins interacting specifically with STAT3, we applied stable isotope labeling with amino acids in cell culture (SILAC). For high-affinity pull-down using streptavidin, we fused STAT3 with a short peptide tag allowing biotinylation in situ (bio-tag), which did not affect STAT3 functions. By this approach, 3642 coprecipitated proteins were detected in human embryonic kidney-293 cells. Filtering using statistical and functional criteria finally extracted 136 proteins as putative interaction partners of STAT3. Both, a physical interaction network analysis and the enrichment of known and predicted interaction partners suggested that our filtering criteria successfully enriched true STAT3 interactors. Our approach identified numerous novel interactors, including ones previously predicted to associate with STAT3. By reciprocal coprecipitation, we were able to verify the physical association between STAT3 and selected interactors, including the novel interaction with TOX4, a member of the TOX high mobility group box family. Applying the same method, we next investigated the activation-dependency of the STAT3 interactome. Again, we identified both known and novel interactions. Thus, our approach allows to study protein-protein interaction effectively and comprehensively. The location, activity, function, degradation, and synthesis of proteins are significantly regulated by interactions of proteins with other proteins, biopolymers and small molecules. Thus, the comprehensive characterization of interactions of proteins in a given proteome is the next milestone on the path to understanding the biochemistry of the cell. In order to generate a comprehensive interactome dataset of proteins specifically interacting with a selected bait protein, we fused our bait protein STAT3 with a short peptide tag allowing biotinylation in situ (bio-tag). This bio-tag allows an affinity pull-down using streptavidin but affected neither the activation of STAT3 by tyrosine phosphorylation nor its transactivating potential. We combined SILAC for accurate relative protein quantification, subcellular fractionation to increase the coverage of interacting proteins, high-affinity pull-down and a stringent filtering method to successfully analyze the interactome of STAT3. With our approach we confirmed several already known and identified numerous novel STAT3 interactors. The approach applied provides a rapid and effective method, which is broadly applicable for studying protein-protein interactions and their dependency on post-translational modifications. © 2013. Published by Elsevier B.V. All rights reserved.

Phosphorylation of Stats at Ser727 in renal proximal tubular epithelial cells exposed to cadmium.

PubMed

Nakagawa, Junko; Nishitai, Gen; Inageda, Kiyoshi; Matsuoka, Masato

2007-11-01

The effects of cadmium exposure on serine phosphorylation of signal transducers and activators of transcription (Stats) and an upstream kinase were examined in renal proximal tubular cells. In porcine LLC-PK1 cells treated with cadmium, Stat1 and Stat3 proteins were phosphorylated at Ser727 without changing total Stat protein levels. While phosphorylated forms of the members of mitogen-activated protein kinases (MAPKs) increased in response to cadmium exposure, treatment with a p38 inhibitor, SB203580 reduced Ser727 phosphorylation of Stat1 and Stat3 markedly in LLC-PK1 cells. The expression of human matrix metalloproteinase-3 (MMP-3), a Stats-inducible gene, was found to be up-regulated in human HK-2 cells exposed to cadmium, and suppressed by preincubation with SB203580. These results suggest that cadmium might induce the phosphorylation of Stat1 and Stat3 at Ser727 via the p38 pathway at least in part, and modulate gene expression in these proximal tubular cells. Copyright © 2007 Elsevier B.V. All rights reserved.

Csf3r mutations in mice confer a strong clonal HSC advantage via activation of Stat5

PubMed Central

Liu, Fulu; Kunter, Ghada; Krem, Maxwell M.; Eades, William C.; Cain, Jennifer A.; Tomasson, Michael H.; Hennighausen, Lothar; Link, Daniel C.

2008-01-01

A fundamental property of leukemic stem cells is clonal dominance of the bone marrow microenvironment. Truncation mutations of CSF3R, which encodes the G-CSF receptor (G-CSFR), are implicated in leukemic progression in patients with severe congenital neutropenia. Here we show that expression of a truncated mutant Csf3r in mice confers a strong clonal advantage at the HSC level that is dependent upon exogenous G-CSF. G-CSF–induced proliferation, phosphorylation of Stat5, and transcription of Stat5 target genes were increased in HSCs isolated from mice expressing the mutant Csf3r. Conversely, the proliferative advantage conferred by the mutant Csf3r was abrogated in myeloid progenitors lacking both Stat5A and Stat5B, and HSC function was reduced in mice expressing a truncated mutant Csf3r engineered to have impaired Stat5 activation. These data indicate that in mice, inappropriate Stat5 activation plays a key role in establishing clonal dominance by stem cells expressing mutant Csf3r. PMID:18292815

A novel de novo activating mutation in STAT3 identified in a patient with common variable immunodeficiency (CVID).

PubMed

Russell, Mark A; Pigors, Manuela; Houssen, Maha E; Manson, Ania; Kelsell, David; Longhurst, Hilary; Morgan, Noel G

2018-02-01

Common variable immunodeficiency (CVID) is characterised by repeated infection associated with primary acquired hypogammaglobulinemia. CVID frequently has a complex aetiology but, in certain cases, it has a monogenic cause. Recently, variants within the gene encoding the transcription factor STAT3 were implicated in monogenic CVID. Here, we describe a patient presenting with symptoms synonymous with CVID, who displayed reduced levels of IgG and IgA, repeated viral infections and multiple additional co-morbidities. Whole-exome sequencing revealed a de novo novel missense mutation in the coiled-coil domain of STAT3 (c.870A>T; p.K290N). Accordingly, the K290N variant of STAT3 was generated, and a STAT3 responsive dual-luciferase reporter assay revealed that the variant strongly enhances STAT3 transcriptional activity both under basal and stimulated (with IL-6) conditions. Overall, these data complement earlier studies in which CVID-associated STAT3 mutations are predicted to enhance transcriptional activity, suggesting that such patients may respond favourably to IL-6 receptor antagonists (e.g. tocilizumab). Copyright © 2017 Elsevier Inc. All rights reserved.

Antagonizing STAT3 dimerization with a rhodium(III) complex.

PubMed

Ma, Dik-Lung; Liu, Li-Juan; Leung, Ka-Ho; Chen, Yen-Ting; Zhong, Hai-Jing; Chan, Daniel Shiu-Hin; Wang, Hui-Min David; Leung, Chung-Hang

2014-08-25

Kinetically inert metal complexes have arisen as promising alternatives to existing platinum and ruthenium chemotherapeutics. Reported herein, to our knowledge, is the first example of a substitutionally inert, Group 9 organometallic compound as a direct inhibitor of signal transducer and activator of transcription 3 (STAT3) dimerization. From a series of cyclometalated rhodium(III) and iridium(III) complexes, a rhodium(III) complex emerged as a potent inhibitor of STAT3 that targeted the SH2 domain and inhibited STAT3 phosphorylation and dimerization. Significantly, the complex exhibited potent anti-tumor activities in an in vivo mouse xenograft model of melanoma. This study demonstrates that rhodium complexes may be developed as effective STAT3 inhibitors with potent anti-tumor activity. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Phosphorylation status determines the opposing functions of Smad2/Smad3 as STAT3 cofactors in TH17 differentiation

PubMed Central

Yoon, Jeong-Hwan; Sudo, Katsuko; Kuroda, Masahiko; Kato, Mitsuyasu; Lee, In-Kyu; Han, Jin Soo; Nakae, Susumu; Imamura, Takeshi; Kim, Juryun; Ju, Ji Hyeon; Kim, Dae-Kee; Matsuzaki, Koichi; Weinstein, Michael; Matsumoto, Isao; Sumida, Takayuki; Mamura, Mizuko

2015-01-01

Transforming growth factor-β (TGF-β) and interleukin-6 (IL-6) are the pivotal cytokines to induce IL-17-producing CD4+ T helper cells (TH17); yet their signalling network remains largely unknown. Here we show that the highly homologous TGF-β receptor-regulated Smads (R-Smads): Smad2 and Smad3 oppositely modify STAT3-induced transcription of IL-17A and retinoic acid receptor-related orphan nuclear receptor, RORγt encoded by Rorc, by acting as a co-activator and co-repressor of STAT3, respectively. Smad2 linker phosphorylated by extracellular signal-regulated kinase (ERK) at the serine 255 residue interacts with STAT3 and p300 to transactivate, whereas carboxy-terminal unphosphorylated Smad3 interacts with STAT3 and protein inhibitor of activated STAT3 (PIAS3) to repress the Rorc and Il17a genes. Our work uncovers carboxy-terminal phosphorylation-independent noncanonical R-Smad–STAT3 signalling network in TH17 differentiation. PMID:26194464

Phosphorylation status determines the opposing functions of Smad2/Smad3 as STAT3 cofactors in TH17 differentiation.

PubMed

Yoon, Jeong-Hwan; Sudo, Katsuko; Kuroda, Masahiko; Kato, Mitsuyasu; Lee, In-Kyu; Han, Jin Soo; Nakae, Susumu; Imamura, Takeshi; Kim, Juryun; Ju, Ji Hyeon; Kim, Dae-Kee; Matsuzaki, Koichi; Weinstein, Michael; Matsumoto, Isao; Sumida, Takayuki; Mamura, Mizuko

2015-07-21

Transforming growth factor-β (TGF-β) and interleukin-6 (IL-6) are the pivotal cytokines to induce IL-17-producing CD4(+) T helper cells (TH17); yet their signalling network remains largely unknown. Here we show that the highly homologous TGF-β receptor-regulated Smads (R-Smads): Smad2 and Smad3 oppositely modify STAT3-induced transcription of IL-17A and retinoic acid receptor-related orphan nuclear receptor, RORγt encoded by Rorc, by acting as a co-activator and co-repressor of STAT3, respectively. Smad2 linker phosphorylated by extracellular signal-regulated kinase (ERK) at the serine 255 residue interacts with STAT3 and p300 to transactivate, whereas carboxy-terminal unphosphorylated Smad3 interacts with STAT3 and protein inhibitor of activated STAT3 (PIAS3) to repress the Rorc and Il17a genes. Our work uncovers carboxy-terminal phosphorylation-independent noncanonical R-Smad-STAT3 signalling network in TH17 differentiation.

STAT3 selectively interacts with Smad3 to antagonize TGF-β signaling

PubMed Central

Wang, Gaohang; Yu, Yi; Sun, Chuang; Liu, Ting; Liang, Tingbo; Zhan, Lixing; Lin, Xia; Feng, Xin-Hua

2015-01-01

Smad and STAT proteins are critical signal transducers and transcription factors in controlling cell growth and tumorigenesis. Here we report that the STAT3 signaling pathway attenuates TGF-β-induced responses through a direct Smad3-STAT3 interplay. Activated STAT3 blunts TGF-β-mediated signaling. Depletion of STAT3 promotes TGF-β-mediated transcriptional and physiological responses, including cell cycle arrest, apoptosis and epithelial-to-mesenchymal transition. STAT3 directly interacts with Smad3 in vivo and in vitro, resulting in attenuation of the Smad3-Smad4 complex formation and suppression of DNA-binding ability of Smad3. The N-terminal region of DNA-binding domain of STAT3 is responsible for the STAT3-Smad3 interaction and also indispensable for STAT3-mediated inhibition of TGF-β signaling. Thus, our finding illustrates a direct crosstalk between the STAT3 and Smad3 signaling pathways that may contribute to tumor development and inflammation. PMID:26616859

Genome-wide STAT3 binding analysis after histone deacetylase inhibition reveals novel target genes in dendritic cells

PubMed Central

Sun, Yaping; Iyer, Matthew; McEachin, Richard; Zhao, Meng; Wu, Yi-Mi; Cao, Xuhong; Oravecz-Wilson, Katherine; Zajac, Cynthia; Mathewson, Nathan; Wu, Shin-Rong Julia; Rossi, Corinne; Toubai, Tomomi; Qin, Zhaohui S.; Chinnaiya, Arul M.; Reddy, Pavan

2016-01-01

STAT3 is a master transcriptional regulator that plays an important role in the induction of both immune activation and immune tolerance in dendritic cells (DCs). The transcriptional targets of STAT3 in promoting DC activation are becoming increasingly understood; however, the mechanisms underpinning its role in causing DC suppression remain largely unknown. To determine the functional gene targets of STAT3, we compared the genome-wide binding of STAT3 using ChIP-seq coupled with gene expression microarrays to determine STAT3-dependent gene regulation in DCs after histone deacetylase (HDAC) inhibition. HDAC inhibition boosted the ability of STAT3 to bind to distinct DNA targets and regulate gene expression. Among the top 500 STAT3 binding sites, the frequency of canonical motifs was significantly higher than that of non-canonical motifs. Functional analysis revealed that after treatment with an HDAC inhibitor, the upregulated STAT3 target genes were those that were primarily the negative regulators of pro-inflammatory cytokines and those in the IL-10 signaling pathway. The downregulated STAT3-dependent targets were those involved in immune effector processes and antigen processing/presentation. The expression and functional relevance of these genes were validated. Specifically, functional studies confirmed that the upregulation of IL-10Ra by STAT3 contributed to the suppressive function of DCs following HDAC inhibition. PMID:27866206

Signal Transducer and Activator of Transcription 1 (STAT1) Knock-down Induces Apoptosis in Malignant Pleural Mesothelioma.

PubMed

Arzt, Lisa; Halbwedl, Iris; Gogg-Kamerer, Margit; Popper, Helmut H

2017-07-01

Malignant pleural mesothelioma (MPM) is the most common primary tumor of the pleura. Its incidence is still increasing in Europe and the prognosis remains poor. We investigated the oncogenic function of signal transducer and activator of transcription 1 (STAT1) in MPM in more detail. A miRNA profiling was performed on 52 MPM tissue samples. Upregulated miRNAs (targeting SOCS1/3) were knocked-down using miRNA inhibitors. mRNA expression levels of STAT1/3, SOCS1/3 were detected in MPM cell lines. STAT1 has been knocked-down using siRNA and qPCR was used to detect mRNA expression levels of all JAK/STAT family members and genes that regulate them. An immunohistochemical staining was performed to detect the expression of caspases. STAT1 was upregulated and STAT3 was downregulated, SOCS1/3 protein was not detected but it was possible to detect SOCS1/3 mRNA in MPM cell lines. The upregulated miRNAs were successfully knocked-down, however the expected effect on SOCS1 expression was not detected. STAT1 knock-down had different effects on STAT3/5 expression. Caspase 3a and 8 expression was found to be increased after STAT1 knock-down. The physiologic regulation of STAT1 via SOCS1 is completely lost in MPM and it does not seem that the miRNAs identified by now, do inhibit the expression of SOCS1. MPM cell lines compensate STAT1 knock-down by increasing the expression of STAT3 or STAT5a, two genes which are generally considered to be oncogenes. And much more important, STAT1 knock-down induces apoptosis in MPM cell lines and STAT1 might therefore be a target for therapeutic intervention.

Antitumor activity of IL-32β through the activation of lymphocytes, and the inactivation of NF-κB and STAT3 signals.

PubMed

Yun, H-M; Oh, J H; Shim, J-H; Ban, J O; Park, K-R; Kim, J-H; Lee, D H; Kang, J-W; Park, Y H; Yu, D; Kim, Y; Han, S B; Yoon, D-Y; Hong, J T

2013-05-23

Cytokine and activation of lymphocytes are critical for tumor growth. We investigated whether interleukin (IL)-32β overexpression changes other cytokine levels and activates cytotoxic lymphocyte, and thus modify tumor growth. Herein, IL-32β inhibited B16 melanoma growth in IL-32β-overexpressing transgenic mice (IL-32β mice), and downregulated the expressions of anti-apoptotic proteins (bcl-2, IAP, and XIAP) and cell growth regulatory proteins (Ki-67 antigen (Ki-67) and proliferating cell nuclear antigen (PCNA)), but upregulated the expressions of pro-apoptotic proteins (bax, cleaved caspase-3, and cleaved caspase-9). IL-32β also inhibited colon and prostate tumor growth in athymic nude mice inoculated with IL-32β-transfected SW620 colon or PC3 prostate cancer cells. The forced expression of IL-32β also inhibited cell growth in cultured colon and prostate cancer cells, and these inhibitory effects were abolished by IL-32 small interfering RNA (siRNA). IL-10 levels were elevated, but IL-1β, IL-6, and tumor necrosis factor-alpha (TNF-α) levels were reduced in the tumor tissues and spleens of IL-32β mice, and athymic nude mice. The number of cytotoxic T (CD8(+)) and natural killer (NK) cells in tumor tissues, spleen, and blood was significantly elevated in IL-32β mice and athymic nude mice inoculated with IL-32β-transfected cancer cells. Constituted activated NF-κB and STAT3 levels were reduced in the tumor tissues of IL-32β mice and athymic nude mice, as well as in IL-32β-transfected cultured cancer cells. These findings suggest that IL-32β inhibits tumor growth by increasing cytotoxic lymphocyte numbers, and by inactivating the NF-κB and STAT3 pathways through changing of cytokine levels in tumor tissues.

Assessing the Role of STAT3 in DC Differentiation and Autologous DC Immunotherapy in Mouse Models of GBM

PubMed Central

Assi, Hikmat; Espinosa, Jaclyn; Suprise, Sarah; Sofroniew, Michael; Doherty, Robert; Zamler, Daniel; Lowenstein, Pedro R.; Castro, Maria G.

2014-01-01

Cellular microenvironments, particularly those found in tumors, elicit a tolerogenic DC phenotype which can attenuate immune responses. Central to this process is the STAT3-mediated signaling cascade. As a transcription factor and oncogene, STAT3 promotes the expression of genes which allow tumor cells to proliferate, migrate and evade apoptosis. More importantly, activation of STAT3 in tumor infiltrating immune cells has been shown to be responsible, in part, for their immune-suppressed phenotype. The ability of STAT3 to orchestrate a diverse set of immunosuppressive instructions has made it an attractive target for cancer vaccines. Using a conditional hematopoietic knockout mouse model of STAT3, we evaluated the impact of STAT3 gene ablation on the differentiation of dendritic cells from bone marrow precursors. We also assessed the impact of STAT3 deletion on phagocytosis, maturation, cytokine secretion and antigen presentation by GM-CSF derived DCs in vitro. In addition to in vitro studies, we compared the therapeutic efficacy of DC vaccination using STAT3 deficient DCs to wild type counterparts in an intracranial mouse model of GBM. Our results indicated the following pleiotropic functions of STAT3: hematopoietic cells which lacked STAT3 were unresponsive to Flt3L and failed to differentiate as DCs. In contrast, STAT3 was not required for GM-CSF induced DC differentiation as both wild type and STAT3 null bone marrow cells gave rise to similar number of DCs. STAT3 also appeared to regulate the response of GM-CSF derived DCs to CpG. STAT3 null DCs expressed high levels of MHC-II, secreted more IL-12p70, IL-10, and TNFα were better antigen presenters in vitro. Although STAT3 deficient DCs displayed an enhanced activated phenotype in culture, they elicited comparable therapeutic efficacy in vivo compared to their wild type counterparts when utilized in vaccination paradigms in mice bearing intracranial glioma tumors. PMID:24806510

Assessing the role of STAT3 in DC differentiation and autologous DC immunotherapy in mouse models of GBM.

PubMed

Assi, Hikmat; Espinosa, Jaclyn; Suprise, Sarah; Sofroniew, Michael; Doherty, Robert; Zamler, Daniel; Lowenstein, Pedro R; Castro, Maria G

2014-01-01

Cellular microenvironments, particularly those found in tumors, elicit a tolerogenic DC phenotype which can attenuate immune responses. Central to this process is the STAT3-mediated signaling cascade. As a transcription factor and oncogene, STAT3 promotes the expression of genes which allow tumor cells to proliferate, migrate and evade apoptosis. More importantly, activation of STAT3 in tumor infiltrating immune cells has been shown to be responsible, in part, for their immune-suppressed phenotype. The ability of STAT3 to orchestrate a diverse set of immunosuppressive instructions has made it an attractive target for cancer vaccines. Using a conditional hematopoietic knockout mouse model of STAT3, we evaluated the impact of STAT3 gene ablation on the differentiation of dendritic cells from bone marrow precursors. We also assessed the impact of STAT3 deletion on phagocytosis, maturation, cytokine secretion and antigen presentation by GM-CSF derived DCs in vitro. In addition to in vitro studies, we compared the therapeutic efficacy of DC vaccination using STAT3 deficient DCs to wild type counterparts in an intracranial mouse model of GBM. Our results indicated the following pleiotropic functions of STAT3: hematopoietic cells which lacked STAT3 were unresponsive to Flt3L and failed to differentiate as DCs. In contrast, STAT3 was not required for GM-CSF induced DC differentiation as both wild type and STAT3 null bone marrow cells gave rise to similar number of DCs. STAT3 also appeared to regulate the response of GM-CSF derived DCs to CpG. STAT3 null DCs expressed high levels of MHC-II, secreted more IL-12p70, IL-10, and TNFα were better antigen presenters in vitro. Although STAT3 deficient DCs displayed an enhanced activated phenotype in culture, they elicited comparable therapeutic efficacy in vivo compared to their wild type counterparts when utilized in vaccination paradigms in mice bearing intracranial glioma tumors.

The Ying and Yang of STAT3 in Human Disease.

PubMed

Vogel, Tiphanie P; Milner, Joshua D; Cooper, Megan A

2015-10-01

The transcription factor signal transducer and activator of transcription 3 (STAT3) is a critical regulator of multiple, diverse cellular processes. Heterozgyous, germline, loss-of-function mutations in STAT3 lead to the primary immune deficiency Hyper-IgE syndrome. Heterozygous, somatic, gain-of-function mutations in STAT3 have been reported in malignancy. Recently, germline, heterozygous mutations in STAT3 that confer a gain-of-function have been discovered and result in early-onset, multi-organ autoimmunity. This review summarizes what is known about the role of STAT3 in human disease.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Unterberger, Claudia; Hanson, Steven; Department of Infection, Immunity and Inflammation, University of Leicester, University Road, Leicester LE1 9HN

Little is known about determinants regulating expression of Mannan-binding lectin associated serine protease-2 (MASP-2), the effector component of the lectin pathway of complement activation. Comparative bioinformatic analysis of the MASP2 promoter regions in human, mouse, and rat, revealed conservation of two putative Stat binding sites, termed StatA and StatB. Site directed mutagenesis specific for these sites was performed. Transcription activity was decreased 5-fold when StatB site was mutated in the wildtype reporter gene construct. Gel retardation and competition assays demonstrated that proteins contained in the nuclear extract prepared from HepG2 specifically bound double-stranded StatB oligonucleotides. Supershift analysis revealed Stat3 tomore » be the major specific binding protein. We conclude that Stat3 binding is important for MASP2 promoter activity.« less

Lithium Suppresses Astrogliogenesis by Neural Stem and Progenitor Cells by Inhibiting STAT3 Pathway Independently of Glycogen Synthase Kinase 3 Beta

PubMed Central

Zhu, Zhenzhong; Kremer, Penny; Tadmori, Iman; Ren, Yi; Sun, Dongming; He, Xijing; Young, Wise

2011-01-01

Transplanted neural stem and progenitor cells (NSCs) produce mostly astrocytes in injured spinal cords. Lithium stimulates neurogenesis by inhibiting GSK3b (glycogen synthetase kinase 3-beta) and increasing WNT/beta catenin. Lithium suppresses astrogliogenesis but the mechanisms were unclear. We cultured NSCs from subventricular zone of neonatal rats and showed that lithium reduced NSC production of astrocytes as well as proliferation of glia restricted progenitor (GRP) cells. Lithium strongly inhibited STAT3 (signal transducer and activator of transcription 3) activation, a messenger system known to promote astrogliogenesis and cancer. Lithium abolished STAT3 activation and astrogliogenesis induced by a STAT3 agonist AICAR (5-aminoimidazole-4-carboxamide 1-beta-D-ribofuranoside), suggesting that lithium suppresses astrogliogenesis by inhibiting STAT3. GSK3β inhibition either by a specific GSK3β inhibitor SB216763 or overexpression of GID5-6 (GSK3β Interaction Domain aa380 to 404) did not suppress astrogliogenesis and GRP proliferation. GSK3β inhibition also did not suppress STAT3 activation. Together, these results indicate that lithium inhibits astrogliogenesis through non-GSK3β-mediated inhibition of STAT. Lithium may increase efficacy of NSC transplants by increasing neurogenesis and reducing astrogliogenesis. Our results also may explain the strong safety record of lithium treatment of manic depression. Millions of people take high-dose (>1 gram/day) lithium carbonate for a lifetime. GSK3b inhibition increases WNT/beta catenin, associated with colon and other cancers. STAT3 inhibition may reduce risk for cancer. PMID:21931595

STAT3 balances myocyte hypertrophy vis-à-vis autophagy in response to Angiotensin II by modulating the AMPKα/mTOR axis.

PubMed

Chen, Lei; Zhao, Lin; Samanta, Anweshan; Mahmoudi, Seyed Morteza; Buehler, Tanner; Cantilena, Amy; Vincent, Robert J; Girgis, Magdy; Breeden, Joshua; Asante, Samuel; Xuan, Yu-Ting; Dawn, Buddhadeb

2017-01-01

Signal transducers and activators of transcription 3 (STAT3) is known to participate in various cardiovascular signal transduction pathways, including those responsible for cardiac hypertrophy and cytoprotection. However, the role of STAT3 signaling in cardiomyocyte autophagy remains unclear. We tested the hypothesis that Angiotensin II (Ang II)-induced cardiomyocyte hypertrophy is effected, at least in part, through STAT3-mediated inhibition of cellular autophagy. In H9c2 cells, Ang II treatment resulted in STAT3 activation and cellular hypertrophy in a dose-dependent manner. Ang II enhanced autophagy, albeit without impacting AMPKα/mTOR signaling or cellular ADP/ATP ratio. Pharmacologic inhibition of STAT3 with WP1066 suppressed Ang II-induced myocyte hypertrophy and mRNA expression of hypertrophy-related genes ANP and β-MHC. These molecular events were recapitulated in cells with STAT3 knockdown. Genetic or pharmacologic inhibition of STAT3 significantly increased myocyte ADP/ATP ratio and enhanced autophagy through AMPKα/mTOR signaling. Pharmacologic activation and inhibition of AMPKα attenuated and exaggerated, respectively, the effects of Ang II on ANP and β-MHC gene expression, while concomitant inhibition of STAT3 accentuated the inhibition of hypertrophy. Together, these data indicate that novel nongenomic effects of STAT3 influence myocyte energy status and modulate AMPKα/mTOR signaling and autophagy to balance the transcriptional hypertrophic response to Ang II stimulation. These findings may have significant relevance for various cardiovascular pathological processes mediated by Ang II signaling.

STAT3 balances myocyte hypertrophy vis-à-vis autophagy in response to Angiotensin II by modulating the AMPKα/mTOR axis

PubMed Central

Samanta, Anweshan; Mahmoudi, Seyed Morteza; Buehler, Tanner; Cantilena, Amy; Vincent, Robert J.; Girgis, Magdy; Breeden, Joshua; Asante, Samuel; Xuan, Yu-Ting

2017-01-01

Signal transducers and activators of transcription 3 (STAT3) is known to participate in various cardiovascular signal transduction pathways, including those responsible for cardiac hypertrophy and cytoprotection. However, the role of STAT3 signaling in cardiomyocyte autophagy remains unclear. We tested the hypothesis that Angiotensin II (Ang II)-induced cardiomyocyte hypertrophy is effected, at least in part, through STAT3-mediated inhibition of cellular autophagy. In H9c2 cells, Ang II treatment resulted in STAT3 activation and cellular hypertrophy in a dose-dependent manner. Ang II enhanced autophagy, albeit without impacting AMPKα/mTOR signaling or cellular ADP/ATP ratio. Pharmacologic inhibition of STAT3 with WP1066 suppressed Ang II-induced myocyte hypertrophy and mRNA expression of hypertrophy-related genes ANP and β-MHC. These molecular events were recapitulated in cells with STAT3 knockdown. Genetic or pharmacologic inhibition of STAT3 significantly increased myocyte ADP/ATP ratio and enhanced autophagy through AMPKα/mTOR signaling. Pharmacologic activation and inhibition of AMPKα attenuated and exaggerated, respectively, the effects of Ang II on ANP and β-MHC gene expression, while concomitant inhibition of STAT3 accentuated the inhibition of hypertrophy. Together, these data indicate that novel nongenomic effects of STAT3 influence myocyte energy status and modulate AMPKα/mTOR signaling and autophagy to balance the transcriptional hypertrophic response to Ang II stimulation. These findings may have significant relevance for various cardiovascular pathological processes mediated by Ang II signaling. PMID:28686615

Nuclear translocation of phosphorylated STAT3 regulates VEGF-A-induced lymphatic endothelial cell migration and tube formation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Okazaki, Hideki; Tokumaru, Sho; Hanakawa, Yasushi

2011-09-02

Highlights: {yields} VEGF-A enhanced lymphatic endothelial cell migration and increased tube formation. {yields} VEGF-A treated lymphatic endothelial cell showed activation of STAT3. {yields} Dominant-negative STAT3 inhibited VEGF-A-induced lymphatic endothelial cell migration and tube formation. -- Abstract: Vascular endothelial growth factor (VEGF) is an endothelial cell-specific growth factor that regulates endothelial functions, and signal transducers and activators of transcription (STATs) are known to be important during VEGF receptor signaling. The aim of this study was to determine whether STAT3 regulates VEGF-induced lymphatic endothelial cell (LEC) migration and tube formation. VEGF-A (33 ng/ml) enhanced LEC migration by 2-fold and increased tube lengthmore » by 25% compared with the control, as analyzed using a Boyden chamber and Matrigel assay, respectively. Western blot analysis and immunostaining revealed that VEGF-A induced the nuclear translocation of phosphorylated STAT3 in LECs, and this translocation was blocked by the transfection of LECs with an adenovirus vector expressing a dominant-negative mutant of STAT3 (Ax-STAT3F). Transfection with Ax-STAT3F also almost completely inhibited VEGF-A-induced LEC migration and tube formation. These results indicate that STAT3 is essential for VEGF-A-induced LEC migration and tube formation and that STAT3 regulates LEC functions.« less

STAT3 Potentiates SIAH-1 Mediated Proteasomal Degradation of β-Catenin in Human Embryonic Kidney Cells.

PubMed

Shin, Minkyung; Yi, Eun Hee; Kim, Byung-Hak; Shin, Jae-Cheon; Park, Jung Youl; Cho, Chung-Hyun; Park, Jong-Wan; Choi, Kang-Yell; Ye, Sang-Kyu

2016-11-30

The β-catenin functions as an adhesion molecule and a component of the Wnt signaling pathway. In the absence of the Wnt ligand, β-catenin is constantly phosphorylated, which designates it for degradation by the APC complex. This process is one of the key regulatory mechanisms of β-catenin. The level of β-catenin is also controlled by the E3 ubiquitin protein ligase SIAH-1 via a phosphorylation-independent degradation pathway. Similar to β-catenin, STAT3 is responsible for various cellular processes, such as survival, proliferation, and differentiation. However, little is known about how these molecules work together to regulate diverse cellular processes. In this study, we investigated the regulatory relationship between STAT3 and β-catenin in HEK293T cells. To our knowledge, this is the first study to report that β-catenin-TCF-4 transcriptional activity was suppressed by phosphorylated STAT3; furthermore, STAT3 inactivation abolished this effect and elevated activated β-catenin levels. STAT3 also showed a strong interaction with SIAH-1, a regulator of active β-catenin via degradation, which stabilized SIAH-1 and increased its interaction with β-catenin. These results suggest that activated STAT3 regulates active β-catenin protein levels via stabilization of SIAH-1 and the subsequent ubiquitin-dependent proteasomal degradation of β-catenin in HEK293T cells.

Comprehensive meta-analysis of Signal Transducers and Activators of Transcription (STAT) genomic binding patterns discerns cell-specific cis-regulatory modules

PubMed Central

2013-01-01

Background Cytokine-activated transcription factors from the STAT (Signal Transducers and Activators of Transcription) family control common and context-specific genetic programs. It is not clear to what extent cell-specific features determine the binding capacity of seven STAT members and to what degree they share genetic targets. Molecular insight into the biology of STATs was gained from a meta-analysis of 29 available ChIP-seq data sets covering genome-wide occupancy of STATs 1, 3, 4, 5A, 5B and 6 in several cell types. Results We determined that the genomic binding capacity of STATs is primarily defined by the cell type and to a lesser extent by individual family members. For example, the overlap of shared binding sites between STATs 3 and 5 in T cells is greater than that between STAT5 in T cells and non-T cells. Even for the top 1,000 highly enriched STAT binding sites, ~15% of STAT5 binding sites in mouse female liver are shared by other STATs in different cell types while in T cells ~90% of STAT5 binding sites are co-occupied by STAT3, STAT4 and STAT6. In addition, we identified 116 cis-regulatory modules (CRM), which are recognized by all STAT members across cell types defining a common JAK-STAT signature. Lastly, in liver STAT5 binding significantly coincides with binding of the cell-specific transcription factors HNF4A, FOXA1 and FOXA2 and is associated with cell-type specific gene transcription. Conclusions Our results suggest that genomic binding of STATs is primarily determined by the cell type and further specificity is achieved in part by juxtaposed binding of cell-specific transcription factors. PMID:23324445

Tofacitinib regulates synovial inflammation in psoriatic arthritis, inhibiting STAT activation and induction of negative feedback inhibitors

PubMed Central

Gao, W; McGarry, T; Orr, C; McCormick, J; Veale, D J; Fearon, U

2016-01-01

Background Psoriatic arthritis (PsA) is a chronic inflammatory disease, characterised by synovitis and destruction of articular cartilage/bone. Janus-kinase and signal transducer and activator of transcription (JAK-STAT) signalling pathway is implicated in the pathogenesis of PsA. Objectives To examine the effect of tofacitinib (JAK inhibitor) on proinflammatory mechanisms in PsA. Methods Primary PsA synovial fibroblasts (PsAFLS) and ex vivo PsA synovial explants were cultured with tofacitinib (1 µM). PhosphoSTAT3 (pSTAT3), phosphoSTAT1 (pSTAT1), suppressor of cytokine signaling-3 (SOCS3), protein inhibitor of activated Stat3 (PIAS3) and nuclear factor kappa B cells (NFκBp65) were quantified by western blot. The effect of tofacitinib on PsAFLS migration, invasion, Matrigel network formation and matrix metallopeptidase (MMP)2/9 was quantified by invasion/migration assays and zymography. Interleukin (IL)-6, IL-8, IFN-gamma-inducible protein 10 (IP-10) monocyte chemoattractant protein (MCP)-1, IL-17, IL-10, MMP3 and tissue inhibitor of metalloproteinases 3 (TIMP3) were assessed by ELISA. Results Tofacitinib significantly decreased pSTAT3, pSTAT1, NFκBp65 and induced SOCS3 and PIAS3 expression in PsAFLS and synovial explant cultures (p<0.05). Functionally, PsAFLS invasion, network formation and migration were inhibited by tofacitinib (all p<0.05). In PsA explant, tofacitinib significantly decreased spontaneous secretion of IL-6, IL-8, MCP-1, MMP9/MMP2, MMP3 (all p<0.05) and decreased the MMP3/TIMP3 ratio (p<0.05), with no effect observed for IP-10 or IL-10. Conclusions This study further supports JAK-STAT inhibition as a therapeutic target for the treatment of PsA. PMID:26353790

Herbal remedy magnolol suppresses IL-6-induced STAT3 activation and gene expression in endothelial cells

PubMed Central

Chen, Shih-Chung; Chang, Ying-Ling; Wang, Danny Ling; Cheng, Jing-Jy

2006-01-01

Magnolol (Mag), an active constituent isolated from the Chinese herb Hou p'u (Magnolia officinalis) has long been used to suppress inflammatory processes. Chronic inflammation is well known to be involved in vascular injuries such as atherosclerosis in which interleukin (IL)-6 may participate. Signal transducer and activator of transcription protein 3 (STAT3), a transcription factor involved in inflammation and the cell cycle, is activated by IL-6. In this study, we evaluated whether Mag can serve as an anti-inflammatory agent during endothelial injuries. The effects of Mag on IL-6-induced STAT3 activation and downstream target gene induction in endothelial cells (ECs) were examined. Pretreatment of ECs with Mag dose dependently inhibited IL-6-induced Tyr705 and Ser727 phosphorylation in STAT3 without affecting the phosphorylation of JAK1, JAK2, and ERK1/2. Mag pretreatment of these ECs dose dependently suppressed IL-6-induced promoter activity of intracellular cell adhesion molecule (ICAM)-1 that contains functional IL-6 response elements (IREs). An electrophoretic mobility shift assay (EMSA) revealed that Mag treatment significantly reduced STAT3 binding to the IRE region. Consistently, Mag treatment markedly inhibited ICAM-1 expression on the endothelial surface. As a result, reduced monocyte adhesion to IL-6-activated ECs was observed. Furthermore, Mag suppressed IL-6-induced promoter activity of cyclin D1 and monocyte chemotactic protein (MCP)-1 for which STAT3 activation plays a role. In conclusion, our results indicate that Mag inhibits IL-6-induced STAT3 activation and subsequently results in the suppression of downstream target gene expression in ECs. These results provide a therapeutic basis for the development of Mag as an anti-inflammatory agent for vascular disorders including atherosclerosis. PMID:16520748

Activation of the GP130-STAT3 axis and its potential implications in nonalcoholic fatty liver disease

PubMed Central

Min, Hae-Ki; Mirshahi, Faridoddin; Verdianelli, Aurora; Pacana, Tommy; Patel, Vaishali; Park, Chun-Geon; Choi, Aejin; Lee, Jeong-Hoon; Park, Chung-Berm; Ren, Shunlin

2015-01-01

The status of the GP130-STAT3 signaling pathway in humans with nonalcoholic fatty liver disease (NAFLD) and its relevance to disease pathogenesis are unknown. The expression of the gp130-STAT3 axis and gp130 cytokine receptors were studied in subjects with varying phenotypes of NAFLD including nonalcoholic steatohepatitis (NASH) and compared with lean and weight-matched controls without NAFLD. Gp130 and its downstream signaling element (Tyk2 and STAT3) expression were inhibited in obese controls whereas they were increased in NAFLD. IL-6 levels were increased in NASH and correlated with gp130 expression (P < 0.01). Palmitate inhibited gp130-STAT3 expression and signaling. IL-6 and palmitate inhibited hepatic insulin signaling via STAT3-dependent and independent mechanisms, respectively. STAT3 overexpression reversed palmitate-induced lipotoxicity by increasing autophagy (ATG7) and decreasing endoplasmic reticulum stress. These data demonstrate that the STAT3 pathway is activated in NAFLD and can worsen insulin resistance while protecting against other lipotoxic mechanisms of disease pathogenesis. PMID:25747354

Chlorogenic acid suppresses lipopolysaccharide‑induced nitric oxide and interleukin‑1β expression by inhibiting JAK2/STAT3 activation in RAW264.7 cells.

PubMed

Kim, Sang-Hun; Park, Sun-Young; Park, Young-Lan; Myung, Dae-Seong; Rew, Jong-Sun; Joo, Young-Eun

2017-12-01

Chlorogenic acid (CA) is a phenolic compound purified from coffee, fruits and their associated beverages, which possess various biological properties, such as antioxidant and anticarcinogenic activities. The present study evaluated the effects of CA on lipopolysaccharide (LPS)‑induced inflammation in RAW264.7 cells and the associated intracellular signaling pathways using reverse transcription‑quantitative polymerase chain reaction, western blotting and enzyme‑linked immunosorbent assays. CA pretreatment inhibited LPS‑induced expression of inducible nitric oxide synthase (iNOS), nitric oxide (NO) and pro‑inflammatory mediators including interleukin (IL)‑6, tumor necrosis factor‑α (TNF‑α), macrophage inflammatory protein‑2 (MIP‑2) and IL‑1β in RAW264.7 cells. In addition, phosphorylation of Janus kinase 2/signal transducer and activator of transcription 3 (JAK2/STAT3) with LPS was inhibited by CA pretreatment. CA and STAT3 inhibitor (STAT3i) pretreatment inhibited LPS‑induced nuclear translocation of phosphorylated STAT3. In addition, STAT3i inhibited the LPS‑induced expression of iNOS, NO and IL‑1β similar to the results of CA pretreatment. By contrast, STAT3i did not inhibit the LPS‑induced increase in IL‑6, TNF‑α and MIP‑2 expression. These results indicate that CA may suppress LPS‑induced NO and IL‑1β expression by inhibiting JAK2/STAT3 activation in RAW264.7 cells.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Han, Seong-Su, E-mail: seong-su-han@uiowa.edu; Han, Sangwoo; Kamberos, Natalie L.

Highlights: • PL inhibits the proliferation of B-ALL cell lines irrespective of GC-resistance. • PL selectively kills B-ALL cells by increasing ROS, but not normal counterpart. • PL does not sensitize majority of B-ALL cells to DEX. • PL represses the network of constitutively activated TFs and modulates their target genes. • PL may serve as a new therapeutic molecule for GC-resistant B-ALL. - Abstract: Piperlongumine (PL), a pepper plant alkaloid from Piper longum, has anti-inflammatory and anti-cancer properties. PL selectively kills both solid and hematologic cancer cells, but not normal counterparts. Here we evaluated the effect of PL onmore » the proliferation and survival of B-cell acute lymphoblastic leukemia (B-ALL), including glucocorticoid (GC)-resistant B-ALL. Regardless of GC-resistance, PL inhibited the proliferation of all B-ALL cell lines, but not normal B cells, in a dose- and time-dependent manner and induced apoptosis via elevation of ROS. Interestingly, PL did not sensitize most of B-ALL cell lines to dexamethasone (DEX). Only UoC-B1 exhibited a weak synergistic effect between PL and DEX. All B-ALL cell lines tested exhibited constitutive activation of multiple transcription factors (TFs), including AP-1, MYC, NF-κB, SP1, STAT1, STAT3, STAT6 and YY1. Treatment of the B-ALL cells with PL significantly downregulated these TFs and modulated their target genes. While activation of AURKB, BIRC5, E2F1, and MYB mRNA levels were significantly downregulated by PL, but SOX4 and XBP levels were increased by PL. Intriguingly, PL also increased the expression of p21 in B-ALL cells through a p53-independent mechanism. Given that these TFs and their target genes play critical roles in a variety of hematological malignancies, our findings provide a strong preclinical rationale for considering PL as a new therapeutic agent for the treatment of B-cell malignancies, including B-ALL and GC-resistant B-ALL.« less

GLI1 inhibitor GANT61 exhibits antitumor efficacy in T-cell lymphoma cells through down-regulation of p-STAT3 and SOCS3

PubMed Central

Geng, Lingyun; Lu, Kang; Li, Peipei; Li, Xinyu; Zhou, Xiangxiang; Li, Ying; Wang, Xin

2017-01-01

T-cell lymphomas are lymphoid malignancies with aggressive clinical course and poor prognosis. Increasing evidences suggest that deregulation of signal transducer and activator of transcription-3 (STAT3) and suppressor of cytokine signaling 3 (SOCS3) is associated with the pathogenesis of T-cell lymphomas. The hedgehog (Hh)/glioma-associated oncogene-1 (GLI1) pathway, aberrantly activated in a number of tumors, has also been extensively studied. We found that protein expressions of GL11, p-STAT3, STAT3, and SOCS3 were up-regulated in T-cell lymphoma tissues and cell lines. Moreover, the protein expressions of p-STAT3 and SOCS3 were positively correlated with GLI1 in T-cell lymphomas. GLI1 inhibitor GANT61 and lentivirus-mediated siGLI1 exhibited inhibitory effects in the three T-cell lines (Jurkat, Karpass299 and Myla3676 cells). The protein expressions of p-STAT3 and SOCS3 were decreased accompanied with the inhibition of GLI1. These findings indicated that GANT61 is a promising agent against T-cell lymphoma and the antitumor activity might be partly mediated by down-regulating p-STAT3 and SOCS3. PMID:27275540

The peroxisome proliferator-activated receptor (PPAR) β/δ agonist GW501516 inhibits IL-6-induced signal transducer and activator of transcription 3 (STAT3) activation and insulin resistance in human liver cells.

PubMed

Serrano-Marco, L; Barroso, E; El Kochairi, I; Palomer, X; Michalik, L; Wahli, W; Vázquez-Carrera, M

2012-03-01

IL-6 induces insulin resistance by activating signal transducer and activator of transcription 3 (STAT3) and upregulating the transcription of its target gene SOCS3. Here we examined whether the peroxisome proliferator-activated receptor (PPAR)β/δ agonist GW501516 prevented activation of the IL-6-STAT3-suppressor of cytokine signalling 3 (SOCS3) pathway and insulin resistance in human hepatic HepG2 cells. Studies were conducted with human HepG2 cells and livers from mice null for Pparβ/δ (also known as Ppard) and wild-type mice. GW501516 prevented IL-6-dependent reduction in insulin-stimulated v-akt murine thymoma viral oncogene homologue 1 (AKT) phosphorylation and in IRS-1 and IRS-2 protein levels. In addition, treatment with this drug abolished IL-6-induced STAT3 phosphorylation of Tyr⁷⁰⁵ and Ser⁷²⁷ and prevented the increase in SOCS3 caused by this cytokine. Moreover, GW501516 prevented IL-6-dependent induction of extracellular-related kinase 1/2 (ERK1/2), a serine-threonine protein kinase involved in serine STAT3 phosphorylation; the livers of Pparβ/δ-null mice showed increased Tyr⁷⁰⁵- and Ser⁷²⁷-STAT3 as well as phospho-ERK1/2 levels. Furthermore, drug treatment prevented the IL-6-dependent reduction in phosphorylated AMP-activated protein kinase (AMPK), a kinase reported to inhibit STAT3 phosphorylation on Tyr⁷⁰⁵. In agreement with the recovery in phospho-AMPK levels observed following GW501516 treatment, this drug increased the AMP/ATP ratio and decreased the ATP/ADP ratio. Overall, our findings show that the PPARβ/δ activator GW501516 prevents IL-6-induced STAT3 activation by inhibiting ERK1/2 phosphorylation and preventing the reduction in phospho-AMPK levels. These effects of GW501516 may contribute to the prevention of cytokine-induced insulin resistance in hepatic cells.

Hierarchy of protein tyrosine kinases in interleukin-2 (IL-2) signaling: activation of syk depends on Jak3; however, neither Syk nor Lck is required for IL-2-mediated STAT activation.

PubMed

Zhou, Y J; Magnuson, K S; Cheng, T P; Gadina, M; Frucht, D M; Galon, J; Candotti, F; Geahlen, R L; Changelian, P S; O'Shea, J J

2000-06-01

Interleukin-2 (IL-2) activates several different families of tyrosine kinases, but precisely how these kinases interact is not completely understood. We therefore investigated the functional relationships among Jak3, Lck, and Syk in IL-2 signaling. We first observed that in the absence of Jak3, both Lck and Syk had the capacity to phosphorylate Stat3 and Stat5a. However, neither supported IL-2-induced STAT activation, nor did dominant negative alleles of these kinases inhibit. Moreover, pharmacological abrogation of Lck activity did not inhibit IL-2-mediated phosphorylation of Jak3 and Stat5a. Importantly, ligand-dependent Syk activation was dependent on the presence of catalytically active Jak3, whereas Lck activation was not. Interestingly, Syk functioned as a direct substrate of Jak1 but not Jak3. Additionally, Jak3 phosphorylated Jak1, whereas the reverse was not the case. Taken together, our data support a model in which Lck functions in parallel with Jak3, while Syk functions as a downstream element of Jaks in IL-2 signaling. Jak3 may regulate Syk catalytic activity indirectly via Jak1. However, IL-2-mediated Jak3/Stat activation is not dependent on Lck or Syk. While the essential roles of Jak1 and Jak3 in signaling by gammac-utilizing cytokines are clear, it will be important to dissect the exact contributions of Lck and Syk in mediating the effects of IL-2 and related cytokines.

IL-15 Activates the Jak3/STAT3 Signaling Pathway to Mediate Glucose Uptake in Skeletal Muscle Cells

PubMed Central

Krolopp, James E.; Thornton, Shantaé M.; Abbott, Marcia J.

2016-01-01

Myokines are specialized cytokines that are secreted from skeletal muscle (SKM) in response to metabolic stimuli, such as exercise. Interleukin-15 (IL-15) is a myokine with potential to reduce obesity and increase lean mass through induction of metabolic processes. It has been previously shown that IL-15 acts to increase glucose uptake in SKM cells. However, the downstream signals orchestrating the link between IL-15 signaling and glucose uptake have not been fully explored. Here we employed the mouse SKM C2C12 cell line to examine potential downstream targets of IL-15-induced alterations in glucose uptake. Following differentiation, C2C12 cells were treated overnight with 100 ng/ml of IL-15. Activation of factors associated with glucose metabolism (Akt and AMPK) and known downstream targets of IL-15 (Jak1, Jak3, STAT3, and STAT5) were assessed with IL-15 stimulation. IL-15 stimulated glucose uptake and GLUT4 translocation to the plasma membrane. IL-15 treatment had no effect on phospho-Akt, phospho-Akt substrates, phospho-AMPK, phospho-Jak1, or phospho-STAT5. However, with IL-15, phospho-Jak3 and phospho-STAT3 levels were increased along with increased interaction of Jak3 and STAT3. Additionally, IL-15 induced a translocation of phospho-STAT3 from the cytoplasm to the nucleus. We have evidence that a mediator of glucose uptake, HIF1α, expression was dependent on IL-15 induced STAT3 activation. Finally, upon inhibition of STAT3 the positive effects of IL-15 on glucose uptake and GLUT4 translocation were abolished. Taken together, we provide evidence for a novel signaling pathway for IL-15 acting through Jak3/STAT3 to regulate glucose metabolism. PMID:28066259

Upregulated STAT3 and RhoA signaling in colorectal cancer (CRC) regulate the invasion and migration of CRC cells.

PubMed

Zhang, G-Y; Yang, W-H; Chen, Z

2016-05-01

We aimed to reveal the expression and activation of signal transducers and activators of transcription 3 (STAT3) and RhoA/Rho-associated coiled-coil forming kinase 1 (ROCK1) signaling in CRC tissues, and to investigate the regulatory role of STAT3 and RhoA signaling in the invasion and migration of colorectal cancer cells. We examined the expression of STAT3, RhoA and ROCK1 in CRC tissues with real-time PCR and Western blotting methods. And then we examined the interaction between STAT3 and RhoA/ROCK1 signaling in CRC HT-29 cells with gain-of-function and loss-of-function strategies. In addition, we determined the regulation by STAT3 and RhoA/ROCK1 on the invasion and migration of CRC HT-29 cells. Our study demonstrated a significant upregulation of RhoA and ROCK1 expression and STAT3-Y705 phosphorylation in 32 CRC specimens, compared to the 17 normal CRC tissues. Further study demonstrated there was a coordination between STAT3 and RhoA/Rock signaling in the HT-29 cells. Moreover, STAT3 knockdown or RhoA knockdown significantly repressed the migration and invasion in HT-29 cells and vice versa. STAT3 and RhoA signaling regulate the invasion and migration of CRC cells, implying the orchestrated and oncogenic roles of STAT3 and RhoA/ROCK1 signaling in CRC.

Acetylation within the N- and C-Terminal Domains of Src Regulates Distinct Roles of STAT3-Mediated Tumorigenesis.

PubMed

Huang, Chao; Zhang, Zhe; Chen, Lihan; Lee, Hank W; Ayrapetov, Marina K; Zhao, Ting C; Hao, Yimei; Gao, Jinsong; Yang, Chunzhang; Mehta, Gautam U; Zhuang, Zhengping; Zhang, Xiaoren; Hu, Guohong; Chin, Y Eugene

2018-06-01

Posttranslational modifications of mammalian c-Src N-terminal and C-terminal domains regulate distinct functions. Myristoylation of G 2 controls its cell membrane association and phosphorylation of Y419/Y527 controls its activation or inactivation, respectively. We provide evidence that Src-cell membrane association-dissociation and catalytic activation-inactivation are both regulated by acetylation. In EGF-treated cells, CREB binding protein (CBP) acetylates an N-terminal lysine cluster (K5, K7, and K9) of c-Src to promote dissociation from the cell membrane. CBP also acetylates the C-terminal K401, K423, and K427 of c-Src to activate intrinsic kinase activity for STAT3 recruitment and activation. N-terminal domain phosphorylation (Y14, Y45, and Y68) of STAT3 by c-Src activates transcriptionally active dimers of STAT3. Moreover, acetyl-Src translocates into nuclei, where it forms the Src-STAT3 enhanceosome for gene regulation and cancer cell proliferation. Thus, c-Src acetylation in the N-terminal and C-terminal domains play distinct roles in Src activity and regulation. Significance: CBP-mediated acetylation of lysine clusters in both the N-terminal and C-terminal regions of c-Src provides additional levels of control over STAT3 transcriptional activity. Cancer Res; 78(11); 2825-38. ©2018 AACR . ©2018 American Association for Cancer Research.

[Knockdown of STAT3 inhibits proliferation and migration of HepG2 hepatoma cells induced by IFN1].

PubMed

Li, Xiaofang; Wang, Yuqi; Yan, Ben; Fang, Peipei; Ma, Chao; Xu, Ning; Fu, Xiaoyan; Liang, Shujuan

2018-02-01

Objective To prepare lentiviruses expressing shRNA sequences targeting human signal transducer and activator of transcription 3 (STAT3) and detect the effect of STAT3 knockdown on type I interferon (IFN1)-induced proliferation and migration in HepG2 cells. Methods Four STAT3-targeting shRNA sequences (shRNA1-shRNA4) and one control sequence (Ctrl shRNA) were selected and cloned respectively into pLKO.1-sp6-pgk-GFP to construct shRNA-expressing vectors. Along with backbone psPAX2 and pMD2.G vectors, they were separately transfected into HEK293T cells to prepare lentiviruses. HepG2 cells were infected with the lentiviruses. Cytoplastic STAT3 level was detected by Western blotting to screen effective shRNA sequence(s) targeting STAT3. Proliferation and migration of HepG2 cells were analyzed by CCK-8 assay and Transwell TM migration and scratching assay, respectively. To detect the effect of IFN1 on cell proliferation and migration of HepG2 cells, the cells were treated with 2000 U/mL IFNα2b for indicated time and the activation of IFN-triggered STAT1 signal transduction was assayed by Western blotting. Results Two most effective STAT3-targeting shRNA sequences shRNA1 and shRNA2 were selected, and the expression of both STAT3 shRNA significantly decreased proliferation and migration of HepG2 cells. When treated with IFNα2b, 2000 U/mL of IFN1 showed more competent in attenuating growth and migration of HepG2 cells. Our data further proved that knockdown of STAT3 increased the phosphorylation of STAT1, and IFNα2b further enhanced the activation of STAT1 signaling in HepG2 cells. Conclusion Knockdown of STAT3 inhibits cell migration and growth, and rescues IFN response through up-regulating STAT1 signal transduction in HepG2 hepatoma cells.

Oncostatin M suppresses metastasis of lung adenocarcinoma by inhibiting SLUG expression through coordination of STATs and PIASs signalings.

PubMed

Pan, Chih-Ming; Wang, Mong-Lien; Chiou, Shih-Hwa; Chen, Hsiao-Yun; Wu, Cheng-Wen

2016-09-13

Oncostatin M (OSM) is linked with multiple biological responses including growth and differentiation. Previous reports showed inhibitory effects of OSM in tumor progression while others showed promoting effects. The dual role of OSM in the development of various cancers is still unclear. We previously described OSM-mediated SLUG suppression, leading to repressed metastasis of lung adenocarcinoma (LAC) cells. However, the underlying mechanism remains elusive. Here, we showed that OSM suppresses SLUG express in LAC cells through a STAT1-dependent transcriptional inhibition. Knockdown of STAT1 reversed the OSM-suppressed SLUG expression and rescued the OSM-mediated inhibition of cell proliferation, migration, and invasion in vitro, as well as pulmonary metastasis in vivo. STAT1 suppressed SLUG transcription through binding to its promoter region in response to OSM. Furthermore, PIAS4, a co-repressor of STAT, and HDAC1 were able to bind to STAT1 on SLUG promoter region, resulting in reduced H3K9 acetylation and suppressed SLUG expression upon OSM treatment. In contrast, PIAS3 bound to activated STAT3, another effector of OSM, in response to OSM and blocked the binding of STAT3 to SLUG promoter region, preventing STAT3-dependent activation of SLUG transcription. Our findings suggested that OSM suppresses SLUG expression and tumor metastasis of LAC through inducing the inhibitory effect of the STAT1-dependent pathway and suppressing the activating effect of STAT3-dependent signaling. These results can serve as a scientific basis for the potential therapeutic intervention of OSM in cancer cells.

Discovery of Tyk2 inhibitors via the virtual site-directed fragment-based drug design.

PubMed

Jang, Woo Dae; Kim, Jun-Tae; Son, Hoon Young; Park, Seung Yeon; Cho, Young Sik; Koo, Tae-sung; Lee, Hyuk; Kang, Nam Sook

2015-09-15

In this study, we synthesized compound 12 with potent Tyk2 inhibitory activity from FBDD study and carried out a cell-based assay for Tyk2/STAT3 signaling activation upon IFNα5 stimulation. Compound 12 completely suppressed the IFNα5-mediated Tyk2/STAT3 signaling pathway as well as the basal levels of pSTAT3. Stimulation with IFNα/β leads to the tyrosine phosphorylation of the JAK1 and Tyk2 receptor-associated kinases with subsequent STATs activation, transmitting signals from the cell surface receptor to the nucleus. In conclusion, the potency of compound 12 to interrupt the signal transmission of Tyk2/STAT3 appeared to be equivalent or superior to that of the reference compound. Copyright © 2015 Elsevier Ltd. All rights reserved.

The STAT3 inhibitor pyrimethamine displays anti-cancer and immune stimulatory effects in murine models of breast cancer.

PubMed

Khan, Mohammad W; Saadalla, Abdulrahman; Ewida, Ahmed H; Al-Katranji, Khalid; Al-Saoudi, Ghadier; Giaccone, Zachary T; Gounari, Fotini; Zhang, Ming; Frank, David A; Khazaie, Khashayarsha

2018-01-01

The transcription factor signal activator and transducer or transcription (STAT3), which regulates genes controlling proliferation, survival, and invasion, is activated inappropriately in many human cancers, including breast cancer. Activation of STAT3 can lead to both malignant cellular behavior and suppression of immune cell function in the tumor microenvironment. Through a chemical-biology screen, pyrimethamine (PYR), an FDA approved anti-microbial drug, was identified as an inhibitor of STAT3 function at concentrations known to be achieved safely in humans. We report that PYR shows therapeutic activity in two independent mouse models of breast cancer, with both direct tumor inhibitory and immune stimulatory effects. PYR-inhibited STAT3 activity in TUBO and TM40D-MB metastatic breast cancer cells in vitro and inhibited tumor cell proliferation and invasion into Matrigel basement membrane matrix. In tumor-transplanted mice, PYR had both direct and indirect tumor inhibitory effects. Tumor-bearing mice treated with PYR showed reduced STAT3 activation in tumor cells, attenuated tumor growth, and reduced tumor-associated inflammation. In addition, expression of Lamp1 by tumor infiltrating CD8 + T cells was elevated, indicating enhanced release of cytotoxic granules. These findings suggest that PYR may have beneficial effects in the treatment of breast cancer.

Cooperation between STAT5 and phosphatidylinositol 3-kinase in the IL-3-dependent survival of a bone marrow derived cell line.

PubMed

Rosa Santos, S C; Dumon, S; Mayeux, P; Gisselbrecht, S; Gouilleux, F

2000-02-24

Cytokine-dependent activation of distinct signaling pathways is a common scheme thought to be required for the subsequent programmation into cell proliferation and survival. The PI 3-kinase/Akt, Ras/MAP kinase, Ras/NFIL3 and JAK/STAT pathways have been shown to participate in cytokine mediated suppression of apoptosis in various cell types. However the relative importance of these signaling pathways seems to depend on the cellular context. In several cases, individual inhibition of each pathway is not sufficient to completely abrogate cytokine mediated cell survival suggesting that cooperation between these pathways is required. Here we showed that individual inhibition of STAT5, PI 3-kinase or MEK activities did not or weakly affected the IL-3 dependent survival of the bone marrow derived Ba/F3 cell line. However, the simultaneous inhibition of STAT5 and PI 3-kinase activities but not that of STAT5 and MEK reduced the IL-3 dependent survival of Ba/F3. Analysis of the expression of the Bcl-2 members indicated that phosphorylation of Bad and Bcl-x expression which are respectively regulated by the PI 3-kinase/Akt pathway and STAT5 probably explain this cooperation. Furthermore, we showed by co-immunoprecipitation studies and pull down experiments with fusion proteins encoding the GST-SH2 domains of p85 that STAT5 in its phosphorylated form interacts with the p85 subunit of the PI 3-kinase. These results indicate that the activations of STAT5 and the PI 3-kinase by IL-3 in Ba/F3 cells are tightly connected and cooperate to mediate IL-3-dependent suppression of apoptosis by modulating Bad phosphorylation and Bcl-x expression.

STAT3 inhibitor WP1066 as a novel therapeutic agent for bCCI neuropathic pain rats.

PubMed

Xue, Zhao-Jing; Shen, Le; Wang, Zhi-Yao; Hui, Shang-Yi; Huang, Yu-Guang; Ma, Chao

2014-10-02

Activation of signal transducer and activator of transcription-3 (STAT3) is suggested to be critically involved in the development of chronic pain, but the complex regulation of STAT3-dependent pathway and the functional significance of inhibiting this pathway during the development of neuropathic pain remain elusive. To evaluate the contribution of the JAK2/STAT3 pathway to neuropathic pain and the potentiality of this pathway as a novel therapeutic target, we examined the effects of the STAT3 inhibitor WP1066 by intrathecal administration in a rat model of bilateral chronic constriction injury (bCCI). The pain behavior tests were performed before the surgery and on postoperative day 3, 7, 14 and 21. L4-L6 dorsal spinal cord were harvested at each time point. Both RT-PCR and Western blot were performed to evaluate the activation of JAK2/STAT3 pathway. To observe the influence of WP1066 on neuropathic pain and its molecular mechanism, WP1066 (10 μl, 10 mmol/L in DMSO) or the same capacity of DMSO as the control were applied through the intrathecal tube on the day before bCCI surgery, and on the postoperative day 3 and 5. Behavioral tests were performed to observe the therapeutic effect on mechanical, thermal and cold hyperalgesia. L4-L6 dorsal spinal cord was harvested on postoperative day fourteen, followed by RT-PCR and Western blot evaluation of the JAK2/STAT3 pathway activation. The mechanical, thermal and cold hyperalgesia of the bCCI rats were significantly decreased when compared with the Sham or the Naïve group at each postoperative time point (P<0.05). JAK2 mRNA and STAT3 mRNA were significantly increased in the bCCI rats, accompanied by SOCS3 mRNA with a similar tendency. Western blot analysis showed that JAK2 and phosphorylated STAT3 increased significantly since 3 days after bCCI. JAK2 peaked on postoperative day 14 while phosphorylated STAT3 peaked on postoperative day 7 and gradually decreased thereafter and SOCS3׳s peak level on postoperative day 3. When WP1066 were administered intrathecally, the pain behaviors of the bCCI rats were significantly improved (P<0.05). WP1066 also inhibited the mRNA level of JAK2, STAT3 and SOCS3 in bCCI rats significantly, together with the protein level of JAK2, phosphorylated STAT3 and SOCS3 on postoperative day 14 as well. Our results found that the JAK2/STAT3 pathway in the spinal cord dorsal horn was significantly activated in the bCCI neuropathic pain rats. WP1066, which inhibited the STAT3 pathway specifically, could partially alleviate the pain behavior of the bCCI rats. So it may serve as a novel therapeutic strategy against neuropathic pain. Copyright © 2014 Elsevier B.V. All rights reserved.

Alternative Splicing of STAT3 Is Affected by RNA Editing.

PubMed

Goldberg, Lior; Abutbul-Amitai, Mor; Paret, Gideon; Nevo-Caspi, Yael

2017-05-01

A-to-I RNA editing, carried out by adenosine deaminase acting on RNA (ADAR) enzymes, is an epigenetic phenomenon of posttranscriptional modifications on pre-mRNA. RNA editing in intronic sequences may influence alternative splicing of flanking exons. We have previously shown that conditions that induce editing result in elevated expression of signal transducer and activator of transcription 3 (STAT3), preferentially the alternatively-spliced STAT3β isoform. Mechanisms regulating alternative splicing of STAT3 have not been elucidated. STAT3 undergoes A-to-I RNA editing in an intron residing in proximity to the alternatively spliced exon. We hypothesized that RNA editing plays a role in regulating alternative splicing toward STAT3β. In this study we extend our observation connecting RNA editing to the preferential induction of STAT3β expression. We study the involvement of ADAR1 in STAT3 editing and reveal the connection between editing and alternative splicing of STAT3. Deferoaxamine treatment caused the induction in STAT3 RNA editing and STAT3β expression. Silencing ADAR1 caused a decrease in STAT3 editing and expression with a preferential decrease in STAT3β. Cells transfected with a mutated minigene showed preferential splicing toward the STAT3β transcript. Editing in the STAT3 intron is performed by ADAR1 and affects STAT3 alternative splicing. These results suggest that RNA editing is one of the molecular mechanisms regulating the expression of STAT3β.

Nicotine protects against DSS colitis through regulating microRNA-124 and STAT3.

PubMed

Qin, Zhen; Wan, Jing-Jing; Sun, Yang; Wu, Tingyu; Wang, Peng-Yuan; Du, Peng; Su, Ding-Feng; Yang, Yili; Liu, Xia

2017-02-01

Although it is generally believed that nicotine accounts for the beneficial effect of smoking on ulcerative colitis, the underlying mechanisms remain not well understood. Our previous finding that nicotine inhibits inflammatory responses through inducing miR-124 prompted us to ask whether the miRNA is involved in the protective action of nicotine against UC. Our present study found that miR-124 expression is upregulated in colon tissues from UC patients and DSS colitis mice. Nicotine treatment further augmented miR-124 expression in lymphocytes isolated from human ulcerative colonic mucosa and ulcerative colon tissues from DSS mice, both in infiltrated lymphocytes and epithelial cells. Moreover, knockdown of miR-124 significantly diminished the beneficial effect of nicotine on murine colitis and IL-6-treated Caco-2 colon epithelial cells. Further analysis indicated that nicotine inhibited STAT3 activation in vivo and in IL-6 treated Caco-2 cells and Jurkat human T lymphocytes, in which miR-124 knockdown led to increased activation of STAT3. Blocking STAT3 activity alone is beneficial for DSS colitis and also abolished nicotine's protective effect in this model. These data indicate that nicotine exerts its protective action in UC through inducing miR-124 and inhibiting STAT3, and suggest that the miR-124/STAT3 system is a potential target for the therapeutic intervention of UC. Nicotine upregulates miR-124 expression in ulcerative colon tissues and cells. MiR-124 is required for the protective role of nicotine in DSS colitis mice and epithelial cells. The protective effect of nicotine in murine DSS colitis depends on blocking STAT3 activation. MiR-124 mediates the inhibitory role of nicotine on STAT3/p-STAT3. Targeting miR-124 and STAT3 represents a novel approach for treating ulcerative colitis.

IL-4 and IL-13 employ discrete signaling pathways for target gene expression in alternatively activated monocytes/macrophages.

PubMed

Bhattacharjee, Ashish; Shukla, Meenakshi; Yakubenko, Valentin P; Mulya, Anny; Kundu, Suman; Cathcart, Martha K

2013-01-01

Monocytes/macrophages are innate immune cells that play a crucial role in the resolution of inflammation. In the presence of the Th2 cytokines interleukin-4 (IL-4) and interleukin-13 (IL-13), they display an anti-inflammatory profile and this activation pathway is known as alternative activation. In this study we compare and differentiate pathways mediated by IL-4 and IL-13 activation of human monocytes/macrophages. Here we report differential regulation of IL-4 and IL-13 signaling in monocytes/macrophages starting from IL-4/IL-13 cytokine receptors to Jak/Stat-mediated signaling pathways that ultimately control expression of several inflammatory genes. Our data demonstrate that although the receptor-associated tyrosine kinases Jak2 and Tyk2 are activated after the recruitment of IL-13 to its receptor (containing IL-4Rα and IL-13Rα1), IL-4 stimulates Jak1 activation. We further show that Jak2 is upstream of Stat3 activation and Tyk2 controls Stat1 and Stat6 activation in response to IL-13 stimulation. In contrast, Jak1 regulates Stat3 and Stat6 activation in IL-4-induced monocytes. Our results further reveal that although IL-13 utilizes both IL-4Rα/Jak2/Stat3 and IL-13Rα1/Tyk2/Stat1/Stat6 signaling pathways, IL-4 can use only the IL-4Rα/Jak1/Stat3/Stat6 cascade to regulate the expression of some critical inflammatory genes, including 15-lipoxygenase, monoamine oxidase A (MAO-A), and the scavenger receptor CD36. Moreover, we demonstrate here that IL-13 and IL-4 can uniquely affect the expression of particular genes such as dual-specificity phosphatase 1 and tissue inhibitor of metalloprotease-3 and do so through different Jaks. As evidence of differential regulation of gene function by IL-4 and IL-13, we further report that MAO-A-mediated reactive oxygen species generation is influenced by different Jaks. Collectively, these results have major implications for understanding the mechanism and function of alternatively activated monocytes/macrophages by IL-4 and IL-13 and add novel insights into the pathogenesis and potential treatment of various inflammatory diseases. Copyright © 2012 Elsevier Inc. All rights reserved.

Molecular integration of HoxB4 and STAT3 for self-renewal of hematopoietic stem cells: a model of molecular convergence for stemness.

PubMed

Hong, Sung-Hyun; Yang, Seung-Jip; Kim, Tae-Min; Shim, Jae-Seung; Lee, Ho-Sun; Lee, Ga-Young; Park, Bo-Bae; Nam, Suk Woo; Ryoo, Zae Young; Oh, Il-Hoan

2014-05-01

The upregulation of HoxB4 promotes self-renewal of hematopoietic stem cells (HSCs) without overriding the normal stem cell pool size. A similar enhancement of HSC self-renewal occurs when signal transducer and activator of transcription 3 (STAT3) is activated in HSCs. In this study, to gain insight into the functional organization of individual transcription factors (TFs) that have similar effects on HSCs, we investigated the molecular interplay between HoxB4 and STAT3 in the regulation of HSC self-renewal. We found that while STAT3-C or HoxB4 similarly enhanced the in vitro self-renewal and in vivo repopulating activities of HSCs, simultaneous transduction of both TFs did not have additive effects, indicating their functional redundancy in HSCs. In addition, activation of STAT3 did not cause changes in the expression levels of HoxB4. In contrast, the inhibition of STAT3 activity in HoxB4-overexpressing hematopoietic cells significantly abrogated the enhancing effects of HoxB4, and the upregulation of HoxB4 caused a ligand-independent Tyr-phosphorylation of STAT3. Microarray analysis revealed a significant overlap of the transcriptomes regulated by STAT3 and HoxB4 in undifferentiated hematopoietic cells. Moreover, a gene set enrichment analysis showed significant overlap in the candidate TFs that can recapitulate the transcriptional changes induced by HoxB4 or STAT3. Interestingly, among these common TFs were the pluripotency-related genes Oct-4 and Nanog. These results indicate that tissue-specific TFs regulating HSC self-renewal are functionally organized to play an equivalent role in transcription and provide insights into the functional convergence of multiple entries of TFs toward a conserved transcription program for the stem cell state. © 2014 AlphaMed Press.

Baicalein reduces lipopolysaccharide-induced inflammation via suppressing JAK/STATs activation and ROS production.

PubMed

Qi, Zhilin; Yin, Fei; Lu, Lina; Shen, Lei; Qi, Shimei; Lan, Lei; Luo, Lan; Yin, Zhimin

2013-09-01

To investigate the precise molecular mechanisms by which baicalein exerts beneficial biochemical activities in RAW264.7 macrophages treated with LPS. RAW264.7 cells were cultured in the absence or presence of baicalein together with or without LPS. iNOS and COX-2 expression were measured by western blot and RT-PCR analyses. TNF-α, IL-1β, and IL-6 were determined by using double-antibody sandwich ELISA. Phosphorylations of JAK1 and JAK2, and of STAT1 and STAT3 were detected by western blotting. Nuclear translocation of STAT1 and STAT3 was visualized by confocal microscopy. ROS production was detected by ROS assay. Baicalein significantly reduced the phosphorylation of STAT1 and STAT3 and the phosphorylation of JAK1 and JAK2, but without affecting MAPKs phosphorylation in LPS-stimulated RAW264.7 cells. Baicalein suppressed the nuclear translocation of STAT1 and STAT3 and inhibited production of iNOS upon LPS-stimulation, resulting in the inhibition of releases of NO and pro-inflammatory cytokines such as IL-1β, IL-6, and TNF-α, in a dose-dependent manner. In addition, we found that baicalein reduced the LPS-induced accumulation of ROS, confirming that baicalein serves as an antioxidant. Our results suggested that suppressing JAK/STATs activation and interfering with ROS production might contribute to the anti-inflammatory action of baicalein in macrophages.

Systemic Administration of a Cyclic Signal Transducer and Activator of Transcription 3 (STAT3) Decoy Oligonucleotide Inhibits Tumor Growth without Inducing Toxicological Effects

PubMed Central

Sen, Malabika; Paul, Kathleen; Freilino, Maria L; Li, Hua; Li, Changyou; Johnson, Daniel E; Wang, Lin; Eiseman, Julie; Grandis, Jennifer R

2014-01-01

Hyperactivation of signal transducer and activator of transcription 3 (STAT3) has been linked to tumorigenesis in most malignancies, including head and neck squamous cell carcinoma. Intravenous delivery of a chemically modified cyclic STAT3 decoy oligonucleotide with improved serum and thermal stability demonstrated antitumor efficacy in conjunction with downmodulation of STAT3 target gene expression such as cyclin D1 and Bcl-XL in a mouse model of head and neck squamous cell carcinoma. The purpose of the present study was to determine the toxicity and dose-dependent antitumor efficacy of the cyclic STAT3 decoy after multiple intravenous doses in Foxn1 nu mice in anticipation of clinical translation. The two doses (5 and 10 mg/kg) of cyclic STAT3 decoy demonstrated a significant decrease in tumor volume compared with the control groups (mutant cyclic STAT3 decoy or saline) in conjunction with downmodulation of STAT3 target gene expression. There was no dose-dependent effect of cyclic STAT3 decoy on tumor volume or STAT3 target gene expression. There were no significant changes in body weights between the groups during the dosing period, after the dosing interval or on the day of euthanasia. No hematology or clinical chemistry parameters suggested toxicity of the cyclic STAT3 decoy compared with saline control. No gross or histological pathological abnormalities were noted at necropsy in any of the animals. These findings suggest a lack of toxicity of intravenous administration of a cyclic STAT3 decoy oligonucleotide. In addition, comparable antitumor effects indicate a lack of dose response at the two dose levels investigated. PMID:24395569

Upregulation of survivin by leptin/STAT3 signaling in MCF-7 cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jiang Haiping; Tianjin Medical University Cancer Hospital, Tianjin; Yu Jinming

2008-03-28

Leptin and its receptors are overexpressed in breast cancer tissues and correlate with poor prognosis. Survivin, a member of the inhibitor of apoptosis protein (IAP) gene family, is generally upregulated in tumor tissues and prevents tumor cells from apoptosis. Here we showed that leptin upregulated survivin mRNA and protein expression in MCF-7 breast cancer cells. Meanwhile, leptin suppressed docetaxel-induced apoptosis by inhibiting caspase activity. Knockdown of signal transducer and activator transcription 3 (STAT3) expression by small interfering RNA (siRNA) blocked leptin-induced upregulation of survivin. TransAM ELISA showed that leptin increased nuclear translocation of active STAT3. In addition, chromatin immunoprecipitation (ChIP)more » assay detected an enhanced binding of STAT3 to survivin promoter in MCF-7 cells after treatment by leptin. Further studies showed that leptin enhanced the transcriptional activity of survivin promoter. Collectively, our findings identify leptin/STAT3 signaling as a novel pathway for survivin expression in breast cancer cells.« less

Expression of JAKs/STATs pathway molecules in rat model of rapid focal segmental glomerulosclerosis.

PubMed

Liang, Yaojun; Jin, Yu; Li, Yuning

2009-09-01

The objective of this study was to investigate the role of the Janus kinase-signal transducers and activators of transcription (JAKs/STATs) pathway in focal segmental glomerulosclerosis. Sixty specific pathogen-free male Wistar rats were randomly divided into two groups: a model group (MG) and a control group (CG). In the MG group, nephropathy was induced by unilateral nephrectomy and a single tail vein injection of adriamycin (5 mg/kg). Ten rats were sacrificed every 2 weeks in each group. The expressions of smooth muscle alpha actin (alpha-SMA), collagen (COL)-IV, STAT1, and STAT3 were examined using histochemical techniques, and Western blotting was used to examine the protein levels of STAT1, STAT3, phosphorylated (P)-STAT1, P-STAT3, and transforming growth factor beta1 (TGFbeta(1)). The expressions of JAK1, JAK2, STAT1, STAT3, suppressors of cytokine signaling (SOCS)1, SOCS3, protein inhibitors of activated STAT (PIAS)1, and PIAS3 were also measured by real-time quantitative reverse transcriptase-PCR. A steady and significant increase in the expressions of alpha-SMA, COL-IV and TGFbeta(1) were observed in MG rats over the whole experimental course. Increased STAT1 and P-STAT1 levels in MG rats were observed by week 6, whereas increased levels of STAT3 and P-STAT3 were noted by week 2. At the mRNA levels, JAK1, STAT1, and PIAS1 were significantly increased in MG rats in week 2, whereas JAK2 mRNA showed a significant decrease by weeks 2 and 4, followed by an significant increase in week 6. Significantly increased STAT3 levels were noted in week 2, followed by a steady and significant decrease in weeks 4 and 6. Significantly reduced levels of SOCS1, SOCS3, and PIAS3 mRNA were noted at all time points. We conclude that the JAKs/STATs signaling pathway may play an important role in the pathological process of rapid focal segmental glomerulosclerosis in the rat model.

Interleukin-6/Stat3 signaling has an essential role in the host antimicrobial response to urinary tract infection.

PubMed

Ching, Christina B; Gupta, Sudipti; Li, Birong; Cortado, Hanna; Mayne, Nicholas; Jackson, Ashley R; McHugh, Kirk M; Becknell, Brian

2018-06-01

The signaling networks regulating antimicrobial activity during urinary tract infection (UTI) are incompletely understood. Interleukin-6 (IL-6) levels increase with UTI severity, but the specific contributions of IL-6 to host immunity against bacterial uropathogens are unknown. To clarify this we tested whether IL-6 activates the Stat3 transcription factor, to drive a program of antimicrobial peptide gene expression in infected urothelium during UTI. Transurethral inoculation of uropathogenic Escherichia coli led to IL-6 secretion, urothelial Stat3 phosphorylation, and activation of antimicrobial peptide transcription, in a Toll-like receptor 4-dependent manner in a murine model of cystitis. Recombinant IL-6 elicited Stat3 phosphorylation in primary urothelial cells in vitro, and systemic IL-6 administration promoted urothelial Stat3 phosphorylation and antimicrobial peptide expression in vivo. IL-6 deficiency led to decreased urothelial Stat3 phosphorylation and antimicrobial peptide mRNA expression following UTI, a finding mirrored by conditional Stat3 deletion. Deficiency in IL-6 or Stat3 was associated with increased formation of intracellular bacterial communities, and exogenous IL-6 reversed this phenotype in IL-6 knockout mice. Moreover, chronic IL-6 depletion led to increased renal bacterial burden and severe pyelonephritis in C3H/HeOuJ mice. Thus, IL-6/Stat3 signaling drives a transcriptional program of antimicrobial gene expression in infected urothelium, with key roles in limiting epithelial invasion and ascending infection. Copyright © 2018 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.

Molecular mechanisms of mucocutaneous immunity against Candida and Staphylococci

PubMed Central

Maródi, László; Cypowyj, Sophie; Tóth, Beáta; Chernyshova, Liudmyla; Puel, Anne; Casanova, Jean-Laurent

2013-01-01

Signal transducer and activator of transcription (STAT) proteins are key components of the innate and adaptive immune responses to pathogenic microorganisms. Recent research on primary immunodeficiency disorders and the identification of patients carrying germline mutations in STAT1, STAT3, and STAT5B have highlighted the role of human STATs in host defense against various viruses, bacteria, and fungi. Mutations in STAT1 and STAT3 may disrupt various cytokine pathways that control mucocutaneous immunity against Candida species, especially Candida albicans, and Staphylococci, especially Staphylococcus aureus. Here, we consider inborn errors of immunity arising from mutations in either STAT1 or STAT3 that affect mucocutaneous immunity to Candida and Staphylococci. PMID:23040277

The role of signal transducer and activator of transcription 3 in Rift Valley fever virus infection.

PubMed

Pinkham, Chelsea; An, Soyeon; Lundberg, Lindsay; Bansal, Neha; Benedict, Ashwini; Narayanan, Aarthi; Kehn-Hall, Kylene

2016-09-01

Rift Valley fever (RVF) is a zoonotic disease that can cause severe illness in humans and livestock, triggering spontaneous abortion in almost 100% of pregnant ruminants. In this study, we demonstrate that signal transducer and activator of transcription 3 (STAT3) is phosphorylated on its conserved tyrosine residue (Y705) following RVFV infection. This phosphorylation was dependent on a major virulence factor, the viral nonstructural protein NSs. Loss of STAT3 had little effect on viral replication, but rather resulted in cells being more susceptible to RVFV-induced cell death. Phosphorylated STAT3 translocated to the nucleus, coinciding with inhibition of fos, jun, and nr4a2 gene expression, and the presence of STAT3 and NSs at the nr4a2 promoter. NSs was found predominantly in the cytoplasm of STAT3 null cells, indicating that STAT3 influences NSs nuclear localization. Collectively, these data demonstrate that STAT3 functions in a pro-survival capacity through modulation of NSs localization. Copyright © 2016 Elsevier Inc. All rights reserved.

Zinc improves learning and memory abilities of fetal growth restriction rats and promotes trophoblast cell invasion and migration via enhancing STAT3-MMP-2/9 axis activity.

PubMed

Zong, Lu; Wei, Xiaohua; Gou, Wenli; Huang, Pu; Lv, Ye

2017-12-29

Fetal growth restriction (FGR) is a well-known risk factor for cognitive dysfunction, especially for learning and memory abilities. However, knowledge about prevention and treatment methods of learning and memory abilities of fetal are limit. Here, Morris water maze and passive avoidance tests showed zinc supplementation could protect the impairment of the learning and memory abilities caused by FGR. As accumulating evidence suggested that insufficiency of placental trophoblast cell invasion was closely related to FGR fetal neurodevelopmental dysplasia, we further explored the relationship between zinc supplementation during pregnancy and placental trophoblast. Microarray identified 346 differently expressed genes in placental tissues with and without zinc supplementation, and GO and KEGG analyses showed these differently expressed genes were highly enriched in cell invasion and migration and STAT3 pathway. Protein-protein interaction(PPI) analysis found that STAT3 interacted with matrix metalloproteinase-2/9 (MMP-2/9). In vivo , western blot results authenticated that the expression levels of phospho-STAT3, STAT3, MMP-2 and MMP-9 were up-regulated in placental tissues after zinc treatment. To validate whether zinc could promotes trophoblast cell invasion and migration via enhancing STAT3-MMP-2/9 activity. In vitro , Transwell assay was performed, and we observed that abilities of invasion and migration were obviously increased in zinc treated trophoblast cells. And phospho-STAT3, STAT3, MMP-2 and MMP-9 expression levels were correspondingly increased in zinc treated trophoblast cells, which were dose-dependent. Moreover, gain-of-function and loss-of-function of STAT3 confirmed that zinc promotes cell invasion and migration via regulating STAT3 mediated up-regulation of MMP-2/9 activity. We propose that activation of MMP-2/9 mediated by STAT3 may contribute to invasion and migration of trophoblast cells, which improved neurodevelopmental impairment of FGR rats probably via contributing to placental development. Our findings are the first to show a possible mechanism of reversing neurodevelopmental impairment of FGR rats by zinc supplementation, holding promise for the development of novel therapeutic modalities for learning and memory abilities impairment caused by FGR.

Cutting edge: A transcriptional repressor and corepressor induced by the STAT3-regulated anti-inflammatory signaling pathway.

PubMed

El Kasmi, Karim C; Smith, Amber M; Williams, Lynn; Neale, Geoffrey; Panopoulos, Athanasia D; Panopolous, Athanasia; Watowich, Stephanie S; Häcker, Hans; Foxwell, Brian M J; Murray, Peter J

2007-12-01

IL-10 regulates anti-inflammatory signaling via the activation of STAT3, which in turn controls the induction of a gene expression program whose products execute inhibitory effects on proinflammatory mediator production. In this study we show that IL-10 induces the expression of an ETS family transcriptional repressor, ETV3, and a helicase family corepressor, Strawberry notch homologue 2 (SBNO2), in mouse and human macrophages. IL-10-mediated induction of ETV3 and SBNO2 expression was dependent upon both STAT3 and a stimulus through the TLR pathway. We also observed that ETV3 expression was strongly induced by the STAT3 pathway regulated by IL-10 but not by STAT3 signaling activated by IL-6, which cannot activate the anti-inflammatory signaling pathway. ETV3 and SBNO2 repressed NF-kappaB- but not IFN regulatory factor 7 (IRF7)-activated transcriptional reporters. Collectively our data suggest that ETV3 and SBNO2 are components of the pathways that contribute to the downstream anti-inflammatory effects of IL-10.

The transcriptional co-activator p/CIP (NCoA-3) is up-regulated by STAT6 and serves as a positive regulator of transcriptional activation by STAT6.

PubMed

Arimura, Akinori; vn Peer, Maartje; Schröder, Andreas J; Rothman, Paul B

2004-07-23

Transcriptional activation by signal transducer and activator of transcription 6 (STAT6) has been shown to require the direct interaction not only with co-activators such as p300 and cAMP-responsive element-binding protein-binding protein (CBP) but also with nuclear co-activator 1, a member of the p160/steroid receptor co-activator family. Among the p160/steroid receptor co-activators, only p/CIP (nuclear co-activator 3) has been shown to be up-regulated by interleukin (IL)-4 in B cells through a STAT-6-dependent mechanism using Gene-Chip analysis. In this study, we have investigated the function of p/CIP in the transcriptional activation by STAT6. We found that p/CIP indirectly interacted with STAT6 via p300, and overexpression of the CBP-interacting domain of p/CIP (p/CIP(947-1084)) prevented the interaction of p/CIP with STAT6 by blocking the binding of p/CIP to p300. Whereas expression of p/CIP(947-1084) resulted in a marked reduction of STAT6-mediated transactivation, overexpression of wild type p/CIP resulted in significant enhancement of it. In addition, p/CIP(947-1084) markedly reduced CD23 expression on B cells stimulated with IL-4, whereas overexpression of wild type p/CIP enhanced it. Chromatin immunoprecipitations demonstrate that IL-4 increases the interaction of p/CIP with the murine immunoglobulin heavy chain germ line epsilon promoter in B cells. These results suggest that p/CIP positively regulates STAT6 transcriptional activation through formation of a STAT6, p300/CBP, and p/CIP complex.

Gastric Cancer Cell Proliferation and Survival Is Enabled by a Cyclophilin B/STAT3/miR-520d-5p Signaling Feedback Loop.

PubMed

Li, Ting; Guo, Hanqing; Zhao, Xiaodi; Jin, Jiang; Zhang, Lifeng; Li, Hong; Lu, Yuanyuan; Nie, Yongzhan; Wu, Kaichun; Shi, Yongquan; Fan, Daiming

2017-03-01

Molecular links between inflammation and cancer remain obscure despite their great pathogenic significance. The JAK2/STAT3 pathway activated by IL6 and other proinflammatory cytokines has garnered attention as a pivotal link in cancer pathogenesis, but the basis for its activation in cancer cells is not understood. Here we report that an IL6-triggered feedback loop involving STAT3-mediated suppression of miR-520d-5p and upregulation of its downstream target cyclophilin B (CypB) regulate the growth and survival of gastric cancer cells. In clinical specimens of gastric cancer, we documented increased expression of CypB and activation of STAT3. Mechanistic investigations identified miR-520d-5p as a regulator of CypB mRNA levels. This signaling axis regulated gastric cancer growth by modulating phosphorylation of STAT3. Furthermore, miR-520d-5p was identified as a direct STAT3 target and IL6-mediated inhibition of miR-520d-5p relied upon STAT3 activity. Our findings define a positive feedback loop that drives gastric carcinogenesis as influenced by H. pylori infections that involve proinflammatory IL6 stimulation. Cancer Res; 77(5); 1227-40. ©2016 AACR . ©2016 American Association for Cancer Research.

Platelet-derived growth factor receptors differentially inform intertumoral and intratumoral heterogeneity

PubMed Central

Kim, Youngmi; Kim, Eunhee; Wu, Qiulian; Guryanova, Olga; Hitomi, Masahiro; Lathia, Justin D.; Serwanski, David; Sloan, Andrew E.; Weil, Robert J.; Lee, Jeongwu; Nishiyama, Akiko; Bao, Shideng; Hjelmeland, Anita B.; Rich, Jeremy N.

2012-01-01

Growth factor-mediated proliferation and self-renewal maintain tissue-specific stem cells and are frequently dysregulated in cancers. Platelet-derived growth factor (PDGF) ligands and receptors (PDGFRs) are commonly overexpressed in gliomas and initiate tumors, as proven in genetically engineered models. While PDGFRα alterations inform intertumoral heterogeneity toward a proneural glioblastoma (GBM) subtype, we interrogated the role of PDGFRs in intratumoral GBM heterogeneity. We found that PDGFRα is expressed only in a subset of GBMs, while PDGFRβ is more commonly expressed in tumors but is preferentially expressed by self-renewing tumorigenic GBM stem cells (GSCs). Genetic or pharmacological targeting of PDGFRβ (but not PDGFRα) attenuated GSC self-renewal, survival, tumor growth, and invasion. PDGFRβ inhibition decreased activation of the cancer stem cell signaling node STAT3, while constitutively active STAT3 rescued the loss of GSC self-renewal caused by PDGFRβ targeting. In silico survival analysis demonstrated that PDGFRB informed poor prognosis, while PDGFRA was a positive prognostic factor. Our results may explain mixed clinical responses of anti-PDGFR-based approaches and suggest the need for integration of models of cancer as an organ system into development of cancer therapies. PMID:22661233

CYLD Enhances Severe Listeriosis by Impairing IL-6/STAT3-Dependent Fibrin Production

PubMed Central

Nishanth, Gopala; Deckert, Martina; Wex, Katharina; Massoumi, Ramin; Schweitzer, Katrin; Naumann, Michael; Schlüter, Dirk

2013-01-01

The facultative intracellular bacterium Listeria monocytogenes (Lm) may cause severe infection in humans and livestock. Control of acute listeriosis is primarily dependent on innate immune responses, which are strongly regulated by NF-κB, and tissue protective factors including fibrin. However, molecular pathways connecting NF-κB and fibrin production are poorly described. Here, we investigated whether the deubiquitinating enzyme CYLD, which is an inhibitor of NF-κB-dependent immune responses, regulated these protective host responses in murine listeriosis. Upon high dose systemic infection, all C57BL/6 Cyld−/− mice survived, whereas 100% of wildtype mice succumbed due to severe liver pathology with impaired pathogen control and hemorrhage within 6 days. Upon in vitro infection with Lm, CYLD reduced NF-κB-dependent production of reactive oxygen species, interleukin (IL)-6 secretion, and control of bacteria in macrophages. Furthermore, Western blot analyses showed that CYLD impaired STAT3-dependent fibrin production in cultivated hepatocytes. Immunoprecipitation experiments revealed that CYLD interacted with STAT3 in the cytoplasm and strongly reduced K63-ubiquitination of STAT3 in IL-6 stimulated hepatocytes. In addition, CYLD diminished IL-6-induced STAT3 activity by reducing nuclear accumulation of phosphorylated STAT3. In vivo, CYLD also reduced hepatic STAT3 K63-ubiquitination and activation, NF-κB activation, IL-6 and NOX2 mRNA production as well as fibrin production in murine listeriosis. In vivo neutralization of IL-6 by anti-IL-6 antibody, STAT3 by siRNA, and fibrin by warfarin treatment, respectively, demonstrated that IL-6-induced, STAT3-mediated fibrin production significantly contributed to protection in Cyld−/− mice. In addition, in vivo Cyld siRNA treatment increased STAT3 phosphorylation, fibrin production, pathogen control and survival of Lm-infected WT mice illustrating that therapeutic inhibition of CYLD augments the protective NF-κB/IL-6/STAT3 pathway and fibrin production. PMID:23825949

CYLD enhances severe listeriosis by impairing IL-6/STAT3-dependent fibrin production.

PubMed

Nishanth, Gopala; Deckert, Martina; Wex, Katharina; Massoumi, Ramin; Schweitzer, Katrin; Naumann, Michael; Schlüter, Dirk

2013-01-01

The facultative intracellular bacterium Listeria monocytogenes (Lm) may cause severe infection in humans and livestock. Control of acute listeriosis is primarily dependent on innate immune responses, which are strongly regulated by NF-κB, and tissue protective factors including fibrin. However, molecular pathways connecting NF-κB and fibrin production are poorly described. Here, we investigated whether the deubiquitinating enzyme CYLD, which is an inhibitor of NF-κB-dependent immune responses, regulated these protective host responses in murine listeriosis. Upon high dose systemic infection, all C57BL/6 Cyld(-/-) mice survived, whereas 100% of wildtype mice succumbed due to severe liver pathology with impaired pathogen control and hemorrhage within 6 days. Upon in vitro infection with Lm, CYLD reduced NF-κB-dependent production of reactive oxygen species, interleukin (IL)-6 secretion, and control of bacteria in macrophages. Furthermore, Western blot analyses showed that CYLD impaired STAT3-dependent fibrin production in cultivated hepatocytes. Immunoprecipitation experiments revealed that CYLD interacted with STAT3 in the cytoplasm and strongly reduced K63-ubiquitination of STAT3 in IL-6 stimulated hepatocytes. In addition, CYLD diminished IL-6-induced STAT3 activity by reducing nuclear accumulation of phosphorylated STAT3. In vivo, CYLD also reduced hepatic STAT3 K63-ubiquitination and activation, NF-κB activation, IL-6 and NOX2 mRNA production as well as fibrin production in murine listeriosis. In vivo neutralization of IL-6 by anti-IL-6 antibody, STAT3 by siRNA, and fibrin by warfarin treatment, respectively, demonstrated that IL-6-induced, STAT3-mediated fibrin production significantly contributed to protection in Cyld(-/-) mice. In addition, in vivo Cyld siRNA treatment increased STAT3 phosphorylation, fibrin production, pathogen control and survival of Lm-infected WT mice illustrating that therapeutic inhibition of CYLD augments the protective NF-κB/IL-6/STAT3 pathway and fibrin production.

Natural and synthetic STAT3 inhibitors reduce hepcidin expression in differentiated mouse hepatocytes expressing the active phosphorylated STAT3 form.

PubMed

Fatih, Nadia; Camberlein, Emilie; Island, Marie Laure; Corlu, Anne; Abgueguen, Emmanuelle; Détivaud, Lénaïck; Leroyer, Patricia; Brissot, Pierre; Loréal, Olivier

2010-05-01

During the inflammatory process, hepcidin overexpression favours the development of anaemia of chronic diseases which represents the second most common form of anaemia worldwide. The identification of therapeutic agents decreasing hepcidin expression is therefore an important goal. The aim of this study was to target the STAT3 signalling involved in the development of increased hepcidin expression related to chronic inflammation. In a co-culture model associating mouse hepatocytes and rat liver epithelial cells, the mRNA levels of hepcidin1, albumin, aldolase B, Cyp3a4, Stat3, Smad4 and iron regulatory genes were measured by real-time PCR. STAT3 and phosphorylated SMAD1/5/8 proteins were analysed by Western blot. At variance of hepatocyte pure culture, co-culture provided high levels of hepcidin1 mRNA, reaching 400% of the freshly isolated hepatocyte values after 6 days of culture. Hepcidin expression was associated with the maintenance of hepatocyte phenotype, STAT3 phosphorylation and functional BMP/SMAD pathway. Stat3 siRNAs inhibited the hepcidin1 mRNA expression. STAT3 inhibitors, including curcumin, AG490 and a peptide (PpYLKTK), reduced hepcidin1 mRNA expression even when cells were additionally exposed to IL-6. Hepcidin1 mRNA was expressed at high levels by hepatocytes in the co-culture model, and STAT3 pathway activation was controlled through STAT3 inhibitors. Such inhibitors could be useful to prevent anaemia related to hepcidin overexpression during chronic inflammation.

pSTAT3 Levels Have Divergent Expression Patterns and Associations with Survival in Squamous Cell Carcinoma and Adenocarcinoma of the Oesophagus.

PubMed

O' Sullivan, Katie E; Michielsen, Adriana J; O' Regan, Esther; Cathcart, Mary C; Moore, Gillian; Breen, Eamon; Segurado, Ricardo; Reynolds, John V; Lysaght, Joanne; O' Sullivan, Jacintha

2018-06-10

Signal transducers and activator of transcription (STAT)-3 is activated in cancers, where it promotes growth, inflammation, angiogenesis, and inhibits apoptosis. Tissue microarrays were generated using tissues from 154 patients, with oesophageal adenocarcinoma (OAC) ( n = 116) or squamous cell carcinoma (SCC) ( n = 38) tumours. The tissues were stained for pSTAT3 and IL-6R using immunohistochemistry. The OE33 (OAC) and OE21 (SCC) cell lines were treated with the STAT3 inhibitor, STATTIC. The Univariate cox regression analysis revealed that a positive pSTAT3 in SCC was adversely associated with survival (Hazard ratio (HR) 6.382, 95% CI 1.266⁻32.184), while a protective effect was demonstrated with the higher pSTAT3 levels in OAC epithelium (HR 0.74, 95% CI 0.574⁻0.953). The IL-6R intensity levels were higher in the SCC tumours compared with the OAC tumours for the core and leading edge tumour tissue. The pSTAT3 levels correlated positively with the IL-6R levels in both the OAC and SCC. The treatment of OE21 and OE33 cells with the STAT3 inhibitor STATTIC in vitro resulted in decreased survival, proliferation, migration, and increased apoptosis. The pSTAT3 expression was associated with adverse survival in SCC, but not in the OAC patients. The inhibition of STAT3 in both of the tumour subtypes resulted in alterations in the survival, proliferation, migration, and apoptosis, suggesting a potential role for therapeutically targeting STAT3.

The role of the inhibitors of interleukin-6 signal transduction SHP2 and SOCS3 for desensitization of interleukin-6 signalling.

PubMed Central

Fischer, Patrick; Lehmann, Ute; Sobota, Radoslaw M; Schmitz, Jochen; Niemand, Claudia; Linnemann, Sonja; Haan, Serge; Behrmann, Iris; Yoshimura, Akihiko; Johnston, James A; Müller-Newen, Gerhard; Heinrich, Peter C; Schaper, Fred

2004-01-01

The immediate early response of cells treated with IL-6 (interleukin-6) is the activation of the signal transducer and activator of transcription (STAT)3. The Src homology domain 2 (SH2)-containing protein tyrosine phosphatase SHP2 and the feedback inhibitor SOCS3 (suppressor of cytokine signalling) are potent inhibitors of IL-6 signal transduction. Impaired function of SOCS3 or SHP2 leads to enhanced and prolonged IL-6 signalling. The inhibitory function of both proteins depends on their recruitment to the tyrosine motif 759 within glycoprotein gp130. In contrast to inactivation, desensitization of signal transduction is regarded as impaired responsiveness due to prestimulation. Usually, after activation the sensing receptor becomes inactivated by modifications such as phosphorylation, internalization or degradation. We designed an experimental approach which allows discrimination between desensitization and inactivation of IL-6 signal transduction. We observed that pre-stimulation with IL-6 renders cells less sensitive to further stimulation with IL-6. After several hours, the cells become sensitive again. We show that not only signal transduction through previously activated receptors is affected by desensitization but signalling through receptors which were not targeted by the first stimulation was also attenuated ( trans -desensitization). Interestingly, in contrast to inhibition, desensitization does not depend on the presence of functional SHP2. Furthermore, cells lacking SOCS3 show constitutive STAT3 activation which is not affected by pre-stimulation with IL-6. All these observations suggest that desensitization and inhibition of signalling are mechanistically distinct. PMID:14611646

A soluble receptor for advanced glycation end-products inhibits myocardial apoptosis induced by ischemia/reperfusion via the JAK2/STAT3 pathway.

PubMed

Jiang, Xue; Guo, Cai-xia; Zeng, Xiang-jun; Li, Hui-hua; Chen, Bu-xing; Du, Feng-he

2015-08-01

sRAGE can protect cardiomyocytes from apoptosis induced by ischemia/reperfusion (I/R). However, the signaling mechanisms in cardioprotection by sRAGE are currently unknown. We investigated the cardioprotective effect and potential molecular mechanisms of sRAGE inhibition on apoptosis in the mouse myocardial I/R as an in vivo model and neonatal rat cardiomyocyte subjected to ischemic buffer as an in vitro model. Cardiac function and myocardial infarct size following by I/R were evaluated with echocardiography and Evans blue/2,3,5-triphenyltetrazolium chloride. Apoptosis was detected by TUNEL staining and caspase-3 activity. Expression of the apoptosis-related proteins p53, Bax, Bcl-2, JAK2/p-JAK2, STAT3/p-STAT3, AKT/p-AKT, ERK/p-ERK, STAT5A/p-STAT5A and STAT6/p-STAT6 were detected by western blot analysis in the presence and absence of the JAK2 inhibitor AG 490. sRAGE (100 µg/day) improved the heart function in mice with I/R: the left ventricular ejection fraction and fractional shortening were increased by 42 and 57%, respectively; the infarct size was decreased by 52%, the TUNEL-positive myocytes by 66%, and activity of caspase-3 by 24%, the protein expression of p53 and ratio of Bax to Bcl-2 by 29 and 88%, respectively; protein expression of the p-JAK2, p-STAT3 and p-AKT were increased by 92, 280 and 31%, respectively. sRAGE have no effect on protein expression of p-ERK1/2, p-STAT5A and p-STAT6 following by I/R. sRAGE (900 nmol/L) exhibited anti-apoptotic effects in cardiomyocytes by decreasing TUNEL-positive myocytes by 67% and caspase-3 activity by 20%, p53 protein level and the Bax/Bcl-2 ratio by 58 and 86%, respectively; increasing protein expression of the p-JAK2 and p-STAT3 by 26 and 156%, respectively, p-AKT protein level by 33%. The anti-apoptotic effects of sRAGE following I/R were blocked by JAK2 inhibitor AG 490. The effect of sRAGE reduction on TUNEL-positive myocytes and caspase-3 activity were abolished by PI3K inhibitor LY294002, but not ERK 1/2 inhibitor PD98059. These results suggest that sRAGE protects cardiomyocytes from apoptosis induced by I/R in vitro and in vivo by activating the JAK2/STAT3 signaling pathway.

Genetic Demonstration of a Role for Stathmin in Adult Hippocampal Neurogenesis, Spinogenesis, and NMDA Receptor-Dependent Memory

PubMed Central

Martel, Guillaume; Uchida, Shusaku; Hevi, Charles; Chévere-Torres, Itzamarie; Fuentes, Ileana; Park, Young Jin; Hafeez, Hannah; Yamagata, Hirotaka; Watanabe, Yoshifumi

2016-01-01

Neurogenesis and memory formation are essential features of the dentate gyrus (DG) area of the hippocampus, but to what extent the mechanisms responsible for both processes overlap remains poorly understood. Stathmin protein, whose tubulin-binding and microtubule-destabilizing activity is negatively regulated by its phosphorylation, is prominently expressed in the DG. We show here that stathmin is involved in neurogenesis, spinogenesis, and memory formation in the DG. tTA/tetO-regulated bitransgenic mice, expressing the unphosphorylatable constitutively active Stathmin4A mutant (Stat4A), exhibit impaired adult hippocampal neurogenesis and reduced spine density in the DG granule neurons. Although Stat4A mice display deficient NMDA receptor-dependent memory in contextual discrimination learning, which is dependent on hippocampal neurogenesis, their NMDA receptor-independent memory is normal. Confirming NMDA receptor involvement in the memory deficits, Stat4A mutant mice have a decrease in the level of synaptic NMDA receptors and a reduction in learning-dependent CREB-mediated gene transcription. The deficits in neurogenesis, spinogenesis, and memory in Stat4A mice are not present in mice in which tTA/tetO-dependent transgene transcription is blocked by doxycycline through their life. The memory deficits are also rescued within 3 d by intrahippocampal infusion of doxycycline, further indicating a role for stathmin expressed in the DG in contextual memory. Our findings therefore point to stathmin and microtubules as a mechanistic link between neurogenesis, spinogenesis, and NMDA receptor-dependent memory formation in the DG. SIGNIFICANCE STATEMENT In the present study, we aimed to clarify the role of stathmin in neuronal and behavioral functions. We characterized the neurogenic, behavioral, and molecular consequences of the gain-of-function stathmin mutation using a bitransgenic mouse expressing a constitutively active form of stathmin. We found that stathmin plays an important role in adult hippocampal neurogenesis and spinogenesis. In addition, stathmin mutation led to impaired NMDA receptor-dependent and neurogenesis-associated memory and did not affect NMDA receptor-independent memory. Moreover, biochemical analysis suggested that stathmin regulates the synaptic transport of NMDA receptors, which in turn influence CREB-mediated gene transcription machinery. Overall, these data suggest that stathmin is an important molecule for neurogenesis, spinogenesis, and NMDA receptor-dependent learning and memory. PMID:26818507

Silibinin-mediated metabolic reprogramming attenuates pancreatic cancer-induced cachexia and tumor growth.

PubMed

Shukla, Surendra K; Dasgupta, Aneesha; Mehla, Kamiya; Gunda, Venugopal; Vernucci, Enza; Souchek, Joshua; Goode, Gennifer; King, Ryan; Mishra, Anusha; Rai, Ibha; Nagarajan, Sangeetha; Chaika, Nina V; Yu, Fang; Singh, Pankaj K

2015-12-01

Pancreatic ductal adenocarcinoma (PDAC) is the fourth leading cause of cancer-related deaths in the US. Cancer-associated cachexia is present in up to 80% of PDAC patients and is associated with aggressive disease and poor prognosis. In the present studies we evaluated an anti-cancer natural product silibinin for its effectiveness in targeting pancreatic cancer aggressiveness and the cachectic properties of pancreatic cancer cells and tumors. Our results demonstrate that silibinin inhibits pancreatic cancer cell growth in a dose-dependent manner and reduces glycolytic activity of cancer cells. Our LC-MS/MS based metabolomics data demonstrates that silibinin treatment induces global metabolic reprogramming in pancreatic cancer cells. Silibinin treatment diminishes c-MYC expression, a key regulator of cancer metabolism. Furthermore, we observed reduced STAT3 signaling in silibinin-treated cancer cells. Overexpression of constitutively active STAT3 was sufficient to substantially revert the silibinin-induced downregulation of c-MYC and the metabolic phenotype. Our in vivo investigations demonstrate that silibinin reduces tumor growth and proliferation in an orthotopic mouse model of pancreatic cancer and prevents the loss of body weight and muscle. It also improves physical activity including grip strength and latency to fall in tumor-bearing mice. In conclusion, silibinin-induced metabolic reprogramming diminishes cell growth and cachectic properties of pancreatic cancer cells and animal models.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Song, Li; Guo, Linlin

Polychlorinated biphenyls (PCBs) are classic persistent organic pollutants (POPs). Many studies have found a positive association between the progression of hepatocellular carcinoma (HCC) and PCBs exposure. However, the influence of PCBs on epithelial-mesenchymal transition (EMT) of HCC remains to be unclear. In this study, we explored the effect of PCB126 on EMT in HCC cells and its underlying mechanisms. The data showed that PCB126, exposing both Bel-7402 and SMMC-7721 cells for 48 h, promoted EMT that was demonstrated by E-cadherin repression, up-regulation of N-cadherin and vimentin, and morphological alteration. We found that signal transducer and activator of transcription 3 (STAT3)/Snail1more » signaling was activated after PCB126 exposure, and the addition of STAT3 inhibitor WP1066 blocked PCB126-induced down-regulation of E-cadherin as well as up-regulation of N-cadherin and vimentin. Moreover, PCB126 exposure increased pyruvate kinase M2 (PKM2) expression and its nuclear translocation, whereas treatment with PKM2 shRNA suppressed the activation of STAT3/Snail1 signaling and the alternation of EMT-related molecules (E-cadherin, N-cadherin and vimentin). Furthermore, this study indicated estrogen receptor (ER) and aryl hydrocarbon receptor (AhR) were involved in PCB126-induced effects on PKM2, STAT3/Snail1 signaling and EMT by according treatment using ER inhibitor ICI and AhR shRNA. Notably, PCB126-increased reactive oxygen species (ROS) production via AhR is associated with activation of PKM2/STAT3/Snail1 cascades and contributes to EMT. Taken together, these results indicated that PCB126 promotes EMT process of HCC cells via PKM2/STAT3/Snail1 signaling which is mediated by ER and AhR. - Highlights: • PCB126 promotes epithelial-mesenchymal transition of HCC cells. • PCB126 regulates EMT through the activation of STAT3/Snail1 signaling. • PKM2 is responsible for PCB126-induced activation of STAT3/Snail1 signaling. • AhR-induced ROS generation regulates PKM2/STAT3/Snail1 signaling. • PCB126-induced EMT is mediated by ER and AhR.« less

Yes-Associated Protein Promotes Angiogenesis via Signal Transducer and Activator of Transcription 3 in Endothelial Cells.

PubMed

He, Jinlong; Bao, Qiankun; Zhang, Yan; Liu, Mingming; Lv, Huizhen; Liu, Yajin; Yao, Liu; Li, Bochuan; Zhang, Chenghu; He, Shuang; Zhai, Guijin; Zhu, Yan; Liu, Xin; Zhang, Kai; Wang, Xiu-Jie; Zou, Ming-Hui; Zhu, Yi; Ai, Ding

2018-02-16

Angiogenesis is a complex process regulating endothelial cell (EC) functions. Emerging lines of evidence support that YAP (Yes-associated protein) plays an important role in regulating the angiogenic activity of ECs. The objective of this study was to specify the effect of EC YAP on angiogenesis and its underlying mechanisms. In ECs, vascular endothelial growth factor reduced YAP phosphorylation time and dose dependently and increased its nuclear accumulation. Using Tie2Cre-mediated YAP transgenic mice, we found that YAP promoted angiogenesis in the postnatal retina and tumor tissues. Mass spectrometry revealed signal transducer and activator of transcription 3 (STAT3) as a potential binding partner of YAP in ECs. Western blot and immunoprecipitation assays indicated that binding with YAP prolonged interleukin 6-induced STAT3 nuclear accumulation by blocking chromosomal maintenance 1-mediated STAT3 nuclear export without affecting its phosphorylation. Moreover, angiopoietin-2 expression induced by STAT3 was enhanced by YAP overexpression in ECs. Finally, a selective STAT3 inhibitor or angiopoietin-2 blockage partly attenuated retinal angiogenesis in Tie2Cre-mediated YAP transgenic mice. YAP binding sustained STAT3 in the nucleus to enhance the latter's transcriptional activity and promote angiogenesis via regulation of angiopoietin-2. © 2018 American Heart Association, Inc.

Viral RNA-Unprimed Rig-I Restrains Stat3 Activation in the Modulation of Regulatory T Cell/Th17 Cell Balance.

PubMed

Yang, Hui; Guo, He-Zhou; Li, Xian-Yang; Lin, Jian; Zhang, Wu; Zhao, Jun-Mei; Zhang, Hong-Xin; Chen, Sai-Juan; Chen, Zhu; Zhu, Jiang

2017-07-01

Innate immunity activation by viral RNA-primed retinoid acid inducible gene-I (Rig-I) in CD4 + T cells antagonizes TGFβ signaling to suppress the differentiation of regulatory T cells (Tregs). However, how viral RNA-unliganded Rig-I (apo-Rig-I) modulates Treg generation remains unclear. In this article, we show that, in the absence of viral infection, Treg differentiation of Rig-I -/- CD4 + T cells was compromised, in the presence of increased generation of Th17 cells and overactivation of Stat3, a critical regulator tilting the Treg/Th17 cell balance. Mechanistically, apo-Rig-I physically associates with Stat3, thereby inhibiting Jak1's association with Stat3 while facilitating Shp2's association to inhibit p-Stat3 levels. Interestingly, inhibition of Stat3 ameliorates the Treg/Th17 imbalance and the colitis observed in Rig-I -/- mice. Collectively, these results uncover an independent functional contribution of the apo-Rig-I/Stat3 interaction in the maintenance of Treg/Th17 cell balance. Copyright © 2017 by The American Association of Immunologists, Inc.

Combination Treatment with Apricoxib and IL-27 Enhances Inhibition of Epithelial-Mesenchymal Transition in Human Lung Cancer Cells through a STAT1 Dominant Pathway

PubMed Central

Lee, Mi-Heon; Kachroo, Puja; Pagano, Paul C; Yanagawa, Jane; Wang, Gerald; Walser, Tonya C; Krysan, Kostyantyn; Sharma, Sherven; John, Maie St.; Dubinett, Steven M; Lee, Jay M

2015-01-01

Background The cyclooxygenase 2 (COX-2) pathway has been implicated in the molecular pathogenesis of many malignancies, including lung cancer. Apricoxib, a selective COX-2 inhibitor, has been described to inhibit epithelial-mesenchymal transition (EMT) in human malignancies. The mechanism by which apricoxib may alter the tumor microenvironment by affecting EMT through other important signaling pathways is poorly defined. IL-27 has been shown to have anti-tumor activity and our recent study showed that IL-27 inhibited EMT through a STAT1 dominant pathway. Objective The purpose of this study is to investigate the role of apricoxib combined with IL-27 in inhibiting lung carcinogenesis by modulation of EMT through STAT signaling. Methods and Results Western blot analysis revealed that IL-27 stimulation of human non-small cell lung cancer (NSCLC) cell lines results in STAT1 and STAT3 activation, decreased Snail protein and mesenchymal markers (N-cadherin and vimentin) and a concomitant increase in expression of epithelial markers (E-cadherin, β-and γ-catenins), and inhibition of cell migration. The combination of apricoxib and IL-27 resulted in augmentation of STAT1 activation. However, IL-27 mediated STAT3 activation was decreased by the addition of apricoxib. STAT1 siRNA was used to determine the involvement of STAT1 pathway in the enhanced inhibition of EMT and cell migration by the combined IL-27 and apricoxib treatment. Pretreatment of cells with STAT1 siRNA inhibited the effect of combined IL-27 and apricoxib in the activation of STAT1 and STAT3. In addition, the augmented expression of epithelial markers, decreased expression mesenchymal markers, and inhibited cell migration by the combination treatment were also inhibited by STAT1 siRNA, suggesting that the STAT1 pathway is important in the enhanced effect from the combination treatment. Conclusion Combined apricoxib and IL-27 has an enhanced effect in inhibition of epithelial-mesenchymal transition and cell migration in human lung cancer cells through a STAT1 dominant pathway. PMID:26523208

Changes in signal transducer and activator of transcription 3 (STAT3) dynamics induced by complexation with pharmacological inhibitors of Src homology 2 (SH2) domain dimerization.

PubMed

Resetca, Diana; Haftchenary, Sina; Gunning, Patrick T; Wilson, Derek J

2014-11-21

The activity of the transcription factor signal transducer and activator of transcription 3 (STAT3) is dysregulated in a number of hematological and solid malignancies. Development of pharmacological STAT3 Src homology 2 (SH2) domain interaction inhibitors holds great promise for cancer therapy, and a novel class of salicylic acid-based STAT3 dimerization inhibitors that includes orally bioavailable drug candidates has been recently developed. The compounds SF-1-066 and BP-1-102 are predicted to bind to the STAT3 SH2 domain. However, given the highly unstructured and dynamic nature of the SH2 domain, experimental confirmation of this prediction was elusive. We have interrogated the protein-ligand interaction of STAT3 with these small molecule inhibitors by means of time-resolved electrospray ionization hydrogen-deuterium exchange mass spectrometry. Analysis of site-specific evolution of deuterium uptake induced by the complexation of STAT3 with SF-1-066 or BP-1-102 under physiological conditions enabled the mapping of the in silico predicted inhibitor binding site to the STAT3 SH2 domain. The binding of both inhibitors to the SH2 domain resulted in significant local decreases in dynamics, consistent with solvent exclusion at the inhibitor binding site and increased rigidity of the inhibitor-complexed SH2 domain. Interestingly, inhibitor binding induced hot spots of allosteric perturbations outside of the SH2 domain, manifesting mainly as increased deuterium uptake, in regions of STAT3 important for DNA binding and nuclear localization. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

Requirement for distinct Janus kinases and STAT proteins in T cell proliferation versus IFN-gamma production following IL-12 stimulation.

PubMed

Ahn, H J; Tomura, M; Yu, W G; Iwasaki, M; Park, W R; Hamaoka, T; Fujiwara, H

1998-12-01

While IL-12 is known to activate JAK2 and TYK2 and induce the phosphorylation of STAT4 and STAT3, little is known regarding how the activation of these signaling molecules is related to the biologic effects of IL-12. Using an IL-12-responsive T cell clone (2D6), we investigated their requirements for proliferation and IFN-gamma production of 2D6 cells. 2D6 cells could be maintained with either IL-12 or IL-2. 2D6 lines maintained with IL-12 (2D6(IL-12)) or IL-2 (2D6(IL-2)) exhibited comparable levels of proliferation, but produced large or only small amounts of IFN-gamma, respectively, when restimulated with IL-12 after starvation of either cytokine. 2D6(IL-12) induced TYK2 and STAT4 phosphorylation. In contrast, their phosphorylation was marginally induced in 2D6(IL-2). The reduced STAT4 phosphorylation was due to a progressive decrease in the amount of STAT4 protein along with the passages in IL-2-containing medium. 2D6(IL-12) and 2D6(IL-2) similarly proliferating in response to IL-12 induced comparable levels of JAK2 activation and STAT5 phosphorylation. JAK2 was associated with STAT5, and IL-12-induced STAT5 phosphorylation was elicited in the absence of JAK3 activation. These results indicate that IL-12 has the capacity to induce/maintain STAT4 and STAT5 proteins, and that TYK2 and JAK2 activation correlate with STAT4 phosphorylation/IFN-gamma induction and STAT5 phosphorylation/cellular proliferation, respectively.

STAT3 Regulates Uterine Epithelial Remodeling and Epithelial-Stromal Crosstalk During Implantation

PubMed Central

Pawar, Sandeep; Starosvetsky, Elina; Orvis, Grant D.; Behringer, Richard R.; Bagchi, Indrani C.

2013-01-01

Embryo implantation is regulated by a variety of endometrial factors, including cytokines, growth factors, and transcription factors. Earlier studies identified the leukemia inhibitory factor (LIF), a cytokine produced by uterine glands, as an essential regulator of implantation. LIF, acting via its cell surface receptor, activates the signal transducer and activator of transcription 3 (STAT3) in the uterine epithelial cells. However, the precise mechanism via which activated STAT3 promotes uterine function during implantation remains unknown. To identify the molecular pathways regulated by STAT3, we created SWd/d mice in which Stat3 gene is conditionally inactivated in uterine epithelium. The SWd/d mice are infertile due to a lack of embryo attachment to the uterine luminal epithelium and consequent implantation failure. Gene expression profiling of uterine epithelial cells of SWd/d mice revealed dysregulated expression of specific components of junctional complexes, including E-cadherin, α- and β-catenin, and several claudins, which critically regulate epithelial junctional integrity and embryo attachment. In addition, uteri of SWd/d mice exhibited markedly reduced stromal proliferation and differentiation, indicating that epithelial STAT3 controls stromal function via a paracrine mechanism. The stromal defect arose from a drastic reduction in the production of several members of the epidermal growth factor family in luminal epithelium of SWd/d uteri and the resulting lack of activation of epidermal growth factor receptor signaling and mitotic activity in the stromal cells. Collectively, our results uncovered an intricate molecular network operating downstream of STAT3 that regulates uterine epithelial junctional reorganization, and stromal proliferation, and differentiation, which are critical determinants of successful implantation. PMID:24100212

The import of the transcription factor STAT3 into mitochondria depends on GRIM-19, a component of the electron transport chain.

PubMed

Tammineni, Prasad; Anugula, Chandrashekhar; Mohammed, Fareed; Anjaneyulu, Murari; Larner, Andrew C; Sepuri, Naresh Babu Venkata

2013-02-15

The signal transducer and activator of transcription 3 (STAT3), a nuclear transcription factor, is also present in mitochondria and regulates cellular respiration in a transcriptional-independent manner. The mechanism of STAT3 import into mitochondria remains obscure. In this report we show that mitochondrial-localized STAT3 resides in the inner mitochondrial membrane. In vitro import studies show that the gene associated with retinoid interferon induced cell mortality 19 (GRIM-19), a complex I subunit that acts as a chaperone to recruit STAT3 into mitochondria. In addition, GRIM-19 enhances the integration of STAT3 into complex I. A S727A mutation in STAT3 reduces its import and assembly even in the presence of GRIM-19. Together, our studies unveil a novel chaperone function for GRIM-19 in the recruitment of STAT3 into mitochondria.

INTERFERON α ACTIVATES NF-κ B IN JAK1-DEFICIENT CELLS THROUGH A TYK2-DEPENDENT PATHWAY

PubMed Central

Yang, Chuan He; Murti, Aruna; Valentine, William J.; Du, Ziyun; Pfeffer, Lawrence M.

2005-01-01

In addition to activating members of the STAT transcription factor family, IFN α/β activates the NF-κ B transcription factor. To determine the role of the JAK-STAT pathway in NF-κ B activation by IFN, we examined NF-κ B activation in JAK1-deficient mutant human fibrosarcoma cells. In wild-type fibrosarcoma cells (2fTGH) IFN activates STAT1, STAT2 and STAT3, as well as NF-κB complexes comprised of p50 and p65. In contrast, in JAK1-deficient cells IFN induces NF-κB activation and NF-κB dependent gene transcription, but does not activate these STAT proteins and has no effect on STAT-dependent gene transcription. Expression of a catalytically-inactive TYK2 tyrosine kinase in JAK1-deficient cells, as well as in the highly IFN-sensitive Daudi lymphoblastoid cell line, abrogates NF-κB activation by IFN. Moreover, IFN does not promote NF-κB activation in TYK2-deficient mutant fibrosarcoma cells. Our results demonstrate a dichotomy between the classical JAK-STAT pathway and the NF-κB signaling pathway. In the IFN signaling pathway leading to STAT activation both JAK1 and TYK2 are essential, while NF-κB activation requires only TYK2. PMID:15883164

Novel signal transducer and activator of transcription 3 (STAT3) mutations, reduced T(H)17 cell numbers, and variably defective STAT3 phosphorylation in hyper-IgE syndrome.

PubMed

Renner, Ellen D; Rylaarsdam, Stacey; Anover-Sombke, Stephanie; Rack, Anita L; Reichenbach, Janine; Carey, John C; Zhu, Qili; Jansson, Annette F; Barboza, Julia; Schimke, Lena F; Leppert, Mark F; Getz, Melissa M; Seger, Reinhard A; Hill, Harry R; Belohradsky, Bernd H; Torgerson, Troy R; Ochs, Hans D

2008-07-01

Hyper-IgE syndrome (HIES) is a rare, autosomal-dominant immunodeficiency characterized by eczema, Staphylococcus aureus skin abscesses, pneumonia with pneumatocele formation, Candida infections, and skeletal/connective tissue abnormalities. Recently it was shown that heterozygous signal transducer and activator of transcription 3 (STAT3) mutations cause autosomal-dominant HIES. To determine the spectrum and functional consequences of heterozygous STAT3 mutations in a cohort of patients with HIES. We sequenced the STAT3 gene in 38 patients with HIES (National Institutes of Health score >40 points) from 35 families, quantified T(H)17 cells in peripheral blood, and evaluated tyrosine phosphorylation of STAT3. Most STAT3 mutations in our cohort were in the DNA-binding domain (DBD; 22/35 families) or Src homology 2 (SH2) domain (10/35) and were missense mutations. We identified 2 intronic mutations resulting in exon skipping and in-frame deletions within the DBD. In addition, we identified 2 mutations located in the transactivation domain downstream of the SH2 domain: a 10-amino acid deletion and an amino acid substitution. In 1 patient, we were unable to identify a STAT3 mutation. T(H)17 cells were absent or low in the peripheral blood of all patients who were evaluated (n = 17). IL-6-induced STAT3-phosphorylation was consistently reduced in patients with SH2 domain mutations but comparable to normal controls in patients with mutations in the DBD. Heterozygous STAT3 mutations were identified in 34 of 35 unrelated HIES families. Patients had impaired T(H)17 cell development, and those with SH2 domain mutations had reduced STAT3 phosphorylation.

Diindolylmethane suppresses ovarian cancer growth and potentiates the effect of cisplatin in tumor mouse model by targeting signal transducer and activator of transcription 3 (STAT3)

PubMed Central

2012-01-01

Background Signal transducer and activator of transcription 3 (STAT3) is activated in majority of ovarian tumors and confers resistance to cisplatin treatment in patients with ovarian cancer. We have reported previously that diindolylmethane (DIM) inhibits the growth of ovarian cancer cells. However, to date the exact mechanism by which DIM induces growth suppressive effects has not been clear. In this report the mode of action of DIM is investigated. Methods Six human ovarian cancer cell lines and an ovarian tumor xenograft animal model were used to study the effect of diindolylmethane alone or in combination with cisplatin. Results Diindolylmethane treatment induced apoptosis in all six ovarian cancer cell lines. Phosphorylation of STAT3 at Tyr-705 and Ser-727 was reduced by DIM in a concentration-dependent manner. In addition, diindolylmethane treatment inhibited nuclear translocation, DNA binding, and transcriptional activity of STAT3. Interleukin (IL)-6-induced phosphorylation of STAT3 at Tyr-705 was significantly blocked by DIM. Overexpression of STAT3 by gene transfection blocked DIM-induced apoptosis. In addition, DIM treatment reduced the levels of IL-6 in ovarian cancer cells and in the tumors. DIM treatment also inhibited cell invasion and angiogenesis by suppressing hypoxia-inducible factor 1α (HIF-1α) and vascular epithelial growth factor (VEGF). Importantly, diindolylmethane treatment potentiated the effects of cisplatin in SKOV-3 cells by targeting STAT3. Oral administration of 3 mg diindolylmethane per day and subsequent administration of cisplatin substantially inhibited in vivo tumor growth. Western blotting analysis of tumor lysates indicated increased apoptosis and reduced STAT3 activation. Conclusions These findings provide a rationale for further clinical investigation of DIM alone or in combination for chemoprevention and/or chemotherapy of ovarian cancer. PMID:22280969

Zoledronic acid overcomes chemoresistance and immunosuppression of malignant mesothelioma

PubMed Central

Kopecka, Joanna; Gazzano, Elena; Sara, Orecchia; Ghigo, Dario; Riganti, Chiara

2015-01-01

The human malignant mesothelioma (HMM) is characterized by a chemoresistant and immunosuppressive phenotype. An effective strategy to restore chemosensitivity and immune reactivity against HMM is lacking. We investigated whether the use of zoledronic acid is an effective chemo-immunosensitizing strategy. We compared primary HMM samples with non-transformed mesothelial cells. HMM cells had higher rate of cholesterol and isoprenoid synthesis, constitutive activation of Ras/extracellular signal-regulated kinase1/2 (ERK1/2)/hypoxia inducible factor-1α (HIF-1α) pathway and up-regulation of the drug efflux transporter P-glycoprotein (Pgp). By decreasing the isoprenoid supply, zoledronic acid down-regulated the Ras/ERK1/2/HIF-1α/Pgp axis and chemosensitized the HMM cells to Pgp substrates. The HMM cells also produced higher amounts of kynurenine, decreased the proliferation of T-lymphocytes and expanded the number of T-regulatory (Treg) cells. Kynurenine synthesis was due to the transcription of the indoleamine 1,2 dioxygenase (IDO) enzyme, consequent to the activation of the signal transducer and activator of transcription-3 (STAT3). By reducing the activity of the Ras/ERK1/2/STAT3/IDO axis, zoledronic acid lowered the kyurenine synthesis and the expansion of Treg cells, and increased the proliferation of T-lymphocytes. Thanks to its ability to decrease Ras/ERK1/2 activity, which is responsible for both Pgp-mediated chemoresistance and IDO-mediated immunosuppression, zoledronic acid is an effective chemo-immunosensitizing agent in HMM cells. PMID:25544757

Two Naturally Occurring Terpenes, Dehydrocostuslactone and Costunolide, Decrease Intracellular GSH Content and Inhibit STAT3 Activation

PubMed Central

Butturini, Elena; Cavalieri, Elisabetta; Carcereri de Prati, Alessandra; Darra, Elena; Rigo, Antonella; Shoji, Kazuo; Murayama, Norie; Yamazaki, Hiroshi; Watanabe, Yasuo; Suzuki, Hisanori; Mariotto, Sofia

2011-01-01

The main purpose of the present study is to envisage the molecular mechanism of inhibitory action ofdehydrocostuslactone (DCE) andcostunolide (CS), two naturally occurring sesquiterpene lactones, towards the activation of signal transducer and activator of transcription 3 (STAT3). We report that, in human THP-1 cell line, they inhibit IL-6-elicited tyrosine phosphorylation of STAT3 and its DNA binding activity with EC50 of 10 µM with concomitantdown-regulation ofthe phosphorylation of the tyrosine Janus kinases JAK1, JAK2 and Tyk2. Furthermore, these compounds that contain an α-β-unsatured carbonyl moiety and function as potent Michael reaction acceptor, induce a rapid drop in intracellular glutathione (GSH) concentration by direct interaction with it, thereby triggering S-glutathionylation of STAT3. Dehydrocostunolide (HCS), the reduced form of CS lacking only the α-β-unsaturated carbonyl group, fails to exert any inhibitory action. Finally, the glutathione ethylene ester (GEE), the cell permeable GSH form, reverts the inhibitory action of DCE and CS on STAT3 tyrosine phosphorylation. We conclude that these two sesquiterpene lactones are able to induce redox-dependent post-translational modification of cysteine residues of STAT3 protein in order to regulate its function. PMID:21625597

Role of proopiomelanocortin neuron Stat3 in regulating arterial pressure and mediating the chronic effects of leptin.

PubMed

Dubinion, John H; do Carmo, Jussara M; Adi, Ahmad; Hamza, Shereen; da Silva, Alexandre A; Hall, John E

2013-05-01

Although signal transducer and activator of transcription 3 (Stat3) is a key second messenger by which leptin regulates appetite and body weight, its role in specific neuronal populations in metabolic regulation and in mediating the chronic effects of leptin on blood pressure is unknown. The current study tested the hypothesis that Stat3 signaling in proopiomelanocortin (POMC) neurons mediates the chronic effects of leptin on mean arterial pressure (MAP), as well as on glucose regulation, energy expenditure, and food intake. Stat3(flox/flox) mice were crossed with POMC-Cre mice to generate mice with Stat3 deletion specifically in POMC neurons (Stat3(flox/flox)/POMC-Cre). Oxygen consumption (Vo2), carbon dioxide respiration (Vco2), motor activity, heat production, food intake, and MAP were measured 24 hours/d. After baseline measurements, leptin was infused (4 μg/kg per min, IP) for 7 days. Stat3(flox/flox)/POMC-Cre mice were hyperphagic, heavier, and had increased respiratory quotients compared with control Stat3(flox/flox) mice. Baseline MAP was not different between the groups, and chronic leptin infusion reduced food intake similarly in both groups (27 versus 29%). Vo2, Vco2, and heat production responses to leptin were not significantly different in control and Stat3(flox/flox)/POMC-Cre mice. However, leptin-mediated increases in MAP were completely abolished, and blood pressure responses to acute air-jet stress were attenuated in male Stat3(flox/flox)/POMC-Cre mice. These results indicate that Stat3 signaling in POMC neurons is essential for leptin-mediated increases in MAP, but not for anorexic or thermogenic effects of leptin.

STAT3/NF-κB interactions determine the level of haptoglobin expression in male rats exposed to dietary restriction and/or acute phase stimuli.

PubMed

Uskoković, Aleksandra; Dinić, Svetlana; Mihailović, Mirjana; Grdović, Nevena; Arambašić, Jelena; Vidaković, Melita; Bogojević, Desanka; Ivanović-Matić, Svetlana; Martinović, Vesna; Petrović, Miodrag; Poznanović, Goran; Grigorov, Ilijana

2012-01-01

Haptoglobin is a constitutively expressed protein which is predominantly synthesized in the liver. During the acute-phase (AP) response haptoglobin is upregulated along with other AP proteins. Its upregulation during the AP response is mediated by cis-trans interactions between the hormone-responsive element (HRE) residing in the haptoglobin gene and inducible transcription factors STAT3 and C/EBP β. In male rats that have been subjected to chronic 50% dietary restriction (DR), the basal haptoglobin serum level is decreased. The aim of this study was to characterize the trans-acting factor(s) responsible for the reduction of haptoglobin expression in male rats subjected to 50% DR for 6 weeks. Protein-DNA interactions between C/EBP and STAT families of transcription factors and the HRE region of the haptoglobin gene were examined in livers of male rats subjected to DR, as well as during the AP response that was induced by turpentine administration. In DR rats, we observed associations between the HRE and C/EBPα/β, STAT5b and NF-κB p50, and the absence of interactions between STAT3 and NF-kB p65. Subsequent induction of the AP response in DR rats by turpentine administration elicited a normal, almost 2-fold increase in the serum haptoglobin level that was accompanied by HRE-binding of C/EBPβ, STAT3/5b and NF-kB p65/p50, and the establishment of interaction between STAT3 and NF-κB p65. These results suggest that STAT3 and NF-κB p65 crosstalk plays a central role while C/EBPβ acquires an accessory role in establishing the level of haptoglobin gene expression in male rats exposed to DR and AP stimuli.

Effects of angiotensin II type 1 receptor antagonist and temperature on prolonged cardioplegic arrest in neonatal rat myocytes.

PubMed

Lucchese, Gianluca; Cambi, Giulia Elisa; De Rita, Fabrizio; Franzoi, Mauro; Faggian, Giuseppe; Mazzucco, Alessandro; Modesti, Pietro Amedeo; Luciani, Giovanni Battista

2013-08-01

Cardioplegic arrest is a model of ischemia/reperfusion injury and results in the death of irreplaceable cardiac myocytes by a programmed cell death or apoptosis. Signal transducers and activators of transcription (STAT) signaling pathways play an important role in the modulation of apoptosis after ischemia and reperfusion. Angiotensin II type 1 (AT1) receptor antagonist added to cardioplegia could represent an additional modality for enhancing myocardial protection during cardioplegic arrest. To test that hypothesis, we studied the effect of AT1 receptor antagonism and cardioplegia temperature perfusion on STATs modulation during cardioplegic arrest in neonatal rat hearts. Isolated, nonworking hearts (n = 4 per group) from neonatal rats were perfused aerobically in the Langendorff mode according to the following scheme: Dulbecco's Modified Eagle's Medium solution (Group 1); cold (4°C) modified St. Thomas' Hospital no. 2 (MSTH2) cardioplegic solution (Group 2); cold (4°C) MSTH2 cardioplegic solution plus AT1 antagonist (Valsartan) (Group 3); and warm (34°C) MSTH2 cardioplegic solution (Group 4). Thus, myocytes were isolated by enzymatic digestion, and STAT1, STAT2, STAT3, and STAT5 were investigated in Western blot studies. Times to arrest after cardioplegia were 6-10 s for all groups with the exception of Group 1 (spontaneous arrest after 12-16 s). Total cardioplegia delivery volume was about 300 mL in 15 min. Perfusion with cold MSTH2 supplemented with AT1 receptor antagonist (Group 3) induced a significant reduction in STAT1, STAT2, and STAT5 tyrosine phosphorylation versus other groups (P < 0.05). The decreased activation of STAT1, STAT2, and STAT5 observed in Group 3 was accompanied by reduction of interleukin-1β (P < 0.05). On the other hand, STAT3 activation was significantly reduced in Groups 1 and 4 (P < 0.05). Only perfusion with AT1 receptor antagonist supplemented with cold MSTH2 significantly decreases the inflammatory response of the neonatal rat cardiomyocytes without affecting antiapoptotic influence provided by activation of STAT3. Therefore, AT1 receptor antagonist could play a pivotal role in cytoprotective effect and cardiac recovery in neonates and infants. © 2013, Copyright the Authors. Artificial Organs © 2013 Wiley Periodicals, Inc. and International Center for Artificial Organs and Transplantation.

4-Phenylbutyrate stimulates Hsp70 expression through the Elp2 component of elongator and STAT-3 in cystic fibrosis epithelial cells.

PubMed

Suaud, Laurence; Miller, Katelyn; Panichelli, Ashley E; Randell, Rachel L; Marando, Catherine M; Rubenstein, Ronald C

2011-12-30

Sodium 4-phenylbutyrate (4PBA) corrects trafficking of ΔF508-CFTR in Cystic Fibrosis (CF) epithelia, which is hypothesized to, at least in part, result from increased expression of Hsp70 (stress-induced 70 kDa heat shock protein). To identify other 4PBA-regulated proteins that may promote correction of ΔF508 trafficking, we performed differential display RT-PCR on mRNA from IB3-1 CF bronchiolar epithelial cells treated for 0-24 h with 1 mM 4PBA. In this screen, a STAT-3 (signal transducer and activator of transcription-3)-interacting protein, StIP-1 that regulates STAT-3 activation had transiently increased expression. StIP-1 is identical to Elongator protein 2 (Elp2), a component of the Elongator complex that regulates RNA polymerase II. Previous studies have suggested that Elongator regulates Hsp70 mRNA transcription, and that the Hsp70 promoter contains functional STAT-3-binding sites. We therefore tested the hypothesis that 4PBA increases Hsp70 expression by an Elongator- and STAT-3-dependent mechanism. 4PBA treatment of IB3-1 CF bronchiolar epithelial cells caused transiently increased expression of Hsp70 protein, as well as Elp2 protein and mRNA. Elp2 depletion by transfection of small interfering RNAs, reduced both Elp2 and Hsp70 protein expression. 4PBA also caused transient activation of STAT-3, and increased abundance of nuclear proteins that bind to the STAT-3-responsive element of the Hsp70 promoter. Luciferase reporter assays demonstrated that both Elp2 overexpression and 4PBA increase Hsp70 promoter activity, while Elp2 depletion blocked the ability of 4PBA to stimulate Hsp70 promoter activity. Together, these data suggest that Elp2 and STAT-3 mediate, at least in part, the stimulation of Hsp70 expression by 4PBA.

4-Phenylbutyrate Stimulates Hsp70 Expression through the Elp2 Component of Elongator and STAT-3 in Cystic Fibrosis Epithelial Cells*

PubMed Central

Suaud, Laurence; Miller, Katelyn; Panichelli, Ashley E.; Randell, Rachel L.; Marando, Catherine M.; Rubenstein, Ronald C.

2011-01-01

Sodium 4-phenylbutyrate (4PBA) corrects trafficking of ΔF508-CFTR in Cystic Fibrosis (CF) epithelia, which is hypothesized to, at least in part, result from increased expression of Hsp70 (stress-induced 70 kDa heat shock protein). To identify other 4PBA-regulated proteins that may promote correction of ΔF508 trafficking, we performed differential display RT-PCR on mRNA from IB3-1 CF bronchiolar epithelial cells treated for 0–24 h with 1 mm 4PBA. In this screen, a STAT-3 (signal transducer and activator of transcription-3)-interacting protein, StIP-1 that regulates STAT-3 activation had transiently increased expression. StIP-1 is identical to Elongator protein 2 (Elp2), a component of the Elongator complex that regulates RNA polymerase II. Previous studies have suggested that Elongator regulates Hsp70 mRNA transcription, and that the Hsp70 promoter contains functional STAT-3-binding sites. We therefore tested the hypothesis that 4PBA increases Hsp70 expression by an Elongator- and STAT-3-dependent mechanism. 4PBA treatment of IB3-1 CF bronchiolar epithelial cells caused transiently increased expression of Hsp70 protein, as well as Elp2 protein and mRNA. Elp2 depletion by transfection of small interfering RNAs, reduced both Elp2 and Hsp70 protein expression. 4PBA also caused transient activation of STAT-3, and increased abundance of nuclear proteins that bind to the STAT-3-responsive element of the Hsp70 promoter. Luciferase reporter assays demonstrated that both Elp2 overexpression and 4PBA increase Hsp70 promoter activity, while Elp2 depletion blocked the ability of 4PBA to stimulate Hsp70 promoter activity. Together, these data suggest that Elp2 and STAT-3 mediate, at least in part, the stimulation of Hsp70 expression by 4PBA. PMID:22069317

Expression and Purification of Soluble STAT5b/STAT3 Proteins for SH2 Domain Binding Assay.

PubMed

Asai, Akira; Takakuma, Kazuyuki

2017-01-01

When a large hydrophobic full-length protein is expressed in bacteria, it is often challenging to obtain recombinant proteins in the soluble fraction. One way to overcome this challenge is expression of deletion mutants that have improved solubility while maintaining biological activity. In this chapter, we describe a protocol for expression of truncated forms of STAT5b and STAT3 proteins that are soluble and retain SH2-mediated activity for phospho-Tyr peptide recognition.

Rhodium(II) proximity-labeling identifies a novel target site on STAT3 for inhibitors with potent anti-leukemia activity

PubMed Central

Minus, Matthew B.; Liu, Wei; Vohidov, Farrukh; Kasembeli, Moses M.; Long, Xin; Krueger, Michael; Stevens, Alexandra; Kolosov, Mikhail I.; Sison, Edward Allen R.; Ball, Zachary T.

2015-01-01

Nearly 40% of children with acute myeloid leukemia (AML) suffer relapse due to chemoresistance, often involving upregulation of the oncoprotein STAT3 (signal transducer and activator of transcription 3). In this paper, rhodium(II)-catalyzed, proximity-driven modification identifies the STAT3 coiled-coil domain (CCD) as a novel ligand-binding site, and we describe a new naphthalene sulfonamide inhibitor that targets the CCD, blocks STAT3 function, and halts its disease-promoting effects in vitro, in tumor growth models, and in a leukemia mouse model, validating this new therapeutic target for resistant AML. PMID:26480340

Gain-of-function human STAT1 mutations impair IL-17 immunity and underlie chronic mucocutaneous candidiasis.

PubMed

Liu, Luyan; Okada, Satoshi; Kong, Xiao-Fei; Kreins, Alexandra Y; Cypowyj, Sophie; Abhyankar, Avinash; Toubiana, Julie; Itan, Yuval; Audry, Magali; Nitschke, Patrick; Masson, Cécile; Toth, Beata; Flatot, Jérome; Migaud, Mélanie; Chrabieh, Maya; Kochetkov, Tatiana; Bolze, Alexandre; Borghesi, Alessandro; Toulon, Antoine; Hiller, Julia; Eyerich, Stefanie; Eyerich, Kilian; Gulácsy, Vera; Chernyshova, Ludmyla; Chernyshov, Viktor; Bondarenko, Anastasia; Grimaldo, Rosa María Cortés; Blancas-Galicia, Lizbeth; Beas, Ileana Maria Madrigal; Roesler, Joachim; Magdorf, Klaus; Engelhard, Dan; Thumerelle, Caroline; Burgel, Pierre-Régis; Hoernes, Miriam; Drexel, Barbara; Seger, Reinhard; Kusuma, Theresia; Jansson, Annette F; Sawalle-Belohradsky, Julie; Belohradsky, Bernd; Jouanguy, Emmanuelle; Bustamante, Jacinta; Bué, Mélanie; Karin, Nathan; Wildbaum, Gizi; Bodemer, Christine; Lortholary, Olivier; Fischer, Alain; Blanche, Stéphane; Al-Muhsen, Saleh; Reichenbach, Janine; Kobayashi, Masao; Rosales, Francisco Espinosa; Lozano, Carlos Torres; Kilic, Sara Sebnem; Oleastro, Matias; Etzioni, Amos; Traidl-Hoffmann, Claudia; Renner, Ellen D; Abel, Laurent; Picard, Capucine; Maródi, László; Boisson-Dupuis, Stéphanie; Puel, Anne; Casanova, Jean-Laurent

2011-08-01

Chronic mucocutaneous candidiasis disease (CMCD) may be caused by autosomal dominant (AD) IL-17F deficiency or autosomal recessive (AR) IL-17RA deficiency. Here, using whole-exome sequencing, we identified heterozygous germline mutations in STAT1 in 47 patients from 20 kindreds with AD CMCD. Previously described heterozygous STAT1 mutant alleles are loss-of-function and cause AD predisposition to mycobacterial disease caused by impaired STAT1-dependent cellular responses to IFN-γ. Other loss-of-function STAT1 alleles cause AR predisposition to intracellular bacterial and viral diseases, caused by impaired STAT1-dependent responses to IFN-α/β, IFN-γ, IFN-λ, and IL-27. In contrast, the 12 AD CMCD-inducing STAT1 mutant alleles described here are gain-of-function and increase STAT1-dependent cellular responses to these cytokines, and to cytokines that predominantly activate STAT3, such as IL-6 and IL-21. All of these mutations affect the coiled-coil domain and impair the nuclear dephosphorylation of activated STAT1, accounting for their gain-of-function and dominance. Stronger cellular responses to the STAT1-dependent IL-17 inhibitors IFN-α/β, IFN-γ, and IL-27, and stronger STAT1 activation in response to the STAT3-dependent IL-17 inducers IL-6 and IL-21, hinder the development of T cells producing IL-17A, IL-17F, and IL-22. Gain-of-function STAT1 alleles therefore cause AD CMCD by impairing IL-17 immunity.

IFN-λ Inhibits MiR-122 Transcription through a Stat3-HNF4α Inflammatory Feedback Loop in an IFN-α Resistant HCV Cell Culture System

PubMed Central

Aboulnasr, Fatma; Hazari, Sidhartha; Nayak, Satyam; Chandra, Partha K.; Panigrahi, Rajesh; Ferraris, Pauline; Chava, Srinivas; Kurt, Ramazan; Song, Kyongsub; Dash, Asha; Balart, Luis A.; Garry, Robert F.; Wu, Tong; Dash, Srikanta

2015-01-01

Background HCV replication in persistently infected cell culture remains resistant to IFN-α/RBV combination treatment, whereas IFN-λ1 induces viral clearance. The antiviral mechanisms by which IFN-λ1 induces sustained HCV clearance have not been determined. Aim To investigate the mechanisms by which IFN-λ clears HCV replication in an HCV cell culture model. Methods IFN-α sensitive (S3-GFP) and resistant (R4-GFP) cells were treated with equivalent concentrations of either IFN-α or IFN-λ. The relative antiviral effects of IFN-α and IFN-λ1 were compared by measuring the HCV replication, quantification of HCV-GFP expression by flow cytometry, and viral RNA levels by real time RT-PCR. Activation of Jak-Stat signaling, interferon stimulated gene (ISG) expression, and miRNA-122 transcription in S3-GFP and R4-GFP cells were examined. Results We have shown that IFN-λ1 induces HCV clearance in IFN-α resistant and sensitive replicon cell lines in a dose dependent manner through Jak-Stat signaling, and induces STAT 1 and STAT 2 activation, ISRE-luciferase promoter activation and ISG expression. Stat 3 activation is also involved in IFN-λ1 induced antiviral activity in HCV cell culture. IFN-λ1 induced Stat 3 phosphorylation reduces the expression of hepatocyte nuclear factor 4 alpha (HNF4α) through miR-24 in R4-GFP cells. Reduced expression of HNF4α is associated with decreased expression of miR-122 resulting in an anti-HCV effect. Northern blot analysis confirms that IFN-λ1 reduces miR-122 levels in R4-GFP cells. Our results indicate that IFN-λ1 activates the Stat 3-HNF4α feedback inflammatory loop to inhibit miR-122 transcription in HCV cell culture. Conclusions In addition to the classical Jak–Stat antiviral signaling pathway, IFN-λ1 inhibits HCV replication through the suppression of miRNA-122 transcription via an inflammatory Stat 3–HNF4α feedback loop. Inflammatory feedback circuits activated by IFNs during chronic inflammation expose non-responders to the risk of hepatocellular carcinoma. PMID:26657215

An SH2 domain model of STAT5 in complex with phospho-peptides define ``STAT5 Binding Signatures''

NASA Astrophysics Data System (ADS)

Gianti, Eleonora; Zauhar, Randy J.

2015-05-01

The signal transducer and activator of transcription 5 (STAT5) is a member of the STAT family of proteins, implicated in cell growth and differentiation. STAT activation is regulated by phosphorylation of protein monomers at conserved tyrosine residues, followed by binding to phospho-peptide pockets and subsequent dimerization. STAT5 is implicated in the development of severe pathological conditions, including many cancer forms. However, nowadays a few STAT5 inhibitors are known, and only one crystal structure of the inactive STAT5 dimer is publicly available. With a view to enabling structure-based drug design, we have: (1) analyzed phospho-peptide binding pockets on SH2 domains of STAT5, STAT1 and STAT3; (2) generated a model of STAT5 bound to phospho-peptides; (3) assessed our model by docking against a class of known STAT5 inhibitors (Müller et al. in ChemBioChem 9:723-727, 2008); (4) used molecular dynamics simulations to optimize the molecular determinants responsible for binding and (5) proposed unique "Binding Signatures" of STAT5. Our results put in place the foundations to address STAT5 as a target for rational drug design, from sequence, structural and functional perspectives.

Target specificity, in vivo pharmacokinetics, and efficacy of the putative STAT3 inhibitor LY5 in osteosarcoma, Ewing's sarcoma, and rhabdomyosarcoma.

PubMed

Yu, Peter Y; Gardner, Heather L; Roberts, Ryan; Cam, Hakan; Hariharan, Seethalakshmi; Ren, Ling; LeBlanc, Amy K; Xiao, Hui; Lin, Jiayuh; Guttridge, Denis C; Mo, Xiaokui; Bennett, Chad E; Coss, Christopher C; Ling, Yonghua; Phelps, Mitch A; Houghton, Peter; London, Cheryl A

2017-01-01

STAT3 is a transcription factor involved in cytokine and receptor kinase signal transduction that is aberrantly activated in a variety of sarcomas, promoting metastasis and chemotherapy resistance. The purpose of this work was to develop and test a novel putative STAT3 inhibitor, LY5. An in silico fragment-based drug design strategy was used to create LY5, a small molecule inhibitor that blocks the STAT3 SH2 domain phosphotyrosine binding site, inhibiting homodimerization. LY5 was evaluated in vitro demonstrating good biologic activity against rhabdomyosarcoma, osteosarcoma and Ewing's sarcoma cell lines at high nanomolar/low micromolar concentrations, as well as specific inhibition of STAT3 phosphorylation without effects on other STAT3 family members. LY5 exhibited excellent oral bioavailability in both mice and healthy dogs, and drug absorption was enhanced in the fasted state with tolerable dosing in mice at 40 mg/kg BID. However, RNAi-mediated knockdown of STAT3 did not phenocopy the biologic effects of LY5 in sarcoma cell lines. Moreover, concentrations needed to inhibit ex vivo metastasis growth using the PuMA assay were significantly higher than those needed to inhibit STAT3 phosphorylation in vitro. Lastly, LY5 treatment did not inhibit the growth of sarcoma xenografts or prevent pulmonary metastasis in mice. LY5 is a novel small molecule inhibitor that effectively inhibits STAT3 phosphorylation and cell proliferation at nanomolar concentrations. LY5 demonstrates good oral bioavailability in mice and dogs. However LY5 did not decrease tumor growth in xenograft mouse models and STAT3 knockdown did not induce concordant biologic effects. These data suggest that the anti-cancer effects of LY5 identified in vitro were not mediated through STAT3 inhibition.

Identification of STAT3 and STAT5 proteins in the rat suprachiasmatic nucleus and the Day/Night difference in astrocytic STAT3 phosphorylation in response to lipopolysaccharide.

PubMed

Moravcová, Simona; Červená, Kateřina; Pačesová, Dominika; Bendová, Zdeňka

2016-01-01

Signal transducers and activators of transcription (STAT) proteins regulate many aspects of cellular physiology from growth and differentiations to immune responses. Using immunohistochemistry, we show the daily rhythm of STAT3 protein in the rat suprachiasmatic nucleus (SCN), with low but significant amplitude peaking in the morning. We also reveal the strong expression of STAT5A in astrocytes of the SCN and the STAT5B signal in nonastrocytic cells. Administration of lipopolysaccharide (LPS) acutely induced phosphorylation of STAT3 on Tyr705 during both the day and the night and induced phosphorylation on Ser727 but only after the daytime application. The LPS-induced phospho-STAT3 (Tyr705) remained elevated for 24 hr after the daytime application but declined within 8 hr when LPS was applied at night. © 2015 Wiley Periodicals, Inc.

Stat5-mediated regulation of the human type II 3beta-hydroxysteroid dehydrogenase/delta5-delta4 isomerase gene: activation by prolactin.

PubMed

Feltus, F A; Groner, B; Melner, M H

1999-07-01

Altered PRL levels are associated with infertility in women. Molecular targets at which PRL elicits these effects have yet to be determined. These studies demonstrate transcriptional regulation by PRL of the gene encoding the final enzymatic step in progesterone biosynthesis: 3beta-hydroxysteroid dehydrogenase/delta5-delta4 isomerase (3beta-HSD). A 9/9 match with the consensus Stat5 response element was identified at -110 to -118 in the human Type II 3beta-HSD promoter. 3beta-HSD chloramphenicol acetyltransferase (CAT) reporter constructs containing either an intact or mutated Stat5 element were tested for PRL activation. Expression vectors for Stat5 and the PRL receptor were cotransfected with a -300 --> +45 3beta-HSD CAT reporter construct into HeLa cells, which resulted in a 21-fold increase in reporter activity in the presence of PRL. Promoter activity showed an increased response with a stepwise elevation of transfected Stat5 expression or by treatment with increasing concentrations of PRL (max, 250 ng/ml). This effect was dramatically reduced when the putative Stat5 response element was removed by 5'-deletion of the promoter or by the introduction of a 3-bp mutation into critical nucleotides in the element. Furthermore, 32P-labeled promoter fragments containing the Stat5 element were shifted in electrophoretic mobility shift assay experiments using nuclear extracts from cells treated with PRL, and this complex was supershifted with antibodies to Stat5. These results demonstrate that PRL has the ability to regulate expression of a key human enzyme gene (type II 3beta-HSD) in the progesterone biosynthetic pathway, which is essential for maintaining pregnancy.

SOCS3 Deficiency in Myeloid Cells Promotes Tumor Development: Involvement of STAT3 Activation and Myeloid-Derived Suppressor Cells

PubMed Central

Yu, Hao; Liu, Yudong; McFarland, Braden C.; Deshane, Jessy S.; Hurst, Douglas R.; Ponnazhagan, Selvarangan; Benveniste, Etty N.; Qin, Hongwei

2015-01-01

Suppressor of cytokine signaling (SOCS) proteins are negative regulators of the JAK/STAT pathway, and generally function as tumor suppressors. The absence of SOCS3 in particular leads to heightened activation of the STAT3 transcription factor, which has a striking ability to promote tumor survival while suppressing antitumor immunity. We report for the first time that genetic deletion of SOCS3 specifically in myeloid cells significantly enhances tumor growth, which correlates with elevated levels of myeloid-derived suppressor cells (MDSC) in the tumor microenvironment, and diminished CD8+ T-cell infiltration in tumors. The importance of MDSCs in promoting tumor growth is documented by reduced tumor growth upon depletion of MDSCs. Furthermore, SOCS3-deficient bone-marrow-derived cells exhibit heightened STAT3 activation and preferentially differentiate into the Gr-1+CD11b+Ly6G+ MDSC phenotype. Importantly, we identify granulocyte colony-stimulating factor (G-CSF) as a critical factor secreted by the tumor microenvironment that promotes development of MDSCs via a STAT3-dependent pathway. Abrogation of tumor-derived G-CSF reduces the proliferation and accumulation of Gr-1+CD11b+ MDSCs and inhibits tumor growth. These findings highlight the critical function of SOCS3 as a negative regulator of MDSC development and function, via inhibition of STAT3 activation. PMID:25649351

Phosphotyrosine enrichment identifies focal adhesion kinase and other tyrosine kinases for targeting in canine hemangiosarcoma.

PubMed

Marley, K; Maier, C S; Helfand, S C

2012-09-01

Canine hemangiosarcoma (HSA) is an endothelial cell malignancy driven, in part, by activating mutations in receptor and non-receptor tyrosine kinases. Proteomics, Western blots and a tyrosine kinase inhibitor were used to elucidate activating mechanisms in HSA cell lines. Phosphotyrosine peptides from focal adhesion kinase (FAK) STAT3, Lyn, Fyn and other signal transduction kinases were identified by mass spectrometry. FAK was constitutively activated at tyrosine 397, the autophosphorylation site, and this was reversible with high concentrations of a FAK inhibitor. FAK inhibitor-14 suppressed migration and phosphorylation of FAK tyrosine 397 and tyrosines 576/577 and was cytotoxic to HSA cells suggesting FAK signalling may be an important contributor to canine HSA survival. © 2012 Blackwell Publishing Ltd.

STAT3 inhibition attenuates the progressive phenotypes of Alport syndrome mouse model.

PubMed

Yokota, Tsubasa; Omachi, Kohei; Suico, Mary Ann; Kamura, Misato; Kojima, Haruka; Fukuda, Ryosuke; Motomura, Keishi; Teramoto, Keisuke; Kaseda, Shota; Kuwazuru, Jun; Takeo, Toru; Nakagata, Naomi; Shuto, Tsuyoshi; Kai, Hirofumi

2018-02-01

Alport syndrome (AS) is a hereditary, progressive nephritis caused by mutation of type IV collagen. Previous studies have shown that activation of signal transducer and activator of transcription 3 (STAT3) exacerbates other renal diseases, but whether STAT3 activation exacerbates AS pathology is still unknown. Here we aim to investigate the involvement of STAT3 in the progression of AS. Phosphorylated STAT3 expression was assessed by immunoblotting analysis of kidneys and glomeruli of an AS mouse model (Col4a5 G5X mutant). To determine the effect of blocking STAT3 signaling, we treated AS mice with the STAT3 inhibitor stattic (10 mg/kg i.p., three times per week for 10 weeks; n = 10). We assessed the renal function [proteinuria, blood urea nitrogen (BUN), serum creatinine] and analyzed the glomerular injury score, fibrosis and inflammatory cell invasion by histological staining. Moreover, we analyzed the gene expression of nephritis-associated molecules. Phosphorylated STAT3 was upregulated in AS kidneys and glomeruli. Treatment with stattic ameliorated the progressive renal dysfunction, such as increased levels of proteinuria, BUN and serum creatinine. Stattic also significantly suppressed the gene expression levels of renal injury markers (Lcn2, Kim-1), pro-inflammatory cytokines (Il-6, KC), pro-fibrotic genes (Tgf-β, Col1a1, α-Sma) and Mmp9. Stattic treatment decreased the renal fibrosis congruently with the decrease of transforming growth factor beta (TGF-β) protein and increase of antifibrosis-associated markers p-Smad1, 5 and 8, which are negative regulators of TGF-β signaling. STAT3 inhibition significantly ameliorated the renal dysfunction in AS mice. Our finding identifies STAT3 as an important regulator in AS progression and provides a promising therapeutic target for AS. © The Author 2017. Published by Oxford University Press on behalf of ERA-EDTA. All rights reserved.

STAT6 deficiency ameliorates Graves' disease severity by suppressing thyroid epithelial cell hyperplasia.

PubMed

Jiang, Xuechao; Zha, Bingbing; Liu, Xiaoming; Liu, Ronghua; Liu, Jun; Huang, Enyu; Qian, Tingting; Liu, Jiajing; Wang, Zhiming; Zhang, Dan; Wang, Luman; Chu, Yiwei

2016-12-01

Signal transducer and activator of transcription 6 (STAT6) is involved in epithelial cell growth. However, little is known regarding the STAT6 phosphorylation status in Graves' disease (GD) and its role in thyroid epithelial cells (TECs). In this study, we found that STAT6 phosphorylation (p-STAT6) was significantly increased in TECs from both GD patients and experimental autoimmune Graves' disease mice and that STAT6 deficiency ameliorated GD symptoms. Autocrine IL-4 signalling in TECs activated the phosphorylation of STAT6 via IL-4 R engagement, and the downstream targets of STAT6 were Bcl-xL and cyclin D1. Thus, the IL-4-STAT6-Bcl-xL/cyclin D1 pathway is crucial for TEC hyperplasia, which aggravates GD. More importantly, in vitro and in vivo experiments demonstrated that STAT6 phosphorylation inhibited by AS1517499 decreased TEC hyperplasia, thereby reducing serum T3 and T4 and ameliorating GD. Thus, our study reveals that in addition to the traditional pathogenesis of GD, in which autoantibody TRAb stimulates thyroid-stimulating hormone receptors and consequently produces T3, T4, TRAb could also trigger TECs producing IL-4, and IL-4 then acts in an autocrine manner to activate p-STAT6 signalling and stimulate unrestricted cell growth, thus aggravating GD. These findings suggest that STAT6 inhibitors could be potent therapeutics for treating GD.

STAT6 deficiency ameliorates Graves' disease severity by suppressing thyroid epithelial cell hyperplasia

PubMed Central

Jiang, Xuechao; Zha, Bingbing; Liu, Xiaoming; Liu, Ronghua; Liu, Jun; Huang, Enyu; Qian, Tingting; Liu, Jiajing; Wang, Zhiming; Zhang, Dan; Wang, Luman; Chu, Yiwei

2016-01-01

Signal transducer and activator of transcription 6 (STAT6) is involved in epithelial cell growth. However, little is known regarding the STAT6 phosphorylation status in Graves' disease (GD) and its role in thyroid epithelial cells (TECs). In this study, we found that STAT6 phosphorylation (p-STAT6) was significantly increased in TECs from both GD patients and experimental autoimmune Graves' disease mice and that STAT6 deficiency ameliorated GD symptoms. Autocrine IL-4 signalling in TECs activated the phosphorylation of STAT6 via IL-4 R engagement, and the downstream targets of STAT6 were Bcl-xL and cyclin D1. Thus, the IL-4-STAT6-Bcl-xL/cyclin D1 pathway is crucial for TEC hyperplasia, which aggravates GD. More importantly, in vitro and in vivo experiments demonstrated that STAT6 phosphorylation inhibited by AS1517499 decreased TEC hyperplasia, thereby reducing serum T3 and T4 and ameliorating GD. Thus, our study reveals that in addition to the traditional pathogenesis of GD, in which autoantibody TRAb stimulates thyroid-stimulating hormone receptors and consequently produces T3, T4, TRAb could also trigger TECs producing IL-4, and IL-4 then acts in an autocrine manner to activate p-STAT6 signalling and stimulate unrestricted cell growth, thus aggravating GD. These findings suggest that STAT6 inhibitors could be potent therapeutics for treating GD. PMID:27906181

Developmental expression of STATs, nuclear factor-κB and inflammatory genes in the jejunum of piglets during weaning.

PubMed

Yi, Hongbo; Jiang, Denghu; Zhang, Lin; Xiong, Haitao; Han, Feifei; Wang, Yizhen

2016-07-01

The signal transducer and activator of transcription (STAT) proteins play essential roles in apoptosis, proliferation and survival. However, the role of STATs in intestinal inflammation during weaning is unclear. This study aimed to investigate developmental expression of STATs, nuclear factor-κB (NF-κB) and inflammatory genes in the jejunum of piglets during weaning. Thirty-two piglets were weaned at 21d and sacrificed at 0, 1, 7, or 14d (n=8) after weaning. Villus height and the villus height/crypt depth ratio were decreased, whereas crypt depth was increased in the jejunum at 7 and 14d after weaning. In addition, the mRNA levels of interferon-γ (IFN-γ), inducible nitric oxide synthase (iNOS), IL-6, IL-8, IL-12 and IL-22 were increased in the jejunum at 7 and 14d after weaning, whereas transforming growth factor-β (TGF-β), suppressor of cytokine signaling 3 (SCOS3) and arginase-1 was decreased. Neutrophil infiltration was increased in the mucosa of the jejunum after weaning. Moreover, phosphorylation of IκB-α, NF-κB, AKT and STAT-3 was increased. However, the phosphorylation of STAT-1 (at 7 and 14d) and STAT-6 (at 1 and 7d) was suppressed in the jejunum after weaning. Treatment of porcine jejunal epithelial (IPEC-J2) cells with the STAT inhibitors fludarabine, niclosamide and teriflunomide, which inhibit the phosphorylation of STAT-1, STAT-3 and STAT-6, respectively, weakened the defense capacity of these cells against bacterial infection. In conclusion, weaning caused severe inflammation associated with activation of the NF-κB and STAT-3 pathways and suppression of STAT-1 and STAT-6 in the jejunum of piglets. Copyright © 2016 Elsevier B.V. All rights reserved.

RANKL downregulates cell surface CXCR6 expression through JAK2/STAT3 signaling pathway during osteoclastogenesis.

PubMed

Li, Changhong; Zhao, Jinxia; Sun, Lin; Yao, Zhongqiang; Liu, Rui; Huang, Jiansheng; Liu, Xiangyuan

2012-12-14

The receptor activator of nuclear factor-κB ligand (RANKL), as a member of the tumor necrosis factor (TNF) family, plays an essential role in osteoclast differentiation and function. Chemokines and their receptors have recently been shown to play critical roles in osteoclastogenesis, however, whether CXCL16-CXCR6 plays role in RANKL-mediated osteoclastogenesis is unknown. In this study, we first reported that RANKL decreased CXCR6 in a dose-dependent manner, which may be through deactivation of Akt and STAT3 signaling induced by CXCL16. Interestingly, RANKL-mediated CXCR6 reduction may be associated to the activation of STAT3 by phosphorylation. When STAT3 activation was blocked by JAK2/STAT3 inhibitor AG490, RANKL failed to shut down CXCR6 expression during osteoclastogenesis. However, CXCL16 alone did not augment RANKL-mediated osteoclast differentiation and did not alter RANKL-receptor RANK mRNA expression. These results demonstrate that reduction of CXCL16-CXCR6 is critical in RANKL-mediated osteoclastogenesis, which is mainly through the activation of JAK2/STAT3 signaling. CXCL16-CXCR6 axis may become a novel target for the therapeutic intervention of bone resorbing diseases such as rheumatoid arthritis and osteoporosis. Copyright © 2012 Elsevier Inc. All rights reserved.

Interleukin-22 induces hepatic stellate cell senescence and restricts liver fibrosis in mice.

PubMed

Kong, Xiaoni; Feng, Dechun; Wang, Hua; Hong, Feng; Bertola, Adeline; Wang, Fu-Sheng; Gao, Bin

2012-09-01

Interleukin (IL)-22 is known to play a key role in promoting antimicrobial immunity, inflammation, and tissue repair at barrier surfaces by binding to the receptors, IL-10R2 and IL-22R1. IL-22R1 is generally thought to be expressed exclusively in epithelial cells. In this study, we identified high levels of IL-10R2 and IL-22R1 expression on hepatic stellate cells (HSCs), the predominant cell type involved in liver fibrogenesis in response to liver damage. In vitro treatment with IL-22 induced the activation of signal transducer and activator of transcription (STAT) 3 in primary mouse and human HSCs. IL-22 administration prevented HSC apoptosis in vitro and in vivo, but surprisingly, the overexpression of IL-22 by either gene targeting (e.g., IL-22 transgenic mice) or exogenous administration of adenovirus expressing IL-22 reduced liver fibrosis and accelerated the resolution of liver fibrosis during recovery. Furthermore, IL-22 overexpression or treatment increased the number of senescence-associated beta-galactosidase-positive HSCs and decreased alpha-smooth muscle actin expression in fibrotic livers in vivo and cultured HSCs in vitro. Deletion of STAT3 prevented IL-22-induced HSC senescence in vitro, whereas the overexpression of a constitutively activated form of STAT3 promoted HSC senescence through p53- and p21-dependent pathways. Finally, IL-22 treatment up-regulated the suppressor of cytokine signaling (SOCS) 3 expression in HSCs. Immunoprecipitation analyses revealed that SOCS3 bound p53 and subsequently increased the expression of p53 and its target genes, contributing to IL-22-mediated HSC senescence. IL-22 induces the senescence of HSCs, which express both IL-10R2 and IL-22R1, thereby ameliorating liver fibrogenesis. The antifibrotic effect of IL-22 is likely mediated by the induction of HSC senescence, in addition to the previously discovered hepatoprotective functions of IL-22. Copyright © 2012 American Association for the Study of Liver Diseases.

Regulatory effect of calcineurin inhibitor, tacrolimus, on IL-6/sIL-6R-mediated RANKL expression through JAK2-STAT3-SOCS3 signaling pathway in fibroblast-like synoviocytes

PubMed Central

2013-01-01

Introduction This study investigated whether the calcineurin inhibitor, tacrolimus, suppresses receptor activator of NF-κB ligand (RANKL) expression in fibroblast-like synoviocytes (FLS) through regulation of IL-6/Janus activated kinase (JAK2)/signal transducer and activator of transcription-3 (STAT3) and suppressor of cytokine signaling (SOCS3) signaling. Methods The expression of RANKL, JAK2, STAT3, and SOCS3 proteins was assessed by western blot analysis, real-time PCR and ELISA in IL-6 combined with soluble IL-6 receptor (sIL-6R)-stimulated rheumatoid arthritis (RA)-FLS with or without tacrolimus treatment. The effects of tacrolimus on synovial inflammation and bone erosion were assessed using mice with arthritis induced by K/BxN serum. Immunofluorescent staining was performed to identify the effect of tacrolimus on RANKL and SOCS3. The tartrate-resistant acid phosphatase staining assay was performed to assess the effect of tacrolimus on osteoclast differentiation. Results We found that RANKL expression in RA FLS is regulated by the IL-6/sIL-6R/JAK2/STAT3/SOCS3 pathway. Inhibitory effects of tacrolimus on RANKL expression in a serum-induced arthritis mice model were identified. Tacrolimus inhibits RANKL expression in IL-6/sIL-6R-stimulated FLS by suppressing STAT3. Among negative regulators of the JAK/STAT pathway, such as CIS1, SOCS1, and SOCS3, only SOCS3 is significantly induced by tacrolimus. As compared to dexamethasone and methotrexate, tacrolimus more potently suppresses RANKL expression in FLS. By up-regulating SOCS3, tacrolimus down-regulates activation of the JAK-STAT pathway by IL-6/sIL-6R trans-signaling, thus decreasing RANKL expression in FLS. Conclusions These data suggest that tacrolimus might affect the RANKL expression in IL-6 stimulated FLS through STAT3 suppression, together with up-regulation of SOCS3. PMID:23406906

ATF3 activates Stat3 phosphorylation through inhibition of p53 expression in skin cancer cells.

PubMed

Hao, Zhen-Feng; Ao, Jun-Hong; Zhang, Jie; Su, You-Ming; Yang, Rong-Ya

2013-01-01

ATF3, a member of the ATF/CREB family of transcription factors, has been found to be selectively induced by calcineurin/NFAT inhibition and to enhance keratinocyte tumor formation, although the precise role of ATF3 in human skin cancer and possible mechanisms remain unknown. In this study, clinical analysis of 30 skin cancer patients and 30 normal donors revealed that ATF3 was accumulated in skin cancer tissues. Functional assays demonstrated that ATF3 significantly promoted skin cancer cell proliferation. Mechanically, ATF3 activated Stat3 phosphorylation in skin cancer cell through regulation of p53 expression. Moreover, the promotion effect of ATF3 on skin cancer cell proliferation was dependent on the p53-Stat3 signaling cascade. Together, the results indicate that ATF3 might promote skin cancer cell proliferation and enhance skin keratinocyte tumor development through inhibiting p53 expression and then activating Stat3 phosphorylation.

Molecular mechanisms of mucocutaneous immunity against Candida and Staphylococcus species.

PubMed

Maródi, László; Cypowyj, Sophie; Tóth, Beáta; Chernyshova, Liudmyla; Puel, Anne; Casanova, Jean-Laurent

2012-11-01

Signal transducer and activator of transcription (STAT) proteins are key components of the innate and adaptive immune responses to pathogenic microorganisms. Recent research on primary immunodeficiency disorders and the identification of patients carrying germline mutations in STAT1, STAT3, and STAT5B have highlighted the role of human STATs in host defense against various viruses, bacteria, and fungi. Mutations in STAT1 and STAT3 disrupt various cytokine pathways that control mucocutaneous immunity against Candida species, especially Candida albicans, and Staphylococcus species, especially Staphylococcus aureus. Here we consider inborn errors of immunity arising from mutations in either STAT1 or STAT3 that affect mucocutaneous immunity to Candida and Staphylococcus species. Copyright © 2012 American Academy of Allergy, Asthma & Immunology. Published by Mosby, Inc. All rights reserved.

Desensitization of the growth hormone-induced Janus kinase 2 (Jak 2)/signal transducer and activator of transcription 5 (Stat5)-signaling pathway requires protein synthesis and phospholipase C.

PubMed

Fernández, L; Flores-Morales, A; Lahuna, O; Sliva, D; Norstedt, G; Haldosén, L A; Mode, A; Gustafsson, J A

1998-04-01

Signal transducers and activators of transcription (Stat) proteins are latent cytoplasmic transcription factors that are tyrosine phosphorylated by Janus kinases (Jak) in response to GH and other cytokines. GH activates Stat5 by a mechanism that involves tyrosine phosphorylation and nuclear translocation. However, the mechanisms that turn off the GH-activated Jak2/Stat5 pathway are unknown. Continuous exposure to GH of BRL-4 cells, a rat hepatoma cell line stably transfected with rat GH receptor, induces a rapid but transient activation of Jak2 and Stat5. GH-induced Stat5 DNA-binding activity was detected after 2 min and reached a maximum at 10 min. Continued exposure to GH resulted in a desensitization characterized by 1) a rapid decrease in Stat5 DNA-binding activity. The rate of decrease of activity was rapid up to 1 h of GH treatment, and the remaining activity declined slowly thereafter. The activity of Stat5 present after 5 h is still higher than the control levels and almost 10-20% with respect to maximal activity at 10 min; and 2) the inability of further GH treatment to reinduce activation of Stat5. In contrast, with transient exposures of BRL-4 cells to GH, Stat5 DNA-binding activity could repeatedly be induced. GH-induced Jak2 and Stat5 activities were independent of ongoing protein synthesis. However, Jak2 tyrosine phosphorylation and Stat5 DNA-binding activity were prolonged for at least 4 h in the presence of cycloheximide, which suggests that the maintenance of desensitization requires ongoing protein synthesis. Furthermore, inhibition of protein synthesis potentiated GH-induced transcriptional activity in BRL-4 cells transiently transfected with SPIGLE1CAT, a reporter plasmid activated by Stat5. GH-induced Jak2 and Stat5 activation were not affected by D609 or mepacrine, both inhibitors of phospholipase C. However, in the presence of D609 and mepacrine, GH maintained prolonged Jak2 and Stat5 activation. Transactivation of SPIGLE1 by GH was potentiated by mepacrine and D609 but not by the phospholipase A2 inhibitor AACOCF3. Thus, a regulatory circuit of GH-induced transcription through the Jak2/Stat5-signaling pathway includes a prompt GH-induced activation of Jak2/Stat5 followed by a negative regulatory response; ongoing protein synthesis and intracellular signaling pathways, where phospholipase C activity is involved, play a critical role to desensitize the GH-activated Jak2/Stat5-signaling pathway.

Isolation and Expression Analysis of STAT Members from Synechogobius hasta and Their Roles in Leptin Affecting Lipid Metabolism

PubMed Central

Wu, Kun; Tan, Xiao-Ying; Wei, Chuan-Chuan; You, Wen-Jing; Zhuo, Mei-Qin; Song, Yu-Feng

2016-01-01

Signal transducers and activators of transcription proteins (STATs) act as important mediators in multiple biological processes induced by a large number of cytokines. In the present study, full-length cDNA sequences of seven STAT members, including some splicing variants different from those in mammals, were obtained from Synechogobius hasta. The phylogenetic analysis revealed that the seven STAT members were derived from paralogous genes that might have arisen by whole genome duplication (WGD) events during vertebrate evolution. All of these members share similar domain structure compared with those of mammals, and were widely expressed across the tested tissues (brain, gill, heart, intestine, liver, muscle and spleen), but at variable levels. Incubation in vitro of recombinant human leptin changed the intracellular triglyceride (TG) content and mRNA levels of several STATs members, as well as expressions and activities of genes involved in lipid metabolism. Furthermore, Tyrphostin B42 (AG490), a specific inhibitor of the Janus Kinase 2(JAK2)-STAT pathway, partially reversed leptin-induced change on STAT3 and its two spliced isoforms expression, as well as expressions and activities of genes involved in lipid metabolism. As a consequence, the decrease of TG content was also reversed. Thus, our study suggests that STAT3 is the requisite for the leptin signal and the activation of the STAT3 member may account for the leptin-induced changes in lipid metabolism in S. hasta. PMID:27011172

Isolation and Expression Analysis of STAT Members from Synechogobius hasta and Their Roles in Leptin Affecting Lipid Metabolism.

PubMed

Wu, Kun; Tan, Xiao-Ying; Wei, Chuan-Chuan; You, Wen-Jing; Zhuo, Mei-Qin; Song, Yu-Feng

2016-03-22

Signal transducers and activators of transcription proteins (STATs) act as important mediators in multiple biological processes induced by a large number of cytokines. In the present study, full-length cDNA sequences of seven STAT members, including some splicing variants different from those in mammals, were obtained from Synechogobius hasta. The phylogenetic analysis revealed that the seven STAT members were derived from paralogous genes that might have arisen by whole genome duplication (WGD) events during vertebrate evolution. All of these members share similar domain structure compared with those of mammals, and were widely expressed across the tested tissues (brain, gill, heart, intestine, liver, muscle and spleen), but at variable levels. Incubation in vitro of recombinant human leptin changed the intracellular triglyceride (TG) content and mRNA levels of several STATs members, as well as expressions and activities of genes involved in lipid metabolism. Furthermore, Tyrphostin B42 (AG490), a specific inhibitor of the Janus Kinase 2(JAK2)-STAT pathway, partially reversed leptin-induced change on STAT3 and its two spliced isoforms expression, as well as expressions and activities of genes involved in lipid metabolism. As a consequence, the decrease of TG content was also reversed. Thus, our study suggests that STAT3 is the requisite for the leptin signal and the activation of the STAT3 member may account for the leptin-induced changes in lipid metabolism in S. hasta.

Chronic sleep fragmentation during the sleep period induces hypothalamic endoplasmic reticulum stress and PTP1b-mediated leptin resistance in male mice.

PubMed

Hakim, Fahed; Wang, Yang; Carreras, Alba; Hirotsu, Camila; Zhang, Jing; Peris, Eduard; Gozal, David

2015-01-01

Sleep fragmentation (SF) is highly prevalent and may constitute an important contributing factor to excessive weight gain and the metabolic syndrome. Increased endoplasmic reticulum (ER) stress and activation of the unfolded protein response (UPR) leading to the attenuation of leptin receptor signaling in the hypothalamus leads to obesity and metabolic dysfunction. Mice were exposed to SF and sleep control (SC) for varying periods of time during which ingestive behaviors were monitored. UPR pathways and leptin receptor signaling were assessed in hypothalami. To further examine the mechanistic role of ER stress, changes in leptin receptor (ObR) signaling were also examined in wild-type mice treated with the ER chaperone tauroursodeoxycholic acid (TUDCA), as well as in CHOP-/+ transgenic mice. Fragmented sleep in male mice induced increased food intake starting day 3 and thereafter, which was preceded by increases in ER stress and activation of all three UPR pathways in the hypothalamus. Although ObR expression was unchanged, signal transducer and activator of transcription 3 (STAT3) phosphorylation was decreased, suggesting reduced ObR signaling. Unchanged suppressor of cytokine signaling-3 (SOCS3) expression and increases in protein-tyrosine phosphatase 1B (PTP1B) expression and activity emerged with SF, along with reduced p-STAT3 responses to exogenous leptin. SF-induced effects were reversed following TUDCA treatment and were absent in CHOP -/+ mice. SF induces hyperphagic behaviors and reduced leptin signaling in hypothalamus that are mediated by activation of ER stress, and ultimately lead to increased PTP1B activity. ER stress pathways are therefore potentially implicated in SF-induced weight gain and metabolic dysfunction, and may represent a viable therapeutic target. © 2014 Associated Professional Sleep Societies, LLC.

Subcellular localization and functional characterization of a fish IRF9 from crucian carp Carassius auratus.

PubMed

Shi, Jun; Zhang, Yi-Bing; Liu, Ting-Kai; Sun, Fan; Gui, Jian-Fang

2012-08-01

Mammalian interferon (IFN) regulatory factor 9 (IRF-9) has long been recognized as the DNA sequence recognition subunit of IFN-stimulated gene factor 3 (ISGF3) complex, which is critical for type I IFN to induce the expression of IFN-stimulated genes (ISGs) against viral infection. Recent studies have shown that fish IFN exerts antiviral effects by induction of a number of ISGs and also of itself; however, little is known about the role of fish IRF9 in IFN signaling. Here we identify a fish IRF9 orthologue (CaIRF9) from IFN-producing cell line, crucian carp Carassius auratus blastulae embryonic (CAB) cells. Analysis of subcellular distribution of CaIRF9-green fluorescent protein indicates that CaIRF9 is constitutively present in the nucleus, which is driven by two nuclear localization signals (NLS), one locating within DNA-binding domain (DBD) of CaIRF9 and the other immediately behind DBD, although human IRF9 contains only one NLS analogous to the former of CaIRF9. Overexpression of CaIRF9 together with CaSTAT2 not only activates ISRE-containing promoter but also upregulates the expression of fish ISGs. Strikingly, CaIRF9 together with CaSTAT2 also exhibits an ability to activate crucian carp IFN promoter, and blockade of cellular CaIRF9 attenuates IFN itself-induced activation of crucian carp IFN promoter. Taken together, these data suggest that crucian carp IFN induces the expression of ISGs and also of itself possibly by the JAK-STAT signaling pathway that is conserved from fish to mammals. Copyright © 2012 Elsevier Ltd. All rights reserved.

Feed-Forward Reciprocal Activation of PAFR and STAT3 Regulates Epithelial-Mesenchymal Transition in Non-Small Cell Lung Cancer.

PubMed

Chen, Jie; Lan, Tian; Zhang, Weimin; Dong, Lijia; Kang, Nan; Zhang, Shumin; Fu, Ming; Liu, Bing; Liu, Kangtai; Zhan, Qimin

2015-10-01

Platelet-activating factor receptor (PAFR), a G-protein-coupled receptor, has been implicated in tumorigenesis, but its contributions to metastatic progression have not been investigated. Here, we show that PAFR is overexpressed in non-small cell lung cancer (NSCLC) as well as in breast, colorectal, and gastric carcinomas. Expression of PAFR correlates closely with clinical stages, survival time, and distant metastasis. In human NSCLC cells, activation of the PAF/PAFR signaling axis accentuated malignant character, including by stimulating epithelial-mesenchymal transition (EMT). In contrast, silencing PAFR in aggressive NSCLC cells inhibited these effects. Mechanistic investigations showed that PAFR stimulated EMT by activating STAT3 via upregulation of G-protein-dependent SRC or JAK2 kinase activity. Notably, STAT3 transcriptionally elevated PAFR expression. Thus, activation of PAFR in NSCLC cells initiated a forward feedback loop responsible for mediating the aggressive malignant character of NSCLC cells in vitro and in vivo. Reinforcing this reciprocal activation loop, PAF/PAFR signaling also upregulated IL6 expression and thereby STAT3 activation. Overall, our results elucidated an important role for PAFR dysregulation in the pathogenicity of NSCLC and unraveled a forward feedback loop between PAFR and STAT3 that acts to drive the malignant progression of NSCLC. ©2015 American Association for Cancer Research.

Withaferin A down-regulates lipopolysaccharide-induced cyclooxygenase-2 expression and PGE2 production through the inhibition of STAT1/3 activation in microglial cells.

PubMed

Min, Kyoung-Jin; Choi, Kyounghwa; Kwon, Taeg Kyu

2011-08-01

Microglia are the major immune effector cells in the brain, and microglia activated by injury and infection can produce inflammatory mediators. A number of studies have reported that withaferin A has anti-inflammatory functions. However, the effects of withaferin A on the microglial inflammatory response have not been investigated. Our results show that withaferin A inhibited lipopolysaccharide (LPS)-induced cyclooxygenase (COX)-2 mRNA and protein expression and prostaglandin E2 (PGE(2)) production in BV2 murine microglial cells. Withaferin A had no effect on LPS-induced Akt and ERK phosphorylation, but phosphorylation of p38 and JNK was slightly decreased by withaferin A. Withaferin A significantly inhibited LPS-induced STAT1 and STAT3 phosphorylation in a dose-dependent manner. Furthermore, withaferin A inhibited nuclear translocation of STAT1 and interferon-gamma activated sequence (GAS)-promoter activity. Taken together, these results suggest that withaferin A inhibits LPS-induced PGE(2) production and COX-2 expression, at least in part, by blocking STAT1 and STAT3 activation. Copyright © 2011 Elsevier B.V. All rights reserved.

Glycogen Synthase Kinase-3 Is an Early Determinant in the Differentiation of Pathogenic Th17 Cells

PubMed Central

Beurel, Eléonore; Yeh, Wen-I; Michalek, Suzanne M.; Harrington, Laurie E.; Jope, Richard S.

2011-01-01

CD4+ T cells are critical for host defense but are also major drivers of immune-mediated diseases. The classical view of Th1 and Th2 subtypes of CD4+ T cells was recently revised by the identification of the Th17 lineage of CD4+ T cells that produce IL-17, which have been found to be critical in the pathogenesis of autoimmune and other diseases. Mechanisms controlling the differentiation of Th17 cells have been well described, but few feasible targets for therapeutically reducing Th17 cells are known. The generation of Th17 cells requires IL-6 and activation of STAT3. During polarization of CD4+ T cells to Th17 cells, we found that inhibition of glycogen synthase kinase-3 (GSK3) blocked IL-6 production, STAT3 activation, and polarization to Th17 cells. Polarization of CD4+ T cells to Th17 cells increased by 10-fold the expression of GSK3β protein levels in Th17 cells, whereas GSK3β was unaltered in regulatory T cells. Diminishing GSK3 activity either pharmacologically or molecularly blocked Th17 cell production, and increasing GSK3 activity promoted polarization to Th17 cells. In vivo inhibition of GSK3 in mice depleted constitutive Th17 cells in intestinal mucosa, blocked Th17 cell generation in the lung after Francisella tularensis infection, and inhibited the increase in spinal cord Th17 cells and disease symptoms in the experimental autoimmune encephalomyelitis mouse model of multiple sclerosis. These findings identify GSK3 as a critical mediator of Th17 cell production and indicate that GSK3 inhibitors provide a potential therapeutic intervention to control Th17-mediated diseases. PMID:21191064

Histidine augments the suppression of hepatic glucose production by central insulin action.

PubMed

Kimura, Kumi; Nakamura, Yusuke; Inaba, Yuka; Matsumoto, Michihiro; Kido, Yoshiaki; Asahara, Shun-Ichiro; Matsuda, Tomokazu; Watanabe, Hiroshi; Maeda, Akifumi; Inagaki, Fuyuhiko; Mukai, Chisato; Takeda, Kiyoshi; Akira, Shizuo; Ota, Tsuguhito; Nakabayashi, Hajime; Kaneko, Shuichi; Kasuga, Masato; Inoue, Hiroshi

2013-07-01

Glucose intolerance in type 2 diabetes is related to enhanced hepatic glucose production (HGP) due to the increased expression of hepatic gluconeogenic enzymes. Previously, we revealed that hepatic STAT3 decreases the expression of hepatic gluconeogenic enzymes and suppresses HGP. Here, we show that increased plasma histidine results in hepatic STAT3 activation. Intravenous and intracerebroventricular (ICV) administration of histidine-activated hepatic STAT3 reduced G6Pase protein and mRNA levels and augmented HGP suppression by insulin. This suppression of hepatic gluconeogenesis by histidine was abolished by hepatic STAT3 deficiency or hepatic Kupffer cell depletion. Inhibition of HGP by histidine was also blocked by ICV administration of a histamine H1 receptor antagonist. Therefore, histidine activates hepatic STAT3 and suppresses HGP via central histamine action. Hepatic STAT3 phosphorylation after histidine ICV administration was attenuated in histamine H1 receptor knockout (Hrh1KO) mice but not in neuron-specific insulin receptor knockout (NIRKO) mice. Conversely, hepatic STAT3 phosphorylation after insulin ICV administration was attenuated in NIRKO but not in Hrh1KO mice. These findings suggest that central histidine action is independent of central insulin action, while both have additive effects on HGP suppression. Our results indicate that central histidine/histamine-mediated suppression of HGP is a potential target for the treatment of type 2 diabetes.

Histidine Augments the Suppression of Hepatic Glucose Production by Central Insulin Action

PubMed Central

Kimura, Kumi; Nakamura, Yusuke; Inaba, Yuka; Matsumoto, Michihiro; Kido, Yoshiaki; Asahara, Shun-ichiro; Matsuda, Tomokazu; Watanabe, Hiroshi; Maeda, Akifumi; Inagaki, Fuyuhiko; Mukai, Chisato; Takeda, Kiyoshi; Akira, Shizuo; Ota, Tsuguhito; Nakabayashi, Hajime; Kaneko, Shuichi; Kasuga, Masato; Inoue, Hiroshi

2013-01-01

Glucose intolerance in type 2 diabetes is related to enhanced hepatic glucose production (HGP) due to the increased expression of hepatic gluconeogenic enzymes. Previously, we revealed that hepatic STAT3 decreases the expression of hepatic gluconeogenic enzymes and suppresses HGP. Here, we show that increased plasma histidine results in hepatic STAT3 activation. Intravenous and intracerebroventricular (ICV) administration of histidine-activated hepatic STAT3 reduced G6Pase protein and mRNA levels and augmented HGP suppression by insulin. This suppression of hepatic gluconeogenesis by histidine was abolished by hepatic STAT3 deficiency or hepatic Kupffer cell depletion. Inhibition of HGP by histidine was also blocked by ICV administration of a histamine H1 receptor antagonist. Therefore, histidine activates hepatic STAT3 and suppresses HGP via central histamine action. Hepatic STAT3 phosphorylation after histidine ICV administration was attenuated in histamine H1 receptor knockout (Hrh1KO) mice but not in neuron-specific insulin receptor knockout (NIRKO) mice. Conversely, hepatic STAT3 phosphorylation after insulin ICV administration was attenuated in NIRKO but not in Hrh1KO mice. These findings suggest that central histidine action is independent of central insulin action, while both have additive effects on HGP suppression. Our results indicate that central histidine/histamine-mediated suppression of HGP is a potential target for the treatment of type 2 diabetes. PMID:23474485

The JAK-STAT signaling pathway: input and output integration.

PubMed

Murray, Peter J

2007-03-01

Universal and essential to cytokine receptor signaling, the JAK-STAT pathway is one of the best understood signal transduction cascades. Almost 40 cytokine receptors signal through combinations of four JAK and seven STAT family members, suggesting commonality across the JAK-STAT signaling system. Despite intense study, there remain substantial gaps in understanding how the cascades are activated and regulated. Using the examples of the IL-6 and IL-10 receptors, I will discuss how diverse outcomes in gene expression result from regulatory events that effect the JAK1-STAT3 pathway, common to both receptors. I also consider receptor preferences by different STATs and interpretive problems in the use of STAT-deficient cells and mice. Finally, I consider how the suppressor of cytokine signaling (SOCS) proteins regulate the quality and quantity of STAT signals from cytokine receptors. New data suggests that SOCS proteins introduce additional diversity into the JAK-STAT pathway by adjusting the output of activated STATs that alters downstream gene activation.

Silibinin downregulates MMP2 expression via Jak2/STAT3 pathway and inhibits the migration and invasive potential in MDA-MB-231 cells.

PubMed

Byun, Hyo Joo; Darvin, Pramod; Kang, Dong Young; Sp, Nipin; Joung, Youn Hee; Park, Jong Hwan; Kim, Sun Jin; Yang, Young Mok

2017-06-01

Worldwide, breast cancer (BCa) is the most common cancer in women. Among its subtypes, triple-negative breast cancer (TNBC) is an aggressive form associated with diminished survival. TNBCs are characterized by their absence, or minimal expression, of the estrogen and progesterone receptors, as well as the human epidermal growth factor receptor 2 (i.e. ER-/-, PR-/-, Her2-/Low). Consequently, treatment for this subtype of BCa remains problematic. Silibinin, a derivative of the flavonoid silymarin, is reported to have anticancer activities against hepatic and non-small cell lung cancers. We hypothesized that silibinin might inhibit cell-extracellular matrix interactions via the regulation, expression, and activation of STAT3 in TNBCs, which could directly inhibit metastasis in silibinin-treated BCa cells. Using proliferation assays, we found that exposure to silibinin at a concentration of 200 µM inhibited the proliferation of breast cancer (BCa) cells; this concentration also inhibited phosphorylation of STAT3 and its principal upstream kinase, Jak2. Furthermore, we found that silibinin inhibited the nuclear translocation of STAT3, as well as its binding to the MMP2 gene promoter. The ability of silibinin to inhibit metastasis was further studied using an in vitro invasion assay. The results confirm the role of STAT3 as a critical mediator in the invasive potential of BCa cells, and STAT3 knock-down resulted in inhibition of invasion. The invasion ability of silibinin-treated BCa cells was studied in detail with the expression of MMP2. Prevention of STAT3 activation also resulted in the inhibition of MMP2 expression. Use of a small interfering RNA to knock down STAT3 (siSTAT3) allowed us to confirm the role of STAT3 in regulating MMP2 expression, as well as the mechanism of action of silibinin in inhibiting MMP2. Taken together, we found that silibinin inhibits the Jak2/STAT3/MMP2 signaling pathway, and inhibits the proliferation, migration, and invasion of triple-negative BCa cells.

Cardiac preconditioning with sphingosine-1-phosphate requires activation of signal transducer and activator of transcription-3

PubMed Central

Kelly-Laubscher, Roisin F; King, Jonathan C; Hacking, Damian; Somers, Sarin; Hastie, Samantha; Stewart, Tessa; Imamdin, Aqeela; Maarman, Gerald; Pedretti, Sarah; Lecour, Sandrine

2014-01-01

Summary Aims Sphingosine-1-phosphate (S1P) is a cardioprotective agent. Signal transducer and activator of transcription 3 (STAT-3) is a key mediator of many cardioprotective agents. We aimed to explore whether STAT-3 is a key mediator in S1P-induced preconditioning. Methods Langendorff-perfused hearts from Wistar rats and wild-type or cardiomyocyte-specific STAT-3 knockout mice were pre-treated with S1P (10 nmol/l), with or without the STAT-3 pathway inhibitor AG490, before an ischaemia–reperfusion insult. Triphenyltetrazolium chloride and Evans blue staining were used for the determination of infarct size. Western blot analysis was carried out on the S1P pre-treated hearts for detection of cytosolic, nuclear and mitochondrial phosphorylated and total STAT-3 proteins. Results Pre-treatment with S1P decreased the infarct size in isolated rat (5 ± 3% vs control 26 ± 8%, p < 0.01) and wild-type mouse hearts (13 ± 1% vs control 33 ± 3%, p < 0.05). This protective effect was abolished in the rat hearts pre-treated with AG490 (30 ± 10%, p = ns vs control) and in the hearts from STAT-3 knockout mice (35 ± 4% vs control 30 ± 3%, p = ns). Levels of phosphorylated STAT-3 were significantly increased in both the nuclear (p < 0.05 vs control) and mitochondrial (p < 0.05 vs control) fractions in the S1P pre-treated hearts, but remained unchanged in the cytosolic fraction (p = ns vs control). Conclusion These novel results demonstrate that pharmacological preconditioning with S1P in the isolated heart is mediated by activation of mitochondrial and nuclear STAT-3, therefore suggesting that S1P may be a novel therapeutic target to modulate mitochondrial and nuclear function in cardiovascular disease in order to protect the heart against ischaemia–reperfusion. PMID:25000441

IL-6-Mediated Activation of Stat3α Prevents Trauma/Hemorrhagic Shock-Induced Liver Inflammation

PubMed Central

Moran, Ana; Thacker, Stephen A.; Arikan, Ayse Akcan; Mastrangelo, Mary-Ann A.; Wu, Yong; Yu, Bi; Tweardy, David J.

2011-01-01

Trauma complicated by hemorrhagic shock (T/HS) is the leading cause of morbidity and mortality in the United States for individuals under the age of 44 years. Initial survivors are susceptible to developing multiple organ failure (MOF), which is thought to be caused, at least in part, by excessive or maladaptive activation of inflammatory pathways. We previously demonstrated in rodents that T/HS results in liver injury that can be prevented by IL-6 administration at the start of resuscitation; however, the contribution of the severity of HS to the extent of liver injury, whether or not resuscitation is required, and the mechanism(s) for the IL-6 protective effect have not been reported. In the experiments described here, we demonstrated that the extent of liver inflammation induced by T/HS depends on the duration of hypotension and requires resuscitation. We established that IL-6 administration at the start of resuscitation is capable of completely reversing liver inflammation and is associated with increased Stat3 activation. Global assessment of the livers showed that the main effect of IL-6 was to normalize the T/HS-induced inflammation transcriptome. Pharmacological inhibition of Stat3 activity within the liver blocked the ability of IL-6 to prevent liver inflammation and to normalize the T/HS-induced liver inflammation transcriptome. Genetic deletion of a Stat3β, a naturally occurring, dominant-negative isoform of the Stat3, attenuated T/HS-induced liver inflammation, confirming a role for Stat3, especially Stat3α, in preventing T/HS-mediated liver inflammation. Thus, T/HS-induced liver inflammation depends on the duration of hypotension and requires resuscitation; IL-6 administration at the start of resuscitation reverses T/HS-induced liver inflammation, through activation of Stat3α, which normalized the T/HS-induced liver inflammation transcriptome. PMID:21738667

STAT3 activation by IL-6 from mesenchymal stem cells promotes the proliferation and metastasis of osteosarcoma.

PubMed

Tu, Bing; Du, Lin; Fan, Qi-Ming; Tang, Ze; Tang, Ting-Ting

2012-12-01

We previously demonstrated that human mesenchymal stem cells (MSCs) promote the growth of osteosarcoma in the bone microenvironment. The aim of the present study was to further determine the effect of IL-6/STAT3 signaling on the progression of osteosarcoma. First, conditioned medium from MSCs was used to stimulate the growth of osteosarcoma cells (Saos-2) in vitro. We found that STAT3 was activated and that the activation could be blocked by an IL-6-neutralizing antibody. The inhibition of STAT3 in Saos-2 cells by siRNA or AG490 decreased cell proliferation, migration and invasion, down-regulated the mRNA expression of Cyclin D, Bcl-xL and Survivin and enhanced the apoptotic response. Furthermore, a nude mouse osteosarcoma model was established by injecting luciferase-labeled Saos-2 cells into the tibia, and the effect of STAT3 on tumor growth was determined by treating the mice with AG490. In vivo bioluminescence images showed that tumor growth was dramatically reduced in the AG490 group. In addition, STAT3 inhibition decreased the lung metastasis rate and prolonged the survival of these mice. After treatment with AG490, the protein levels of IL-6, p-STAT3 and PCNA were decreased, and the level of apoptosis in the tumor was increased. Altogether, these data indicate that MSCs in the bone microenvironment might promote the progression of osteosarcoma and protect tumor cells from drug-induced apoptosis through IL-6/STAT3 signaling. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Quercetin abrogates IL-6/STAT3 signaling and inhibits glioblastoma cell line growth and migration

DOE Office of Scientific and Technical Information (OSTI.GOV)

Michaud-Levesque, Jonathan; Bousquet-Gagnon, Nathalie; Beliveau, Richard, E-mail: oncomol@nobel.si.uqam.ca

Evidence has suggested that STAT3 functions as an oncogene in gliomagenesis. As a consequence, changes in the inflammatory microenvironment are thought to promote tumor development. Regardless of its origin, cancer-related inflammation has many tumor-promoting effects, such as the promotion of cell cycle progression, cell proliferation, cell migration and cell survival. Given that IL-6, a major cancer-related inflammatory cytokine, regulates STAT3 activation and is upregulated in glioblastoma, we sought to investigate the inhibitory effects of the chemopreventive flavonoid quercetin on glioblastoma cell proliferation and migration triggered by IL-6, and to determine the underlying mechanisms of action. In this study, we showmore » that quercetin is a potent inhibitor of the IL-6-induced STAT3 signaling pathway in T98G and U87 glioblastoma cells. Exposure to quercetin resulted in the reduction of GP130, JAK1 and STAT3 activation by IL-6, as well as a marked decrease of the proliferative and migratory properties of glioblastoma cells induced by IL-6. Interestingly, quercetin also modulated the expression of two target genes regulated by STAT3, i.e. cyclin D1 and matrix metalloproteinase-2 (MMP-2). Moreover, quercetin reduced the recruitment of STAT3 at the cyclin D1 promoter and inhibited Rb phosphorylation in the presence of IL-6. Overall, these results provide new insight into the role of quercetin as a blocker of the STAT3 activation pathway stimulated by IL-6, with a potential role in the prevention and treatment of glioblastoma.« less

Interleukin 6 inhibits proliferation and, in cooperation with an epidermal growth factor receptor autocrine loop, increases migration of T47D breast cancer cells.

PubMed

Badache, A; Hynes, N E

2001-01-01

Interleukin (IL)-6, a multifunctional regulator of immune response, hematopoiesis, and acute phase reactions, has also been shown to regulate cancer cell proliferation. We have investigated IL-6 signaling pathways and cellular responses in the T47D breast carcinoma cell line. The IL-6-type cytokines, IL-6 and oncostatin M, simultaneously inhibited cell proliferation and increased cell migration. In T47D cells, IL-6 stimulated the activation of Janus-activated kinase 1 tyrosine kinase and signal transducers and activators of transcription (STAT) 1 and STAT3 transcription factors. Expression of dominant negative STAT3 in the cells strongly reduced IL-6-mediated growth inhibition but did not prevent IL-6-induced cell migration. IL-6 treatment led to activation of the mitogen-activated protein kinase (MAPK) and the phosphatidylinositol 3'-kinase (PI3K) pathways. Inhibition of MAPK or PI3K activity reversed IL-6- and oncostatin M-stimulated migration. Because cross-talk between cytokine receptors and members of the ErbB family of receptor tyrosine kinases has been described previously, we have examined their interaction in T47D cells. Down-regulation of ErbB receptor activity, through the use of specific pharmacological inhibitors or dominant negative receptor constructs, revealed that IL-6-induced MAPK activation was largely dependent on epidermal growth factor (EGF) receptor activity, but not on ErbB-2 activity. Using a monoclonal antibody that interferes with EGF receptor-ligand interaction, we have shown that in T47D cells, IL-6 cooperates with an EGF receptor autocrine activity loop for signaling through the MAPK and PI3K pathways and for cell migration. Both the tyrosine phosphatase SHP-2 and the multisubstrate docking molecule Gab1, which are potential links between IL-6 and the MAPK/PI3K pathways, were constitutively associated with the active EGF receptor. On IL-6 stimulation, SHP-2 and Gab1 were recruited to the gp130 subunit of the IL-6 receptor and tyrosine phosphorylated, allowing downstream signaling to the MAPK and PI3K pathways. Thus, in T47D breast carcinoma cells, IL-6 acts in synergy with EGF receptor autocrine activity to signal through the MAPK/PI3K pathways. Cooperation between IL-6 and the EGF receptor in T47D breast carcinoma cells illustrates how a combination of multiple stimuli, either exogenous or endogenous, may result in synergistic cellular responses.

Serine protease allergen favours Th2 responses via PAR-2 and STAT3 activation in murine model.

PubMed

Agrawal, K; Arora, N

2018-03-01

Protease activity of Per a 10 favours Th2 responses by differential regulation of IL-12p70 and IL-23 cytokine subunits. This study aimed to elucidate the underlying mechanism of differential regulation of IL-12p70 and IL-23. PAR-2 activation was blocked in murine model by administering SAM11 before each sensitization. CD11c + p-STAT3 + cells were measured in lungs by flow cytometry. BMDCs were pretreated with SAM11 or isotype control or stattic and stimulated with Per a 10. p-STAT3 levels were measured using Western blot. Transcript levels of IL-12p35, IL-12/23p40 and IL-23p19 were measured using RT-PCR. Cytokine levels were analysed using ELISA. Protease activity of Per a 10 increased p-STAT3 levels in mouse lungs, which was reduced upon PAR-2 blockage. Percentage of p-STAT3 + CD11c + cells was higher in Per a 10-administered mice and was reduced upon PAR-2 blockage. IL-12p35 and IL-12p70 levels were higher, and IL-23p19 and IL-23 levels were lower in both SAM11-treated mice and BMDCs indicating a role of PAR-2-mediated signalling. IL-4, TSLP, IL-17A, EPO activity, total cell count and specific IgE and IgG1 levels were lower in SAM11-administered mice. Inhibiting STAT3 activation via stattic also leads to lower levels of IL-23p19 and IL-23 and higher levels of IL-12p35. Per a 10 leads to PAR-2 activation on BMDCs resulting in downstream activation of STAT3 to regulate the balance between IL-12/IL-23 subunits causing a cytokine milieu rich in IL-23 to favour Th2 polarization. © 2017 EAACI and John Wiley and Sons A/S. Published by John Wiley and Sons Ltd.

The regulation of inflammation by interferons and their STATs.

PubMed

Rauch, Isabella; Müller, Mathias; Decker, Thomas

2013-01-01

Interferons (IFN) are subdivided into type I IFN (IFN-I, here synonymous with IFN-α/β), type II (IFN-γ) and type III IFN (IFN-III/IFN-λ) that reprogram nuclear gene expression through STATs 1 and 2 by forming STAT1 dimers (mainly IFN-γ) or the ISGF3 complex, a STAT1-STAT2-IRF9 heterotrimer (IFN-I and IFN-III). Dominant IFN activities in the immune system are to protect cells from viral replication and to activate macrophages for enhanced effector function. However, the impact of IFN and their STATs on the immune system stretches far beyond these activities and includes the control of inflammation. The goal of this review is to give an overview of the different facets of the inflammatory process that show regulatory input by IFN/STAT.

Rhodium(II) Proximity-Labeling Identifies a Novel Target Site on STAT3 for Inhibitors with Potent Anti-Leukemia Activity.

PubMed

Minus, Matthew B; Liu, Wei; Vohidov, Farrukh; Kasembeli, Moses M; Long, Xin; Krueger, Michael J; Stevens, Alexandra; Kolosov, Mikhail I; Tweardy, David J; Sison, Edward Allan R; Redell, Michele S; Ball, Zachary T

2015-10-26

Nearly 40 % of children with acute myeloid leukemia (AML) suffer relapse arising from chemoresistance, often involving upregulation of the oncoprotein STAT3 (signal transducer and activator of transcription 3). Herein, rhodium(II)-catalyzed, proximity-driven modification identifies the STAT3 coiled-coil domain (CCD) as a novel ligand-binding site, and we describe a new naphthalene sulfonamide inhibitor that targets the CCD, blocks STAT3 function, and halts its disease-promoting effects in vitro, in tumor growth models, and in a leukemia mouse model, validating this new therapeutic target for resistant AML. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Gain-of-function human STAT1 mutations impair IL-17 immunity and underlie chronic mucocutaneous candidiasis

PubMed Central

Liu, Luyan; Okada, Satoshi; Kong, Xiao-Fei; Kreins, Alexandra Y.; Cypowyj, Sophie; Abhyankar, Avinash; Toubiana, Julie; Itan, Yuval; Audry, Magali; Nitschke, Patrick; Masson, Cécile; Toth, Beata; Flatot, Jérome; Migaud, Mélanie; Chrabieh, Maya; Kochetkov, Tatiana; Bolze, Alexandre; Borghesi, Alessandro; Toulon, Antoine; Hiller, Julia; Eyerich, Stefanie; Eyerich, Kilian; Gulácsy, Vera; Chernyshova, Ludmyla; Chernyshov, Viktor; Bondarenko, Anastasia; María Cortés Grimaldo, Rosa; Blancas-Galicia, Lizbeth; Madrigal Beas, Ileana Maria; Roesler, Joachim; Magdorf, Klaus; Engelhard, Dan; Thumerelle, Caroline; Burgel, Pierre-Régis; Hoernes, Miriam; Drexel, Barbara; Seger, Reinhard; Kusuma, Theresia; Jansson, Annette F.; Sawalle-Belohradsky, Julie; Belohradsky, Bernd; Jouanguy, Emmanuelle; Bustamante, Jacinta; Bué, Mélanie; Karin, Nathan; Wildbaum, Gizi; Bodemer, Christine; Lortholary, Olivier; Fischer, Alain; Blanche, Stéphane; Al-Muhsen, Saleh; Reichenbach, Janine; Kobayashi, Masao; Rosales, Francisco Espinosa; Lozano, Carlos Torres; Kilic, Sara Sebnem; Oleastro, Matias; Etzioni, Amos; Traidl-Hoffmann, Claudia; Renner, Ellen D.; Abel, Laurent; Picard, Capucine; Maródi, László; Boisson-Dupuis, Stéphanie

2011-01-01

Chronic mucocutaneous candidiasis disease (CMCD) may be caused by autosomal dominant (AD) IL-17F deficiency or autosomal recessive (AR) IL-17RA deficiency. Here, using whole-exome sequencing, we identified heterozygous germline mutations in STAT1 in 47 patients from 20 kindreds with AD CMCD. Previously described heterozygous STAT1 mutant alleles are loss-of-function and cause AD predisposition to mycobacterial disease caused by impaired STAT1-dependent cellular responses to IFN-γ. Other loss-of-function STAT1 alleles cause AR predisposition to intracellular bacterial and viral diseases, caused by impaired STAT1-dependent responses to IFN-α/β, IFN-γ, IFN-λ, and IL-27. In contrast, the 12 AD CMCD-inducing STAT1 mutant alleles described here are gain-of-function and increase STAT1-dependent cellular responses to these cytokines, and to cytokines that predominantly activate STAT3, such as IL-6 and IL-21. All of these mutations affect the coiled-coil domain and impair the nuclear dephosphorylation of activated STAT1, accounting for their gain-of-function and dominance. Stronger cellular responses to the STAT1-dependent IL-17 inhibitors IFN-α/β, IFN-γ, and IL-27, and stronger STAT1 activation in response to the STAT3-dependent IL-17 inducers IL-6 and IL-21, hinder the development of T cells producing IL-17A, IL-17F, and IL-22. Gain-of-function STAT1 alleles therefore cause AD CMCD by impairing IL-17 immunity. PMID:21727188

STAT3 targeting by polyphenols: Novel therapeutic strategy for melanoma.

PubMed

Momtaz, Saeideh; Niaz, Kamal; Maqbool, Faheem; Abdollahi, Mohammad; Rastrelli, Luca; Nabavi, Seyed Mohammad

2017-05-06

Melanoma or malignant melanocytes appear with the low incidence rate, but very high mortality rate worldwide. Epidemiological studies suggest that polyphenolic compounds contribute for prevention or treatment of several cancers particularly melanoma. Such findings motivate to dig out novel therapeutic strategies against melanoma, including research toward the development of new chemotherapeutic and biologic agents that can target the tumor cells by different mechanisms. Recently, it has been found that signal transducer and activator of transcription 3 (STAT3) is activated in many cancer cases surprisingly. Different evidences supply the aspect that STAT3 activation plays a vital role in the metastasis, including proliferation of cells, survival, invasion, migration, and angiogenesis. This significant feature plays a vital role in various cellular processes, such as cell proliferation and survival. Here, we reviewed the mechanisms of the STAT3 pathway regulation and their role in promoting melanoma. Also, we have evaluated the emerging data on polyphenols (PPs) specifically their contribution in melanoma therapies with an emphasis on their regulatory/inhibitory actions in relation to STAT3 pathway and current progress in the development of phytochemical therapeutic techniques. An understanding of targeting STAT3 by PPs brings an opportunity to melanoma therapy. © 2016 BioFactors, 43(3):347-370, 2017. © 2016 International Union of Biochemistry and Molecular Biology.

IbeA and OmpA of Escherichia coli K1 Exploit Rac1 Activation for Invasion of Human Brain Microvascular Endothelial Cells

PubMed Central

Maruvada, Ravi

2012-01-01

Meningitis-causing Escherichia coli K1 internalization of the blood-brain barrier is required for penetration into the brain, but the host-microbial interactions involved in E. coli entry of the blood-brain barrier remain incompletely understood. We show here that a meningitis-causing E. coli K1 strain RS218 activates Rac1 (GTP-Rac1) of human brain microvascular endothelial cells (HBMEC) in a time-dependent manner. Both activation and bacterial invasion were significantly inhibited in the presence of a Rac1 inhibitor. We further showed that the guanine nucleotide exchange factor Vav2, not β-Pix, was involved in E. coli K1-mediated Rac1 activation. Since activated STAT3 is known to bind GTP-Rac1, the relationship between STAT3 and Rac1 was examined in E. coli K1 invasion of HBMEC. Downregulation of STAT3 resulted in significantly decreased E. coli invasion compared to control HBMEC, as well as a corresponding decrease in GTP-Rac1, suggesting that Rac1 activation in response to E. coli is under the control of STAT3. More importantly, two E. coli determinants contributing to HBMEC invasion, IbeA and OmpA, were shown to affect both Rac1 activation and their association with STAT3. These findings demonstrate for the first time that specific E. coli determinants regulate a novel mechanism of STAT3 cross talk with Rac1 in E. coli K1 invasion of HBMEC. PMID:22451524

IbeA and OmpA of Escherichia coli K1 exploit Rac1 activation for invasion of human brain microvascular endothelial cells.

PubMed

Maruvada, Ravi; Kim, Kwang Sik

2012-06-01

Meningitis-causing Escherichia coli K1 internalization of the blood-brain barrier is required for penetration into the brain, but the host-microbial interactions involved in E. coli entry of the blood-brain barrier remain incompletely understood. We show here that a meningitis-causing E. coli K1 strain RS218 activates Rac1 (GTP-Rac1) of human brain microvascular endothelial cells (HBMEC) in a time-dependent manner. Both activation and bacterial invasion were significantly inhibited in the presence of a Rac1 inhibitor. We further showed that the guanine nucleotide exchange factor Vav2, not β-Pix, was involved in E. coli K1-mediated Rac1 activation. Since activated STAT3 is known to bind GTP-Rac1, the relationship between STAT3 and Rac1 was examined in E. coli K1 invasion of HBMEC. Downregulation of STAT3 resulted in significantly decreased E. coli invasion compared to control HBMEC, as well as a corresponding decrease in GTP-Rac1, suggesting that Rac1 activation in response to E. coli is under the control of STAT3. More importantly, two E. coli determinants contributing to HBMEC invasion, IbeA and OmpA, were shown to affect both Rac1 activation and their association with STAT3. These findings demonstrate for the first time that specific E. coli determinants regulate a novel mechanism of STAT3 cross talk with Rac1 in E. coli K1 invasion of HBMEC.

Prevention of Hypovolemic Circulatory Collapse by IL-6 Activated Stat3

PubMed Central

Tsimelzon, Anna I.; Mastrangelo, Mary-Ann A.; Hilsenbeck, Susan G.; Poli, Valeria; Tweardy, David J.

2008-01-01

Half of trauma deaths are attributable to hypovolemic circulatory collapse (HCC). We established a model of HCC in rats involving minor trauma plus severe hemorrhagic shock (HS). HCC in this model was accompanied by a 50% reduction in peak acceleration of aortic blood flow and cardiomyocyte apoptosis. HCC and apoptosis increased with increasing duration of hypotension. Apoptosis required resuscitation, which provided an opportunity to intervene therapeutically. Administration of IL-6 completely reversed HCC, prevented cardiac dysfunction and cardiomyocyte apoptosis, reduced mortality 5-fold and activated intracardiac signal transducer and activator of transcription (STAT) 3. Pre-treatment of rats with a selective inhibitor of Stat3, T40214, reduced the IL-6-mediated increase in cardiac Stat3 activity, blocked successful resuscitation by IL-6 and reversed IL-6-mediated protection from cardiac apoptosis. The hearts of mice deficient in the naturally occurring dominant negative isoform of Stat3, Stat3β, were completely resistant to HS-induced apoptosis. Microarray analysis of hearts focusing on apoptosis related genes revealed that expression of 29% of apoptosis related genes was altered in HS vs. sham rats. IL-6 treatment normalized the expression of these genes, while T40214 pretreatment prevented IL-6-mediated normalization. Thus, cardiac dysfunction, cardiomyocyte apoptosis and induction of apoptosis pathway genes are important components of HCC; IL-6 administration prevented HCC by blocking cardiomyocyte apoptosis and induction of apoptosis pathway genes via Stat3 and warrants further study as a resuscitation adjuvant for prevention of HCC and death in trauma patients. PMID:18270592

RANKL downregulates cell surface CXCR6 expression through JAK2/STAT3 signaling pathway during osteoclastogenesis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li, Changhong; Zhao, Jinxia; Sun, Lin

Highlights: Black-Right-Pointing-Pointer CXCR6 is down-regulated during RANKL-induced osteoclastogenesis in RAW264.7 cells. Black-Right-Pointing-Pointer CXCR6 reduction was nearly reversed by inhibition of JAK2/STAT3 signaling pathway. Black-Right-Pointing-Pointer CXCL16 alone does not positively regulate osteoclastogenesis. -- Abstract: The receptor activator of nuclear factor-{kappa}B ligand (RANKL), as a member of the tumor necrosis factor (TNF) family, plays an essential role in osteoclast differentiation and function. Chemokines and their receptors have recently been shown to play critical roles in osteoclastogenesis, however, whether CXCL16-CXCR6 plays role in RANKL-mediated osteoclastogenesis is unknown. In this study, we first reported that RANKL decreased CXCR6 in a dose-dependent manner, which maymore » be through deactivation of Akt and STAT3 signaling induced by CXCL16. Interestingly, RANKL-mediated CXCR6 reduction may be associated to the activation of STAT3 by phosphorylation. When STAT3 activation was blocked by JAK2/STAT3 inhibitor AG490, RANKL failed to shut down CXCR6 expression during osteoclastogenesis. However, CXCL16 alone did not augment RANKL-mediated osteoclast differentiation and did not alter RANKL-receptor RANK mRNA expression. These results demonstrate that reduction of CXCL16-CXCR6 is critical in RANKL-mediated osteoclastogenesis, which is mainly through the activation of JAK2/STAT3 signaling. CXCL16-CXCR6 axis may become a novel target for the therapeutic intervention of bone resorbing diseases such as rheumatoid arthritis and osteoporosis.« less

Targeting STAT3 with silibinin to improve cancer therapeutics.

PubMed

Bosch-Barrera, Joaquim; Queralt, Bernardo; Menendez, Javier A

2017-07-01

Signal transducer and activator of transcription 3 (STAT3) has a prominent role in mediating resistance to conventional chemo-/radio-therapies and modern targeted drugs. While a number of STAT3 inhibitors have been shown to enhance the efficacy of therapeutic agents in vitro, the majority of them have yet to enter clinical evaluation mostly because of lack of efficacy issues. Silibinin is the main component of the silymarin complex, a standardized extract obtained from the seeds of the milk thistle herb Silybum marianum. This review summarizes current evidence supporting the ability of silibinin to function as a natural down-modulator of STAT3 activity. We examine the reported capacity of silibinin to reduce the toxicity of cancer treatments and to reverse tumor cell resistance via STAT3 inhibition. We also briefly review our clinical data in cancer patients treated with oral nutraceutical products containing silibinin. The beneficial effects of silibinin might accelerate the design of strategies aimed to overcome and prevent the emergence of STAT3-mediated cancer drug resistance in clinical settings. Copyright © 2017 Elsevier Ltd. All rights reserved.

Exogenous hydrogen sulfide promotes hepatocellular carcinoma cell growth by activating the STAT3-COX-2 signaling pathway

PubMed Central

Zhen, Yulan; Wu, Qiaomei; Ding, Yiqian; Zhang, Wei; Zhai, Yuansheng; Lin, Xiaoxiong; Weng, Yunxia; Guo, Ruixian; Zhang, Ying; Feng, Jianqiang; Lei, Yiyan; Chen, Jingfu

2018-01-01

The effects of hydrogen sulfide (H2S) on cancer are controversial. Our group previously demonstrated that exogenous H2S promotes the development of cancer via amplifying the activation of the nuclear factor-κB signaling pathway in hepatocellular carcinoma (HCC) cells (PLC/PRF/5). The present study aimed to further investigate the hypothesis that exogenous H2S promotes PLC/PRF/5 cell proliferation and migration, and inhibits apoptosis by activating the signal transducer and activator of transcription 3 (STAT3)-cyclooxygenase-2 (COX-2) signaling pathway. PLC/PRF/5 cells were treated with 500 µmol/l NaHS (a donor of H2S) for 24 h. The expression levels of phosphorylated (p)-STAT3, STAT3, cleaved caspase-3 and COX-2 were measured by western blot assay. Cell viability was detected by Cell Counting kit-8 assay. Apoptotic cells were observed by Hoechst 33258 staining. The expression of STAT3 and COX-2 messenger RNA (mRNA) was detected by semiquantitative reverse transcription-polymerase chain reaction. The production of vascular endothelial growth factor (VEGF) was evaluated by ELISA. The results indicated that treatment of PLC/PRF/5 cells with 500 µmol/l NaHS for 24 h markedly increased the expression levels of p-STAT3 and STAT3 mRNA, leading to COX-2 and COX-2 mRNA overexpression, VEGF induction, decreased cleaved caspase-3 production, increased cell viability and migration, and decreased number of apoptotic cells. However, co-treatment of PLC/PRF/5 cells with 500 µmol/l NaHS and 30 µmol/l AG490 (an inhibitor of STAT3) or 20 µmol/l NS-398 (an inhibitor of COX-2) for 24 h significantly reverted the effects induced by NaHS. Furthermore, co-treatment of PLC/PRF/5 cells with 500 µmol/l NaHS and 30 µmol/l AG490 markedly decreased the NaHS-induced increase in the expression level of COX-2. By contrast, co-treatment of PLC/PRF/5 cells with 500 µmol/l NaHS and 20 µmol/l NS-398 inhibited the NaHS-induced increase in the expression level of p-STAT3. In conclusion, the findings of the present study provide evidence that the STAT3-COX-2 signaling pathway is involved in NaHS-induced cell proliferation, migration, angiogenesis and anti-apoptosis in PLC/PRF/5 cells, and suggest that the positive feedback between STAT3 and COX-2 may serve a crucial role in hepatocellular carcinoma carcinogenesis. PMID:29725404

Toxoplasma gondii induces FAK-Src-STAT3 signaling during infection of host cells that prevents parasite targeting by autophagy.

PubMed

Portillo, Jose-Andres C; Muniz-Feliciano, Luis; Lopez Corcino, Yalitza; Lee, So Jung; Van Grol, Jennifer; Parsons, Sarah J; Schiemman, William P; Subauste, Carlos S

2017-10-01

Targeting of Toxoplasma gondii by autophagy is an effective mechanism by which host cells kill the protozoan. Thus, the parasite must avoid autophagic targeting to survive. Here we show that the mammalian cytoplasmic molecule Focal Adhesion Kinase (FAK) becomes activated during invasion of host cells. Activated FAK appears to accompany the formation of the moving junction (as assessed by expression the parasite protein RON4). FAK activation was inhibited by approaches that impaired β1 and β3 integrin signaling. FAK caused activation of Src that in turn mediated Epidermal Growth Factor Receptor (EGFR) phosphorylation at the unique Y845 residue. Expression of Src-resistant Y845F EGFR mutant markedly inhibited ROP16-independent activation of STAT3 in host cells. Activation of FAK, Y845 EGFR or STAT3 prevented activation of PKR and eIF2α, key stimulators of autophagy. Genetic or pharmacologic inhibition of FAK, Src, EGFR phosphorylation at Y845, or STAT3 caused accumulation of the autophagy protein LC3 and LAMP-1 around the parasite and parasite killing dependent on autophagy proteins (ULK1 and Beclin 1) and lysosomal enzymes. Parasite killing was inhibited by expression of dominant negative PKR. Thus, T. gondii activates a FAK→Src→Y845-EGFR→STAT3 signaling axis within mammalian cells, thereby enabling the parasite to survive by avoiding autophagic targeting through a mechanism likely dependent on preventing activation of PKR and eIF2α.

STAT3-targeted treatment with silibinin overcomes the acquired resistance to crizotinib in ALK-rearranged lung cancer.

PubMed

Cuyàs, Elisabet; Pérez-Sánchez, Almudena; Micol, Vicente; Menendez, Javier A; Bosch-Barrera, Joaquim

2016-12-16

The signal transducer and activator of transcription 3 (STAT3) has been suggested to play a prominent role in mediating non-small-cell lung cancer (NSCLC) resistance to some tyrosine kinase inhibitor (TKI)-mediated therapies. Using a model of anaplastic lymphoma kinase gene (ALK)-translocated NSCLC with acquired resistance to the ALK TKI crizotinib, but lacking amplifications or mutations in the kinase domain of ALK, we herein present evidence that STAT3 activation is a novel mechanism of crizotinib resistance that involves the upregulation of immune escape and epithelial to mesenchymal transition (EMT) signaling pathways. Taking advantage of the flavonolignan silibinin as a naturally occurring STAT3-targeted pharmacological inhibitor, we confirmed that STAT3 activation protects ALK-translocated NSCLC from crizotinib. Accordingly, silibinin-induced inhibition of STAT3 worked synergistically with crizotinib to reverse acquired resistance and restore sensitivity in crizotinib-resistant cells. Moreover, silibinin treatment significantly inhibited the upregulation of the immune checkpoint regulator PD-L1 and also EMT regulators (e.g., SLUG, VIM, CD44) in crizotinib-refractory cells. These findings provide a valuable strategy to potentially improve the efficacy of ALK inhibition by cotreatment with silibinin-based therapeutics, which merit clinical investigation for ALK TKI-resistant NSCLC patients.

Spatio-temporal kinetics of growth hormone receptor signaling in single cells using FRET microscopy.

PubMed

Biener-Ramanujan, Eva; Ramanujan, V Krishnan; Herman, Brian; Gertler, Arieh

2006-08-01

The growth hormone (GH) receptor (R)-mediated JAK2 (Janus kinase-2)-STAT5 (signaling transducer and activator of transcription-5) pathway involves a cascade of protein-protein interactions and tyrosine phosphorylations that occur in a spatially and temporally sensitive manner in cells. To study GHR dimerization or GH-induced conformational change of predimerized GHRs and STAT5 activation kinetics in intact cells, fluorescence resonance energy transfer (FRET) and live-cell imaging methods were employed. FRET measurements at the membrane of HEK-293T cells co-expressing GHRs tagged at the C-terminus with cyan (C) and yellow (Y) fluorescent proteins (FPs) revealed transient GHR dimerization lasting 2-3 min, with a maximum at 3 min after GH stimulation, which was sufficient to induce STAT5 activation. The transient nature of the dimerization or GH-induced conformational change of predimerized GHRs kinetics was not a result of GHR internalization, as neither potassium- nor cholesterol-depletion treatments prolonged the FRET signal. YFP-tagged STAT5 recruitment to the membrane, binding to GHR-CFP, and phosphorylation, occurred within minutes of GH stimulation. Activated STAT5a-YFP did not show nuclear accumulation, despite nuclear pSTAT5 increase, suggesting high turnover of STAT5 nuclear shuttling. Although GHR dimerization and STAT5 activation have been reported previously, this is the first spatially resolved demonstration of GHR-signaling kinetics in intact cells.

Leptin Protects Host Cells from Entamoeba histolytica Cytotoxicity by a STAT3-Dependent Mechanism

PubMed Central

Verkerke, Hans P.; Paul, Shom N.; Mackey, Aaron J.; Petri, William A.

2012-01-01

The adipocytokine leptin links nutritional status to immune function. Leptin signaling protects from amebiasis, but the molecular mechanism is not understood. We developed an in vitro model of ameba-host cell interaction to test the hypothesis that leptin prevents ameba-induced apoptosis in host epithelial cells. We demonstrated that activation of mammalian leptin signaling increased cellular resistance to amebic cytotoxicity, including caspase-3 activation. Exogenous expression of the leptin receptor conferred resistance in susceptible cells, and leptin stimulation enhanced protection. A series of leptin receptor signaling mutants showed that resistance to amebic cytotoxicity was dependent on activation of STAT3 but not the Src homology-2 domain-containing tyrosine phosphatase (SHP-2) or STAT5. A common polymorphism in the leptin receptor (Q223R) that increases susceptibility to amebiasis in humans and mice was found to increase susceptibility to amebic cytotoxicity in single cells. The Q223R polymorphism also decreased leptin-dependent STAT3 activation by 21% relative to that of the wild-type (WT) receptor (P = 0.035), consistent with a central role of STAT3 signaling in protection. A subset of genes uniquely regulated by STAT3 in response to leptin was identified. Most notable were the TRIB1 and suppressor of cytokine signaling 3 (SOCS3) genes, which have opposing roles in the regulation of apoptosis. Overall apoptotic genes were highly enriched in this gene set (P < 1E−05), supporting the hypothesis that leptin regulation of host apoptotic genes via STAT3 is responsible for protection. This is the first demonstration of a mammalian signaling pathway that restricts amebic pathogenesis and represents an important advance in our mechanistic understanding of how leptin links nutrition and susceptibility to infection. PMID:22331430

Methanethiosulfonate derivatives as ligands of the STAT3-SH2 domain.

PubMed

Gabriele, Elena; Ricci, Chiara; Meneghetti, Fiorella; Ferri, Nicola; Asai, Akira; Sparatore, Anna

2017-12-01

With the aim to discover new STAT3 direct inhibitors, potentially useful as anticancer agents, a set of methanethiosulfonate drug hybrids were synthesized. The in vitro tests showed that all the thiosulfonic compounds were able to strongly and selectively bind STAT3-SH2 domain, whereas the parent drugs were completely devoid of this ability. In addition, some of them showed a moderate antiproliferative activity on HCT-116 cancer cell line. These results suggest that methanethiosulfonate moiety can be considered a useful scaffold in the preparation of new direct STAT3 inhibitors. Interestingly, an unusual kind of organo-sulfur derivative, endowed with valuable antiproliferative activity, was occasionally isolated. [Formula: see text].

Survival response of hippocampal neurons under low oxygen conditions induced by Hippophae rhamnoides is associated with JAK/STAT signaling.

PubMed

Manickam, Manimaran; Tulsawani, Rajkumar

2014-01-01

Janus activated kinase/signal transducers and activators of transcription (JAK/STATs) pathway are associated with various neuronal functions including cell survival and inflammation. In the present study, it is hypothesized that protective action of aqueous extract of Hippophae rhamnoides in hippocampal neurons against hypoxia is mediated via JAK/STATs. Neuronal cells exposed to hypoxia (0.5% O2) display higher reactive oxygen species with compromised antioxidant status compared to unexposed control cells. Further, these cells had elevated levels of pro-inflammatory cytokines; tumor necrosis factor α and interleukin 6 and nuclear factor κappa B. Moreover, the expression of JAK1 was found to be highly expressed with phosphorylation of STAT3 and STAT5. Cells treated with JAK1, STAT3 and STAT5 specific inhibitors resulted in more cell death compared to hypoxic cells. Treatment of cells with extract prevented oxidative stress and inflammatory response associated with hypoxia. The extract treated cells had more cell survival than hypoxic cells with induction of JAK1 and STAT5b. Cells treated with extract having suppressed JAK1 or STAT3 or STAT5 expression showed reduced cell viability than the cell treated with extract alone. Overall, the findings from these studies indicate that the aqueous extract of Hippophae rhamnoides treatment inhibited hypoxia induced oxidative stress by altering cellular JAK1, STAT3 and STAT5 levels thereby enhancing cellular survival response to hypoxia and provide a basis for possible use of aqueous extract of Hippophae rhamnoides in facilitating tolerance to hypoxia.

Survival Response of Hippocampal Neurons under Low Oxygen Conditions Induced by Hippophae rhamnoides is Associated with JAK/STAT Signaling

PubMed Central

Manickam, Manimaran; Tulsawani, Rajkumar

2014-01-01

Janus activated kinase/signal transducers and activators of transcription (JAK/STATs) pathway are associated with various neuronal functions including cell survival and inflammation. In the present study, it is hypothesized that protective action of aqueous extract of Hippophae rhamnoides in hippocampal neurons against hypoxia is mediated via JAK/STATs. Neuronal cells exposed to hypoxia (0.5% O2) display higher reactive oxygen species with compromised antioxidant status compared to unexposed control cells. Further, these cells had elevated levels of pro-inflammatory cytokines; tumor necrosis factor α and interleukin 6 and nuclear factor κappa B. Moreover, the expression of JAK1 was found to be highly expressed with phosphorylation of STAT3 and STAT5. Cells treated with JAK1, STAT3 and STAT5 specific inhibitors resulted in more cell death compared to hypoxic cells. Treatment of cells with extract prevented oxidative stress and inflammatory response associated with hypoxia. The extract treated cells had more cell survival than hypoxic cells with induction of JAK1 and STAT5b. Cells treated with extract having suppressed JAK1 or STAT3 or STAT5 expression showed reduced cell viability than the cell treated with extract alone. Overall, the findings from these studies indicate that the aqueous extract of Hippophae rhamnoides treatment inhibited hypoxia induced oxidative stress by altering cellular JAK1, STAT3 and STAT5 levels thereby enhancing cellular survival response to hypoxia and provide a basis for possible use of aqueous extract of Hippophae rhamnoides in facilitating tolerance to hypoxia. PMID:24516559

The regulation of inflammation by interferons and their STATs

PubMed Central

Rauch, Isabella; Müller, Mathias; Decker, Thomas

2013-01-01

Interferons (IFN) are subdivided into type I IFN (IFN-I, here synonymous with IFN-α/β), type II (IFN-γ) and type III IFN (IFN-III/IFN-λ) that reprogram nuclear gene expression through STATs 1 and 2 by forming STAT1 dimers (mainly IFN-γ) or the ISGF3 complex, a STAT1-STAT2-IRF9 heterotrimer (IFN-I and IFN-III). Dominant IFN activities in the immune system are to protect cells from viral replication and to activate macrophages for enhanced effector function. However, the impact of IFN and their STATs on the immune system stretches far beyond these activities and includes the control of inflammation. The goal of this review is to give an overview of the different facets of the inflammatory process that show regulatory input by IFN/STAT. PMID:24058799

Hormonal Resistance And Metastasis ER-Coregulartor-Src Signaling Targeted Therapy

DTIC Science & Technology

2011-09-01

with STAT3 and cross-talk between ER~ and STAT3 play an important role in leptin -in- duced STAT3 activation’’ The II.Kl axis is the major signaling...cancer. These patients could be treated with an aromatase inhibitor (Letrozole) that ablates peripheral estrogen synthesis and ER degraders /signaling

The differentiation and plasticity of Tc17 cells are regulated by CTLA-4-mediated effects on STATs.

PubMed

Arra, Aditya; Lingel, Holger; Kuropka, Benno; Pick, Jonas; Schnoeder, Tina; Fischer, Thomas; Freund, Christian; Pierau, Mandy; Brunner-Weinzierl, Monika C

2017-01-01

As the blockade of inhibitory surface-molecules such as CTLA-4 on T cells has led to recent advances in antitumor immune therapy, there is great interest in identifying novel mechanisms of action of CD8 + T cells to evoke effective cytotoxic antitumor responses. Using in vitro and in vivo models, we investigated the molecular pathways underlying the CTLA-4-mediated differentiation of IL-17-producing CD8 + T cells (Tc17 cells) that strongly impairs cytotoxicity. Our studies demonstrate that Tc17 cells lacking CTLA-4 signaling have limited production of STAT3-target gene products such as IL-17, IL-21, IL-23R and RORγt. Upon re-stimulation with IL-12, these cells display fast downregulation of Tc17 hallmarks and acquire Tc1 characteristics such as IFNγ and TNF-α co-expression, which is known to correlate with tumor control. Indeed, upon adoptive transfer, these cells were highly efficient in the antigen-specific rejection of established OVA-expressing B16 melanoma in vivo . Mechanistically, in primary and re-stimulated Tc17 cells, STAT3 binding to the IL-17 promoter was strongly augmented by CTLA-4, associated with less binding of STAT5 and reduced relative activation of STAT1 which is known to block STAT3 activity. Inhibiting CTLA-4-induced STAT3 activity reverses enhancement of signature Tc17 gene products, rendering Tc17 cells susceptible to conversion to Tc1-like cells with enhanced cytotoxic potential. Thus, CTLA-4 critically shapes the characteristics of Tc17 cells by regulating relative STAT3 activation, which provides new perspectives to enhance cytotoxicity of antitumor responses.

A multi-level model accounting for the effects of JAK2-STAT5 signal modulation in erythropoiesis.

PubMed

Lai, Xin; Nikolov, Svetoslav; Wolkenhauer, Olaf; Vera, Julio

2009-08-01

We develop a multi-level model, using ordinary differential equations, based on quantitative experimental data, accounting for murine erythropoiesis. At the sub-cellular level, the model includes a description of the regulation of red blood cell differentiation through Epo-stimulated JAK2-STAT5 signalling activation, while at the cell population level the model describes the dynamics of (STAT5-mediated) red blood cell differentiation from their progenitors. Furthermore, the model includes equations depicting the hypoxia-mediated regulation of hormone erythropoietin blood levels. Take all together, the model constitutes a multi-level, feedback loop-regulated biological system, involving processes in different organs and at different organisational levels. We use our model to investigate the effect of deregulation in the proteins involved in the JAK2-STAT5 signalling pathway in red blood cells. Our analysis results suggest that down-regulation in any of the three signalling system components affects the hematocrit level in an individual considerably. In addition, our analysis predicts that exogenous Epo injection (an already existing treatment for several blood diseases) may compensate the effects of single down-regulation of Epo hormone level, STAT5 or EpoR/JAK2 expression level, and that it may be insufficient to counterpart a combined down-regulation of all the elements in the JAK2-STAT5 signalling cascade.

Hyperactivation of Ha-ras oncogene, but not Ink4a/Arf deficiency, triggers bladder tumorigenesis

PubMed Central

Mo, Lan; Zheng, Xiaoyong; Huang, Hong-Ying; Shapiro, Ellen; Lepor, Herbert; Cordon-Cardo, Carlos; Sun, Tung-Tien; Wu, Xue-Ru

2007-01-01

Although ras is a potent mitogenic oncogene, its tumorigenicity depends on cellular context and cooperative events. Here we show that low-level expression of a constitutively active Ha-ras in mouse urothelium induces simple urothelial hyperplasia that is resistant to progression to full-fledged bladder tumors even in the absence of Ink4a/Arf. In stark contrast, doubling of the gene dosage of the activated Ha-ras triggered early-onset, rapidly growing, and 100% penetrant tumors throughout the urinary tract. Tumor initiation required superseding a rate-limiting step between simple and nodular hyperplasia, the latter of which is marked by the emergence of mesenchymal components and the coactivation of AKT and STAT pathways as well as PTEN inactivation. These results indicate that overactivation of Ha-ras is both necessary and sufficient to induce bladder tumors along a low-grade, noninvasive papillary pathway, and they shed light on the recent findings that ras activation, via point mutation, overexpression, or intensified signaling from FGF receptor 3, occurs in 70%–90% of these tumors in humans. Our results highlight the critical importance of the dosage/strength of Ha-ras activation in dictating its tumorigenicity — a mechanism of oncogene activation not fully appreciated to date. Finally, our results have clinical implications, as inhibiting ras and/or its downstream effectors, such as AKT and STAT3/5, could provide alternative means to treat low-grade, superficial papillary bladder tumors, the most common tumor in the urinary system. PMID:17256055

The Akt1/IL-6/STAT3 pathway regulates growth of lung tumor initiating cells.

PubMed

Malanga, Donatella; De Marco, Carmela; Guerriero, Ilaria; Colelli, Fabiana; Rinaldo, Nicola; Scrima, Marianna; Mirante, Teresa; De Vitis, Claudia; Zoppoli, Pietro; Ceccarelli, Michele; Riccardi, Miriam; Ravo, Maria; Weisz, Alessandro; Federico, Antonella; Franco, Renato; Rocco, Gaetano; Mancini, Rita; Rizzuto, Antonia; Gulletta, Elio; Ciliberto, Gennaro; Viglietto, Giuseppe

2015-12-15

Here we report that the PI3K/Akt1/IL-6/STAT3 signalling pathway regulates generation and stem cell-like properties of Non-Small Cell Lung Cancer (NSCLC) tumor initiating cells (TICs). Mutant Akt1, mutant PIK3CA or PTEN loss enhances formation of lung cancer spheroids (LCS), self-renewal, expression of stemness markers and tumorigenic potential of human immortalized bronchial cells (BEAS-2B) whereas Akt inhibition suppresses these activities in established (NCI-H460) and primary NSCLC cells. Matched microarray analysis of Akt1-interfered cells and LCSs identified IL-6 as a critical target of Akt signalling in NSCLC TICs. Accordingly, suppression of Akt in NSCLC cells decreases IL-6 levels, phosphorylation of IkK and IkB, NF-kB transcriptional activity, phosphorylation and transcriptional activity of STAT3 whereas active Akt1 up-regulates them. Exposure of LCSs isolated from NSCLC cells to blocking anti-IL-6 mAbs, shRNA to IL-6 receptor or to STAT3 markedly reduces the capability to generate LCSs, to self-renew and to form tumors, whereas administration of IL-6 to Akt-interfered cells restores the capability to generate LCSs. Finally, immunohistochemical studies in NSCLC patients demonstrated a positive correlative trend between activated Akt, IL-6 expression and STAT3 phosphorylation (n = 94; p < 0.05). In conclusion, our data indicate that aberrant Akt signalling contributes to maintaining stemness in lung cancer TICs through a NF-kB/IL-6/STAT3 pathway and provide novel potential therapeutic targets for eliminating these malignant cells in NSCLC.

The Akt1/IL-6/STAT3 pathway regulates growth of lung tumor initiating cells

PubMed Central

Malanga, Donatella; De Marco, Carmela; Guerriero, Ilaria; Colelli, Fabiana; Rinaldo, Nicola; Scrima, Marianna; Mirante, Teresa; De Vitis, Claudia; Zoppoli, Pietro; Ceccarelli, Michele; Riccardi, Miriam; Ravo, Maria; Weisz, Alessandro; Federico, Antonella; Franco, Renato; Rocco, Gaetano; Mancini, Rita; Rizzuto, Antonia; Gulletta, Elio; Ciliberto, Gennaro; Viglietto, Giuseppe

2015-01-01

Here we report that the PI3K/Akt1/IL-6/STAT3 signalling pathway regulates generation and stem cell-like properties of Non-Small Cell Lung Cancer (NSCLC) tumor initiating cells (TICs). Mutant Akt1, mutant PIK3CA or PTEN loss enhances formation of lung cancer spheroids (LCS), self-renewal, expression of stemness markers and tumorigenic potential of human immortalized bronchial cells (BEAS-2B) whereas Akt inhibition suppresses these activities in established (NCI-H460) and primary NSCLC cells. Matched microarray analysis of Akt1-interfered cells and LCSs identified IL-6 as a critical target of Akt signalling in NSCLC TICs. Accordingly, suppression of Akt in NSCLC cells decreases IL-6 levels, phosphorylation of IkK and IkB, NF-kB transcriptional activity, phosphorylation and transcriptional activity of STAT3 whereas active Akt1 up-regulates them. Exposure of LCSs isolated from NSCLC cells to blocking anti-IL-6 mAbs, shRNA to IL-6 receptor or to STAT3 markedly reduces the capability to generate LCSs, to self-renew and to form tumors, whereas administration of IL-6 to Akt-interfered cells restores the capability to generate LCSs. Finally, immunohistochemical studies in NSCLC patients demonstrated a positive correlative trend between activated Akt, IL-6 expression and STAT3 phosphorylation (n = 94; p < 0.05). In conclusion, our data indicate that aberrant Akt signalling contributes to maintaining stemness in lung cancer TICs through a NF-kB/IL-6/STAT3 pathway and provide novel potential therapeutic targets for eliminating these malignant cells in NSCLC. PMID:26486080

Spatiotemporal regulation of cell fusion by JNK and JAK/STAT signaling during Drosophila wound healing.

PubMed

Lee, Ji-Hyun; Lee, Chan-Wool; Park, Si-Hyoung; Choe, Kwang-Min

2017-06-01

Cell-cell fusion is widely observed during development and disease, and imposes a dramatic change on participating cells. Cell fusion should be tightly controlled, but the underlying mechanism is poorly understood. Here, we found that the JAK/STAT pathway suppressed cell fusion during wound healing in the Drosophila larval epidermis, restricting cell fusion to the vicinity of the wound. In the absence of JAK/STAT signaling, a large syncytium containing a 3-fold higher number of nuclei than observed in wild-type tissue formed in wounded epidermis. The JAK/STAT ligand-encoding genes upd2 and upd3 were transcriptionally induced by wounding, and were required for suppressing excess cell fusion. JNK (also known as Basket in flies) was activated in the wound vicinity and activity peaked at ∼8 h after injury, whereas JAK/STAT signaling was activated in an adjoining concentric ring and activity peaked at a later stage. Cell fusion occurred primarily in the wound vicinity, where JAK/STAT activation was suppressed by fusion-inducing JNK signaling. JAK/STAT signaling was both necessary and sufficient for the induction of βPS integrin (also known as Myospheroid) expression, suggesting that the suppression of cell fusion was mediated at least in part by integrin protein. © 2017. Published by The Company of Biologists Ltd.

SOCS3 promotes apoptosis of mammary differentiated cells.

PubMed

Le Provost, Fabienne; Miyoshi, Keiko; Vilotte, Jean-Luc; Bierie, Brian; Robinson, Gertraud W; Hennighausen, Lothar

2005-12-30

Growth and function of the mammary gland is regulated by cytokines and modulated by suppressor of cytokine signalling (SOCS) proteins. In vitro experiments demonstrated that SOCS3 can inhibit PRL induction of milk protein gene expression and STAT5 activation. We explored the SOCS3 expression pattern during mouse mammary development and its regulation by PRL and GH in wild-type and STAT5a-null mammary tissue. Our results suggest that, in vivo, PRL stimulates SOCS3 expression in stromal adipocytes, independently of STAT5a stimulation. In mammary epithelial cells, SOCS3 expression appears to be related to STAT3 activation. Together, our results are consistent with a role of SOCS3 in the mammary gland by promoting apoptosis of differentiated cells (adipocytes during gestation and epithelial cells during involution).

Targeting Stat3 in the myeloid compartment drastically improves the in vivo antitumor functions of adoptively transferred T cells

PubMed Central

Herrmann, Andreas; Kortylewski, Marcin; Kujawski, Maciej; Zhang, Chunyan; Reckamp, Karen; Armstrong, Brian; Wang, Lin; Kowolik, Claudia; Deng, Jiehui; Robert, Figlin; Yu, Hua

2010-01-01

Improving effector T cell functions is highly desirable for preventive or therapeutic interventions of diverse diseases. Stat3 in the myeloid compartment constrains Th-1 type immunity, dampening natural and induced antitumor immune responses. We have recently developed an in vivo siRNA delivery platform by conjugating a TLR9 agonist with siRNA that efficiently targets myeloid and B cells. Here we show that either ablating the Stat3 alleles in the myeloid compartment and B cells combined with CpG triggering or administrating the CpG-Stat3siRNA conjugates drastically augments effector functions of adoptively transferred CD8+ T cells. Specifically, we demonstrate that both approaches are capable of increasing dendritic cell and CD8+ T cell engagement in tumor draining lymph nodes. Furthermore, both approaches can significantly activate the transferred CD8+ T cells in vivo, upregulating effector molecules such as perforin, granzyme B and IFN-γ. Intravital multiphoton microscopy reveals that Stat3 silencing combined with CpG triggering greatly increases killing activity and tumor infiltration of transferred T cells. These results suggest the use of CpG-Stat3siRNA, and possibly other Stat3 inhibitors, as a potent adjuvant to improve T cell therapies. PMID:20841481

Silibinin-mediated metabolic reprogramming attenuates pancreatic cancer-induced cachexia and tumor growth

PubMed Central

Shukla, Surendra K.; Dasgupta, Aneesha; Mehla, Kamiya; Gunda, Venugopal; Vernucci, Enza; Souchek, Joshua; Goode, Gennifer; King, Ryan; Mishra, Anusha; Rai, Ibha; Nagarajan, Sangeetha; Chaika, Nina V.; Yu, Fang; Singh, Pankaj K.

2015-01-01

Pancreatic ductal adenocarcinoma (PDAC) is the fourth leading cause of cancer-related deaths in the US. Cancer-associated cachexia is present in up to 80% of PDAC patients and is associated with aggressive disease and poor prognosis. In the present studies we evaluated an anti-cancer natural product silibinin for its effectiveness in targeting pancreatic cancer aggressiveness and the cachectic properties of pancreatic cancer cells and tumors. Our results demonstrate that silibinin inhibits pancreatic cancer cell growth in a dose-dependent manner and reduces glycolytic activity of cancer cells. Our LC-MS/MS based metabolomics data demonstrates that silibinin treatment induces global metabolic reprogramming in pancreatic cancer cells. Silibinin treatment diminishes c-MYC expression, a key regulator of cancer metabolism. Furthermore, we observed reduced STAT3 signaling in silibinin-treated cancer cells. Overexpression of constitutively active STAT3 was sufficient to substantially revert the silibinin-induced downregulation of c-MYC and the metabolic phenotype. Our in vivo investigations demonstrate that silibinin reduces tumor growth and proliferation in an orthotopic mouse model of pancreatic cancer and prevents the loss of body weight and muscle. It also improves physical activity including grip strength and latency to fall in tumor-bearing mice. In conclusion, silibinin-induced metabolic reprogramming diminishes cell growth and cachectic properties of pancreatic cancer cells and animal models. PMID:26510913

p38 Mitogen-Activated Protein Kinase/Signal Transducer and Activator of Transcription-3 Pathway Signaling Regulates Expression of Inhibitory Molecules in T Cells Activated by HIV-1–Exposed Dendritic Cells

PubMed Central

Che, Karlhans Fru; Shankar, Esaki Muthu; Muthu, Sundaram; Zandi, Sasan; Sigvardsson, Mikael; Hinkula, Jorma; Messmer, Davorka; Larsson, Marie

2012-01-01

Human immunodeficiency virus type 1 (HIV-1) infection enhances the expression of inhibitory molecules on T cells, leading to T-cell impairment. The signaling pathways underlying the regulation of inhibitory molecules and subsequent onset of T-cell impairment remain elusive. We showed that both autologous and allogeneic T cells exposed to HIV-pulsed dendritic cells (DCs) upregulated cytotoxic T-lymphocyte antigen (CTLA-4), tumor-necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL), lymphocyte-activation gene-3 (LAG3), T-cell immunoglobulin mucin-3 (TIM-3), CD160 and certain suppression-associated transcription factors, such as B-lymphocyte induced maturation protein-1 (BLIMP-1), deltex homolog 1 protein (DTX1) and forkhead box P3 (FOXP3), leading to T-cell suppression. This induction was regulated by p38 mitogen-activated protein kinase/signal transducer and activator of transcription-3 (P38MAPK/STAT3) pathways, because their blockade significantly abrogated expression of all the inhibitory molecules studied and a subsequent recovery in T-cell proliferation. Neither interleukin-6 (IL-6) nor IL-10 nor growth factors known to activate STAT3 signaling events were responsible for STAT3 activation. Involvement of the P38MAPK/STAT3 pathways was evident because these proteins had a higher level of phosphorylation in the HIV-1–primed cells. Furthermore, blockade of viral CD4 binding and fusion significantly reduced the negative effects DCs imposed on primed T cells. In conclusion, HIV-1 interaction with DCs modulated their functionality, causing them to trigger the activation of the P38MAPK/STAT3 pathway in T cells, which was responsible for the upregulation of inhibitory molecules. PMID:22777388

High Glucose-Mediated STAT3 Activation in Endometrial Cancer Is Inhibited by Metformin: Therapeutic Implications for Endometrial Cancer

PubMed Central

Wallbillich, John J.; Josyula, Srirama; Saini, Uksha; Zingarelli, Roman A.; Dorayappan, Kalpana Deepa Priya; Riley, Maria K.; Wanner, Ross A.; Cohn, David E.; Selvendiran, Karuppaiyah

2017-01-01

Objectives STAT3 is over-expressed in endometrial cancer, and diabetes is a risk factor for the development of type 1 endometrial cancer. We therefore investigated whether glucose concentrations influence STAT3 expression in type 1 endometrial cancer, and whether such STAT3 expression might be inhibited by metformin. Methods In Ishikawa (grade 1) endometrial cancer cells subjected to media with low, normal, or high concentrations of glucose, expression of STAT3 and its target proteins was evaluated by real-time quantitative PCR (qPCR). Ishikawa cells were treated with metformin and assessed with cell proliferation, survival, migration, and ubiquitin assays, as well as Western blot and qPCR. Expression of apoptosis proteins was evaluated with Western blot in Ishikawa cells transfected with a STAT3 overexpression plasmid and treated with metformin. A xenograft tumor model was used for studying the in vivo efficacy of metformin. Results Expression of STAT3 and its target proteins was increased in Ishikawa cells cultured in high glucose media. In vitro, metformin inhibited cell proliferation, survival and migration but induced apoptosis. Metformin reduced expression levels of pSTAT3 ser727, total STAT3, and its associated cell survival and anti-apoptotic proteins. Additionally, metformin treatment was associated with increased degradation of pSTAT3 ser727. No change in apoptotic protein expression was noticed with STAT3 overexpression in Ishikawa cells. In vivo, metformin treatment led to a decrease in tumor weight as well as reductions of STAT3, pSTAT3 ser727, its target proteins. Conclusions These results suggest that STAT3 expression in type 1 endometrial cancer is stimulated by a high glucose environment and inhibited by metformin. PMID:28114390

Sophoraflavanone G induces apoptosis of human cancer cells by targeting upstream signals of STATs.

PubMed

Kim, Byung-Hak; Won, Cheolhee; Lee, Yun-Han; Choi, Jung Sook; Noh, Kum Hee; Han, Songhee; Lee, Haeri; Lee, Chang Seok; Lee, Dong-Sup; Ye, Sang-Kyu; Kim, Myoung-Hwan

2013-10-01

Aberrantly activated signal transducer and activator of transcription (STAT) proteins are implicated with human cancers and represent essential roles for cancer cell survival and proliferation. Therefore, the development of small-molecule inhibitors of STAT signaling bearing pharmacological activity has therapeutic potential for the treatment of human cancers. In this study, we identified sophoraflavanone G as a novel small-molecule inhibitor of STAT signaling in human cancer cells. Sophoraflavanone G inhibited tyrosine phosphorylation of STAT proteins in Hodgkin's lymphoma and tyrosine phosphorylation of STAT3 in solid cancer cells by inhibiting phosphorylation of the Janus kinase (JAK) proteins, Src family tyrosine kinases, such as Lyn and Src, Akt, and ERK1/2. In addition, sophoraflavanone G inhibited STAT5 phosphorylation in murine-bone-marrow-derived pro-B cells transfected with translocated Ets Leukemia (TEL)-JAKs and cytokine-induced rat pre-T lymphoma cells, as well as STAT5b reporter activity in TEL-JAKs and STAT5b reporter systems. Sophoraflavanone G also inhibited nuclear factor-κB (NF-κB) signaling in multiple myeloma cells. Furthermore, sophoraflavanone G inhibited cancer cell proliferation and induced apoptosis by regulating the expression of apoptotic and anti-apoptotic proteins. Our data suggest that sophoraflavanone G is a novel small-molecule inhibitor of STAT signaling by targeting upstream signals of STATs that may have therapeutic potential for cancers caused by persistently activated STAT proteins. Copyright © 2013 Elsevier Inc. All rights reserved.

[Study on the correlation between EGFR-STAT3 signal pathway and laryngeal papilloma].

PubMed

Wang, Xinhua; Sun, Jingwu

2009-09-01

To explore the relationship between the expression of EGFR and STAT3 in human laryngeal papilloma and its biological behavior. Reverse transcription polymerase chain reaction(RT-PCR), immunohistochemical staining and Western blot were used to evaluate the mRNA and protein expression of EGFR and STAT3 (p-STAT3) in 42 laryngeal papilloma tissues and 15 samples of normal laryngeal tissue, and the relationship between the protein expression of them and clinic pathological parameters was also analyzed. The mRNA expression levels of EGFR and STAT3 in laryngeal papilloma tissue were significantly higher than that in normal laryngeal tissue (P < 0.05, P < 0.01). Protein positive expression of EGFR and p-STAT3 were also detected in a significantly greater proportion of laryngeal papilloma than normal laryngeal tissue by immunohistochemistry and western blot (P < 0.01, P < 0.05). There was relationship between EGFR and p-STAT3 overexpression in laryngeal papilloma (P < 0.05). The expression p-STAT3 was correlated with the recurrence and canceration of laryngeal papilloma (P < 0.05). The EGFR-STAT3 signal transduction pathway may be involved in the pathogenesis of laryngeal papilloma,, and the persistent activation of STAT3 gene plays an important role in the recurrence and canceration of laryngeal papilloma.

Identification of STAT target genes in adipocytes

PubMed Central

Zhao, Peng; Stephens, Jacqueline M.

2013-01-01

Adipocytes play important roles in lipid storage, energy homeostasis and whole body insulin sensitivity. Studies in the last two decades have identified the hormones and cytokines that activate specific STATs in adipocytes in vitro and in vivo. Five of the seven STAT family members are expressed in adipocyte (STATs 1, 3, 5A, 5B and 6). Many transcription factors, including STATs, have been shown to play an important role in adipose tissue development and function. This review will summarize the importance of adipocytes, indicate the cytokines and hormones that utilize the JAK-STAT signaling pathway in fat cells and focus on the identification of STAT target genes in mature adipocytes. To date, specific target genes have been identified for STATs, 1, 5A and 5B, but not for STATs 3 and 6. PMID:24058802

Identification of JAK2 as a Mediator of FIP1L1-PDGFRA-Induced Eosinophil Growth and Function in CEL

PubMed Central

Li, Bin; Zhang, Guangsen; Li, Cui; He, Dan; Li, Xinying; Zhang, Chunfang; Tang, Faqing; Deng, Xiyun; Lu, Jingchen; Tang, Youhong; Li, Ruijuan; Chen, Zhuchu; Duan, Chaojun

2012-01-01

The Fip1-like1 (FIP1L1)-platelet-derived growth factor receptor alpha fusion gene (F/P) arising in the pluripotent hematopoietic stem cell (HSC),causes 14% to 60% of patients with hypereosinophilia syndrome (HES). These patients, classified as having F/P (+) chronic eosinophilic leukemia (CEL), present with clonal eosinophilia and display a more aggressive disease phenotype than patients with F/P (–) HES patients. The mechanisms underlying predominant eosinophil lineage targeting and the cytotoxicity of eosinophils in this leukemia remain unclear. Given that the Janus tyrosine kinase (JAK)/signal transducers and activators of transcription (Stat) signaling pathway is key to cytokine receptor-mediated eosinophil development and activated Stat3 and Stat5 regulate the expression of genes involved in F/P malignant transformation, we investigated whether and how JAK proteins were involved in the pathogenesis of F/P-induced CEL. F/P activation of JAK2, Stat3 and Stat5, were confirmed in all the 11 F/P (+) CEL patients examined. In vitro inhibition of JAK2 in EOL-1, primary F/P(+) CEL cells (PC) and T674I F/P Imatinib resistant cells(IR) by either JAK2-specific short interfering RNA (siRNA) or the tryphostin derivative AG490(AG490), significantly reduced cellular proliferation and induced cellular apoptosis. The F/P can enhance the IL-5-induced JAK2 activation, and further results indicated that JAK2 inhibition blocked IL-5-induced cellular migration and activation of the EOL-1 and PC cells in vitro. F/P-stimulation of the JAK2 suppressed cells led to a significantly reduction in Stat3 activation, but relatively normal induction of Stat5 activation. Interestingly, JAK2 inhibition also reduced PI3K, Akt and NF-κB activity in a dose-dependent manner, and suppressed expression levels of c-Myc and Survivin. These results strongly suggest that JAK2 is activated by F/P and is required for F/P stimulation of cellular proliferation and infiltration, possibly through induction of c-Myc and Survivin expression via activation of multiple signaling pathways, including NF-κB, Stat3, and PI3K/Akt. PMID:22523564

Discrete roles and bifurcation of PTEN signaling and mTORC1-mediated anabolic metabolism underlie IL-7–driven B lymphopoiesis

PubMed Central

Zeng, Hu; Yu, Mei; Tan, Haiyan; Li, Yuxin; Su, Wei; Shi, Hao; Dhungana, Yogesh; Guy, Cliff; Neale, Geoffrey; Cloer, Caryn; Peng, Junmin; Wang, Demin; Chi, Hongbo

2018-01-01

Interleukin-7 (IL-7) drives early B lymphopoiesis, but the underlying molecular circuits remain poorly understood, especially how Stat5 (signal transducer and activator of transcription 5)–dependent and Stat5-independent pathways contribute to this process. Combining transcriptome and proteome analyses and mouse genetic models, we show that IL-7 promotes anabolic metabolism and biosynthetic programs in pro-B cells. IL-7–mediated activation of mTORC1 (mechanistic target of rapamycin complex 1) supported cell proliferation and metabolism in a Stat5-independent, Myc-dependent manner but was largely dispensable for cell survival or Rag1 and Rag2 gene expression. mTORC1 was also required for Myc-driven lymphomagenesis. PI3K (phosphatidylinositol 3-kinase) and mTORC1 had discrete effects on Stat5 signaling and independently controlled B cell development. PI3K was actively suppressed by PTEN (phosphatase and tensin homolog) in pro-B cells to ensure proper IL-7R expression, Stat5 activation, heavy chain rearrangement, and cell survival, suggesting the unexpected bifurcation of the classical PI3K-mTOR signaling. Together, our integrative analyses establish IL-7R–mTORC1–Myc and PTEN-mediated PI3K suppression as discrete signaling axes driving B cell development, with differential effects on IL-7R–Stat5 signaling. PMID:29399633

Arctigenin promotes apoptosis in ovarian cancer cells via the iNOS/NO/STAT3/survivin signalling.

PubMed

Huang, Ke; Li, Li-an; Meng, Yuan-guang; You, Yan-qin; Fu, Xiao-yu; Song, Lei

2014-12-01

Arctigenin is a biologically active lignan extracted from the seeds of Arctium lappa and shows anticancer activity against a variety of human cancers. The aim of this study was to determine the effects of arctigenin on ovarian cancer cell proliferation and survival and associated molecular mechanisms. Human ovarian cancer OVCAR3 and SKOV3 cells were treated with arctigenin, and cell proliferation and apoptosis were assessed. Western blot analysis was used to examine signal transducer and activator of transcription-3 (STAT3) phosphorylation and survivin and inducible nitric oxide synthase (iNOS) expression. The involvement of STAT3/survivin/iNOS/NO signalling in arctigenin action was checked. Arctigenin treatment resulted in a significant and dose-dependent inhibition of cell proliferation. Arctigenin-treated cells showed a 4-6 times increase in the percentage of apoptosis, compared with control cells. Pre-treatment with Ac-DEVD-CHO, a specific inhibitor of caspase-3, counteracted the induction of apoptosis by arctigenin. Arctigenin treatment significantly inhibited STAT3 phosphorylation and survivin and iNOS expression. Arctigenin-induced apoptosis was impaired by pre-transfection with survivin-expressing plasmid or addition of chemical nitric oxide (NO) donors. Additionally, exogenous NO prevented the suppression of STAT3 phosphorylation and survivin expression by arctigenin. Arctigenin treatment inhibits the proliferation and induces caspase-3-dependent apoptosis of ovarian cancer cells. Suppression of iNOS/NO/STAT3/survivin signalling is causally linked to the anticancer activity of arctigenin. Therefore, arctigenin may be applicable to anticancer therapy for ovarian cancer. © 2014 Nordic Association for the Publication of BCPT (former Nordic Pharmacological Society).

Sphingosylphosphorylcholine promotes the differentiation of resident Sca-1 positive cardiac stem cells to cardiomyocytes through lipid raft/JNK/STAT3 and β-catenin signaling pathways.

PubMed

Li, Wenjing; Liu, Honghong; Liu, Pingping; Yin, Deling; Zhang, Shangli; Zhao, Jing

2016-07-01

Resident cardiac Sca-1-positive (+) stem cells may differentiate into cardiomyocytes to improve the function of damaged hearts. However, little is known about the inducers and molecular mechanisms underlying the myogenic conversion of Sca-1(+) stem cells. Here we report that sphingosylphosphorylcholine (SPC), a naturally occurring bioactive lipid, induces the myogenic conversion of Sca-1(+) stem cells, as evidenced by the increased expression of cardiac transcription factors (Nkx2.5 and GATA4), structural proteins (cardiac Troponin T), transcriptional enhancer (Mef2c) and GATA4 nucleus translocation. First, SPC activated JNK and STAT3, and the JNK inhibitor SP600125 or STAT3 inhibitor stattic impaired the SPC-induced expression of cardiac transcription factors and GATA4 nucleus translocation, which suggests that JNK and STAT3 participated in SPC-promoted cardiac differentiation. Moreover, STAT3 activation was inhibited by SP600125, whereas JNK was inhibited by β-cyclodextrin as a lipid raft breaker, which indicates a lipid raft/JNK/STAT3 pathway involved in SPC-induced myogenic transition. β-Catenin, degraded by activated GSK3β, was inhibited by SPC. Furthermore, GSK3β inhibitors weakened but the β-catenin inhibitor promoted SPC-induced differentiation. We found no crosstalk between the lipid raft/JNK/STAT3 and β-catenin pathway. Our study describes a lipid, SPC, as an endogenic inducer of myogenic conversion in Sca-1(+) stem cells with low toxicity and high efficiency for uptake. Copyright © 2016 Elsevier B.V. All rights reserved.

Prognostic significance of nuclear pSTAT3 in oral cancer.

PubMed

Macha, Muzafar A; Matta, Ajay; Kaur, Jatinder; Chauhan, S S; Thakar, Alok; Shukla, Nootan K; Gupta, Siddhartha Datta; Ralhan, Ranju

2011-04-01

Aberrant nuclear accumulation of proteins influences tumor development and may predict biologic aggressiveness and disease prognosis. This study determined the prognostic significance of pSTAT3 (phosphorylayed signal transducer and activator of transcription 3) in oral squamous cell carcinomas (OSCCs). Using immunohistochemistry, a significant increase in nuclear accumulation of pSTAT3 was observed in 49 of 90 leukoplakias (54.4%) and 63/94 OSCCs (67%) (p(trend) < .001). Increased pSTAT3 was associated with tumor stage (p = .01), nodal metastasis (p = .0018), and tobacco consumption (p = .004). Kaplan-Meier analysis demonstrated that OSCC with increased nuclear pSTAT3 showed significantly reduced disease-free survival (13 months), compared with the patients with no nuclear pSTAT3 expression (64 months, p = .019). Cox regression analysis revealed nuclear pSTAT3 as the most significant predictor of poor prognosis (p = .024, hazard ratio [HR] = 2.7). Increased nuclear accumulation of pSTAT3 occurs in early premalignant stages and is a marker for poor prognosis of OSCC. Copyright © 2010 Wiley Periodicals, Inc.

Targeting mitochondrial STAT3 with the novel phospho-valproic acid (MDC-1112) inhibits pancreatic cancer growth in mice.

PubMed

Mackenzie, Gerardo G; Huang, Liqun; Alston, Ninche; Ouyang, Nengtai; Vrankova, Kvetoslava; Mattheolabakis, George; Constantinides, Panayiotis P; Rigas, Basil

2013-01-01

New agents are needed to treat pancreatic cancer, one of the most lethal human malignancies. We synthesized phospho-valproic acid, a novel valproic acid derivative, (P-V; MDC-1112) and evaluated its efficacy in the control of pancreatic cancer. P-V inhibited the growth of human pancreatic cancer xenografts in mice by 60%-97%, and 100% when combined with cimetidine. The dominant molecular target of P-V was STAT3. P-V inhibited the phosphorylation of JAK2 and Src, and the Hsp90-STAT3 association, suppressing the activating phosphorylation of STAT3, which in turn reduced the expression of STAT3-dependent proteins Bcl-xL, Mcl-1 and survivin. P-V also reduced STAT3 levels in the mitochondria by preventing its translocation from the cytosol, and enhanced the mitochondrial levels of reactive oxygen species, which triggered apoptosis. Inhibition of mitochondrial STAT3 by P-V was required for its anticancer effect; mitochondrial STAT3 overexpression rescued animals from the tumor growth inhibition by P-V. Our results indicate that P-V is a promising candidate drug against pancreatic cancer and establish mitochondrial STAT3 as its key molecular target.

Autophagy Facilitates IFN-γ-induced Jak2-STAT1 Activation and Cellular Inflammation*

PubMed Central

Chang, Yu-Ping; Tsai, Cheng-Chieh; Huang, Wei-Ching; Wang, Chi-Yun; Chen, Chia-Ling; Lin, Yee-Shin; Kai, Jui-In; Hsieh, Chia-Yuan; Cheng, Yi-Lin; Choi, Pui-Ching; Chen, Shun-Hua; Chang, Shih-Ping; Liu, Hsiao-Sheng; Lin, Chiou-Feng

2010-01-01

Autophagy is regulated for IFN-γ-mediated antimicrobial efficacy; however, its molecular effects for IFN-γ signaling are largely unknown. Here, we show that autophagy facilitates IFN-γ-activated Jak2-STAT1. IFN-γ induces autophagy in wild-type but not in autophagy protein 5 (Atg5−/−)-deficient mouse embryonic fibroblasts (MEFs), and, autophagy-dependently, IFN-γ induces IFN regulatory factor 1 and cellular inflammatory responses. Pharmacologically inhibiting autophagy using 3-methyladenine, a known inhibitor of class III phosphatidylinositol 3-kinase, confirms these effects. Either Atg5−/− or Atg7−/− MEFs are, independent of changes in IFN-γ receptor expression, resistant to IFN-γ-activated Jak2-STAT1, which suggests that autophagy is important for IFN-γ signal transduction. Lentivirus-based short hairpin RNA for Atg5 knockdown confirmed the importance of autophagy for IFN-γ-activated STAT1. Without autophagy, reactive oxygen species increase and cause SHP2 (Src homology-2 domain-containing phosphatase 2)-regulated STAT1 inactivation. Inhibiting SHP2 reversed both cellular inflammation and the IFN-γ-induced activation of STAT1 in Atg5−/− MEFs. Our study provides evidence that there is a link between autophagy and both IFN-γ signaling and cellular inflammation and that autophagy, because it inhibits the expression of reactive oxygen species and SHP2, is pivotal for Jak2-STAT1 activation. PMID:20592027

A chimeric cyclic interferon-α2b peptide induces apoptosis by sequential activation of phosphatidylinositol 3-kinase, protein kinase Cδ and p38 MAP kinase.

PubMed

Blank, V C; Bertucci, L; Furmento, V A; Peña, C; Marino, V J; Roguin, L P

2013-06-10

We have previously demonstrated that tyrosine phosphorylation of STAT1/3 and p38 mitogen-activated protein kinase (p38 MAPK) activation are involved in the apoptotic response triggered by a chimeric cyclic peptide of the interferon-α2b (IFN-α2b) in WISH cells. Since the peptide also induced serine phosphorylation of STAT proteins, in the present study we examined the kinase involved in serine STAT1 phosphorylation and the signaling effectors acting upstream such activation. We first found that p38 MAPK is involved in serine STAT1 phosphorylation, since a reduction of phophoserine-STAT1 levels was evident after incubating WISH cells with cyclic peptide in the presence of a p38 pharmacological inhibitor or a dominant-negative p38 mutant. Next, we demonstrated that the peptide induced activation of protein kinase Cδ (PKCδ). Based on this finding, the role of this kinase was then evaluated. After incubating WISH cells with a PKCδ inhibitor or after decreasing PKCδ expression levels by RNA interference, both peptide-induced serine STAT1 and p38 phosphorylation levels were significantly decreased, indicating that PKCδ functions as an upstream regulator of p38. We also showed that PKCδ and p38 activation stimulated by the peptide was inhibited by a specific pharmacological inhibitor of phosphatidylinositol 3-kinase (PI3K) or by a dominant-negative p85 PI3K-regulatory subunit, suggesting that PI3K is upstream in the signaling cascade. In addition, the role of PI3K and PKCδ in cyclic peptide-induced apoptosis was examined. Both signaling effectors were found to regulate the antiproliferative activity and the apoptotic response triggered by the cyclic peptide in WISH cells. In conclusion, we herein demonstrated that STAT1 serine phosphorylation is mediated by the sequential activation of PI3K, PKCδ and p38 MAPK. This signaling cascade contributes to the antitumor effect induced by the chimeric IFN-α2b cyclic peptide in WISH cells. Copyright © 2013 Elsevier Inc. All rights reserved.

Sevoflurane postconditioning protects the myocardium against ischemia/reperfusion injury via activation of the JAK2–STAT3 pathway

PubMed Central

Wu, Jianjiang; Yu, Jin; Xie, Peng; Maimaitili, Yiliyaer; Wang, Jiang; Yang, Long; Ma, Haiping; Zhang, Xing; Yang, Yining

2017-01-01

Background Sevoflurane postconditioning (S-post) has similar cardioprotective effects as ischemic preconditioning. However, the underlying mechanism of S-post has not been fully elucidated. Janus kinase signaling transduction/transcription activator (JAK2–STAT3) plays an important role in cardioprotection. The purpose of this study was to determine whether the cardioprotective effects of S-post are associated with activation of the JAK2–STAT3 signal pathway. Methods An adult male Sprague–Dawley (SD) rat model of myocardial ischemia/reperfusion (I/R) injury was established using the Langendorff isolated heart perfusion apparatus. At the beginning of reperfusion, 2.4% sevoflurane alone or in combination with AG490 (a JAK2 selective inhibitor) was used as a postconditioning treatment. The cardiac function indicators, myocardial infarct size, lactic dehydrogenase (LDH) release, mitochondrial ultrastructure, mitochondrial reactive oxygen species (ROS) generation rates, ATP content, protein expression of p-JAK, p-STAT3, Bcl-2 and Bax were measured. Results Compared with the I/R group, S-post significantly increased the expression of p-JAK, p-STAT3 and Bcl-2 and reduced the protein expression of Bax, which markedly decreased the myocardial infarction areas, improved the cardiac function indicators and the mitochondrial ultrastructure, decreased the mitochondrial ROS and increased the ATP content. However, the cardioprotective effects of S-post were abolished by treatment with a JAK2 selective inhibitor (p < 0.05). Conclusion This study demonstrates that the cardioprotective effects of S-post are associated with the activation of JAK2–STAT3. The mechanism may be related to an increased expression of p-JAK2 and p-STAT3 after S-post, which reduced mitochondrial ROS generation and increased mitochondrial ATP content, thereby reducing apoptosis and myocardial infarct size. PMID:28392989

Caffeic Acid Inhibits Chronic UVB-Induced Cellular Proliferation Through JAK-STAT3 Signaling in Mouse Skin.

PubMed

Agilan, Balupillai; Rajendra Prasad, N; Kanimozhi, Govindasamy; Karthikeyan, Ramasamy; Ganesan, Muthusamy; Mohana, Shanmugam; Velmurugan, Devadasan; Ananthakrishnan, Dhanapalan

2016-05-01

Signal transducers and activators of transcription 3 (STAT3) play a critical role in inflammation, proliferation and carcinogenesis. Inhibition of JAK-STAT3 signaling is proved to be a novel target for prevention of UVB-induced skin carcinogenesis. In this study, chronic UVB irradiation (180 mJ cm(-2) ; weekly thrice for 30 weeks) induces the expression of IL-10 and JAK1 that eventually activates the STAT3 which leads to the transcription of proliferative and antiapoptotic markers such as PCNA, Cyclin-D1, Bcl2 and Bcl-xl, respectively. Caffeic acid (CA) inhibits JAK-STAT3 signaling, thereby induces apoptotic cell death by upregulating Bax, Cytochrome-C, Caspase-9 and Caspase-3 expression in mouse skin. Furthermore, TSP-1 is an antiangiogeneic protein, which is involved in the inhibition of angiogenesis and proliferation. Chronic UVB exposure decreased the expression of TSP-1 and pretreatment with CA prevented the UVB-induced loss of TSP-1 in UVB-irradiated mouse skin. Thus, CA offers protection against UVB-induced photocarcinogenesis probably through modulating the JAK-STAT3 in the mouse skin. © 2016 The American Society of Photobiology.

Early-onset lymphoproliferation and autoimmunity caused by germline STAT3 gain-of-function mutations

PubMed Central

Vogel, Tiphanie P.; Forbes, Lisa; Ma, Chi A.; Stray-Pedersen, Asbjørg; Niemela, Julie E.; Lyons, Jonathan J.; Engelhardt, Karin R.; Zhang, Yu; Topcagic, Nermina; Roberson, Elisha D. O.; Matthews, Helen; Verbsky, James W.; Dasu, Trivikram; Vargas-Hernandez, Alexander; Varghese, Nidhy; McClain, Kenneth L.; Karam, Lina B.; Nahmod, Karen; Makedonas, George; Mace, Emily M.; Sorte, Hanne S.; Perminow, Gøri; Rao, V. Koneti; O’Connell, Michael P.; Price, Susan; Su, Helen C.; Butrick, Morgan; McElwee, Joshua; Hughes, Jason D.; Willet, Joseph; Swan, David; Xu, Yaobo; Santibanez-Koref, Mauro; Slowik, Voytek; Dinwiddie, Darrell L.; Ciaccio, Christina E.; Saunders, Carol J.; Septer, Seth; Kingsmore, Stephen F.; White, Andrew J.; Cant, Andrew J.; Hambleton, Sophie

2015-01-01

Germline loss-of-function mutations in the transcription factor signal transducer and activator of transcription 3 (STAT3) cause immunodeficiency, whereas somatic gain-of-function mutations in STAT3 are associated with large granular lymphocytic leukemic, myelodysplastic syndrome, and aplastic anemia. Recently, germline mutations in STAT3 have also been associated with autoimmune disease. Here, we report on 13 individuals from 10 families with lymphoproliferation and early-onset solid-organ autoimmunity associated with 9 different germline heterozygous mutations in STAT3. Patients exhibited a variety of clinical features, with most having lymphadenopathy, autoimmune cytopenias, multiorgan autoimmunity (lung, gastrointestinal, hepatic, and/or endocrine dysfunction), infections, and short stature. Functional analyses demonstrate that these mutations confer a gain-of-function in STAT3 leading to secondary defects in STAT5 and STAT1 phosphorylation and the regulatory T-cell compartment. Treatment targeting a cytokine pathway that signals through STAT3 led to clinical improvement in 1 patient, suggesting a potential therapeutic option for such patients. These results suggest that there is a broad range of autoimmunity caused by germline STAT3 gain-of-function mutations, and that hematologic autoimmunity is a major component of this newly described disorder. Some patients for this study were enrolled in a trial registered at www.clinicaltrials.gov as #NCT00001350. PMID:25359994

Elevated levels of STAT1 in Fanconi anemia group A lymphoblasts correlate with the cells’ sensitivity to DNA interstrand crosslinking drugs

PubMed Central

Prieto-Remón, Inés; Sánchez-Carrera, Dámaso; López-Duarte, Mónica; Richard, Carlos; Pipaón, Carlos

2013-01-01

Progressive bone marrow failure starting in the first decade of life is one of the main characteristics of Fanconi anemia. Along with the bone marrow failure, this pathology is characterized by congenital malformations, endocrine dysfunction and an extraordinary predisposition to develop cancer. The fact that hematopoietic progenitor cells from subjects with Fanconi anemia are sensitive to both DNA-interstrand crosslinking agents and inflammatory cytokines, which are aberrantly overproduced in these patients, has led to different explanations for the causes of the bone marrow failure. We analyzed STAT1 expression in lymphoblastoid cell lines derived from patients with Fanconi anemia group A and correlated this with aspects of the Fanconi anemia phenotype such as sensitivity to genotoxic agents or to inhibitory cytokines. We provide evidence of overexpression of STAT1 in FANCA-deficient cells which has both transcriptional and post-translational components, and is related to the constitutive activation of ERK in Fanconi anemia group A cells, since it can be reverted by treatment with U0126. STAT1 phosphorylation was not defective in the lymphoblasts, so these cells accumulated higher levels of active STAT1 in response to interferon gamma, probably in relation to their greater sensitivity to this cytokine. On the other hand, inhibition of STAT1 by genetic or chemical means reverted the hypersensitivity of Fanconi anemia group A lymphoblasts to DNA interstrand crosslinking agents. Our data provide an explanation for the mixed sensitivity of Fanconi anemia group A cells to both genotoxic stress and inflammatory cytokines and indicate new targets for the treatment of bone marrow failure in these patients. PMID:23585528

STAT3 Target Genes Relevant to Human Cancers

PubMed Central

Carpenter, Richard L.; Lo, Hui-Wen

2014-01-01

Since its discovery, the STAT3 transcription factor has been extensively studied for its function as a transcriptional regulator and its role as a mediator of development, normal physiology, and pathology of many diseases, including cancers. These efforts have uncovered an array of genes that can be positively and negatively regulated by STAT3, alone and in cooperation with other transcription factors. Through regulating gene expression, STAT3 has been demonstrated to play a pivotal role in many cellular processes including oncogenesis, tumor growth and progression, and stemness. Interestingly, recent studies suggest that STAT3 may behave as a tumor suppressor by activating expression of genes known to inhibit tumorigenesis. Additional evidence suggested that STAT3 may elicit opposing effects depending on cellular context and tumor types. These mixed results signify the need for a deeper understanding of STAT3, including its upstream regulators, parallel transcription co-regulators, and downstream target genes. To help facilitate fulfilling this unmet need, this review will be primarily focused on STAT3 downstream target genes that have been validated to associate with tumorigenesis and/or malignant biology of human cancers. PMID:24743777

A combined omics study on activated macrophages--enhanced role of STATs in apoptosis, immunity and lipid metabolism.

PubMed

Dinasarapu, Ashok Reddy; Gupta, Shakti; Ram Maurya, Mano; Fahy, Eoin; Min, Jun; Sud, Manish; Gersten, Merril J; Glass, Christopher K; Subramaniam, Shankar

2013-11-01

Macrophage activation by lipopolysaccharide and adenosine triphosphate (ATP) has been studied extensively because this model system mimics the physiological context of bacterial infection and subsequent inflammatory responses. Previous studies on macrophages elucidated the biological roles of caspase-1 in post-translational activation of interleukin-1β and interleukin-18 in inflammation and apoptosis. However, the results from these studies focused only on a small number of factors. To better understand the host response, we have performed a high-throughput study of Kdo2-lipid A (KLA)-primed macrophages stimulated with ATP. The study suggests that treating mouse bone marrow-derived macrophages with KLA and ATP produces 'synergistic' effects that are not seen with treatment of KLA or ATP alone. The synergistic regulation of genes related to immunity, apoptosis and lipid metabolism is observed in a time-dependent manner. The synergistic effects are produced by nuclear factor kappa-light-chain-enhancer of activated B cells (NF-kB) and activator protein (AP)-1 through regulation of their target cytokines. The synergistically regulated cytokines then activate signal transducer and activator of transcription (STAT) factors that result in enhanced immunity, apoptosis and lipid metabolism; STAT1 enhances immunity by promoting anti-microbial factors; and STAT3 contributes to downregulation of cell cycle and upregulation of apoptosis. STAT1 and STAT3 also regulate glycerolipid and eicosanoid metabolism, respectively. Further, western blot analysis for STAT1 and STAT3 showed that the changes in transcriptomic levels were consistent with their proteomic levels. In summary, this study shows the synergistic interaction between the toll-like receptor and purinergic receptor signaling during macrophage activation on bacterial infection. Time-course data of transcriptomics and lipidomics can be queried or downloaded from http://www.lipidmaps.org. shankar@ucsd.edu. Supplementary data are available at Bioinformatics online.

Allosteric pathway identification through network analysis: from molecular dynamics simulations to interactive 2D and 3D graphs.

PubMed

Allain, Ariane; Chauvot de Beauchêne, Isaure; Langenfeld, Florent; Guarracino, Yann; Laine, Elodie; Tchertanov, Luba

2014-01-01

Allostery is a universal phenomenon that couples the information induced by a local perturbation (effector) in a protein to spatially distant regulated sites. Such an event can be described in terms of a large scale transmission of information (communication) through a dynamic coupling between structurally rigid (minimally frustrated) and plastic (locally frustrated) clusters of residues. To elaborate a rational description of allosteric coupling, we propose an original approach - MOdular NETwork Analysis (MONETA) - based on the analysis of inter-residue dynamical correlations to localize the propagation of both structural and dynamical effects of a perturbation throughout a protein structure. MONETA uses inter-residue cross-correlations and commute times computed from molecular dynamics simulations and a topological description of a protein to build a modular network representation composed of clusters of residues (dynamic segments) linked together by chains of residues (communication pathways). MONETA provides a brand new direct and simple visualization of protein allosteric communication. A GEPHI module implemented in the MONETA package allows the generation of 2D graphs of the communication network. An interactive PyMOL plugin permits drawing of the communication pathways between chosen protein fragments or residues on a 3D representation. MONETA is a powerful tool for on-the-fly display of communication networks in proteins. We applied MONETA for the analysis of communication pathways (i) between the main regulatory fragments of receptors tyrosine kinases (RTKs), KIT and CSF-1R, in the native and mutated states and (ii) in proteins STAT5 (STAT5a and STAT5b) in the phosphorylated and the unphosphorylated forms. The description of the physical support for allosteric coupling by MONETA allowed a comparison of the mechanisms of (a) constitutive activation induced by equivalent mutations in two RTKs and (b) allosteric regulation in the activated and non-activated STAT5 proteins. Our theoretical prediction based on results obtained with MONETA was validated for KIT by in vitro experiments. MONETA is a versatile analytical and visualization tool entirely devoted to the understanding of the functioning/malfunctioning of allosteric regulation in proteins - a crucial basis to guide the discovery of next-generation allosteric drugs.

Suppression of microRNA-135b-5p protects against myocardial ischemia/reperfusion injury by activating JAK2/STAT3 signaling pathway in mice during sevoflurane anesthesia.

PubMed

Xie, Xiao-Juan; Fan, Dong-Mei; Xi, Kai; Chen, Ya-Wei; Qi, Peng-Wei; Li, Qian-Hui; Fang, Liang; Ma, Li-Gang

2017-06-30

The study aims to explore the effects of miR-135b-5p on myocardial ischemia/reperfusion (I/R) injuries by regulating Janus protein tyrosine kinase 2 (JAK2)/signal transducer and activator of transcription (STAT) signaling pathway by mediating inhalation anesthesia with sevoflurane. A sum of 120 healthy Wistar male mice was assigned into six groups. Left ventricular ejection fraction (LVEF) and left ventricular shortening fraction (LVSF) were detected. Cardiomyocyte apoptosis was determined by terminal dexynucleotidyl transferase mediated dUTP-biotin nick end labeling (TUNEL) assay. MiR-135b-5p expression, mRNA and protein expression of p-STAT3, p-JAK2, STAT3, JAK2, B-cell lymphoma-2 (Bcl-2) and Bcl-2 associated X protein B (Bax) were detected by quantitative real-time PCR (qRT-PCR) and Western blotting. Target relationship between miR-135b-5p and JAK2 was confirmed by dual-luciferase reporter assay. The other five groups exhibited increased cardiomyocyte necrosis, apoptosis, miR-135b-5p and Bax expression, mRNA expression of JAK2 and STAT3, and protein expression of p-STAT3 and p-JAK2 compared with the sham group, but showed decreased LVEF, LVFS, and Bcl-2 expression. Compared with the model and AG490 + Sevo groups, the Sevo, inhibitor + Sevo and inhibitor + AG490 + Sevo groups displayed reduced cardiomyocyte necrosis, apoptosis, miR-135b-5p and Bax expression, but displayed elevated mRNA expression of JAK2 and STAT3, protein expression of p-STAT3 and p-JAK2, LVEF, LVFS and Bcl-2 expression. Compared with the Sevo and inhibitor + AG490 + Sevo groups, the AG490 + Sevo group showed decreased LVEF, LVFS, Bcl-2 expression, mRNA expressions of JAK2 and STAT3, and protein expressions of p-STAT3 and p-JAK2, but increased cardiomyocyte necrosis, apoptosis, and Bax expressions. MiR-135b-5p negatively targetted JAK2. Inhibition of miR-135b-5p can protect against myocardial I/R injury by activating JAK2/STAT3 signaling pathway through mediation of inhalation anesthesia with sevoflurane. © 2017 The Author(s).

Saw Palmetto Extract Inhibits Metastasis and Antiangiogenesis through STAT3 Signal Pathway in Glioma Cell.

PubMed

Ding, Hong; Shen, Jinglian; Yang, Yang; Che, Yuqin

2015-01-01

Signal transducer and activator of transcription factor 3 (STAT3) plays an important role in the proliferation and angiogenesis in human glioma. Previous research indicated that saw palmetto extract markedly inhibited the proliferation of human glioma cells through STAT3 signal pathway. But its effect on tumor metastasis and antiangiogenesis is not clear. This study is to further clear the impact of saw palmetto extract on glioma cell metastasis, antiangiogenesis, and its mechanism. TUNEL assay indicated that the apoptotic cells in the saw palmetto treated group are higher than that in the control group (p < 0.05). The apoptosis related protein is detected and the results revealed that saw palmetto extract inhibits the proliferation of human glioma. Meanwhile pSTAT3 is lower in the experimental group and CD34 is also inhibited in the saw palmetto treated group. This means that saw palmetto extract could inhibit the angiogenesis in glioma. We found that saw palmetto extract was an important phytotherapeutic drug against the human glioma through STAT3 signal pathway. Saw palmetto extract may be useful as an adjunctive therapeutic agent for treatment of individuals with glioma and other types of cancer in which STAT3 signaling is activated.

Three STATs are involved in the regulation of the expression of antimicrobial peptides in the triangle sail mussel, Hyriopsis cumingii.

PubMed

Dai, Yun-Jia; Hui, Kai-Min; Zhang, Ying-Hao; Liu, Yan; Wang, Yu-Qing; Zhao, Li-Juan; Lin, Li; Chai, Lian-Qin; Wei, Shun; Lan, Jiang-Feng

2017-04-01

Janus kinase (Jak) and signal transducers and activators of transcription (STAT) signaling pathway is associated in antiviral and antibacterial immune response. Previous studies primarily investigated the function of STATs in mammals. For most invertebrates, only one STAT was found in each species, such as STAT92E was found in Drosophila melanogaster. The studies, which focus on the functional difference between various STATs in the same species of invertebrate, are limited. In the present study, three STATs (HcSTAT1, HcSTAT2 and HcSTAT3) were identified in triangle shell pearl mussel, Hyriopsis cumingii. Phylogenetic analysis showed that HcSTAT1 and HcSTAT3 were clustered with Homo sapiens STAT5, and HcSTAT2 was clustered with Pinctada fucata STAT and Crassostea gigas STAT6. All three STATs could be detected in all tested tissues (hemocytes, hepatopancreas, gill, mantle and foot), and were induced expression when challenged with Staphylococcus aureus or Aeromonas hydrophilia in hemocytes and hepatopancreas. HcSTAT1 regulated the expression of HcDef, HcWAP, HcThe and HcTNF. The expression of HcWAP and HcTNF was down-regulated in HcSTAT2-RNAi mussel. And HcSTAT3 affected the expression of HcTNF. The study is the first report of different functions in antibacterial immune responses between STATs in mollusks. Copyright © 2017 Elsevier Ltd. All rights reserved.

Astaxanthin Inhibits JAK/STAT-3 Signaling to Abrogate Cell Proliferation, Invasion and Angiogenesis in a Hamster Model of Oral Cancer

PubMed Central

Kowshik, J.; Baba, Abdul Basit; Giri, Hemant; Deepak Reddy, G.; Dixit, Madhulika; Nagini, Siddavaram

2014-01-01

Identifying agents that inhibit STAT-3, a cytosolic transcription factor involved in the activation of various genes implicated in tumour progression is a promising strategy for cancer chemoprevention. In the present study, we investigated the effect of dietary astaxanthin on JAK-2/STAT-3 signaling in the 7,12-dimethylbenz[a]anthracene (DMBA)-induced hamster buccal pouch (HBP) carcinogenesis model by examining the mRNA and protein expression of JAK/STAT-3 and its target genes. Quantitative RT-PCR, immunoblotting and immunohistochemical analyses revealed that astaxanthin supplementation inhibits key events in JAK/STAT signaling especially STAT-3 phosphorylation and subsequent nuclear translocation of STAT-3. Furthermore, astaxanthin downregulated the expression of STAT-3 target genes involved in cell proliferation, invasion and angiogenesis, and reduced microvascular density, thereby preventing tumour progression. Molecular docking analysis confirmed inhibitory effects of astaxanthin on STAT signaling and angiogenesis. Cell culture experiments with the endothelial cell line ECV304 substantiated the role of astaxanthin in suppressing angiogenesis. Taken together, our data provide substantial evidence that dietary astaxanthin prevents the development and progression of HBP carcinomas through the inhibition of JAK-2/STAT-3 signaling and its downstream events. Thus, astaxanthin that functions as a potent inhibitor of tumour development and progression by targeting JAK/STAT signaling may be an ideal candidate for cancer chemoprevention. PMID:25296162

The antihypertension drug doxazosin suppresses JAK/STATs phosphorylation and enhances the effects of IFN-α/γ-induced apoptosis.

PubMed

Park, Mi Sun; Kim, Boh-Ram; Kang, Sokbom; Kim, Dae-Yong; Rho, Seung Bae

2014-11-01

Doxazosin, a commonly prescribed treatment for patients with benign prostatic hyperplasia, serves as an α1-blocker of the adrenergic receptors. In this study, we calculated its effect on the ovarian carcinoma cells. Doxazosin induces dose-dependent growth suppression and is additively activated through IFN-α or IFN-γ stimulation. They both enhanced G1 phase arrest, as well as the activity of caspase-3, and the reduction of cyclin D1 and CDK4 protein levels. Doxazosin growth suppression was abolished either by the Janus family of tyrosine kinase (JAK) or the signal transducer and activator of transcription (STAT) inhibitor treatment. The activity of JAK/STAT was dependent on the level of doxazosin, suggesting a requirement of doxazosin for the activation of JAK/STAT. Furthermore, doxazosin plus IFN-α or doxazosin plus IFN-γ additively suppressed the activation of the JAK/STAT signals through phosphorylation of JAK and STAT, thus affecting the activation of subsequent downstream signaling components PI3K, mTOR, 70S6K, and PKCδ. In vivo study demonstrated that doxazosin significantly suppressed tumor growth in an ovarian cancer cell xenograft mouse model, inducing apoptotic cell death by up-regulating the expression of p53, whereas c-Myc expression was markedly reduced. Our data indicate that doxazosin can modulate the apoptotic effects of IFN-α- and IFN-γ through the JAK/STAT signaling pathways. Collectively, we indicate that this action may be a potent chemotherapeutic property against ovarian carcinoma.

Cryptotanshinone suppresses the proliferation and induces the apoptosis of pancreatic cancer cells via the STAT3 signaling pathway.

PubMed

Ge, Yuqing; Yang, Bo; Chen, Zhe; Cheng, Rubin

2015-11-01

Pancreatic cancer remains a challenging disease worldwide. Cryptotanshinone (CPT) is one of the active constituents of Salvia miltiorrhiza Bunge and exhibits significant antitumor activities in several human cancer cells. However, the efficacy and molecular mechanism of CPT in pancreatic cancer remains to be elucidated. In the present study, the effect of CPT on the proliferation, apoptosis and cell cycle of human pancreatic cancer cell BxPC‑3 cells was evaluated. The results demonstrated that CPT inhibited proliferation of the BxPC‑3 cells in a concentration‑dependent manner, and significantly induced cell apoptosis and cell cycle arrest. The protein levels of cleaved caspase‑3, caspase‑9 and poly ADP ribose polymerase were upregulated, while the levels of c‑myc, survivin and cyclin D1 were downregulated following treatment with CPT. In addition, CPT decreased the activities of signal transducer and activator of transcription 3 (STAT3) and several upstream regulatory signaling pathways after 24 h. However, CPT only inhibited the phosphorylation of STAT3 Tyr705 within 30 min, without marked effects on the phosphorylation of the other proteins. These results suggested that the inhibition of STAT3 activity by CPT was directly and independent of the upstream regulators in human pancreatic cancer. The present study demonstrated that CPT exerts anticancer effects by inducing apoptosis and cell cycle arrest via inhibition of the STAT3 signaling pathway in human BxPC-3 cells.

CTLA4 Promotes Tyk2-STAT3-Dependent B-cell Oncogenicity.

PubMed

Herrmann, Andreas; Lahtz, Christoph; Nagao, Toshikage; Song, Joo Y; Chan, Wing C; Lee, Heehyoung; Yue, Chanyu; Look, Thomas; Mülfarth, Ronja; Li, Wenzhao; Jenkins, Kurt; Williams, John; Budde, Lihua E; Forman, Stephen; Kwak, Larry; Blankenstein, Thomas; Yu, Hua

2017-09-15

CTL-associated antigen 4 (CTLA4) is a well-established immune checkpoint for antitumor immune responses. The protumorigenic function of CTLA4 is believed to be limited to T-cell inhibition by countering the activity of the T-cell costimulating receptor CD28. However, as we demonstrate here, there are two additional roles for CTLA4 in cancer, including via CTLA4 overexpression in diverse B-cell lymphomas and in melanoma-associated B cells. CTLA4-CD86 ligation recruited and activated the JAK family member Tyk2, resulting in STAT3 activation and expression of genes critical for cancer immunosuppression and tumor growth and survival. CTLA4 activation resulted in lymphoma cell proliferation and tumor growth, whereas silencing or antibody-blockade of CTLA4 in B-cell lymphoma tumor cells in the absence of T cells inhibits tumor growth. This inhibition was accompanied by reduction of Tyk2/STAT3 activity, tumor cell proliferation, and induction of tumor cell apoptosis. The CTLA4-Tyk2-STAT3 signal pathway was also active in tumor-associated nonmalignant B cells in mouse models of melanoma and lymphoma. Overall, our results show how CTLA4-induced immune suppression occurs primarily via an intrinsic STAT3 pathway and that CTLA4 is critical for B-cell lymphoma proliferation and survival. Cancer Res; 77(18); 5118-28. ©2017 AACR . ©2017 American Association for Cancer Research.

Thymoquinone Augments Cisplatin-Induced Apoptosis on Esophageal Carcinoma Through Mitigating the Activation of JAK2/STAT3 Pathway.

PubMed

Hu, Xue; Ma, Jingjing; Vikash, Vikash; Li, Jiao; Wu, Dandan; Liu, Ya; Zhang, Jixiang; Dong, Weiguo

2018-01-01

Thymoquinone (TQ) is the major constituent of Nigella sativa seed and has shown biological activity in various human carcinomas. However, few studies have reported its effect on esophageal carcinoma (EC). To explore the chemosensitive effect and mechanism of TQ in augmentation of cisplatin (DDP)-induced apoptosis of EC, both in vitro and in vivo. The viability and apoptosis of esophageal carcinoma cells were detected by the Cell Counting Kit-8 assay, flow cytometry, and Hoechst 33258 staining. The expression levels of JAK2, p-JAK2, STAT3, p-STAT3, Bax, Bcl-2, Cyclin D1, Survivin, and caspase-3, 7, 9 were evaluated by western blot analysis. The histological changes were examined by TUNEL technique and immunohistochemical analysis. TQ enhanced the proapoptotic effect of DDP in human esophageal carcinoma cell line Eca-109, while blocking the activation of JAK2/STAT3 signaling pathway. The apoptosis of esophageal carcinoma cells was induced via blocking the activation of JAK2/STAT3 by using a molecular inhibitor (WP1066). Consistent with the in vivo and in vitro results, TQ increased cellular apoptosis and enriched the chemosensitivity of DDP. TQ along with DDP may regulate the progression of EC and has potential to be a chemotherapeutic agent in EC.

The Thyroid Hormone Receptors Inhibit Hepatic Interleukin-6 Signaling During Endotoxemia.

PubMed

Contreras-Jurado, Constanza; Alonso-Merino, Elvira; Saiz-Ladera, Cristina; Valiño, Arturo José; Regadera, Javier; Alemany, Susana; Aranda, Ana

2016-08-03

Decreased thyroidal hormone production is found during lipopolysaccharide (LPS)-induced endotoxic shock in animals as well as in critically ill patients. Here we studied the role of the thyroid hormone receptors (TRs) in activation of STAT3, NF-κB and ERK, which play a key role in the response to inflammatory cytokines during sepsis. TR knockout mice showed down-regulation of hepatic inflammatory mediators, including interleukin 6 (IL-6) in response to LPS. Paradoxically, STAT3 and ERK activity were higher, suggesting that TRs could act as endogenous repressors of these pathways. Furthermore, hyperthyroidism increased cytokine production and mortality in response to LPS, despite decreasing hepatic STAT3 and ERK activity. This suggested that TRs could directly repress the response of the cells to inflammatory mediators. Indeed, we found that the thyroid hormone T3 suppresses IL-6 signalling in macrophages and hepatocarcinoma cells, inhibiting STAT3 activation. Consequently, the hormone strongly antagonizes IL-6-stimulated gene transcription, reducing STAT3 recruitment and histone acetylation at IL-6 target promoters. In conclusion, TRs are potent regulators of inflammatory responses and immune homeostasis during sepsis. Reduced responses to IL-6 should serve as a negative feedback mechanism for preventing deleterious effects of excessive hormone signaling during infections.

Protective effects of calycosin against CCl4-induced liver injury with activation of FXR and STAT3 in mice.

PubMed

Chen, Xinli; Meng, Qiang; Wang, Changyuan; Liu, Qi; Sun, Huijun; Huo, Xiaokui; Sun, Pengyuan; Yang, Xiaobo; Peng, Jinyong; Liu, Kexin

2015-02-01

Investigating the hepatoprotective effect of calycosin against acute liver injury in association with FXR activation and STAT3 phosphorylation. The acute liver injury model was established by intraperitoneal injection of CCl4 in C57BL/6 mice. Serum alanine aminotransferase, aspartate aminotransferase, HE staining and TUNEL assay were used to identify the amelioration of the liver histopathological changes and hepatocytes apoptosis after calycosin treatment. ELISA kit and 5-bromo-2-deoxyuridine immunohistochemistry were used to measure the liver bile acid concentration and hepatocyte mitotic rate in vivo. The relation between calycosin and activation of FXR and STAT3 was comfirmed using the Luciferase assay, Molecular docking, Real-time PCR and Western Blot in vitro. The liver histopathological changes, hepatocytes apoptosis, liver bile acid overload and hepatocyte mitosis showed significant changes after calycosin treatment. Calycosin promoted the expression of FXR target genes such as FoxM1B and SHP but the effect was reversed by FXR suppressor guggulsterone. Molecular docking results indicated that calycosin could be embedded into the binding pocket of FXR, thereby increasing the expressions of STAT3 tyrosine phosphorylation and its target genes, Bcl-xl and SOCS3. Calycosin plays a critical role in hepatoprotection against liver injury in association with FXR activation and STAT3 phosphorylation.

The Thyroid Hormone Receptors Inhibit Hepatic Interleukin-6 Signaling During Endotoxemia

PubMed Central

Contreras-Jurado, Constanza; Alonso-Merino, Elvira; Saiz-Ladera, Cristina; Valiño, Arturo José; Regadera, Javier; Alemany, Susana; Aranda, Ana

2016-01-01

Decreased thyroidal hormone production is found during lipopolysaccharide (LPS)-induced endotoxic shock in animals as well as in critically ill patients. Here we studied the role of the thyroid hormone receptors (TRs) in activation of STAT3, NF-κB and ERK, which play a key role in the response to inflammatory cytokines during sepsis. TR knockout mice showed down-regulation of hepatic inflammatory mediators, including interleukin 6 (IL-6) in response to LPS. Paradoxically, STAT3 and ERK activity were higher, suggesting that TRs could act as endogenous repressors of these pathways. Furthermore, hyperthyroidism increased cytokine production and mortality in response to LPS, despite decreasing hepatic STAT3 and ERK activity. This suggested that TRs could directly repress the response of the cells to inflammatory mediators. Indeed, we found that the thyroid hormone T3 suppresses IL-6 signalling in macrophages and hepatocarcinoma cells, inhibiting STAT3 activation. Consequently, the hormone strongly antagonizes IL-6-stimulated gene transcription, reducing STAT3 recruitment and histone acetylation at IL-6 target promoters. In conclusion, TRs are potent regulators of inflammatory responses and immune homeostasis during sepsis. Reduced responses to IL-6 should serve as a negative feedback mechanism for preventing deleterious effects of excessive hormone signaling during infections. PMID:27484112

Activation of Antitumorigenic Stat3beta in Breast Cancer by Splicing Redirection

DTIC Science & Technology

2013-07-01

putative mapped ESEs (shown in green). (B) (Top) RT-PCR and (Bottom) Western Blot analysis of STAT3 a/b levels in MDA-MB-435s cells treated with...codon (PTC), ultimately causing RNA degradation following nonsense mediated decay (NMD). (B) RT-PCR and Western Blot analysis of STAT3 α/β levels in MDA...MB-435s cells treated with 16µM of ST6, ST7 or INV for 4 days. α-tubulin was used as loading control. (C) RT-PCR and Western Blot analysis of STAT3

Activation of T-cell Protein-tyrosine Phosphatase Suppresses Keratinocyte Survival and Proliferation following UVB Irradiation*

PubMed Central

Lee, Hyunseung; Morales, Liza D.; Slaga, Thomas J.; Kim, Dae Joon

2015-01-01

Chronic exposure to UV radiation can contribute to the development of skin cancer by promoting protein-tyrosine kinase (PTK) signaling. Studies show that exposure to UV radiation increases the ligand-independent activation of PTKs and induces protein-tyrosine phosphatase (PTP) inactivation. In the present work, we report that T-cell PTP (TC-PTP) activity is stimulated during the initial response to UVB irradiation, which leads to suppression of keratinocyte cell survival and proliferation via the down-regulation of STAT3 signaling. Our results show that TC-PTP-deficient keratinocyte cell lines expressed a significantly increased level of phosphorylated STAT3 after exposure to low dose UVB. This increase corresponded with increased cell proliferation in TC-PTP-deficient keratinocytes following UVB irradiation. Loss of TC-PTP also reduced UVB-induced apoptosis. Corroborating with these results, overexpression of TC-PTP in keratinocyte cell lines yielded a decrease in phosphorylated STAT3 levels, which corresponded with a significant decrease in cell proliferation in response to low dose UVB. We demonstrate that TC-PTP activity was increased upon UVB exposure, and overexpression of TC-PTP in keratinocyte cell lines further increased its activity in the presence of UVB. Treatment of TC-PTP-deficient keratinocytes with the STAT3 inhibitor STA21 significantly reduced cell viability following UVB exposure in comparison with untreated TC-PTP-deficient keratinocytes, confirming that the effect of TC-PTP on cell viability is mediated by STAT3 dephosphorylation. Combined, our results indicate that UVB-mediated activation of TC-PTP plays an important role in the STAT3-dependent regulation of keratinocyte cell proliferation and survival. Furthermore, these results suggest that TC-PTP may be a novel potential target for the prevention of UVB-induced skin cancer. PMID:25406309

New sulfurated derivatives of cinnamic acids and rosmaricine as inhibitors of STAT3 and NF-κB transcription factors.

PubMed

Gabriele, Elena; Brambilla, Dario; Ricci, Chiara; Regazzoni, Luca; Taguchi, Kyoko; Ferri, Nicola; Asai, Akira; Sparatore, Anna

2017-12-01

A set of new sulfurated drug hybrids, mainly derived from caffeic and ferulic acids and rosmaricine, has been synthesized and their ability to inhibit both STAT3 and NF-κB transcription factors have been evaluated. Results showed that most of the new hybrid compounds were able to strongly and selectively bind to STAT3, whereas the parent drugs were devoid of this ability at the tested concentrations. Some of them were also able to inhibit the NF-κB transcriptional activity in HCT-116 cell line and inhibited HCT-116 cell proliferation in vitro with IC 50 in micromolar range, thus suggesting a potential anticancer activity. Taken together, our study described the identification of new derivatives with dual STAT3/NF-κB inhibitory activity, which may represent hit compounds for developing multi-target anticancer agents.

Kurarinol induces hepatocellular carcinoma cell apoptosis through suppressing cellular signal transducer and activator of transcription 3 signaling

DOE Office of Scientific and Technical Information (OSTI.GOV)

Shu, Guangwen; Yang, Jing; Zhao, Wenhao

Kurarinol is a flavonoid isolated from roots of the medical plant Sophora flavescens. However, its cytotoxic activity against hepatocellular carcinoma (HCC) cells and toxic effects on mammalians remain largely unexplored. Here, the pro-apoptotic activities of kurarinol on HCC cells and its toxic impacts on tumor-bearing mice were evaluated. The molecular mechanisms underlying kurarinol-induced HCC cell apoptosis were also investigated. We found that kurarinol dose-dependently provoked HepG2, Huh-7 and H22 HCC cell apoptosis. In addition, kurarinol gave rise to a considerable decrease in the transcriptional activity of signal transducer and activator of transcription 3 (STAT3) in HCC cells. Suppression of STAT3more » signaling is involved in kurarinol-induced HCC cell apoptosis. In vivo studies showed that kurarinol injection substantially induced transplanted H22 cell apoptosis with low toxic impacts on tumor-bearing mice. Similarly, the transcriptional activity of STAT3 in transplanted tumor tissues was significantly suppressed after kurarinol treatment. Collectively, our current research demonstrated that kurarinol has the capacity of inducing HCC cell apoptosis both in vitro and in vivo with undetectable toxic impacts on the host. Suppressing STAT3 signaling is implicated in kurarinol-mediated HCC cell apoptosis. - Highlights: • Kurarinol induces hepatocellular carcinoma (HCC) cell apoptosis. • Kurarinol induces HCC cell apoptosis via inhibiting STAT3. • Kurarinol exhibits low toxic effects on tumor-bearing animals.« less

Understanding STAT3 signaling in cardiac ischemia.

PubMed

O'Sullivan, K E; Breen, E P; Gallagher, H C; Buggy, D J; Hurley, J P

2016-05-01

Cardiovascular disease is the leading cause of death worldwide. It remains one of the greatest challenges to global health and will continue to dominate mortality trends in the future. Acute myocardial infarction results in 7.4 million deaths globally per annum. Current management strategies are centered on restoration of coronary blood flow via percutaneous coronary intervention, coronary artery bypass grafting and administration of anti-platelet agents. Such myocardial reperfusion accounts for 40-50 % of the final infarct size in most cases. Signaling transducer and activator of transcription 3 (STAT3) has been shown to have cardioprotective effects via canonical and non-canonical activation and modulation of mitochondrial and transcriptional responses. A significant body of in vitro and in vivo evidence suggests that activation of the STAT3 signal transduction pathway results in a cardio protective response to ischemia and attempts have been made to modulate this with therapeutic effect. Not only is STAT3 important for cardiomyocyte function, but it also modulates the cardiac microenvironment and communicates with cardiac fibroblasts. To this end, we here review the current evidence supporting the manipulation of STAT3 for therapeutic benefit in cardiac ischemia and identify areas for future research.

IL-2 activation of STAT5 enhances production of IL-10 from human cytotoxic regulatory T cells, HOZOT.

PubMed

Tsuji-Takayama, Kazue; Suzuki, Motoyuki; Yamamoto, Mayuko; Harashima, Akira; Okochi, Ayumi; Otani, Takeshi; Inoue, Toshiya; Sugimoto, Akira; Motoda, Ryuichi; Yamasaki, Fumiyuki; Nakamura, Shuji; Kibata, Masayoshi

2008-02-01

Interleukin (IL)-10 is an immunosuppressive cytokine produced by many cell types, including T cells. We previously reported that a novel type of regulatory T (Treg) cells, termed HOZOT, which possesses a FOXP3+CD4+CD8+CD25+ phenotype and dual suppressor/cytotoxic activities, produced high levels of IL-10. In this study, we examined the mechanisms of high IL-10 production by HOZOT, focusing on Janus activating kinase (JAK)/signal transducers and activators of transcription (STAT) signaling pathway. We prepared five different types of T cells, including HOZOT from human umbilical cord blood. Cytokine productions of IL-10, interferon-gamma (IFN-gamma), and tumor necrosis factor-alpha (TNF-alpha) were compared among these T cells after anti-CD3/CD28 antibody stimulation in the presence or absence of IL-2. Specific inhibitors for JAK/STAT, nuclear factor-kappaB (NF-kappaB), and nuclear factor for activated T cell (NFAT) were used to analyze signal transduction mechanisms. IL-10 production by HOZOTs was greatly enhanced by the addition of IL-2. Little or no enhancement of IFN-gamma and TNF-alpha production was observed under the same conditions. The enhancing effect of IL-2 was specific for both HOZOT and IL-10-secreting Treg cells. T helper type 2 cells, whose IL-10 production mechanisms involve GATA-3, failed to show IL-2-mediated enhancement of IL-10. Similar enhancing effects of IL-15 and IFN-alpha suggested a major role of JAK/STAT activation pathway for high IL-10 production. Further inhibitor experiments demonstrated that STAT5 rather than STAT3 was critically involved in this mechanism. Our results demonstrated that IL-2 selectively enhanced production of IL-10 in HOZOT primarily through activation of STAT5, which synergistically acts with NF-kappaB/NFAT activation, implying a novel regulatory mechanism of IL-10 production in Treg cells.

Genotype tunes pancreatic ductal adenocarcinoma tissue tension to induce matricellular-fibrosis and tumor progression

PubMed Central

Laklai, Hanane; Miroshnikova, Yekaterina A.; Pickup, Michael W.; Collisson, Eric A.; Kim, Grace E.; Barrett, Alex S.; Hill, Ryan C.; Lakins, Johnathon N.; Schlaepfer, David D.; Mouw, Janna K.; LeBleu, Valerie S.; Roy, Nilotpal; Novitskiy, Sergey V.; Johansen, Julia S.; Poli, Valeria; Kalluri, Raghu; Iacobuzio-Donahue, Christine A.; Wood, Laura D.; Hebrok, Matthias; Hansen, Kirk; Moses, Harold L.; Weaver, Valerie M.

2016-01-01

Fibrosis compromises pancreatic ductal carcinoma (PDAC) treatment and contributes to patient mortality yet anti-stromal therapies are controversial. We found that human PDACs with impaired epithelial transforming growth factor β (TGF-β) signaling have elevated epithelial Stat3 activity and develop a stiffer, matricellular-enriched fibrosis associated with high epithelial tension and shorter patient survival. In several Kras-driven mouse models, both the loss of TGF-β signaling and elevated β1-integrin mechanosignaling engaged a positive feedback loop whereby Stat3 signaling promotes tumor progression by increasing matricellular fibrosis and tissue tension. In contrast, epithelial Stat3 ablation attenuated tumor progression by reducing the stromal stiffening and epithelial contractility induced by loss of TGF-β signaling. In PDAC patient biopsies, higher matricellular protein and activated Stat3 associated with SMAD4 mutation and shorter survival. The findings implicate epithelial tension and matricellular fibrosis in the aggressiveness of SMAD4 mutant pancreatic tumors, and highlight Stat3 and mechanics as key drivers of this phenotype. PMID:27089513

JAK/STAT3 and Smad3 activities are required for the wound healing properties of Periplaneta americana extracts.

PubMed

Song, Qin; Xie, Yuxin; Gou, Qiheng; Guo, Xiaoqiang; Yao, Qian; Gou, Xiaojun

2017-08-01

Periplaneta americana extracts (PAEs) play a crucial role in skin wound healing. However, their molecular effects and signaling pathways in regenerating tissues and cells are not clear. In this study, we refined the PAE from Periplaneta americana to investigate the mechanisms underlying skin wound healing. The human keratinocyte line HaCaT was selected and a mouse model of deep second-degree thermal burn was established for in vitro and in vivo studies, respectively. PAE treatment induced the proliferation and migration of HaCaT cells and wound healing in the burn model. Furthermore, the effects of PAE on wound healing were found to depend on the Janus-activated kinase/signal transducer and activator of transcription 3 (JAK/STAT3) pathway and Smad3 activities, according to western blot analysis and immunohistochemical (IHC) assays in vitro and in vivo. Pretreatment with a STAT3 inhibitor blocked the cell proliferation and migration induced by PAE. The results indicate the wound-healing function of PAE via enhanced JAK/STAT3 signaling and Smad3 activities. Our studies provide a theoretical basis underlying the role of PAE in cutaneous wound healing.

ROCK2 signaling is required to induce a subset of T follicular helper cells through opposing effects on STATs in autoimmune settings.

PubMed

Weiss, Jonathan M; Chen, Wei; Nyuydzefe, Melanie S; Trzeciak, Alissa; Flynn, Ryan; Tonra, James R; Marusic, Suzana; Blazar, Bruce R; Waksal, Samuel D; Zanin-Zhorov, Alexandra

2016-07-19

Rho-associated kinase 2 (ROCK2) determines the balance between human T helper 17 (TH17) cells and regulatory T (Treg) cells. We investigated its role in the generation of T follicular helper (TFH) cells, which help to generate antibody-producing B cells under normal and autoimmune conditions. Inhibiting ROCK2 in normal human T cells or peripheral blood mononuclear cells from patients with active systemic lupus erythematosus (SLE) decreased the number and function of TFH cells induced by activation ex vivo. Moreover, inhibition of ROCK2 activity decreased the abundance of the transcriptional regulator Bcl6 (B cell lymphoma 6) and increased that of Blimp1 by reducing the binding of signal transducer and activator of transcription 3 (STAT3) and increasing that of STAT5 to the promoters of the genes Bcl6 and PRDM1, respectively. In the MRL/lpr murine model of SLE, oral administration of the selective ROCK2 inhibitor KD025 resulted in a twofold reduction in the numbers of TFH cells and antibody-producing plasma cells in the spleen, as well as a decrease in the size of splenic germinal centers, which are the sites of interaction between TFH cells and B cells. KD025-treated mice showed a substantial improvement in both histological and clinical scores compared to those of untreated mice and had reduced amounts of Bcl6 and phosphorylated STAT3, as well as increased STAT5 phosphorylation. Together, these data suggest that ROCK2 signaling plays a critical role in controlling the development of TFH cells induced by autoimmune conditions through reciprocal regulation of STAT3 and STAT5 activation. Copyright © 2016, American Association for the Advancement of Science.

Epstein-Barr Virus Latent Membrane Protein 2A (LMP2A) enhances IL-10 production through the activation of Bruton's tyrosine kinase and STAT3.

PubMed

Incrocci, Ryan; Barse, Levi; Stone, Amanda; Vagvala, Sai; Montesano, Michael; Subramaniam, Vijay; Swanson-Mungerson, Michelle

2017-01-01

Previous data demonstrate that Epstein-Barr Virus Latent Membrane Protein 2A (LMP2A) enhances IL-10 to promote the survival of LMP2A-expressing B cell lymphomas. Since STAT3 is an important regulator of IL-10 production, we hypothesized that LMP2A activates a signal transduction cascade that increases STAT3 phosphorylation to enhance IL-10. Using LMP2A-negative and -positive B cell lines, the data indicate that LMP2A requires the early signaling molecules of the Syk/RAS/PI3K pathway to increase IL-10. Additional studies indicate that the PI3K-regulated kinase, BTK, is responsible for phosphorylating STAT3, which ultimately mediates the LMP2A-dependent increase in IL-10. These data are the first to show that LMP2A signaling results in STAT3 phosphorylation in B cells through a PI3K/BTK-dependent pathway. With the use of BTK and STAT3 inhibitors to treat B cell lymphomas in clinical trials, these findings highlight the possibility of using new pharmaceutical approaches to treat EBV-associated lymphomas that express LMP2A. Copyright © 2016 Elsevier Inc. All rights reserved.

Cellular STAT3 functions via PCBP2 to restrain Epstein-Barr Virus lytic activation in B lymphocytes.

PubMed

Koganti, Siva; Clark, Carissa; Zhi, Jizu; Li, Xiaofan; Chen, Emily I; Chakrabortty, Sharmistha; Hill, Erik R; Bhaduri-McIntosh, Sumita

2015-05-01

A major hurdle to killing Epstein-Barr virus (EBV)-infected tumor cells using oncolytic therapy is the presence of a substantial fraction of EBV-infected cells that does not support the lytic phase of EBV despite exposure to lytic cycle-promoting agents. To determine the mechanism(s) underlying this refractory state, we developed a strategy to separate lytic from refractory EBV-positive (EBV(+)) cells. By examining the cellular transcriptome in separated cells, we previously discovered that high levels of host STAT3 (signal transducer and activator of transcription 3) curtail the susceptibility of latently infected cells to lytic cycle activation signals. The goals of the present study were 2-fold: (i) to determine the mechanism of STAT3-mediated resistance to lytic activation and (ii) to exploit our findings to enhance susceptibility to lytic activation. We therefore analyzed our microarray data set, cellular proteomes of separated lytic and refractory cells, and a publically available STAT3 chromatin immunoprecipitation sequencing (ChIP-Seq) data set to identify cellular PCBP2 [poly(C)-binding protein 2], an RNA-binding protein, as a transcriptional target of STAT3 in refractory cells. Using Burkitt lymphoma cells and EBV(+) cell lines from patients with hypomorphic STAT3 mutations, we demonstrate that single cells expressing high levels of PCBP2 are refractory to spontaneous and induced EBV lytic activation, STAT3 functions via cellular PCBP2 to regulate lytic susceptibility, and suppression of PCBP2 levels is sufficient to increase the number of EBV lytic cells. We expect that these findings and the genome-wide resources that they provide will accelerate our understanding of a longstanding mystery in EBV biology and guide efforts to improve oncolytic therapy for EBV-associated cancers. Most humans are infected with Epstein-Barr virus (EBV), a cancer-causing virus. While EBV generally persists silently in B lymphocytes, periodic lytic (re)activation of latent virus is central to its life cycle and to most EBV-related diseases. However, a substantial fraction of EBV-infected B cells and tumor cells in a population is refractory to lytic activation. This resistance to lytic activation directly and profoundly impacts viral persistence and the effectiveness of oncolytic therapy for EBV(+) cancers. To identify the mechanisms that underlie susceptibility to EBV lytic activation, we used host gene and protein expression profiling of separated lytic and refractory cells. We find that STAT3, a transcription factor overactive in many cancers, regulates PCBP2, a protein important in RNA biogenesis, to regulate susceptibility to lytic cycle activation signals. These findings advance our understanding of EBV persistence and provide important leads on devising methods to improve viral oncolytic therapies. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Gut-derived commensal bacterial products inhibit liver dendritic cell maturation by stimulating hepatic interleukin-6/signal transducer and activator of transcription 3 activity.

PubMed

Lunz, John G; Specht, Susan M; Murase, Noriko; Isse, Kumiko; Demetris, Anthony J

2007-12-01

Intraorgan dendritic cells (DCs) monitor the environment and help translate triggers of innate immunity into adaptive immune responses. Liver-based DCs are continually exposed, via gut-derived portal venous blood, to potential antigens and bacterial products that can trigger innate immunity. However, somehow the liver avoids a state of perpetual inflammation and protects central immune organs from overstimulation. In this study, we tested the hypothesis that hepatic interleukin-6 (IL-6)/signal transducer and activator of transcription 3 (STAT3) activity increases the activation/maturation threshold of hepatic DCs toward innate immune signals. The results show that the liver nuclear STAT3 activity is significantly higher than that of other organs and is IL-6-dependent. Hepatic DCs in normal IL-6 wild-type (IL-6(+/+)) mice are phenotypically and functionally less mature than DCs from IL-6-deficient (IL-6(-/-)) or STAT3-inhibited IL-6(+/+) mice, as determined by surface marker expression, proinflammatory cytokine secretion, and allogeneic T-cell stimulation. IL-6(+/+) liver DCs produce IL-6 in response to exposure to lipopolysaccharide (LPS) and cytidine phosphate guanosine oligonucleotides (CpG) but are resistant to maturation compared with IL-6(-/-) liver DCs. Conversely, exogenous IL-6 inhibits LPS-induced IL-6(-/-) liver DC maturation. IL-6/STAT3 signaling influences the liver DC expression of toll-like receptor 9 and IL-1 receptor associated kinase-M. The depletion of gut commensal bacteria in IL-6(+/+) mice with oral antibiotics decreased portal blood endotoxin levels, lowered the expression of IL-6 and phospho-STAT3, and significantly increased liver DC maturation. Gut-derived bacterial products, by stimulating hepatic IL-6/STAT3 signaling, inhibit hepatic DC activation/maturation and thereby elevate the threshold needed for translating triggers of innate immunity into adaptive immune responses. Manipulating gut bacteria may therefore be an effective strategy for altering intrahepatic immune responses.

Cytokine receptor signaling is required for the survival of ALK− anaplastic large cell lymphoma, even in the presence of JAK1/STAT3 mutations

PubMed Central

Chen, Jing; Zhang, Yong; Petrus, Michael N.; Xiao, Wenming; Nicolae, Alina; Raffeld, Mark; Pittaluga, Stefania; Bamford, Richard N.; Nakagawa, Masao; Ouyang, Sunny Tianyi; Epstein, Alan L.; Kadin, Marshall E.; Del Mistro, Annarose; Woessner, Richard; Jaffe, Elaine S.; Waldmann, Thomas A.

2017-01-01

Activating Janus kinase (JAK) and signal transducer and activator of transcription (STAT) mutations have been discovered in many T-cell malignancies, including anaplastic lymphoma kinase (ALK)− anaplastic large cell lymphomas (ALCLs). However, such mutations occur in a minority of patients. To investigate the clinical application of targeting JAK for ALK− ALCL, we treated ALK− cell lines of various histological origins with JAK inhibitors. Interestingly, most exogenous cytokine-independent cell lines responded to JAK inhibition regardless of JAK mutation status. JAK inhibitor sensitivity correlated with the STAT3 phosphorylation status of tumor cells. Using retroviral shRNA knockdown, we have demonstrated that these JAK inhibitor-sensitive cells are dependent on both JAK1 and STAT3 for survival. JAK1 and STAT3 gain-of-function mutations were found in some, but not all, JAK inhibitor-sensitive cells. Moreover, the mutations alone cannot explain the JAK1/STAT3 dependency, given that wild-type JAK1 or STAT3 was sufficient to promote cell survival in the cells that had either JAK1or STAT3 mutations. To investigate whether other mechanisms were involved, we knocked down upstream receptors GP130 or IL-2Rγ. Knockdown of GP130 or IL-2Rγ induced cell death in selected JAK inhibitor-sensitive cells. High expression levels of cytokines, including IL-6, were demonstrated in cell lines as well as in primary ALK− ALCL tumors. Finally, ruxolitinib, a JAK1/2 inhibitor, was effective in vivo in a xenograft ALK− ALCL model. Our data suggest that cytokine receptor signaling is required for tumor cell survival in diverse forms of ALK− ALCL, even in the presence of JAK1/STAT3 mutations. Therefore, JAK inhibitor therapy might benefit patients with ALK− ALCL who are phosphorylated STAT3+. PMID:28356514

STAT3 activation in skeletal muscle links muscle wasting and the acute phase response in cancer cachexia.

PubMed

Bonetto, Andrea; Aydogdu, Tufan; Kunzevitzky, Noelia; Guttridge, Denis C; Khuri, Sawsan; Koniaris, Leonidas G; Zimmers, Teresa A

2011-01-01

Cachexia, or weight loss despite adequate nutrition, significantly impairs quality of life and response to therapy in cancer patients. In cancer patients, skeletal muscle wasting, weight loss and mortality are all positively associated with increased serum cytokines, particularly Interleukin-6 (IL-6), and the presence of the acute phase response. Acute phase proteins, including fibrinogen and serum amyloid A (SAA) are synthesized by hepatocytes in response to IL-6 as part of the innate immune response. To gain insight into the relationships among these observations, we studied mice with moderate and severe Colon-26 (C26)-carcinoma cachexia. Moderate and severe C26 cachexia was associated with high serum IL-6 and IL-6 family cytokines and highly similar patterns of skeletal muscle gene expression. The top canonical pathways up-regulated in both were the complement/coagulation cascade, proteasome, MAPK signaling, and the IL-6 and STAT3 pathways. Cachexia was associated with increased muscle pY705-STAT3 and increased STAT3 localization in myonuclei. STAT3 target genes, including SOCS3 mRNA and acute phase response proteins, were highly induced in cachectic muscle. IL-6 treatment and STAT3 activation both also induced fibrinogen in cultured C2C12 myotubes. Quantitation of muscle versus liver fibrinogen and SAA protein levels indicates that muscle contributes a large fraction of serum acute phase proteins in cancer. These results suggest that the STAT3 transcriptome is a major mechanism for wasting in cancer. Through IL-6/STAT3 activation, skeletal muscle is induced to synthesize acute phase proteins, thus establishing a molecular link between the observations of high IL-6, increased acute phase response proteins and muscle wasting in cancer. These results suggest a mechanism by which STAT3 might causally influence muscle wasting by altering the profile of genes expressed and translated in muscle such that amino acids liberated by increased proteolysis in cachexia are synthesized into acute phase proteins and exported into the blood.

STAT3 Activation in Skeletal Muscle Links Muscle Wasting and the Acute Phase Response in Cancer Cachexia

PubMed Central

Kunzevitzky, Noelia; Guttridge, Denis C.; Khuri, Sawsan; Koniaris, Leonidas G.; Zimmers, Teresa A.

2011-01-01

Background Cachexia, or weight loss despite adequate nutrition, significantly impairs quality of life and response to therapy in cancer patients. In cancer patients, skeletal muscle wasting, weight loss and mortality are all positively associated with increased serum cytokines, particularly Interleukin-6 (IL-6), and the presence of the acute phase response. Acute phase proteins, including fibrinogen and serum amyloid A (SAA) are synthesized by hepatocytes in response to IL-6 as part of the innate immune response. To gain insight into the relationships among these observations, we studied mice with moderate and severe Colon-26 (C26)-carcinoma cachexia. Methodology/Principal Findings Moderate and severe C26 cachexia was associated with high serum IL-6 and IL-6 family cytokines and highly similar patterns of skeletal muscle gene expression. The top canonical pathways up-regulated in both were the complement/coagulation cascade, proteasome, MAPK signaling, and the IL-6 and STAT3 pathways. Cachexia was associated with increased muscle pY705-STAT3 and increased STAT3 localization in myonuclei. STAT3 target genes, including SOCS3 mRNA and acute phase response proteins, were highly induced in cachectic muscle. IL-6 treatment and STAT3 activation both also induced fibrinogen in cultured C2C12 myotubes. Quantitation of muscle versus liver fibrinogen and SAA protein levels indicates that muscle contributes a large fraction of serum acute phase proteins in cancer. Conclusions/Significance These results suggest that the STAT3 transcriptome is a major mechanism for wasting in cancer. Through IL-6/STAT3 activation, skeletal muscle is induced to synthesize acute phase proteins, thus establishing a molecular link between the observations of high IL-6, increased acute phase response proteins and muscle wasting in cancer. These results suggest a mechanism by which STAT3 might causally influence muscle wasting by altering the profile of genes expressed and translated in muscle such that amino acids liberated by increased proteolysis in cachexia are synthesized into acute phase proteins and exported into the blood. PMID:21799891

Smad7 enables STAT3 activation and promotes pluripotency independent of TGF-β signaling

PubMed Central

Yu, Yi; Gu, Shuchen; Li, Wenjian; Sun, Chuang; Chen, Fenfang; Xiao, Mu; Wang, Lei; Xu, Dewei; Li, Ye; Ding, Chen; Xia, Zongping; Li, Yi; Ye, Sheng; Xu, Pinglong; Zhao, Bin; Qin, Jun; Chen, Ye-Guang; Lin, Xia; Feng, Xin-Hua

2017-01-01

Smad7 is a negative feedback product of TGF-β superfamily signaling and fine tunes a plethora of pleiotropic responses induced by TGF-β ligands. However, its noncanonical functions independent of TGF-β signaling remain to be elucidated. Here, we show that Smad7 activates signal transducers and activators of transcription 3 (STAT3) signaling in maintaining mouse embryonic stem cell pluripotency in a manner independent of the TGF-β receptors, yet dependent on the leukemia inhibitory factor (LIF) coreceptor glycoprotein 130 (gp130). Smad7 directly binds to the intracellular domain of gp130 and disrupts the SHP2–gp130 or SOCS3–gp130 complex, thereby amplifying STAT3 activation. Consequently, Smad7 facilitates LIF-mediated self-renewal of mouse ESCs and is also critical for induced pluripotent stem cell reprogramming. This finding illustrates an uncovered role of the Smad7–STAT3 interplay in maintaining cell pluripotency and also implicates a mechanism involving Smad7 underlying cytokine-dependent regulation of cancer and inflammation. PMID:28874583

The role of the JAK/STAT signal pathway in rheumatoid arthritis

PubMed Central

Malemud, Charles J.

2018-01-01

Proinflammatory cytokine activation of the Janus kinase/signal transducers and activators of transcription (JAK/STAT) signal transduction pathway is a critical event in the pathogenesis and progression of rheumatoid arthritis. Under normal conditions, JAK/STAT signaling reflects the influence of negative regulators of JAK/STAT, exemplified by the suppressor of cytokine signaling and protein inhibitor of activated STAT. However, in rheumatoid arthritis (RA) both of these regulators are dysfunctional. Thus, continuous activation of JAK/STAT signaling in RA synovial joints results in the elevated level of matrix metalloproteinase gene expression, increased frequency of apoptotic chondrocytes and most prominently ‘apoptosis resistance’ in the inflamed synovial tissue. Tofacitinib, a JAK small molecule inhibitor, with selectivity for JAK2/JAK3 was approved by the United States Food and Drug Administration (US FDA) for the therapy of RA. Importantly, tofacitinib has demonstrated significant clinical efficacy for RA in the post-US FDA-approval surveillance period. Of note, the success of tofacitinib has spurred the development of JAK1, JAK2 and other JAK3-selective small molecule inhibitors, some of which have also entered the clinical setting, whereas other JAK inhibitors are currently being evaluated in RA clinical trials. PMID:29942363

Imbalanced gp130 signalling in ApoE-deficient mice protects against atherosclerosis.

PubMed

Jones, Gareth W; McLeod, Louise; Kennedy, Catherine L; Bozinovski, Steven; Najdovska, Meri; Jenkins, Brendan J

2015-02-01

Interleukin (IL)-6 is a key modulator of the acute phase response (APR), and while both are implicated in atherosclerosis, the pathological role of specific IL-6 signalling cascades is ill-defined. Since IL-6 employs the cytokine receptor gp130 to primarily activate the STAT3 pathway, here we evaluate whether gp130-dependent STAT3 activation modulates atherosclerosis. High-fat diet-induced atherosclerosis was established in ApoE(-/-) mice crossed with gp130(F/F) knock-in mice displaying elevated gp130-dependent STAT3 activation and production of the APR protein, serum amyloid A (SAA). Also generated were gp130(F/F):Stat3(-/+):ApoE(-/-) mice displaying genetically-normalised STAT3 activation and SAA levels, and bone marrow chimeras involving ApoE(-/-) and gp130(F/F):ApoE(-/-) mice. At 10 weeks post high-fat diet, aortic atherosclerotic lesions, including the presence of CD68(+) macrophages, and plasma lipid and SAA profiles, were assessed. Aortic plaque development and plasma triglyceride levels in gp130(F/F):ApoE(-/-) mice were significantly reduced (3-fold, P < 0.001) compared to ApoE(-/-) littermates. By contrast, in gp130(F/F):ApoE(-/-) mice, atherosclerotic plaques contained augmented CD68(+) macrophage infiltrates, and plasma SAA levels were elevated, compared to ApoE(-/-) mice. Atherosclerotic lesion development and plasma triglyceride levels in gp130(F/F):ApoE(-/-) and gp130(F/F):Stat3(-/+):ApoE(-/-) mice were comparable, despite a significant (P < 0.05) reduction in macrophage numbers in lesions, and also plasma SAA levels, in gp130(F/F):Stat3(-/+):ApoE(-/-) mice. Aortic plaque development and plasma triglyceride levels were comparable in ApoE(-/-) mice reconstituted with gp130(F/F):ApoE(-/-) (ApoE(F/F:ApoE)) or ApoE(-/-) (ApoE(ApoE)) bone marrow cells. Deregulation of gp130/STAT3 signalling augments the APR and macrophage infiltration during atherosclerosis without impacting on the development of aortic plaques. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

SOCS3, a Major Regulator of Infection and Inflammation

PubMed Central

Carow, Berit; Rottenberg, Martin E.

2014-01-01

In this review, we describe the role of suppressor of cytokine signaling-3 (SOCS3) in modulating the outcome of infections and autoimmune diseases as well as the underlying mechanisms. SOCS3 regulates cytokine or hormone signaling usually preventing, but in some cases aggravating, a variety of diseases. A main role of SOCS3 results from its binding to both the JAK kinase and the cytokine receptor, which results in the inhibition of STAT3 activation. Available data also indicate that SOCS3 can regulate signaling via other STATs than STAT3 and also controls cellular pathways unrelated to STAT activation. SOCS3 might either act directly by hampering JAK activation or by mediating the ubiquitination and subsequent proteasome degradation of the cytokine/growth factor/hormone receptor. Inflammation and infection stimulate SOCS3 expression in different myeloid and lymphoid cell populations as well as in diverse non-hematopoietic cells. The accumulated data suggest a relevant program coordinated by SOCS3 in different cell populations, devoted to the control of immune homeostasis in physiological and pathological conditions such as infection and autoimmunity. PMID:24600449

Genetic partitioning of interleukin-6 signalling in mice dissociates Stat3 from Smad3-mediated lung fibrosis

PubMed Central

O'Donoghue, Robert J J; Knight, Darryl A; Richards, Carl D; Prêle, Cecilia M; Lau, Hui Ling; Jarnicki, Andrew G; Jones, Jessica; Bozinovski, Steven; Vlahos, Ross; Thiem, Stefan; McKenzie, Brent S; Wang, Bo; Stumbles, Philip; Laurent, Geoffrey J; McAnulty, Robin J; Rose-John, Stefan; Zhu, Hong Jian; Anderson, Gary P; Ernst, Matthias R; Mutsaers, Steven E

2012-01-01

Idiopathic pulmonary fibrosis (IPF) is a fatal disease that is unresponsive to current therapies and characterized by excessive collagen deposition and subsequent fibrosis. While inflammatory cytokines, including interleukin (IL)-6, are elevated in IPF, the molecular mechanisms that underlie this disease are incompletely understood, although the development of fibrosis is believed to depend on canonical transforming growth factor (TGF)-β signalling. We examined bleomycin-induced inflammation and fibrosis in mice carrying a mutation in the shared IL-6 family receptor gp130. Using genetic complementation, we directly correlate the extent of IL-6-mediated, excessive Stat3 activity with inflammatory infiltrates in the lung and the severity of fibrosis in corresponding gp130757F mice. The extent of fibrosis was attenuated in B lymphocyte-deficient gp130757F;µMT−/− compound mutant mice, but fibrosis still occurred in their Smad3−/− counterparts consistent with the capacity of excessive Stat3 activity to induce collagen 1α1 gene transcription independently of canonical TGF-β/Smad3 signalling. These findings are of therapeutic relevance, since we confirmed abundant STAT3 activation in fibrotic lungs from IPF patients and showed that genetic reduction of Stat3 protected mice from bleomycin-induced lung fibrosis. PMID:22684844

Scutellarin suppresses migration and invasion of human hepatocellular carcinoma by inhibiting the STAT3/Girdin/Akt activity.

PubMed

Ke, Yang; Bao, Tianhao; Wu, Xuesong; Tang, Haoran; Wang, Yan; Ge, Jiayun; Fu, Bimang; Meng, Xu; Chen, Li; Zhang, Cheng; Tan, Yuqi; Chen, Haotian; Guo, Zhitang; Ni, Fan; Lei, Xuefen; Shi, Zhitian; Wei, Dong; Wang, Lin

2017-01-29

Scutellarin is an active flavone from Erigeron breviscapine (vant) Hand Mass. This study aimed to investigate the potential role of scutellarin in migration and invasion of human hepatocellular carcinoma (HCC) cells and its possible mechanism. In comparison with the vehicle-treated controls, treatment with scutellarin (50 mg/kg/day) for 35 days significantly mitigated the lung and intrahepatic metastasis of HCC tumors in vivo. Scutellarin treatment significantly reduced HepG2 cell viability in a dose-dependent manner, and inhibited migration and invasion of HCC cells in vitro. Scutellarin treatment significantly reduced STAT3 and Girders of actin filaments (Girdin) expression, STAT3 and Akt phosphorylation in HCC cells. Introduction of STAT3 overexpression restored the scutellarin-downregulated Girdin expression, Akt activation, migration and invasion of HCC cells. Furthermore, induction of Girdin overexpression completely abrogated the inhibition of scutellarin on the Akt phosphorylation, migration and invasion of HCC cells. Scutellarin can inhibit HCC cell metastasis in vivo, and migration and invasion in vitro by down-regulating the STAT3/Girdin/Akt signaling. Copyright © 2016 Elsevier Inc. All rights reserved.

IL-6 secreted by cancer-associated fibroblasts promotes epithelial-mesenchymal transition and metastasis of gastric cancer via JAK2/STAT3 signaling pathway

PubMed Central

Li, Jie; Yu, Zhenjia; Wang, Xiaofeng; Li, Jiaanfang; Li, Chen; Yan, Min; Zhu, Zhenggang; Liu, Bingya; Su, Liping

2017-01-01

Cancer-associated fibroblasts (CAFs), as the activated fibroblasts in tumor stroma, are important modifiers of tumor progression. However, the molecular mechanisms underlying the tumor-promoting properties of CAFs in gastric cancer remain unclear. Here, we show that CAFs isolated from gastric cancer produce significant amounts of interleukin-6 (IL-6). CAFs enhances the migration and EMT of gastric cancer cells through the secretion of IL-6 that activates Janus kinase 2/signal transducers and activators of transcription (JAK2/STAT3) pathway in gastric cancer cells, while deprivation of IL-6 using a neutralizing antibody or inhibition of JAK/STAT3 pathway with specific inhibitor AG490 markedly attenuates these phenotypes in gastric cancer cells induced by CAFs. Moreover, silencing IL-6 expression in CAFs or inhibiting JAK2/STAT3 pathway in gastric cancer cells impairs tumor peritoneal metastasis induced by CAFs in vivo. Taken together, these results suggest that CAFs in the tumor microenvironment promote the progression of gastric cancer through IL-6/JAK2/STAT3 signaling, and IL-6 targeted therapy could be a complementary approach against gastric cancer by exerting their action on stromal fibroblasts. PMID:28186964

DOE Office of Scientific and Technical Information (OSTI.GOV)

Park, Yu-Kyoung; Lee, Tae-Yoon; Choi, Jong-Soon

Highlights: • Compound 7b, a meridianin C derivative, inhibits adipogenesis. • Compound 7b inhibits C/EBP-α, PPAR-γ, FAS, STAT-3, and STAT-5 in 3T3-L1 adipocytes. • Compound 7b inhibits leptin, but not adiponectin, expression in 3T3-L1 adipocytes. • Compound 7b thus may have therapeutic potential against obesity. - Abstract: Meridianin C, a marine alkaloid, is a potent protein kinase inhibitor and has anti-cancer activity. We have recently developed a series of meridianin C derivatives (compound 7a–7j) and reported their proviral integration Moloney Murine Leukemia Virus (pim) kinases’ inhibitory and anti-proliferative effects on human leukemia cells. Here we investigated the effect of thesemore » meridianin C derivatives on adipogenesis. Strikingly, among the derivatives tested, compound 7b most strongly inhibited lipid accumulation during the differentiation of 3T3-L1 preadipocytes into adipocytes. However, meridianin C treatment was largely cytotoxic to 3T3-L1 adipocytes. On mechanistic levels, compound 7b reduced not only the expressions of CCAAT/enhancer-binding protein-α (C/EBP-α), peroxisome proliferator-activated receptor-γ (PPAR-γ), and fatty acid synthase (FAS) but also the phosphorylation levels of signal transducer and activator of transcription-3 (STAT-3) and STAT-5 during adipocyte differentiation. Moreover, compound 7b repressed leptin, but not adiponectin, expression during adipocyte differentiation. Collectively, these findings demonstrate that a meridianin C derivative inhibits adipogenesis by down-regulating expressions and/or phosphorylations of C/EBP-α, PPAR-γ, FAS, STAT-3 and STAT-5.« less

C/EBPβ Promotes STAT3 Expression and Affects Cell Apoptosis and Proliferation in Porcine Ovarian Granulosa Cells.

PubMed

Yuan, Xiaolong; Zhou, Xiaofeng; He, Yingting; Zhong, Yuyi; Zhang, Ailing; Zhang, Zhe; Zhang, Hao; Li, Jiaqi

2018-06-13

Previous studies suggest that signal transducer and activator of transcription 3 (STAT3) and CCAAT/enhancer binding protein beta (C/EBPβ) play an essential role in ovarian granulosa cells (GCs) for mammalian follicular development. Several C/EBPβ putative binding sites were previously predicted on the STAT3 promoter in mammals. However, the molecular regulation of C/EBPβ on STAT3 and their effects on cell proliferation and apoptosis remain virtually unexplored in GCs. Using porcine GCs as a model, the 5′-deletion, luciferase report assay, mutation, chromatin immunoprecipitation, Annexin-V/PI staining and EdU assays were applied to investigate the molecular mechanism for C/EBPβ regulating the expression of STAT3 and their effects on the cell proliferation and apoptosis ability. We found that over and interfering with the expression of C/EBPβ significantly increased and decreased the messenger RNA (mRNA) and protein levels of STAT3 , respectively. The dual luciferase reporter assay showed that C/EBPβ directly bound at −1397/−1387 of STAT3 to positively regulate the mRNA and protein expressions of STAT3 . Both C/EBPβ and STAT3 were observed to inhibit cell apoptosis and promote cell proliferation. Furthermore, C/EBPβ might enhance the antiapoptotic and pro-proliferative effects of STAT3 . These results would be of great insight in further exploring the molecular mechanism of C/EBPβ and STAT3 on the function of GCs and the development of ovarian follicles in mammals.

Cardiac-Specific SOCS3 Deletion Prevents In Vivo Myocardial Ischemia Reperfusion Injury through Sustained Activation of Cardioprotective Signaling Molecules.

PubMed

Nagata, Takanobu; Yasukawa, Hideo; Kyogoku, Sachiko; Oba, Toyoharu; Takahashi, Jinya; Nohara, Shoichiro; Minami, Tomoko; Mawatari, Kazutoshi; Sugi, Yusuke; Shimozono, Koutatsu; Pradervand, Sylvain; Hoshijima, Masahiko; Aoki, Hiroki; Fukumoto, Yoshihiro; Imaizumi, Tsutomu

2015-01-01

Myocardial ischemia reperfusion injury (IRI) adversely affects cardiac performance and the prognosis of patients with acute myocardial infarction. Although myocardial signal transducer and activator of transcription (STAT) 3 is potently cardioprotective during IRI, the inhibitory mechanism responsible for its activation is largely unknown. The present study aimed to investigate the role of the myocardial suppressor of cytokine signaling (SOCS)-3, an intrinsic negative feedback regulator of the Janus kinase (JAK)-STAT signaling pathway, in the development of myocardial IRI. Myocardial IRI was induced in mice by ligating the left anterior descending coronary artery for 1 h, followed by different reperfusion times. One hour after reperfusion, the rapid expression of JAK-STAT-activating cytokines was observed. We precisely evaluated the phosphorylation of cardioprotective signaling molecules and the expression of SOCS3 during IRI and then induced myocardial IRI in wild-type and cardiac-specific SOCS3 knockout mice (SOCS3-CKO). The activation of STAT3, AKT, and ERK1/2 rapidly peaked and promptly decreased during IRI. This decrease correlated with the induction of SOCS3 expression up to 24 h after IRI in wild-type mice. The infarct size 24 h after reperfusion was significantly reduced in SOCS3-CKO compared with wild-type mice. In SOCS3-CKO mice, STAT3, AKT, and ERK1/2 phosphorylation was sustained, myocardial apoptosis was prevented, and the expression of anti-apoptotic Bcl-2 family member myeloid cell leukemia-1 (Mcl-1) was augmented. Cardiac-specific SOCS3 deletion led to the sustained activation of cardioprotective signaling molecules including and prevented myocardial apoptosis and injury during IRI. Our findings suggest that SOCS3 may represent a key factor that exacerbates the development of myocardial IRI.

Excess thyroid hormone inhibits embryonic neural stem/progenitor cells proliferation and maintenance through STAT3 signalling pathway.

PubMed

Chen, Chunhai; Zhou, Zhou; Zhong, Min; Li, Maoquan; Yang, Xuesen; Zhang, Yanwen; Wang, Yuan; Wei, Aimin; Qu, Mingyue; Zhang, Lei; Xu, Shangcheng; Chen, Shude; Yu, Zhengping

2011-07-01

Hyperthyroidism is prevalent during pregnancy, but little is known about the effects of excess thyroid hormone on the development of embryonic neural stem/progenitor cells (NSCs), and the mechanisms underlying these effects. Previous studies indicate that STAT3 plays a crucial role in determining NSC fate during neurodevelopment. In this study, we investigated the effects of a supraphysiological dose of 3,5,3'-L-triiodothyronine (T3) on the proliferation and maintenance of NSCs derived from embryonic day 13.5 mouse neocortex, and the involvement of STAT3 in this process. Our results suggest that excess T3 treatment inhibits NSC proliferation and maintenance. T3 decreased tyrosine phosphorylation of JAK1, JAK2 and STAT3, and subsequently inhibited STAT3-DNA binding activity. Furthermore, proliferation and maintenance of NSCs were decreased by inhibitors of JAKs and STAT3, indicating that the STAT3 signalling pathway is involved in the process of NSC proliferation and maintenance. Taken together, these results suggest that the STAT3 signalling pathway is involved in the process of T3-induced inhibition of embryonic NSC proliferation and maintenance. These findings provide data for understanding the effects of hyperthyroidism during pregnancy on fetal brain development, and the mechanisms underlying these effects.

Phosphoinositide 3-kinase/protein kinase B (PI3K/AKT) and Janus kinase/signal transducer and activator of transcription (JAK/STAT) follicular signalling is conserved in the mare ovary.

PubMed

Hall, Sally E; Upton, Rose M O; McLaughlin, Eileen A; Sutherland, Jessie M

2017-09-26

The mare ovary is unique in its anatomical structure; however, the signalling pathways responsible for physiological processes, such as follicular activation, remain uncharacterised. This provided us with the impetus to explore whether signalling molecules from important folliculogenesis pathways, phosphoinositide 3-kinase/protein kinase B (PI3K/AKT) and Janus kinase/signal transducer and activator of transcription (JAK/STAT), are conserved in the mare ovary. Messenger RNA expression of six genes important in follicle development was measured using quantitative polymerase chain reaction and protein localisation of key pathway members (PI3K, AKT1, phosphatase and tensin homologue (PTEN), JAK1, STAT3 and suppressor of cytokine signalling 4 (SOCS4)) was compared in tissue from fetal and adult mare ovaries. Tissue from adult ovaries exhibited significantly increased levels of mRNA expression of PI3K, AKT1, PTEN, JAK1, STAT3 and SOCS4 compared with tissue from fetal ovaries. PI3K, AKT1, JAK1 and STAT3 demonstrated redistributed localisation, from pregranulosa cells in fetal development, to both the oocyte and granulosa cells of follicles in the adult ovary, whilst negative feedback molecules PTEN and SOCS4 were only localised to the granulosa cells in the adult ovary. These findings suggest that the PI3K/AKT and JAK/STAT signalling pathways are utilised during folliculogenesis in the mare, similarly to previously studied mammalian species, and may serve as useful biomarkers for assessment of ovary development in the horse.

Differential Pro-Inflammatory Responses of Astrocytes and Microglia Involve STAT3 Activation in Response to 1800 MHz Radiofrequency Fields

PubMed Central

Lu, Yonghui; He, Mindi; Zhang, Yang; Xu, Shangcheng; Zhang, Lei; He, Yue; Chen, Chunhai; Liu, Chuan; Pi, Huifeng; Yu, Zhengping; Zhou, Zhou

2014-01-01

Microglia and astrocytes play important role in maintaining the homeostasis of central nervous system (CNS). Several CNS impacts have been postulated to be associated with radiofrequency (RF) electromagnetic fields exposure. Given the important role of inflammation in neural physiopathologic processes, we investigated the pro-inflammatory responses of microglia and astrocytes and the involved mechanism in response to RF fields. Microglial N9 and astroglial C8-D1A cells were exposed to 1800 MHz RF for different time with or without pretreatment with STAT3 inhibitor. Microglia and astrocytes were activated by RF exposure indicated by up-regulated CD11b and glial fibrillary acidic protein (GFAP). However, RF exposure induced differential pro-inflammatory responses in astrocytes and microglia, characterized by different expression and release profiles of IL-1β, TNF-α, IL-6, PGE2, nitric oxide (NO), inducible nitric oxide synthase (iNOS) and cyclooxygenase 2 (COX2). Moreover, the RF exposure activated STAT3 in microglia but not in astrocytes. Furthermore, the STAT3 inhibitor Stattic ameliorated the RF-induced release of pro-inflammatory cytokines in microglia but not in astrocytes. Our results demonstrated that RF exposure differentially induced pro-inflammatory responses in microglia and astrocytes, which involved differential activation of STAT3 in microglia and astrocytes. Our data provide novel insights into the potential mechanisms of the reported CNS impacts associated with mobile phone use and present STAT3 as a promising target to protect humans against increasing RF exposure. PMID:25275372

Differential pro-inflammatory responses of astrocytes and microglia involve STAT3 activation in response to 1800 MHz radiofrequency fields.

PubMed

Lu, Yonghui; He, Mindi; Zhang, Yang; Xu, Shangcheng; Zhang, Lei; He, Yue; Chen, Chunhai; Liu, Chuan; Pi, Huifeng; Yu, Zhengping; Zhou, Zhou

2014-01-01

Microglia and astrocytes play important role in maintaining the homeostasis of central nervous system (CNS). Several CNS impacts have been postulated to be associated with radiofrequency (RF) electromagnetic fields exposure. Given the important role of inflammation in neural physiopathologic processes, we investigated the pro-inflammatory responses of microglia and astrocytes and the involved mechanism in response to RF fields. Microglial N9 and astroglial C8-D1A cells were exposed to 1800 MHz RF for different time with or without pretreatment with STAT3 inhibitor. Microglia and astrocytes were activated by RF exposure indicated by up-regulated CD11b and glial fibrillary acidic protein (GFAP). However, RF exposure induced differential pro-inflammatory responses in astrocytes and microglia, characterized by different expression and release profiles of IL-1β, TNF-α, IL-6, PGE2, nitric oxide (NO), inducible nitric oxide synthase (iNOS) and cyclooxygenase 2 (COX2). Moreover, the RF exposure activated STAT3 in microglia but not in astrocytes. Furthermore, the STAT3 inhibitor Stattic ameliorated the RF-induced release of pro-inflammatory cytokines in microglia but not in astrocytes. Our results demonstrated that RF exposure differentially induced pro-inflammatory responses in microglia and astrocytes, which involved differential activation of STAT3 in microglia and astrocytes. Our data provide novel insights into the potential mechanisms of the reported CNS impacts associated with mobile phone use and present STAT3 as a promising target to protect humans against increasing RF exposure.

Transcriptional regulation of the novel monoamine oxidase renalase: Crucial roles of transcription factors Sp1, STAT3, and ZBP89.

PubMed

Sonawane, Parshuram J; Gupta, Vinayak; Sasi, Binu K; Kalyani, Ananthamohan; Natarajan, Bhargavi; Khan, Abrar A; Sahu, Bhavani S; Mahapatra, Nitish R

2014-11-11

Renalase, a novel monoamine oxidase, is emerging as an important regulator of cardiovascular, metabolic, and renal diseases. However, the mechanism of transcriptional regulation of this enzyme remains largely unknown. We undertook a systematic analysis of the renalase gene to identify regulatory promoter elements and transcription factors. Computational analysis coupled with transfection of human renalase promoter/luciferase reporter plasmids (5'-promoter-deletion constructs) into various cell types (HEK-293, IMR32, and HepG2) identified two crucial promoter domains at base pairs -485 to -399 and -252 to -150. Electrophoretic mobility shift assays using renalase promoter oligonucleotides with and without potential binding sites for transcription factors Sp1, STAT3, and ZBP89 displayed formation of specific complexes with HEK-293 nuclear proteins. Consistently, overexpression of Sp1, STAT3, and ZBP89 augmented renalase promoter activity; additionally, siRNA-mediated downregulation of Sp1, STAT3, and ZBP89 reduced the level of endogenous renalase transcription as well as the transfected renalase promoter activity. In addition, chromatin immunoprecipitation assays showed in vivo interactions of these transcription factors with renalase promoter. Interestingly, renalase promoter activity was augmented by nicotine and catecholamines; while Sp1 and STAT3 synergistically activated the nicotine-induced effect, Sp1 appeared to enhance epinephrine-evoked renalase transcription. Moreover, renalase transcript levels in mouse models of human essential hypertension were concomitantly associated with endogenous STAT3 and ZBP89 levels, suggesting crucial roles for these transcription factors in regulating renalase gene expression in cardiovascular pathological conditions.

Inhibition of STAT-3 Results in Radiosensitization of Human Squamous Cell Carcinoma

PubMed Central

Bonner, James A.; Trummell, Hoa Q.; Willey, Christopher D.; Plants, Brian A.; Raisch, Kevin P.

2009-01-01

Background Signal Transducer and Activator of Transcription – 3 (STAT-3) is a downstream component of the Epidermal Growth Factor Receptor (EGFr) signaling process that may facilitate the resistance of tumor cells to conventional cancer treatments. Studies were performed to determine if inhibition of this downstream protein may produce radiosensitization. Methods/Results A431 cells (human squamous cell carcinoma cells with EGFr overexpression) were found to be sensitized to radiation after treatment with STAT-3 small interfering RNA (siRNA). Therefore, a short hairpin RNA (shRNA) against STAT-3 was designed and cloned into a pBABE vector system modified for shRNA expression. Following transfection, clone 2.1 was selected for further study as it showed a dramatic reduction of STAT-3 protein (and mRNA) when compared to A431 parental cells or a negative control shRNA cell line (transfected with STAT-3 shRNA with 2 base pairs mutated). A431 2.1 showed doubling times of 25-31 h as compared to 18-24 h for the parental cell line. The A431 shRNA knockdown STAT-3 cells A431 were more sensitive to radiation than A431 parental or negative STAT-3 control cells. Conclusion A431 cells stably transfected with shRNA against STAT-3 resulted in enhanced radiosensitivity. Further work will be necessary to determine whether inhibition of STAT-3 phosphorylation is a necessary step for the radiosensitization that is induced by inhibition of EGFr. PMID:19616333

Staphylococcal enterotoxin A (SEA) stimulates STAT3 activation and IL-17 expression in cutaneous T-cell lymphoma.

PubMed

Willerslev-Olsen, Andreas; Krejsgaard, Thorbjørn; Lindahl, Lise M; Litvinov, Ivan V; Fredholm, Simon; Petersen, David L; Nastasi, Claudia; Gniadecki, Robert; Mongan, Nigel P; Sasseville, Denis; Wasik, Mariusz A; Bonefeld, Charlotte M; Geisler, Carsten; Woetmann, Anders; Iversen, Lars; Kilian, Mogens; Koralov, Sergei B; Odum, Niels

2016-03-10

Cutaneous T-cell lymphoma (CTCL) is characterized by proliferation of malignant T cells in a chronic inflammatory environment. With disease progression, bacteria colonize the compromised skin barrier and half of CTCL patients die of infection rather than from direct organ involvement by the malignancy. Clinical data indicate that bacteria play a direct role in disease progression, but little is known about the mechanisms involved. Here, we demonstrate that bacterial isolates containing staphylococcal enterotoxin A (SEA) from the affected skin of CTCL patients, as well as recombinant SEA, stimulate activation of signal transducer and activator of transcription 3 (STAT3) and upregulation of interleukin (IL)-17 in immortalized and primary patient-derived malignant and nonmalignant T cells. Importantly, SEA induces STAT3 activation and IL-17 expression in malignant T cells when cocultured with nonmalignant T cells, indicating an indirect mode of action. In accordance, malignant T cells expressing an SEA-nonresponsive T-cell receptor variable region β chain are nonresponsive to SEA in monoculture but display strong STAT3 activation and IL-17 expression in cocultures with SEA-responsive nonmalignant T cells. The response is induced via IL-2 receptor common γ chain cytokines and a Janus kinase 3 (JAK3)-dependent pathway in malignant T cells, and blocked by tofacitinib, a clinical-grade JAK3 inhibitor. In conclusion, we demonstrate that SEA induces cell cross talk-dependent activation of STAT3 and expression of IL-17 in malignant T cells, suggesting a mechanism whereby SEA-producing bacteria promote activation of an established oncogenic pathway previously implicated in carcinogenesis. © 2016 by The American Society of Hematology.

Inhibition of STAT3 activity delays obesity-induced thyroid carcinogenesis in a mouse model

PubMed Central

Park, Jeong Won; Han, Cho Rong; Zhao, Li; Willingham, Mark C.; Cheng, Sheue-yann

2015-01-01

Compelling epidemiologic studies indicate that obesity is a risk factor for many human cancers, including thyroid cancer. In recent decades, the incidence of thyroid cancer has dramatically increased along with a marked rise in obesity prevalence. We previously demonstrated that a high fat diet (HFD) effectively induced the obese phenotype in a mouse model of thyroid cancer (ThrbPV/PVPten+/− mice). Moreover, HFD activates the STAT3 signal pathway to promote more aggressive tumor phenotypes. The aim of the present study was to evaluate the effect of S3I-201, a specific inhibitor of STAT3 activity, on HFD-induced aggressive cancer progression in the mouse model of thyroid cancer. Wild type and ThrbPV/PVPten+/− mice were treated with HFD together with S3I-201 or vehicle-only as controls. We assessed the effects of S3I-201 on HFD-induced thyroid cancer progression, the leptin-JAK2-STAT3 signaling pathway, and key regulators of epithelial-mesenchymal transition. S3I-201 effectively inhibited HFD-induced aberrant activation of STAT3 and its downstream targets to markedly inhibit thyroid tumor growth and to prolong survival. Decreased protein levels of cyclins D1 and B1, cyclin dependent kinase (CDK) 4, CDK 6, and phosphorylated retinoblastoma protein led to the inhibition of tumor cell proliferation in S3I-201-treated ThrbPV/PVPten+/− mice. Reduced occurrence of vascular invasion and blocking of anaplasia and lung metastasis in thyroid tumors of S3I-201-treated ThrbPV/PVPten+/− mice were mediated via decreased expression of vimentin and matrix metalloproteinases, two key effectors of epithelial-mesenchymal transition. The present findings suggest that inhibition of the STAT3 activity would be a novel treatment strategy for obesity-induced thyroid cancer. PMID:26552408

The Growth Hormone Receptor: Mechanism of Receptor Activation, Cell Signaling, and Physiological Aspects

PubMed Central

Dehkhoda, Farhad; Lee, Christine M. M.; Medina, Johan; Brooks, Andrew J.

2018-01-01

The growth hormone receptor (GHR), although most well known for regulating growth, has many other important biological functions including regulating metabolism and controlling physiological processes related to the hepatobiliary, cardiovascular, renal, gastrointestinal, and reproductive systems. In addition, growth hormone signaling is an important regulator of aging and plays a significant role in cancer development. Growth hormone activates the Janus kinase (JAK)–signal transducer and activator of transcription (STAT) signaling pathway, and recent studies have provided a new understanding of the mechanism of JAK2 activation by growth hormone binding to its receptor. JAK2 activation is required for growth hormone-mediated activation of STAT1, STAT3, and STAT5, and the negative regulation of JAK–STAT signaling comprises an important step in the control of this signaling pathway. The GHR also activates the Src family kinase signaling pathway independent of JAK2. This review covers the molecular mechanisms of GHR activation and signal transduction as well as the physiological consequences of growth hormone signaling. PMID:29487568

Signal Transducer and Activator of Transcription 3, Mediated Remodeling of the Tumor Microenvironment Results in Enhanced Tumor Drug Delivery in a Mouse Model of Pancreatic Cancer.

PubMed

Nagathihalli, Nagaraj S; Castellanos, Jason A; Shi, Chanjuan; Beesetty, Yugandhar; Reyzer, Michelle L; Caprioli, Richard; Chen, Xi; Walsh, Alex J; Skala, Melissa C; Moses, Harold L; Merchant, Nipun B

2015-12-01

A hallmark of pancreatic ductal adenocarcinoma (PDAC) is the presence of a dense desmoplastic reaction (stroma) that impedes drug delivery to the tumor. Attempts to deplete the tumor stroma have resulted in formation of more aggressive tumors. We have identified signal transducer and activator of transcription (STAT) 3 as a biomarker of resistance to cytotoxic and molecularly targeted therapy in PDAC. The purpose of this study is to investigate the effects of targeting STAT3 on the PDAC stroma and on therapeutic resistance. Activated STAT3 protein expression was determined in human pancreatic tissues and tumor cell lines. In vivo effects of AZD1480, a JAK/STAT3 inhibitor, gemcitabine or the combination were determined in Ptf1a(cre/+);LSL-Kras(G12D/+);Tgfbr2(flox/flox) (PKT) mice and in orthotopic tumor xenografts. Drug delivery was analyzed by matrix-assisted laser desorption/ionization imaging mass spectrometry. Collagen second harmonic generation imaging quantified tumor collagen alignment and density. STAT3 activation correlates with decreased survival and advanced tumor stage in patients with PDAC. STAT3 inhibition combined with gemcitabine significantly inhibits tumor growth in both an orthotopic and the PKT mouse model of PDAC. This combined therapy attenuates in vivo expression of SPARC, increases microvessel density, and enhances drug delivery to the tumor without depletion of stromal collagen or hyaluronan. Instead, the PDAC tumors demonstrate vascular normalization, remodeling of the tumor stroma, and down-regulation of cytidine deaminase. Targeted inhibition of STAT3 combined with gemcitabine enhances in vivo drug delivery and therapeutic response in PDAC. These effects occur through tumor stromal remodeling and down-regulation of cytidine deaminase without depletion of tumor stromal content. Copyright © 2015 AGA Institute. Published by Elsevier Inc. All rights reserved.

The potency of STAT (signal transducers and activators of transcription) 3 protein as growth promoter for chicken

NASA Astrophysics Data System (ADS)

Ma'ruf, Anwar; Iswati, Sri; Hidajati, Nove; Damayanti, Ratna

2017-09-01

The long-term objective of this study was to produce STAT synthetic protein in chicken during growth period resulting from the increase of growth hormone (GH) as growth promoter. This study used ten male chicken Lohman from PT. Multibreeder Indonesia. The chicken were kept within batteried cage, with a capacity of one chicken in each cage. The chickens were fed twice a day, at 6 a.m. and 6 p.m. with the amount of feed 10% less than standard. On day 21 the chicken were slaughtered to obtain the samples, i.e., adipose, liver and muscles for the following examinations (1) isolation of STAT-3 signaling protein from adipose, liver and muscles of the chicken, (2) analysis of STAT-3 signaling protein using SDS-PAGE method, and (3) identification of STAT-3 signaling protein using Western blot method by means of protein detection using electrophoresis with polyacrylamide gels. Results of examination on protein in hepatic, muscle and adipose of chickens in growth period revealed that STAT protein was positively present in those tissues. This finding was followed-up with SDS-PAGE examination, from which we found the presence of protein band between the markers of 116 kDa and 14.4 kDa. The protein band was supposedly the STAT-3 protein. To prove that protein band formed was the STAT-3, Western blot examination was conducted using rabbit polyclonal antibody STAT-3. The result showed the formation of the protein band, indicating the presence of reaction between antigen (STAT-3 protein) and STAT-3 protein antibody. In conclusion, STAT-3 protein is present in hepatic, muscular, and adipose tissues, with molecular weight of 59.4 kDa.