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Sample records for aberrant notch signaling

  1. Immunohistochemical expression of aberrant Notch-1 signaling in vitiligo: an implication for pathogenesis.

    PubMed

    Seleit, Iman; Bakry, Ola Ahmed; Abdou, Asmaa Gaber; Dawoud, Noha Mohammed

    2014-06-01

    The etiopathogenetic mechanisms leading to pigment loss in vitiligo are not fully understood. Notch signaling is required for development and maintenance of melanocyte lineage and acts as a key component among keratinocyte-melanocyte interactions. The current study aimed to investigate the possible role of Notch signaling and its effect on the whole melanocyte lineage in vitiligo and correlating it with the different clinicopathologic parameters. Using immunohistochemical technique, Notch-1 expression was evaluated in 50 lesional and 20 perilesional biopsies of patients with vitiligo in comparison with 20 normal skin biopsies as a control group. Lesional biopsies were stained with human melanoma black-45 and tyrosinase-related protein-2 to demonstrate the melanocyte lineage. Membranous and/or nuclear expression of Notch-1 was in favor of control and perilesional skin, whereas cytoplasmic expression appeared only in vitiliginous lesions (P < .05). Membranous and/or nuclear expression of Notch-1 was significantly associated with epidermal human melanoma black-45 positivity (P = .01) and percentage of expression in both epidermis (P = .02) and hair follicles (P = .03) of lesional skin. Cytoplasmic pattern of Notch-1 expression in epidermis was significantly found in lesions with white hair (P = .04) and in cases with marked keratinocyte vacuolization (P = .03). Segmental and acrofacial vitiligo were associated with mild to moderate Notch-1 intensity, whereas generalized vitiligo was associated with strong intensity of expression (P = .02). In conclusion, Notch-1 signaling is inactivated in vitiligo with consequent loss of epidermal and/or follicular active melanocytes. Aberrant Notch signaling in vitiliginous white hair and acral and segmental vitiligo may be the cause of their treatment resistance.

  2. Aberrant activation of canonical Notch1 signaling in the mouse uterus decreases progesterone receptor by hypermethylation and leads to infertility

    PubMed Central

    Su, Ren-Wei; Strug, Michael R.; Jeong, Jae-Wook; Miele, Lucio; Fazleabas, Asgerally T.

    2016-01-01

    In mammalian reproduction, implantation is one of the most critical events. Failure of implantation and the subsequent decidualization contribute to more than 75% of pregnancy losses in women. Our laboratory has previously reported that inhibition of Notch signaling results in impaired decidualization in both women and a transgenic mouse model. In this study, we generated a Notch gain-of-function transgenic mouse by conditionally overexpressing the Notch1 intracellular domain (N1ICD) in the reproductive tract driven by a progesterone receptor (Pgr) -Cre. We show that the overexpression of N1ICD in the uterus results in complete infertility as a consequence of multiple developmental and physiological defects, including the absence of uterine glands and dysregulation of progesterone and estrogen signaling by a Recombination Signal Binding Protein Jκ-dependent signaling mechanism. We further show that the inhibition of progesterone signaling is caused by hypermethylation of its receptor Pgr by Notch1 overexpression through the transcription factor PU.1 and DNA methyltransferase 3b (Dnmt3b). We have generated a mouse model to study the consequence of increased Notch signaling in female reproduction and provide the first evidence, to our knowledge, that Notch signaling can regulate epigenetic modification of the Pgr. PMID:26858409

  3. Notch Signaling Components

    PubMed Central

    Liu, Zhi-Yan; Wu, Tao; Li, Qing; Wang, Min-Cong; Jing, Li; Ruan, Zhi-Ping; Yao, Yu; Nan, Ke-Jun; Guo, Hui

    2016-01-01

    Abstract Non-small-cell lung cancer (NSCLC) is a lethal and aggressive malignancy. Currently, the identities of prognostic and predictive makers of NSCLC have not been fully established. Dysregulated Notch signaling has been implicated in many human malignancies, including NSCLC. However, the prognostic value of measuring Notch signaling and the utility of developing Notch-targeted therapies in NSCLC remain inconclusive. The present study investigated the association of individual Notch receptor and ligand levels with lung adenocarcinoma (ADC) and squamous cell carcinoma (SCC) prognosis using the Kaplan-Meier plotte database. This online database encompasses 2437 lung cancer samples. Hazard ratios with 95% confidence intervals were calculated. The results showed that higher Notch1, Notch2, JAG1, and DLL1 mRNA expression predicted better overall survival (OS) in lung ADC, but showed no significance in SCC patients. Elevated Notch3, JAG2, and DLL3 mRNA expression was associated with poor OS of ADC patients, but not in SCC patients. There was no association between Notch4 and OS in either lung ADC or SCC patients. In conclusion, the set of Notch1, Notch2, JAG1, DLL1 and that of Notch3, JAG2, DLL3 played opposing prognostic roles in lung ADC patients. Neither set of Notch receptors and ligands was indicative of lung SCC prognosis. Notch signaling could serve as promising marker to predict outcomes in lung ADC patients. The distinct features of lung cancer subtypes and Notch components should be considered when developing future Notch-targeted therapies. PMID:27196489

  4. Notch Signaling in Pancreatic Development

    PubMed Central

    Li, Xu-Yan; Zhai, Wen-Jun; Teng, Chun-Bo

    2015-01-01

    The Notch signaling pathway plays a significant role in embryonic cell fate determination and adult tissue homeostasis. Various studies have demonstrated the deep involvement of Notch signaling in the development of the pancreas and the lateral inhibition of Notch signaling in pancreatic progenitor differentiation and maintenance. The targeted inactivation of the Notch pathway components promotes premature differentiation of the endocrine pancreas. However, there is still the contrary opinion that Notch signaling specifies the endocrine lineage. Here, we review the current knowledge of the Notch signaling pathway in pancreatic development and its crosstalk with the Wingless and INT-1 (Wnt) and fibroblast growth factor (FGF) pathways. PMID:26729103

  5. Notch Signaling in Vascular Smooth Muscle Cells.

    PubMed

    Baeten, J T; Lilly, B

    2017-01-01

    The Notch signaling pathway is a highly conserved pathway involved in cell fate determination in embryonic development and also functions in the regulation of physiological processes in several systems. It plays an especially important role in vascular development and physiology by influencing angiogenesis, vessel patterning, arterial/venous specification, and vascular smooth muscle biology. Aberrant or dysregulated Notch signaling is the cause of or a contributing factor to many vascular disorders, including inherited vascular diseases, such as cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy, associated with degeneration of the smooth muscle layer in cerebral arteries. Like most signaling pathways, the Notch signaling axis is influenced by complex interactions with mediators of other signaling pathways. This complexity is also compounded by different members of the Notch family having both overlapping and unique functions. Thus, it is vital to fully understand the roles and interactions of each Notch family member in order to effectively and specifically target their exact contributions to vascular disease. In this chapter, we will review the Notch signaling pathway in vascular smooth muscle cells as it relates to vascular development and human disease.

  6. Recent advances on NOTCH signaling in T-ALL.

    PubMed

    Tzoneva, Gannie; Ferrando, Adolfo A

    2012-01-01

    NOTCH1 receptor signaling plays a central role in T-cell lineage specification and in supporting the growth and proliferation of immature T-cell progenitors in the thymus during lymphoid development. In T-cell acute lymphoblastic leukemia (T-ALL), a tumor resulting from the malignant transformation of T-cell progenitors, aberrant and constitutively active NOTCH1 signaling triggered by activating mutations in the NOTCH1 gene contributes to oncogenic transformation and is a hallmark of this disease. Most notably, small molecule γ-secretase inhibitors (GSIs) can effectively block NOTCH1 signaling in T-ALL, and could be exploited as a targeted therapy in this disease. In addition, a number of emerging anti-NOTCH therapeutic strategies including anti-NOTCH1 inhibitory antibodies, small peptide inhibitors of NOTCH signaling and combination therapies with GSIs and glucocorticoids, have recently been proposed. Finally, the identification of NOTCH1 mutations in solid tumors and chronic lymphocytic leukemias has increased even further the clinical relevance of NOTCH signaling as a therapeutic target in human cancer. Here we review our current understanding of NOTCH1-induced transformation, the mechanisms of action of oncogenic NOTCH1 in T-ALL and the therapeutic and prognostic implications of NOTCH1 mutations in T-ALL.

  7. Notch signaling: its roles and therapeutic potential in hematological malignancies.

    PubMed

    Gu, Yisu; Masiero, Massimo; Banham, Alison H

    2016-05-17

    Notch is a highly conserved signaling system that allows neighboring cells to communicate, thereby controlling their differentiation, proliferation and apoptosis, with the outcome of its activation being highly dependent on signal strength and cell type. As such, there is growing evidence that disturbances in physiological Notch signaling contribute to cancer development and growth through various mechanisms. Notch was first reported to contribute to tumorigenesis in the early 90s, through identification of the involvement of the Notch1 gene in the chromosomal translocation t(7;9)(q34;q34.3), found in a small subset of T-cell acute lymphoblastic leukemia. Since then, Notch mutations and aberrant Notch signaling have been reported in numerous other precursor and mature hematological malignancies, of both myeloid and lymphoid origin, as well as many epithelial tumor types. Of note, Notch has been reported to have both oncogenic and tumor suppressor roles, dependent on the cancer cell type. In this review, we will first give a general description of the Notch signaling pathway, and its physiologic role in hematopoiesis. Next, we will review the role of aberrant Notch signaling in several hematological malignancies. Finally, we will discuss current and potential future therapeutic approaches targeting this pathway.

  8. Notch signaling: its roles and therapeutic potential in hematological malignancies

    PubMed Central

    Gu, Yisu

    2016-01-01

    Notch is a highly conserved signaling system that allows neighboring cells to communicate, thereby controlling their differentiation, proliferation and apoptosis, with the outcome of its activation being highly dependent on signal strength and cell type. As such, there is growing evidence that disturbances in physiological Notch signaling contribute to cancer development and growth through various mechanisms. Notch was first reported to contribute to tumorigenesis in the early 90s, through identification of the involvement of the Notch1 gene in the chromosomal translocation t(7;9)(q34;q34.3), found in a small subset of T-cell acute lymphoblastic leukemia. Since then, Notch mutations and aberrant Notch signaling have been reported in numerous other precursor and mature hematological malignancies, of both myeloid and lymphoid origin, as well as many epithelial tumor types. Of note, Notch has been reported to have both oncogenic and tumor suppressor roles, dependent on the cancer cell type. In this review, we will first give a general description of the Notch signaling pathway, and its physiologic role in hematopoiesis. Next, we will review the role of aberrant Notch signaling in several hematological malignancies. Finally, we will discuss current and potential future therapeutic approaches targeting this pathway. PMID:26934331

  9. Notch Signaling in Neuroendocrine Tumors

    PubMed Central

    Crabtree, Judy S.; Singleton, Ciera S.; Miele, Lucio

    2016-01-01

    Carcinoids and neuroendocrine tumors (NETs) are a heterogeneous group of tumors that arise from the neuroendocrine cells of the GI tract, endocrine pancreas, and the respiratory system. NETs remain significantly understudied with respect to molecular mechanisms of pathogenesis, particularly the role of cell fate signaling systems such as Notch. The abundance of literature on the Notch pathway is a testament to its complexity in different cellular environments. Notch receptors can function as oncogenes in some contexts and tumor suppressors in others. The genetic heterogeneity of NETs suggests that to fully understand the roles and the potential therapeutic implications of Notch signaling in NETs, a comprehensive analysis of Notch expression patterns and potential roles across all NET subtypes is required. PMID:27148486

  10. Notch Signaling Inhibits Axon Regeneration

    PubMed Central

    Bejjani, Rachid El; Hammarlund, Marc

    2013-01-01

    Summary Many neurons have limited capacity to regenerate their axons after injury. Neurons in the mammalian CNS do not regenerate, and even neurons in the PNS often fail to regenerate to their former targets. This failure is likely due in part to pathways that actively restrict regeneration; however, only a few factors that limit regeneration are known. Here, using single-neuron analysis of regeneration in vivo, we show that Notch/lin-12 signaling inhibits the regeneration of mature C. elegans neurons. Notch signaling suppresses regeneration by acting autonomously in the injured cell to prevent growth cone formation. The metalloprotease and gamma-secretase cleavage events that lead to Notch activation during development are also required for its activity in regeneration. Furthermore, blocking Notch activation immediately after injury improves regeneration. Our results define a novel, post-developmental role for the Notch pathway as a repressor of axon regeneration in vivo. PMID:22284182

  11. Notch signaling and new therapeutic options in liver disease.

    PubMed

    Morell, Carola Maria; Strazzabosco, Mario

    2014-04-01

    Notch signaling is a crucial determinant of cell fate decision during development and disease in several organs. Notch effects are strictly dependent on the cellular context in which it is activated. In the liver, Notch signaling is involved in biliary tree development and tubulogenesis. Recent advances have shed light on Notch as a critical player in liver regeneration and repair, as well as in liver metabolism and inflammation and cancer. Notch signaling is finely regulated at several levels. The complexity of the pathway provides several possible targets for development of therapeutic agents able to inhibit Notch. Recent reports have shown that persistent activation of Notch signaling is associated with liver malignancies, particularly hepatocellular with stem cell features and cholangiocarcinoma. These novel findings suggest that interfering with the aberrant activation of the Notch pathway may have therapeutic relevance. However, further studies are needed to clarify the mechanisms regulating physiologic and pathologic Notch activation in the adult liver, to better understand the mechanistic role(s) of Notch in liver diseases and to develop safe and specific therapeutic agents.

  12. Role of Notch and its oncogenic signaling crosstalk in breast cancer

    PubMed Central

    Guo, Shanchun; Liu, Mingli; Gonzalez-Perez, Ruben R.

    2011-01-01

    The Notch signaling plays a key role in cell differentiation, survival, and proliferation through diverse mechanisms. Notch signaling is also involved in vasculogenesis and angiogenesis. Moreover, Notch expression is regulated by hypoxia and inflammatory cytokines (IL-1, IL-6 and leptin). Entangled crosstalk between Notch and other developmental signaling (Hedgehog and Wnt), and signaling triggered by growth factors, estrogens and oncogenic kinases, could impact on Notch targeted genes. Thus, alterations of the Notch signaling can lead to a variety of disorders, including human malignancies. Notch signaling is activated by ligand binding, followed by ADAM/Tumor necrosis factor-α-converting enzyme (TACE) metalloprotease and γ-secretase cleavages that produce the Notch intracellular domain (NICD). Translocation of NICD into the nucleus induces the transcriptional activation of Notch target genes. The relationships between Notch deregulated signaling, cancer stem cells and the carcinogenesis process reinforced by Notch crosstalk with many oncogenic signaling pathways suggest that Notch signaling may be a critical drug target for breast and other cancers. Since current status of knowledge in this field changes quickly, our insight should be continuously revised. In this review, we will focus on recent advancements in identification of aberrant Notch signaling in breast cancer and the possible underlying mechanisms, including potential role of Notch in breast cancer stem cells, tumor angiogenesis, as well as its crosstalk with other oncogenic signaling pathways in breast cancer. We will also discuss the prognostic value of Notch proteins and therapeutic potential of targeting Notch signaling for cancer treatment. PMID:21193018

  13. Notch signaling deregulation in multiple myeloma: A rational molecular target

    PubMed Central

    Garavelli, Silvia; Platonova, Natalia; Paoli, Alessandro; Basile, Andrea; Taiana, Elisa; Neri, Antonino; Chiaramonte, Raffaella

    2015-01-01

    Despite recent therapeutic advances, multiple myeloma (MM) is still an incurable neoplasia due to intrinsic or acquired resistance to therapy. Myeloma cell localization in the bone marrow milieu allows direct interactions between tumor cells and non-tumor bone marrow cells which promote neoplastic cell growth, survival, bone disease, acquisition of drug resistance and consequent relapse. Twenty percent of MM patients are at high-risk of treatment failure as defined by tumor markers or presentation as plasma cell leukemia. Cumulative evidences indicate a key role of Notch signaling in multiple myeloma onset and progression. Unlike other Notch-related malignancies, where the majority of patients carry gain-of-function mutations in Notch pathway members, in MM cell Notch signaling is aberrantly activated due to an increased expression of Notch receptors and ligands; notably, this also results in the activation of Notch signaling in surrounding stromal cells which contributes to myeloma cell proliferation, survival and migration, as well as to bone disease and intrinsic and acquired pharmacological resistance. Here we review the last findings on the mechanisms and the effects of Notch signaling dysregulation in MM and provide a rationale for a therapeutic strategy aiming at inhibiting Notch signaling, along with a complete overview on the currently available Notch-directed approaches. PMID:26308486

  14. Notch signaling regulates gastric antral LGR5 stem cell function.

    PubMed

    Demitrack, Elise S; Gifford, Gail B; Keeley, Theresa M; Carulli, Alexis J; VanDussen, Kelli L; Thomas, Dafydd; Giordano, Thomas J; Liu, Zhenyi; Kopan, Raphael; Samuelson, Linda C

    2015-10-14

    The major signaling pathways regulating gastric stem cells are unknown. Here we report that Notch signaling is essential for homeostasis of LGR5(+) antral stem cells. Pathway inhibition reduced proliferation of gastric stem and progenitor cells, while activation increased proliferation. Notch dysregulation also altered differentiation, with inhibition inducing mucous and endocrine cell differentiation while activation reduced differentiation. Analysis of gastric organoids demonstrated that Notch signaling was intrinsic to the epithelium and regulated growth. Furthermore, in vivo Notch manipulation affected the efficiency of organoid initiation from glands and single Lgr5-GFP stem cells, suggesting regulation of stem cell function. Strikingly, constitutive Notch activation in LGR5(+) stem cells induced tissue expansion via antral gland fission. Lineage tracing using a multi-colored reporter demonstrated that Notch-activated stem cells rapidly generate monoclonal glands, suggesting a competitive advantage over unmanipulated stem cells. Notch activation was associated with increased mTOR signaling, and mTORC1 inhibition normalized NICD-induced increases in proliferation and gland fission. Chronic Notch activation induced undifferentiated, hyper-proliferative polyps, suggesting that aberrant activation of Notch in gastric stem cells may contribute to gastric tumorigenesis.

  15. Small molecule activation of NOTCH signaling inhibits acute myeloid leukemia

    PubMed Central

    Ye, Qi; Jiang, Jue; Zhan, Guanqun; Yan, Wanyao; Huang, Liang; Hu, Yufeng; Su, Hexiu; Tong, Qingyi; Yue, Ming; Li, Hua; Yao, Guangmin; Zhang, Yonghui; Liu, Hudan

    2016-01-01

    Aberrant activation of the NOTCH signaling pathway is crucial for the onset and progression of T cell leukemia. Yet recent studies also suggest a tumor suppressive role of NOTCH signaling in acute myeloid leukemia (AML) and reactivation of this pathway offers an attractive opportunity for anti-AML therapies. N-methylhemeanthidine chloride (NMHC) is a novel Amaryllidaceae alkaloid that we previously isolated from Zephyranthes candida, exhibiting inhibitory activities in a variety of cancer cells, particularly those from AML. Here, we report NMHC not only selectively inhibits AML cell proliferation in vitro but also hampers tumor development in a human AML xenograft model. Genome-wide gene expression profiling reveals that NMHC activates the NOTCH signaling. Combination of NMHC and recombinant human NOTCH ligand DLL4 achieves a remarkable synergistic effect on NOTCH activation. Moreover, pre-inhibition of NOTCH by overexpression of dominant negative MAML alleviates NMHC-mediated cytotoxicity in AML. Further mechanistic analysis using structure-based molecular modeling as well as biochemical assays demonstrates that NMHC docks in the hydrophobic cavity within the NOTCH1 negative regulatory region (NRR), thus promoting NOTCH1 proteolytic cleavage. Our findings thus establish NMHC as a potential NOTCH agonist that holds great promises for future development as a novel agent beneficial to patients with AML. PMID:27211848

  16. Notch signaling in cerebrovascular diseases (Review)

    PubMed Central

    Cai, Zhiyou; Zhao, Bin; Deng, Yanqing; Shangguan, Shouqin; Zhou, Faming; Zhou, Wenqing; Li, Xiaoli; Li, Yanfeng; Chen, Guanghui

    2016-01-01

    The Notch signaling pathway is a crucial regulator of numerous fundamental cellular processes. Increasing evidence suggests that Notch signaling is involved in inflammation and oxidative stress, and thus in the progress of cerebrovascular diseases. In addition, Notch signaling in cerebrovascular diseases is associated with apoptosis, angiogenesis and the function of blood-brain barrier. Despite the contradictory results obtained to date as to whether Notch signaling is harmful or beneficial, the regulation of Notch signaling may provide a novel strategy for the treatment of cerebrovascular diseases. PMID:27574001

  17. Notch signalling in placental development and gestational diseases.

    PubMed

    Haider, S; Pollheimer, J; Knöfler, M

    2017-01-16

    Activation of Notch signalling upon cell-cell contact of neighbouring cells controls a plethora of cellular processes such as stem cell maintenance, cell lineage determination, cell proliferation, and survival. Accumulating evidence suggests that the pathway also critically regulates these events during placental development and differentiation. Herein, we summarize our present knowledge about Notch signalling in murine and human placentation and discuss its potential role in the pathophysiology of gestational disorders. Studies in mice suggest that Notch controls trophectoderm formation, decidualization, placental branching morphogenesis and endovascular trophoblast invasion. In humans, the particular signalling cascade promotes formation of the extravillous trophoblast lineage and regulates trophoblast proliferation, survival and differentiation. Expression patterns as well as functional analyses indicate distinct roles of Notch receptors in different trophoblast subtypes. Altered effects of Notch signalling have been detected in choriocarcinoma cells, consistent with its role in cancer development and progression. Moreover, deregulation of Notch signalling components were observed in pregnancy disorders such as preeclampsia and fetal growth restriction. In summary, Notch plays fundamental roles in different developmental processes of the placenta. Abnormal signalling through this pathway could contribute to the pathogenesis of gestational diseases with aberrant placentation and trophoblast function.

  18. Hypoxia, notch signalling, and prostate cancer.

    PubMed

    Marignol, Laure; Rivera-Figueroa, Karla; Lynch, Thomas; Hollywood, Donal

    2013-07-01

    The notch signalling pathway is involved in differentiation, proliferation, angiogenesis, vascular remodelling, and apoptosis. Deregulated expression of notch receptors, ligands, and targets is observed in many solid tumours, including prostate cancer. Hypoxia is a common feature of prostate tumours, leading to increased gene instability, reduced treatment response, and increased tumour aggressiveness. The notch signalling pathway is known to regulate vascular cell fate and is responsive to hypoxia-inducible factors. Evidence to date suggests similar, therapeutically exploitable, behaviour of notch-activated and hypoxic prostate cancer cells.

  19. Phosphorylation-dependent regulation of Notch1 signaling: the fulcrum of Notch1 signaling.

    PubMed

    Lee, Hye-Jin; Kim, Mi-Yeon; Park, Hee-Sae

    2015-08-01

    Notch signaling plays a pivotal role in cell fate determination, cellular development, cellular self-renewal, tumor progression, and has been linked to developmental disorders and carcinogenesis. Notch1 is activated through interactions with the ligands of neighboring cells, and acts as a transcriptional activator in the nucleus. The Notch1 intracellular domain (Notch1-IC) regulates the expression of target genes related to tumor development and progression. The Notch1 protein undergoes modification after translation by posttranslational modification enzymes. Phosphorylation modification is critical for enzymatic activation, complex formation, degradation, and subcellular localization. According to the nuclear cycle, Notch1-IC is degraded by E3 ligase, FBW7 in the nucleus via phosphorylation-dependent degradation. Here, we summarize the Notch signaling pathway, and resolve to understand the role of phosphorylation in the regulation of Notch signaling as well as to understand its relation to cancer.

  20. Notch Signaling Functions in Retinal Pericyte Survival

    PubMed Central

    Arboleda-Velasquez, Joseph F.; Primo, Vincent; Graham, Mark; James, Alexandra; Manent, Jan; D'Amore, Patricia A.

    2014-01-01

    Purpose. Pericytes, the vascular cells that constitute the outer layer of capillaries, have been shown to have a crucial role in vascular development and stability. Loss of pericytes precedes endothelial cell dysfunction and vascular degeneration in small-vessel diseases, including diabetic retinopathy. Despite their clinical relevance, the cellular pathways controlling survival of retinal pericytes remain largely uncharacterized. Therefore, we investigated the role of Notch signaling, a master regulator of cell fate decisions, in retinal pericyte survival. Methods. A coculture system of ligand-dependent Notch signaling was developed using primary cultured retinal pericytes and a mesenchymal cell line derived from an inducible mouse model expressing the Delta-like 1 Notch ligand. This model was used to examine the effect of Notch activity on pericyte survival using quantitative PCR (qPCR) and a light-induced cell death assay. The effect of Notch gain- and loss-of-function was analyzed in monocultures of retinal pericytes using antibody arrays to interrogate the expression of apoptosis-related proteins. Results. Primary cultured retinal pericytes differentially expressed key molecules of the Notch pathway and displayed strong expression of canonical Notch/RBPJK (recombination signal-binding protein 1 for J-kappa) downstream targets. A gene expression screen using gain- and loss-of-function approaches identified genes relevant to cell survival as downstream targets of Notch activity in retinal pericytes. Ligand-mediated Notch activity protected retinal pericytes from light-induced cell death. Conclusions. Our results have identified signature genes downstream of Notch activity in retinal pericytes and suggest that tight regulation of Notch signaling is crucial for pericyte survival. PMID:25015359

  1. Aberrant expression of Notch1, HES1, and DTX1 genes in glioblastoma formalin-fixed paraffin-embedded tissues.

    PubMed

    Narayanappa, Rajeswari; Rout, Pritilata; Aithal, Madhuri G S; Chand, Ashis Kumar

    2016-05-01

    Glioblastoma is the most common malignant brain tumor accounting for more than 54 % of all gliomas. Despite aggressive treatments, median survival remains less than 1 year. This might be due to the unavailability of effective molecular diagnostic markers and targeted therapy. Thus, it is essential to discover molecular mechanisms underlying disease by identifying dysregulated pathways involved in tumorigenesis. Notch signaling is one such pathway which plays an important role in determining cell fates. Since it is found to play a critical role in many cancers, we investigated the role of Notch genes in glioblastoma with an aim to identify biomarkers that can improve diagnosis. Using real-time PCR, we assessed the expression of Notch genes including receptors (Notch1, Notch2, Notch3, and Notch4), ligands (JAG1, JAG2, and DLL3), downstream targets (HES1 and HEY2), regulator Deltex1 (DTX1), inhibitor NUMB along with transcriptional co-activator MAML1, and a component of gamma-secretase complex APH1A in 15 formalin-fixed paraffin-embedded (FFPE) patient samples. Relative quantification was done by the 2(-ΔΔCt) method; the data are presented as fold change in gene expression normalized to an internal control gene and relative to the calibrator. The data revealed aberrant expression of Notch genes in glioblastoma compared to normal brain. More than 85 % of samples showed high Notch1 (P = 0.0397) gene expression and low HES1 (P = 0.011) and DTX1 (P = 0.0001) gene expression. Our results clearly show aberrant expression of Notch genes in glioblastoma which can be used as putative biomarkers together with histopathological observation to improve diagnosis, therapeutic strategies, and patient prognosis.

  2. [Abnormal Notch-Hes Signaling Pathways and Acute Leukemia -Review].

    PubMed

    Gu, Zhen-Yang; Wang, Li; Gao, Chun-Ji

    2017-02-01

    The abnormal activation of Notch signaling is closely related to the development of acute leukemia (AL). The core elements of the Notch signaling system include Notch receptors, Notch ligands, CSL DNA-binding proteins, and effectors like target genes. Any factors, which affect ligands, receptors, signal transducers and effectors, can influence the signal transduction of Notch signaling greatly. Based on the role of Notch signaling in AL, several targeted drugs against Notch upstream signaling have been developed. However, due to the complexity and pleiotropic effects of Notch upstream signaling, these targeted drugs display strong side effects. Thus, Hes (Hairy Enhancer of Split) factors as a primary Notch effector, also play an important role in the pathogenesis of AL. This review summarizes recent progresses on Notch-Hes signaling in AL, hopping to provide references for further excavation of the Notch-Hes signaling, and lay foundations for developing the next generation of targeted drugs.

  3. Non-degradative ubiquitination of the Notch1 receptor by the E3 ligase MDM2 activates the Notch signalling pathway.

    PubMed

    Pettersson, Susanne; Sczaniecka, Matylda; McLaren, Lorna; Russell, Fiona; Gladstone, Karen; Hupp, Ted; Wallace, Maura

    2013-03-15

    The Notch receptor is necessary for modulating cell fate decisions throughout development, and aberrant activation of Notch signalling has been associated with many diseases, including tumorigenesis. The E3 ligase MDM2 (murine double minute 2) plays a role in regulating the Notch signalling pathway through its interaction with NUMB. In the present study we report that MDM2 can also exert its oncogenic effects on the Notch signalling pathway by directly interacting with the Notch 1 receptor through dual-site binding. This involves both the N-terminal and acidic domains of MDM2 and the RAM [RBP-Jκ (recombination signal-binding protein 1 for Jκ)-associated molecule] and ANK (ankyrin) domains of Notch 1. Although the interaction between Notch1 and MDM2 results in ubiquitination of Notch1, this does not result in degradation of Notch1, but instead leads to activation of the intracellular domain of Notch1. Furthermore, MDM2 can synergize with Notch1 to inhibit apoptosis and promote proliferation. This highlights yet another target for MDM2-mediated ubiquitination that results in activation of the protein rather than degradation and makes MDM2 an attractive target for drug discovery for both the p53 and Notch signalling pathways.

  4. Notch signaling in the developing cardiovascular system.

    PubMed

    Niessen, Kyle; Karsan, Aly

    2007-07-01

    The Notch proteins encompass a family of transmembrane receptors that have been highly conserved through evolution as mediators of cell fate. Recent findings have demonstrated a critical role of Notch in the developing cardiovascular system. Notch signaling has been implicated in the endothelial-to-mesenchymal transition during development of the heart valves, in arterial-venous differentiation, and in remodeling of the primitive vascular plexus. Mutations of Notch pathway components in humans are associated with congenital defects of the cardiovascular system such as Alagille syndrome, cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL), and bicuspid aortic valves. This article focuses on the role of the Notch pathway in the developing cardiovascular system and congenital human cardiovascular diseases.

  5. Overexpression of protein O-fucosyltransferase 1 accelerates hepatocellular carcinoma progression via the Notch signaling pathway.

    PubMed

    Ma, Lijie; Dong, Pingping; Liu, Longzi; Gao, Qiang; Duan, Meng; Zhang, Si; Chen, She; Xue, Ruyi; Wang, Xiaoying

    2016-04-29

    Aberrant activation of Notch signaling frequently occurs in liver cancer, and is associated with liver malignancies. However, the mechanisms regulating pathologic Notch activation in hepatocellular carcinoma (HCC) remain unclear. Protein O-fucosyltransferase 1 (Pofut1) catalyzes the addition of O-linked fucose to the epidermal growth factor-like repeats of Notch. In the present study, we detected the expression of Pofut1 in 8 HCC cell lines and 253 human HCC tissues. We reported that Pofut1 was overexpressed in HCC cell lines and clinical HCC tissues, and Pofut1 overexpression clinically correlated with the unfavorable survival and high disease recurrence in HCC. The in vitro assay demonstrated that Pofut1 overexpression accelerated the cell proliferation and migration in HCC cells. Furthermore, Pofut1 overexpression promoted the binding of Notch ligand Dll1 to Notch receptor, and hence activated Notch signaling pathway in HCC cells, indicating that Pofut1 overexpression could be a reason for the aberrant activation of Notch signaling in HCC. Taken together, our findings indicated that an aberrant activated Pofut1-Notch pathway was involved in HCC progression, and blockage of this pathway could be a promising strategy for the therapy of HCC.

  6. Regulation of angiogenesis via Notch signaling in breast cancer and cancer stem cells.

    PubMed

    Zhou, Weiqiang; Wang, Guangdi; Guo, Shanchun

    2013-12-01

    Breast cancer angiogenesis is elicited and regulated by a number of factors including the Notch signaling. Notch receptors and ligands are expressed in breast cancer cells as well as in the stromal compartment and have been implicated in carcinogenesis. Signals exchanged between neighboring cells through the Notch pathway can amplify and consolidate molecular differences, which eventually dictate cell fates. Notch signaling and its crosstalk with many signaling pathways play an important role in breast cancer cell growth, migration, invasion, metastasis and angiogenesis, as well as cancer stem cell (CSC) self-renewal. Therefore, significant attention has been paid in recent years toward the development of clinically useful antagonists of Notch signaling. Better understanding of the structure, function and regulation of Notch intracellular signaling pathways, as well as its complex crosstalk with other oncogenic signals in breast cancer cells will be essential to ensure rational design and application of new combinatory therapeutic strategies. Novel opportunities have emerged from the discovery of Notch crosstalk with inflammatory and angiogenic cytokines and their links to CSCs. Combinatory treatments with drugs designed to prevent Notch oncogenic signal crosstalk may be advantageous over λ secretase inhibitors (GSIs) alone. In this review, we focus on the more recent advancements in our knowledge of aberrant Notch signaling contributing to breast cancer angiogenesis, as well as its crosstalk with other factors contributing to angiogenesis and CSCs.

  7. Notch Signaling in Bone Regeneration

    DTIC Science & Technology

    2011-10-01

    Notch delivery, a large animal model (e.g., sheep ) could be developed and tested in another 2 years. Within five years, we will have a product to...phenotype ( macrophage , endothelial cell, osteoblast, chondrocyte) using dual antibody labeling with antibodies for specific cell types. Additionally, we

  8. Overview of genetic tools and techniques to study Notch signaling in mice.

    PubMed

    Gridley, Thomas; Groves, Andrew K

    2014-01-01

    Aberrations of Notch signaling in humans cause both congenital and acquired defects and cancers. Genetically engineered mice provide the most efficient and cost-effective models to study Notch signaling in a mammalian system. Here, we review the various types of genetic models, tools, and strategies to study Notch signaling in mice, and provide examples of their use. We also provide advice on breeding strategies for conditional mutant mice, and a protocol for tamoxifen administration to mouse strains expressing inducible Cre recombinase-estrogen receptor fusion proteins.

  9. The role of Notch signaling in kidney podocytes.

    PubMed

    Asanuma, Katsuhiko; Oliva Trejo, Juan Alejandro; Tanaka, Eriko

    2017-02-01

    The Notch signaling pathway is a basic cell-to-cell communication mechanism. This pathway is activated by the interaction between Notch receptors and the ligands of adjacent cells. Once activated, Notch receptors are cleaved and the intracellular domains translocate into the nucleus, where the transcription of target genes starts. In the mammalian kidney, Notch receptors are activated during nephrogenesis. Afterwards, in the mature glomeruli, the Notch pathway becomes silent. However, many researchers have reported the activation of Notch receptors in mature podocytes under pathological conditions. In this review, we discuss the role of Notch signaling in podocytes.

  10. Complex regulation of HSC emergence by the Notch signaling pathway

    PubMed Central

    Butko, Emerald; Pouget, Claire; Traver, David

    2016-01-01

    Hematopoietic stem cells are formed during embryonic development, and serve as the foundation of the definitive blood program for life. Notch signaling has been well established as an essential direct contributor to HSC specification. However, several recent studies have indicated that the contribution of Notch signaling is complex. HSC specification requires multiple Notch signaling inputs, some received directly by hematopoietic precursors, and others that occur indirectly within neighboring somites. Of note, proinflammatory signals provided by primitive myeloid cells are needed for HSC specification via upregulation of the Notch pathway in hemogenic endothelium. In addition to multiple requirements for Notch activation, recent studies indicate that Notch signaling must subsequently be repressed to permit HSC emergence. Finally, Notch must then be reactivated to maintain HSC fate. In this review, we discuss the growing understanding of the dynamic contributions of Notch signaling to the establishment of hematopoiesis during development. PMID:26586199

  11. Kinase active Misshapen regulates Notch signaling in Drosophila melanogaster.

    PubMed

    Mishra, Abhinava K; Sachan, Nalani; Mutsuddi, Mousumi; Mukherjee, Ashim

    2015-11-15

    Notch signaling pathway represents a principal cellular communication system that plays a pivotal role during development of metazoans. Drosophila misshapen (msn) encodes a protein kinase, which is related to the budding yeast Ste20p (sterile 20 protein) kinase. In a genetic screen, using candidate gene approach to identify novel kinases involved in Notch signaling, we identified msn as a novel regulator of Notch signaling. Data presented here suggest that overexpression of kinase active form of Msn exhibits phenotypes similar to Notch loss-of-function condition and msn genetically interacts with components of Notch signaling pathway. Kinase active form of Msn associates with Notch receptor and regulate its signaling activity. We further show that kinase active Misshapen leads to accumulation of membrane-tethered form of Notch. Moreover, activated Msn also depletes Armadillo and DE-Cadherin from adherens junctions. Thus, this study provides a yet unknown mode of regulation of Notch signaling by Misshapen.

  12. The Notch signaling pathway as a mediator of tumor survival.

    PubMed

    Capaccione, Kathleen M; Pine, Sharon R

    2013-07-01

    The Notch signaling pathway is evolutionarily conserved and responsible for cell fate determination in the developing embryo and mature tissue. At the molecular level, ligand binding activates Notch signaling by liberating the Notch intracellular domain, which then translocates into the nucleus and activates gene transcription. Despite the elegant simplicity of this pathway, which lacks secondary messengers or a signaling cascade, Notch regulates gene expression in a highly context- and cell-type-dependent manner. Notch signaling is frequently dysregulated, most commonly by overactivation, across many cancers and confers a survival advantage on tumors, leading to poorer outcomes for patients. Recent studies demonstrate how Notch signaling increases tumor cell proliferation and provide evidence that active Notch signaling maintains the cancer stem-cell pool, induces epithelial-mesenchymal transition and promotes chemoresistance. These studies imply that pharmacological inhibition of Notch signaling may refine control of cancer therapy and improve patient survival. Gamma secretase inhibitors (GSIs) are drugs that inhibit Notch signaling and may be successful in controlling cancer cell growth in conjunction with standard chemotherapy, but substantial side effects have hampered their widespread use. Recent efforts have been aimed at the development of antibodies against specific Notch receptors and ligands with the hope of limiting side effects while providing the same therapeutic benefit as GSIs. Together, studies characterizing Notch signaling and modulation have offered hope that refined methods targeting Notch may become powerful tools in anticancer therapeutics.

  13. The Role of Adams in Notch Signaling

    PubMed Central

    Groot, Arjan J.; Vooijs, Marc A.

    2014-01-01

    Regulated intramembrane proteolysis (RIP) is a highly conserved signaling paradigm whereby membrane-bound signaling proteins are cleaved in their transmembrane region and then released into the cytoplasm to act as signaling molecules. In most if not all cases intramembrane cleavage is preceded and regulated by a membrane proximal cleavage step called “ectodomain shedding”. Here we will review the role of ectodomain shedding in RIP of the NOTCH signaling pathway, a highly conserved cell-cell communication pathway that mediates cell fate decisions during development and in adult tissues. PMID:22399336

  14. Identification of Domains for Efficient Notch Signaling Activity in Immobilized Notch Ligand Proteins.

    PubMed

    Liu, Ledi; Wada, Hiroe; Matsubara, Natsuki; Hozumi, Katsuto; Itoh, Motoyuki

    2017-04-01

    Notch is a critical signaling pathway that controls cell fate and tissue homeostasis, but the functional characterization of Notch ligand domains that activate Notch receptors remains incomplete. Here, we established a method for immobilizing Notch ligand proteins onto beads to measure time-dependent Notch activity after the addition of Notch ligand-coated beads. A comparison between activities by the Notch ligand found on the cell surface to that of the ligand immobilized on beads showed that immobilized Notch ligand protein produces comparable signal activity during the first 10 h. Follow-up truncation studies showed that the N-terminal epidermal growth factor (EGF) repeat three region of delta like canonical Notch ligand 4 (DLL4) or jagged 1 (JAG1) is the minimum region for activating Notch signaling, and the DLL4 EGF repeat three domain may have a role in activation through a mechanism other than by increasing binding affinity. In addition, we found that reconstruction of the DLL4 delta and OSM-11 (DOS) motif (N257P) resulted in an increase in both binding affinity and signaling activity, which suggests that the role of the DOS motif is conserved among Notch ligands. Furthermore, active DLL4 protein on beads promoted T cell differentiation or inhibited B cell differentiation in vitro, whereas JAG1 proteins on beads did not have any effect. Taken together, our findings provide unambiguous evidence for the role of different Notch ligands and their domains in Notch signal activation, and may be potential tools for controlling Notch signaling activation. J. Cell. Biochem. 118: 785-796, 2017. © 2016 Wiley Periodicals, Inc.

  15. Rme-8 depletion perturbs Notch recycling and predisposes to pathogenic signaling

    PubMed Central

    Gomez-Lamarca, Maria J.; Snowdon, Laura A.; Seib, Ekatarina; Klein, Thomas

    2015-01-01

    Notch signaling is a major regulator of cell fate, proliferation, and differentiation. Like other signaling pathways, its activity is strongly influenced by intracellular trafficking. Besides contributing to signal activation and down-regulation, differential fluxes between trafficking routes can cause aberrant Notch pathway activation. Investigating the function of the retromer-associated DNAJ protein Rme-8 in vivo, we demonstrate a critical role in regulating Notch receptor recycling. In the absence of Rme-8, Notch accumulated in enlarged tubulated Rab4-positive endosomes, and as a consequence, signaling was compromised. Strikingly, when the retromer component Vps26 was depleted at the same time, Notch no longer accumulated and instead was ectopically activated. Likewise, depletion of ESCRT-0 components Hrs or Stam in combination with Rme-8 also led to high levels of ectopic Notch activity. Together, these results highlight the importance of Rme-8 in coordinating normal endocytic recycling route and reveal that its absence predisposes toward conditions in which pathological Notch signaling can occur. PMID:26169355

  16. Notching on Cancer’s Door: Notch Signaling in Brain Tumors

    PubMed Central

    Teodorczyk, Marcin; Schmidt, Mirko H. H.

    2015-01-01

    Notch receptors play an essential role in the regulation of central cellular processes during embryonic and postnatal development. The mammalian genome encodes for four Notch paralogs (Notch 1–4), which are activated by three Delta-like (Dll1/3/4) and two Serrate-like (Jagged1/2) ligands. Further, non-canonical Notch ligands such as epidermal growth factor like protein 7 (EGFL7) have been identified and serve mostly as antagonists of Notch signaling. The Notch pathway prevents neuronal differentiation in the central nervous system by driving neural stem cell maintenance and commitment of neural progenitor cells into the glial lineage. Notch is therefore often implicated in the development of brain tumors, as tumor cells share various characteristics with neural stem and progenitor cells. Notch receptors are overexpressed in gliomas and their oncogenicity has been confirmed by gain- and loss-of-function studies in vitro and in vivo. To this end, special attention is paid to the impact of Notch signaling on stem-like brain tumor-propagating cells as these cells contribute to growth, survival, invasion, and recurrence of brain tumors. Based on the outcome of ongoing studies in vivo, Notch-directed therapies such as γ-secretase inhibitors and blocking antibodies have entered and completed various clinical trials. This review summarizes the current knowledge on Notch signaling in brain tumor formation and therapy. PMID:25601901

  17. Notch signal integration in the vasculature during remodeling

    PubMed Central

    Rostama, Bahman; Peterson, Sarah M.; Vary, Calvin P. H.; Liaw, Lucy

    2014-01-01

    Notch signaling plays many important roles in homeostasis and remodeling in the vessel wall, and serves a critical role in the communication between endothelial cells and smooth muscle cells. Within blood vessels, Notch signaling integrates with multiple pathways by mechanisms including direct protein-protein interaction, cooperative or synergistic regulation of signal cascades, and co-regulation of transcriptional targets. After establishment of the mature blood vessel, the spectrum and intensity of Notch signaling changes during phases of active remodeling or disease progression. These changes can be mediated by regulation via microRNAs and protein stability or signaling, and corresponding changes in complementary signaling pathways. Notch also affects endothelial cells on a systems level by regulating key metabolic components. This review will outline the most recent findings of Notch activity in blood vessels, with a focus on how Notch signals integrate with other molecular signaling pathways controlling vascular phenotype. PMID:25464152

  18. HES6 promotes prostate cancer aggressiveness independently of Notch signalling.

    PubMed

    Carvalho, Filipe L F; Marchionni, Luigi; Gupta, Anuj; Kummangal, Basheer A; Schaeffer, Edward M; Ross, Ashley E; Berman, David M

    2015-07-01

    Notch signalling is implicated in the pathogenesis of a variety of cancers, but its role in prostate cancer is poorly understood. However, selected Notch pathway members are overrepresented in high-grade prostate cancers. We comprehensively profiled Notch pathway components in prostate cells and found prostate cancer-specific up-regulation of NOTCH3 and HES6. Their expression was particularly high in androgen responsive lines. Up- and down-regulating Notch in these cells modulated expression of canonical Notch targets, HES1 and HEY1, which could also be induced by androgen. Surprisingly, androgen treatment also suppressed Notch receptor expression, suggesting that androgens can activate Notch target genes in a receptor-independent manner. Using a Notch-sensitive Recombination signal binding protein for immunoglobulin kappa J region (RBPJ) reporter assay, we found that basal levels of Notch signalling were significantly lower in prostate cancer cells compared to benign cells. Accordingly pharmacological Notch pathway blockade did not inhibit cancer cell growth or viability. In contrast to canonical Notch targets, HES6, a HES family member known to antagonize Notch signalling, was not regulated by Notch signalling, but relied instead on androgen levels, both in cultured cells and in human cancer tissues. When engineered into prostate cancer cells, reduced levels of HES6 resulted in reduced cancer cell invasion and clonogenic growth. By molecular profiling, we identified potential roles for HES6 in regulating hedgehog signalling, apoptosis and cell migration. Our results did not reveal any cell-autonomous roles for canonical Notch signalling in prostate cancer. However, the results do implicate HES6 as a promoter of prostate cancer progression.

  19. Not(ch) just development: Notch signalling in the adult brain

    PubMed Central

    Ables, Jessica L.; Breunig, Joshua J.; Eisch, Amelia J.; Rakic, Pasko

    2011-01-01

    The Notch pathway is often regarded as a developmental pathway, but components of Notch signalling are expressed and active in the adult brain. With the advent of more sophisticated genetic manipulations, evidence has emerged that suggests both conserved and novel roles for Notch signalling in the adult brain. Not surprisingly, Notch is a key regulator of adult neural stem cells, but it is increasingly clear that Notch signalling also has roles in the regulation of migration, morphology, synaptic plasticity and survival of immature and mature neurons. Understanding the many functions of Notch signalling in the adult brain, and its dysfunction in neurodegenerative disease and malignancy, is crucial to the development of new therapeutics that are centred around this pathway. PMID:21505516

  20. Notch signaling in cancer stem cells.

    PubMed

    Wang, Jialiang; Sullenger, Bruce A; Rich, Jeremy N

    2012-01-01

    Subpopulations of cancer cells with stem cell-like characteristics, termed cancer stem cells, have been identified in a wide range of human cancers. Cancer stem cells are defined by their ability to self-renew as well as recapitulate the original heterogeneity of cancer cells in culture and in serial xenotransplants. Not only are cancer stem cells highly tumorigenic, but these cells are implicated in tumor resistance to conventional chemotherapy and radiotherapy, thus highlighting their significance as therapeutic targets. Considerable similarities have been found between cancer stem cells and normal stem cells on their dependence on certain signaling pathways. More specifically, the core stem cell signaling pathways, such as the Wnt, Notch and Hedgehog pathways, also critically regulate the self-renewal and survival of cancer stem cells. While the oncogenic functions of Notch pathway have been well documented, its role in cancer stem cells is just emerging. In this chapter, we will discuss recent advances in cancer stem cell research and highlight the therapeutic potential of targeting Notch in cancer stem cells.

  1. The Role of Notch Signaling Pathway in Breast Cancer Pathogenesis

    DTIC Science & Technology

    2005-07-01

    proteins are activated upon binding to ligands of the Delta/Serrate family. In previous experiments I had found that activated allele of Notch1 cooperates...breast cancer cells, I tested whether ErbB2 overexpression will cooperate with Notch in HMLE cells. While overexpression of activated Notch1 failed to...tumorigenic behavior. 15. SUBJECT TERMS Notch, Ras, signaling, transformation, tumorigenesis 16. SECURITY CLASSIFICATION OF: 17. LIMITATION OF

  2. Notch signaling and the determination of appendage identity

    PubMed Central

    Kurata, Shoichiro; Go, Masahiro J.; Artavanis-Tsakonas, Spyros; Gehring, Walter J.

    2000-01-01

    The Notch signaling pathway defines an evolutionarily conserved cell–cell interaction mechanism that throughout development controls the ability of precursor cells to respond to developmental signals. Here we show that Notch signaling regulates the expression of the master control genes eyeless, vestigial, and Distal-less, which in combination with homeotic genes induce the formation of eyes, wings, antennae, and legs. Therefore, Notch is involved in a common regulatory pathway for the determination of the various Drosophila appendages. PMID:10681430

  3. Notch signaling in prostate cancer: refining a therapeutic opportunity

    PubMed Central

    Su, Qingtai; Xin, Li

    2016-01-01

    Summary Notch is an evolutionarily conserved signaling pathway that plays a critical role in specifying cell fate and regulating tissue homeostasis and carcinogenesis. Studies using organ cultures and genetically engineered mouse models have demonstrated that Notch signaling regulates prostate development and homeostasis. However, the role of the Notch signaling pathway in prostate cancer remains inconclusive. Many published studies have documented consistent deregulation of major Notch signaling components in human prostate cancer cell lines, mouse models for prostate cancers, and human prostate cancer specimens at both the mRNA and the protein levels. However, functional studies in human cancer cells by modulation of Notch pathway elements suggest both tumor suppressive and oncogenic roles of Notch. These controversies may originate from our inadequate understanding of the regulation of Notch signaling under versatile genetic contexts, and reflect the multifaceted and pleiotropic roles of Notch in regulating different aspects of prostate cancer cell biology, such as proliferation, metastasis, and chemo-resistance. Future comprehensive studies using various mouse models for prostate cancer may help clarify the role of Notch signaling in prostate cancer and provide a solid basis for determining whether and how Notch should be employed as a therapeutic target for prostate cancer. PMID:26521657

  4. Emerging roles of Notch signaling in liver disease

    PubMed Central

    Geisler, Fabian; Strazzabosco, Mario

    2014-01-01

    This review critically discusses the most recent advances on the role of Notch signaling in liver development, homeostasis and disease. It is now clear that the significance of Notch in determining mammalian cell fates and functions extends beyond development, and Notch is a major regular of organ homeostasis. Moreover, Notch signaling is reactivated upon injury and regulates the complex interactions between the distinct cellular types involved in the repair process. Notch is also involved in the regulation of liver metabolism, inflammation and cancer. The net effects of Notch signaling are highly variable and finely regulated at multiple levels, but also depend on the specific cellular context in which Notch is activated. Persistent activation of Notch signaling is associated with liver malignancies, such as hepatocellular carcinoma with stem cell features and intrahepatic cholangiocarcinoma. The complexity of the pathway provides several possible targets for agents able to inhibit Notch. However, further cell- and context-specific in depth understanding of Notch signaling in liver homeostasis and disease will be essential to translate these concepts into the clinical practice and be able to predict benefits and risks of evolving therapies. PMID:24930574

  5. Involvement of Notch signaling in early chick ovarian follicle development.

    PubMed

    Li, Jun; Zhao, Dan; Guo, Changquan; Li, Jian; Mi, Yuling; Zhang, Caiqiao

    2016-01-01

    The formation of primordial follicles is a crucial process in the establishment of follicle pools required for the female's reproductive life span. For laying hens, ample follicles are a prerequisite for high laying performance. Notch signaling plays critical roles in germ cell cysts breakdown and in the formation of primordial follicles. Here, we investigated the role of Notch signaling in the ovarian development of post-hatch chicks. Results showed that around post-hatch day 4 (H4), the germ cell cysts broke apart, oocytes became surrounded by squamous pregranulosa cells, and the primordial follicles were then formed. Subsequently, we detected the expression of Notch signaling-related genes including Notch receptors (Notch1, 2), ligands (Jag1, 2 and Dll1, 4), and target genes (Hes1, Hey1). These genes all showed expression at H4 and some of these genes were up-regulated during primordial follicle formation. To evaluate the Notch signaling requirement for early follicular development, we adopted an in vitro ovary culture system. Suppression of Notch signaling by γ-secretase inhibitor induced a decrease of primordial follicles and an increase of germ cells in cysts. Attenuating Notch signaling also inhibited the phosphatidylinositol 3-kinase/protein kinase B pathways and suppressed cadherin expression. These results suggest that Notch signaling is endowed with an indispensable role in primordial follicle formation in post-hatch chicks.

  6. Olfactory Sensory Neurons Control Dendritic Complexity of Mitral Cells via Notch Signaling

    PubMed Central

    Saito, Tetsuichiro

    2016-01-01

    Mitral cells (MCs) of the mammalian olfactory bulb have a single primary dendrite extending into a single glomerulus, where they receive odor information from olfactory sensory neurons (OSNs). Molecular mechanisms for controlling dendritic arbors of MCs, which dynamically change during development, are largely unknown. Here we found that MCs displayed more complex dendritic morphologies in mouse mutants of Maml1, a crucial gene in Notch signaling. Similar phenotypes were observed by conditionally misexpressing a dominant negative form of MAML1 (dnMAML1) in MCs after their migration. Conversely, conditional misexpression of a constitutively active form of Notch reduced their dendritic complexity. Furthermore, the intracellular domain of Notch1 (NICD1) was localized to nuclei of MCs. These findings suggest that Notch signaling at embryonic stages is involved in the dendritic complexity of MCs. After the embryonic misexpression of dnMAML1, many MCs aberrantly extended dendrites to more than one glomerulus at postnatal stages, suggesting that Notch signaling is essential for proper formation of olfactory circuits. Moreover, dendrites in cultured MCs were shortened by Jag1-expressing cells. Finally, blocking the activity of Notch ligands in OSNs led to an increase in dendritic complexity as well as a decrease in NICD1 signals in MCs. These results demonstrate that the dendritic complexity of MCs is controlled by their presynaptic partners, OSNs. PMID:28027303

  7. A novel anticancer therapy that simultaneously targets aberrant p53 and Notch activities in tumors.

    PubMed

    Yao, Yuting; Wang, Li; Zhang, He; Wang, Haibo; Zhao, Xiaoping; Zhang, Yidan; Zhang, Leilei; Fan, Xianqun; Qian, Guanxiang; Hu, Ji-Fan; Ge, Shengfang

    2012-01-01

    Notch signaling pathway plays an important role in tumorigenesis by maintaining the activity of self-renewal of cancer stem cells, and therefore, it is hypothesized that interference of Notch signaling may inhibit tumor formation and progression. H101 is a recombinant oncolytic adenovirus that is cytolytic in cells lacking intact p53, but it is unable to eradicate caner stem cells. In this study, we tested a new strategy of tumor gene therapy by combining a Notch1-siRNA with H101 oncolytic adenovirus. In HeLa-S3 tumor cells, the combined therapy blocked the Notch pathway and induced apoptosis in tumors that are p53-inactive. In nude mice bearing xenograft tumors derived from HeLa-S3 cells, the combination of H101/Notch1-siRNA therapies inhibited tumor growth. Moreover, Notch1-siRNA increased Hexon gene expression at both the transcriptional and the translational levels, and promoted H101 replication in tumors, thereby enhancing the oncolytic activity of H101. These data demonstrate the feasibility to combine H101 p53-targted oncolysis and anti-Notch siRNA activities as a novel anti-cancer therapy.

  8. Notch-Nrf2 Axis: Regulation of Nrf2 Gene Expression and Cytoprotection by Notch Signaling

    PubMed Central

    Skoko, John J.; Chartoumpekis, Dionysios V.; Kimura, Shoko; Slocum, Stephen L.; Noda, Kentaro; Palliyaguru, Dushani L.; Fujimuro, Masahiro; Boley, Patricia A.; Tanaka, Yugo; Shigemura, Norihisa; Biswal, Shyam; Yamamoto, Masayuki; Kensler, Thomas W.

    2014-01-01

    The Notch signaling pathway enables regulation and control of development, differentiation, and homeostasis through cell-cell communication. Our investigation shows that Notch signaling directly activates the Nrf2 stress adaptive response pathway through recruitment of the Notch intracellular domain (NICD) transcriptosome to a conserved Rbpjκ site in the promoter of Nrf2. Stimulation of Notch signaling through Notch ligand expression in cells and by overexpression of the NICD in RosaNICD/−::AlbCre mice in vivo induces expression of Nrf2 and its target genes. Continuous and transient NICD expression in the liver produces a Notch-dependent cytoprotective response through direct transcriptional activation of Nrf2 signaling to rescue mice from acute acetaminophen toxicity. This response can be reversed upon genetic disruption of Nrf2. Morphological studies showed that the characteristic phenotype of high-density intrahepatic bile ducts and enlarged liver in RosaNICD/−::AlbCre mice could be at least partially reversed after Nrf2 disruption. Furthermore, the liver and bile duct phenotypes could be recapitulated with constitutive activation of Nrf2 signaling in Keap1F/F::AlbCre mice. It appears that Notch-to-Nrf2 signaling is another important determinant in liver development and function and promotes cell-cell cytoprotective signaling responses. PMID:24298019

  9. Stra13 regulates satellite cell activation by antagonizing Notch signaling

    PubMed Central

    Sun, Hong; Li, Li; Vercherat, Cécile; Gulbagci, Neriman Tuba; Acharjee, Sujata; Li, Jiali; Chung, Teng-Kai; Thin, Tin Htwe; Taneja, Reshma

    2007-01-01

    Satellite cells play a critical role in skeletal muscle regeneration in response to injury. Notch signaling is vital for satellite cell activation and myogenic precursor cell expansion but inhibits myogenic differentiation. Thus, precise spatial and temporal regulation of Notch activity is necessary for efficient muscle regeneration. We report that the basic helix-loop-helix transcription factor Stra13 modulates Notch signaling in regenerating muscle. Upon injury, Stra13−/− mice exhibit increased cellular proliferation, elevated Notch signaling, a striking regeneration defect characterized by degenerated myotubes, increased mononuclear cells, and fibrosis. Stra13−/− primary myoblasts also exhibit enhanced Notch activity, increased proliferation, and defective differentiation. Inhibition of Notch signaling ex vivo and in vivo ameliorates the phenotype of Stra13−/− mutants. We demonstrate in vitro that Stra13 antagonizes Notch activity and reverses the Notch-imposed inhibition of myogenesis. Thus, Stra13 plays an important role in postnatal myogenesis by attenuating Notch signaling to reduce myoblast proliferation and promote myogenic differentiation. PMID:17502421

  10. Emerin suppresses Notch signaling by restricting the Notch intracellular domain to the nuclear membrane.

    PubMed

    Lee, Byongsun; Lee, Tae-Hee; Shim, Jaekyung

    2017-02-01

    Emerin is an inner nuclear membrane protein that is involved in maintaining the mechanical integrity of the nuclear membrane. Increasing evidence supports the involvement of emerin in the regulation of gene expression; however, its precise function remains to be elucidated. Here, we show that emerin downregulated genes downstream of Notch signaling, which are activated exclusively by the Notch intracellular domain (NICD). Deletion mutant experiments revealed that the transmembrane domain of emerin is important for the inhibition of Notch signaling. Emerin interacted directly and colocalized with the NICD at the nuclear membrane. Emerin knockdown induced the phosphorylation of ERK and AKT, increased endogenous Notch signaling, and inhibited hydrogen peroxide-induced apoptosis in HeLa cells. Notably, the downregulation of barrier-to-autointegration factor (BAF) or lamin A/C increased Notch signaling by inducing the release of emerin into the cytosol, implying that nuclear membrane-bound emerin acts as an endogenous inhibitor of Notch signaling. Taken together, our results indicate that emerin negatively regulates Notch signaling by promoting the retention of the NICD at the nuclear membrane. This mechanism could constitute a new therapeutic target for the treatment of emerin-related diseases.

  11. MAGP2 controls Notch via interactions with RGD binding integrins: Identification of a novel ECM-integrin-Notch signaling axis.

    PubMed

    Deford, Peter; Brown, Kasey; Richards, Rae Lee; King, Aric; Newburn, Kristin; Westover, Katherine; Albig, Allan R

    2016-02-01

    Canonical Notch signaling involves Notch receptor activation via interaction with cell surface bound Notch ligand. Recent findings also indicate that Notch signaling may be modulated by cross-talk with other signaling mechanisms. The ECM protein MAGP2 was previously shown to regulate Notch in a cell type dependent manner, although the molecular details of this interaction have not been dissected. Here, we report that MAGP2 cell type specific control of Notch is independent of individual Notch receptor-ligand combinations but dependent on interaction with RGD binding integrins. Overexpressed MAGP2 was found to suppress transcriptional activity from the Notch responsive Hes1 promoter activity in endothelial cells, while overexpression of a RGD→RGE MAGP2 mutant increased Notch signaling in the same cell type. This effect was not unique to MAGP2 since the RGD domain of the ECM protein EGFL7 was also found to be an important modulator of Hes1 promoter activity. Independently of MAGP2 or EGFL7, inhibition of RGD-binding integrins with soluble RGD peptides also increased accumulation of active N1ICD fragments and Notch responsive promoter activity independently of changes in Notch1, Jag1, or Dll4 expression. Finally, β1 or β3 integrin blocking antibodies also enhanced Notch signaling. Collectively, these results answer the question of how MAGP2 controls cell type dependent Notch signaling, but more importantly uncover a new mechanism to understand how extracellular matrices and cellular environments impact Notch signaling.

  12. Role of Notch Signaling in Human Breast Cancer Pathogenesis

    DTIC Science & Technology

    2006-11-01

    allele of Notch1 cooperates with low levels of oncogenic Ras expressing HMLE cells(termed HMLER). Further investigations revealed that Notch-IC...transformation, tumorigenesis , 16. SECURITY CLASSIFICATION OF: 17. LIMITATION OF ABSTRACT 18. NUMBER OF PAGES 19a. NAME OF RESPONSIBLE PERSON USAMRMC...15. Number of Pages (count all pages including appendices) 9 Notch, Ras, signaling, transformation, tumorigenesis , 16. Price Code (Leave Blank) 17

  13. Targeting Notch signaling as a novel therapy for retinoblastoma

    PubMed Central

    Asnaghi, Laura; Tripathy, Arushi; Yang, Qian; Kaur, Harpreet; Hanaford, Allison; Yu, Wayne; Eberhart, Charles G.

    2016-01-01

    Retinoblastoma is the most common intraocular malignancy of childhood. Notch plays a key role in retinal cells from which retinoblastomas arise, and we therefore studied the role of Notch signaling in promoting retinoblastoma proliferation. Moderate or strong nuclear expression of Hes1 was found in 10 of 11 human retinoblastoma samples analyzed immunohistochemically, supporting a role for Notch in retinoblastoma growth. Notch pathway components were present in WERI Rb1 and Y79 retinoblastoma lines, with Jag2 and DLL4 more highly expressed than other ligands, and Notch1 and Notch2 more abundant than Notch3. The cleaved/active form of Notch1 was detectable in both lines. Inhibition of the pathway, achieved using a γ-secretase inhibitor (GSI) or by downregulating Jag2, DLL4 or CBF1 using short hairpin RNA, potently reduced growth, proliferation and clonogenicity in both lines. Upregulation of CXCR4 and CXCR7 and downregulation of PI3KC2β were identified by microarray upon Jag2 suppression. The functional importance of PI3KC2β was confirmed using shRNA. Synergy was found by combining GSI with Melphalan at their IC50. These findings indicate that Notch pathway is active in WERI Rb1 and Y79, and in most human retinoblastoma samples, and suggest that Notch antagonists may represent a new approach to more effectively treat retinoblastoma. PMID:27661116

  14. Mammary stem cells and breast cancer--role of Notch signalling.

    PubMed

    Farnie, Gillian; Clarke, Robert B

    2007-06-01

    Adult stem cells are found in numerous tissues of the body and play a role in tissue development, replacement and repair. Evidence shows that breast stem cells are multipotent and can self renew, which are key characteristics of stem cells, and a single cell enriched with cell surface markers has the ability to grow a fully functional mammary gland in vivo. Many groups have extrapolated the cancer stem cell hypothesis from the haematopoietic system to solid cancers, where using in vitro culture techniques and in vivo transplant models have established evidence of cancer stem cells in colon, pancreas, prostate, brain and breast cancers. In the report we describe the evidence for breast cancer stem cells; studies consistently show that stem cell like and breast cancer initiating populations can be enriched using cell surface makers CD44+/CD24- and have upregulated genes which include Notch. Notch signalling has been highlighted as a pathway involved in the development of the breast and is frequently dysregulated in invasive breast cancer. We have investigated the role of Notch in a pre-invasive breast lesion, ductal carcinoma in situ (DCIS), and have found that aberrant activation of Notch signalling is an early event in breast cancer. High expression of Notch 1 intracellular domain (NICD) in DCIS also predicted a reduced time to recurrence 5 years after surgery. Using a non-adherent sphere culture technique we have grown DCIS mammospheres from primary DCIS tissue, where self-renewal capacity, measured by the number of mammosphere initiating cells, were increased from normal breast tissue. A gamma-secretase inhibitor, DAPT, which inhibits all four Notch receptors and a Notch 4 neutralising antibody were shown to reduce DCIS mammosphere formation, indicating that Notch signalling and other stem cell self-renewal pathways may represent novel therapeutic targets to prevent recurrence of pre-invasive and invasive breast cancer.

  15. Notch signaling promotes osteoclast maturation and resorptive activity

    PubMed Central

    Ashley, Jason W; Ahn, Jaimo; Hankenson, Kurt D

    2015-01-01

    The role of Notch signaling in osteoclast differentiation is controversial with conflicting experimental evidence indicating both stimulatory and inhibitory roles. Differences in experimental protocols and in vivo versus in vitro models may explain the discrepancies between studies. In this study, we investigated cell autonomous roles of Notch signaling in osteoclast differentiation and function by altering Notch signaling during osteoclast differentiation using stimulation with immobilized ligands Jagged1 or Delta-like1 or by suppression with γ-secretase inhibitor DAPT or transcriptional inhibitor SAHM1. Stimulation of Notch signaling in committed osteoclast precursors resulted in larger osteoclasts with a greater number of nuclei and resorptive activity whereas suppression resulted in smaller osteoclasts with fewer nuclei and suppressed resorptive activity. Conversely, stimulation of Notch signaling in osteoclast precursors prior to induction of osteoclastogenesis resulted in fewer osteoclasts. Our data support a mechanism of context-specific Notch signaling effects wherein Notch stimulation inhibits commitment to osteoclast differentiation, but enhances the maturation and function of committed precursors. PMID:25914241

  16. Targeting the Notch signaling pathway in cancer therapeutics.

    PubMed

    Guo, Huajiao; Lu, Yi; Wang, Jianhua; Liu, Xia; Keller, Evan T; Liu, Qian; Zhou, Qinghua; Zhang, Jian

    2014-11-01

    Despite advances in surgery, imaging, chemotherapy, and radiotherapy, the poor overall cancer-related death rate remains unacceptable. Novel therapeutic strategies are desperately needed. Nowadays, targeted therapy has become the most promising therapy and a welcome asset to the cancer therapeutic arena. There is a large body of evidence demonstrating that the Notch signaling pathway is critically involved in the pathobiology of a variety of malignancies. In this review, we provide an overview of emerging data, highlight the mechanism of the Notch signaling pathway in the development of a wide range of cancers, and summarize recent progress in therapeutic targeting of the Notch signaling pathway.

  17. Notch signaling regulates cardiomyocyte proliferation during zebrafish heart regeneration.

    PubMed

    Zhao, Long; Borikova, Asya L; Ben-Yair, Raz; Guner-Ataman, Burcu; MacRae, Calum A; Lee, Richard T; Burns, C Geoffrey; Burns, Caroline E

    2014-01-28

    The human heart's failure to replace ischemia-damaged myocardium with regenerated muscle contributes significantly to the worldwide morbidity and mortality associated with coronary artery disease. Remarkably, certain vertebrate species, including the zebrafish, achieve complete regeneration of amputated or injured myocardium through the proliferation of spared cardiomyocytes. Nonetheless, the genetic and cellular determinants of natural cardiac regeneration remain incompletely characterized. Here, we report that cardiac regeneration in zebrafish relies on Notch signaling. Following amputation of the zebrafish ventricular apex, Notch receptor expression becomes activated specifically in the endocardium and epicardium, but not the myocardium. Using a dominant negative approach, we discovered that suppression of Notch signaling profoundly impairs cardiac regeneration and induces scar formation at the amputation site. We ruled out defects in endocardial activation, epicardial activation, and dedifferentiation of compact myocardial cells as causative for the regenerative failure. Furthermore, coronary endothelial tubes, which we lineage traced from preexisting endothelium in wild-type hearts, formed in the wound despite the myocardial regenerative failure. Quantification of myocardial proliferation in Notch-suppressed hearts revealed a significant decrease in cycling cardiomyocytes, an observation consistent with a noncell autonomous requirement for Notch signaling in cardiomyocyte proliferation. Unexpectedly, hyperactivation of Notch signaling also suppressed cardiomyocyte proliferation and heart regeneration. Taken together, our data uncover the exquisite sensitivity of regenerative cardiomyocyte proliferation to perturbations in Notch signaling.

  18. Notch Signaling Pathway Regulates Progesterone Secretion in Murine Luteal Cells.

    PubMed

    Wang, Jing; Liu, Shuangmei; Peng, Lichao; Dong, Qiming; Bao, Riqiang; Lv, Qiulan; Tang, Min; Hu, Chuan; Li, Gang; Liang, Shangdong; Zhang, Chunping

    2015-10-01

    Notch signaling is an evolutionarily conserved pathway, which involves in various cell life activities. Other studies and our report showed that the Notch signaling plays very important role in follicle development in mammalian ovaries. In luteal cells, Notch ligand, delta-like ligand 4, is involved in normal luteal vasculature. In this study, murine luteal cells were cultured in vitro and treated with Notch signaling inhibitors, L-658,458 and N-[N-(3,5-difluorophenacetyl)-l-alanyl]-S-phenylglycinet-butyl ester (DAPT). We found that L-658,458 and DAPT treatment decrease basal and human chorionic gonadotropin (hCG)-stimulated progesterone secretion. On the contrary, overexpression of intracellular domain of Notch3 increased basal and hCG-stimulated progesterone secretion. Further studies demonstrated that Notch signaling regulated the expression of steroidogenic acute regulatory protein and CYP11A, 2 key enzymes for progesterone synthesis. In conclusion, Notch signaling plays important role in regulating progesterone secretion in murine luteal cells.

  19. Cyclosporin A Disrupts Notch Signaling and Vascular Lumen Maintenance

    PubMed Central

    Pandey, Raghav; Botros, Mark A.; Nacev, Benjamin A.; Albig, Allan R.

    2015-01-01

    Cyclosporin A (CSA) suppresses immune function by blocking the cyclophilin A and calcineurin/NFAT signaling pathways. In addition to immunosuppression, CSA has also been shown to have a wide range of effects in the cardiovascular system including disruption of heart valve development, smooth muscle cell proliferation, and angiogenesis inhibition. Circumstantial evidence has suggested that CSA might control Notch signaling which is also a potent regulator of cardiovascular function. Therefore, the goal of this project was to determine if CSA controls Notch and to dissect the molecular mechanism(s) by which CSA impacts cardiovascular homeostasis. We found that CSA blocked JAG1, but not Dll4 mediated Notch1 NICD cleavage in transfected 293T cells and decreased Notch signaling in zebrafish embryos. CSA suppression of Notch was linked to cyclophilin A but not calcineurin/NFAT inhibition since N-MeVal-4-CsA but not FK506 decreased Notch1 NICD cleavage. To examine the effect of CSA on vascular development and function, double transgenic Fli1-GFP/Gata1-RFP zebrafish embryos were treated with CSA and monitored for vasculogenesis, angiogenesis, and overall cardiovascular function. Vascular patterning was not obviously impacted by CSA treatment and contrary to the anti-angiogenic activity ascribed to CSA, angiogenic sprouting of ISV vessels was normal in CSA treated embryos. Most strikingly, CSA treated embryos exhibited a progressive decline in blood flow that was associated with eventual collapse of vascular luminal structures. Vascular collapse in zebrafish embryos was partially rescued by global Notch inhibition with DAPT suggesting that disruption of normal Notch signaling by CSA may be linked to vascular collapse. However, multiple signaling pathways likely cause the vascular collapse phenotype since both cyclophilin A and calcineurin/NFAT were required for normal vascular function. Collectively, these results show that CSA is a novel inhibitor of Notch signaling and

  20. The role of NOTCH1 signaling in T-ALL.

    PubMed

    Ferrando, Adolfo A

    2009-01-01

    The identification of activating mutations in NOTCH1 in over 50% of T-cell acute lymphoblastic leukemias (T-ALL) has generated major interest in the elucidation of the mechanisms of transformation downstream of oncogenic NOTCH and in the targeting of the NOTCH signaling pathway in this disease. Small molecule gamma-secretase inhibitors (GSIs) block NOTCH1 signaling in T-ALL lymphoblasts, yet the clinical development of GSIs has been held back by the development of gastrointestinal toxicity and their weak antileukemic effects against human T-ALL. However, new therapeutic strategies aiming to optimize the use of anti-NOTCH1 therapies for T-ALL, including combination therapies with molecularly targeted drugs and glucocorticoids, have started to emerge as a result of improved understanding of the molecular mechanisms that mediate the effects of GSIs in leukemic cells and the intestinal epithelium. This review focuses on the molecular basis of NOTCH1-induced transformation, the mechanisms of action of oncogenic NOTCH1 and clinical significance of NOTCH1 mutations in T-ALL.

  1. Notch signaling promotes nephrogenesis by downregulating Six2

    PubMed Central

    Chung, Eunah; Deacon, Patrick; Marable, Sierra; Shin, Juhyun

    2016-01-01

    During nephrogenesis, multipotent mesenchymal nephron progenitors develop into distinct epithelial segments. Each nephron segment has distinct cell types and physiological function. In the current model of kidney development, Notch signaling promotes the formation of proximal tubules and represses the formation of distal tubules. Here, we present a novel role of Notch in nephrogenesis. We show in mice that differentiation of nephron progenitors requires downregulation of Six2, a transcription factor required for progenitor maintenance, and that Notch signaling is necessary and sufficient for Six2 downregulation. Furthermore, we find that nephron progenitors lacking Notch signaling fail to differentiate into any nephron segments, not just proximal tubules. Our results demonstrate how cell fates of progenitors are regulated by a transcription factor governing progenitor status and by a differentiation signal in nephrogenesis. PMID:27633993

  2. Notch signaling during cell fate determination in the inner ear

    PubMed Central

    Kiernan, Amy

    2013-01-01

    In the inner ear, Notch signaling has been proposed to specify the sensory regions, as well as regulate the differentiation of hair cells and supporting cell within those regions. In addition, Notch plays an important role in otic neurogenesis, by determining which cells differentiate as neurons, sensory cells and non-sensory cells. Here, I review the evidence for the complex and myriad roles Notch participates in during inner ear development. A particular challenge for those studying ear development and Notch is to decipher how activation of a single pathway can lead to different outcomes within the ear, which may include changes in the intrinsic properties of the cell, Notch modulation, and potential non-canonical pathways. PMID:23578865

  3. Notch signalling mediates reproductive constraint in the adult worker honeybee

    PubMed Central

    Duncan, Elizabeth J.; Hyink, Otto; Dearden, Peter K.

    2016-01-01

    The hallmark of eusociality is the reproductive division of labour, in which one female caste reproduces, while reproduction is constrained in the subordinate caste. In adult worker honeybees (Apis mellifera) reproductive constraint is conditional: in the absence of the queen and brood, adult worker honeybees activate their ovaries and lay haploid male eggs. Here, we demonstrate that chemical inhibition of Notch signalling can overcome the repressive effect of queen pheromone and promote ovary activity in adult worker honeybees. We show that Notch signalling acts on the earliest stages of oogenesis and that the removal of the queen corresponds with a loss of Notch protein in the germarium. We conclude that the ancient and pleiotropic Notch signalling pathway has been co-opted into constraining reproduction in worker honeybees and we provide the first molecular mechanism directly linking ovary activity in adult worker bees with the presence of the queen. PMID:27485026

  4. Role of Delta-Notch signaling in cerebral cavernous malformations.

    PubMed

    Kar, Souvik; Baisantry, Arpita; Nabavi, Arya; Bertalanffy, Helmut

    2016-10-01

    Cerebral cavernous malformations (CCM) commonly known as cavernous hemangioma are associated with abnormally enlarged thin-walled blood vessels. As a result, these dilated capillaries are prone to leakage and result in hemorrhages. Clinically, such hemorrhages lead to severe headaches, focal neurological deficits, and epileptic seizures. CCM is caused by loss of function mutations in one of the three well-known CCM genes: Krev interaction trapped 1 (KRIT1), OSM, and programmed cell death 10 (PDCD10). Loss of CCM genes have been shown to be synergistically related to decreased Notch signaling and excessive angiogenesis. Despite recent evidences indicating that Notch signaling plays a pivotal role in regulating angiogenesis, the role of Notch in CCM development and progression is still not clear. Here, we provide an update literature review on the current knowledge of the structure of Notch receptor and its ligands, its relevance to angiogenesis and more precisely to CCM pathogenesis. In addition to reviewing the current literatures, this review will also focus on the cross talk between Delta-Notch and vascular endothelial growth factor (VEGF) signaling in angiogenesis and in CCM pathogenesis. Understanding the role of Notch signaling in CCM development and progression might help provide a better insight for novel anti-angiogenic therapies.

  5. Nrf2 and Notch Signaling in Lung Cancer: Near the Crossroad

    PubMed Central

    Sparaneo, Angelo; Fabrizio, Federico Pio

    2016-01-01

    The transcription factor Nrf2 (NF-E2 related factor 2) is a master regulator of the cell antioxidant response associated with tumor growth and resistance to cytotoxic treatments. In particular, Nrf2 induces upregulation of cytoprotective genes by interacting with the closely situated AREs (Antioxidant Response Elements) in response to endogenous or exogenous stress stimuli and takes part to several oncogenic signaling pathways. Among these, the crosstalk with Notch pathway has been shown to enhance cytoprotection and maintenance of cellular homeostasis, tissue organization by modulating cell proliferation kinetics, and stem cell self-renewal in several organs. The role of Notch and Nrf2 related pathways in tumorigenesis is highly variable and when they are both abnormally activated they can synergistically cause neoplastic proliferation by promoting cell survival, differentiation, invasion, and metastases. NFE2L2, KEAP1, and NOTCH genes family appear in the list of significantly mutated genes in tumors in both combined and individual sets, supporting the crucial role that the aberrant Nrf2-Notch crosstalk might have in cancerogenesis. In this review, we summarize current knowledge about the alterations of Nrf2 and Notch pathways and their reciprocal transcriptional regulation throughout tumorigenesis and progression of lung tumors, supporting the potentiality of putative biomarkers and therapeutic targets. PMID:27847554

  6. Notch3 Interactome Analysis Identified WWP2 as a Negative Regulator of Notch3 Signaling in Ovarian Cancer

    PubMed Central

    Guan, Bin; Wu, Ren-Chin; Zhu, Heng; Blackshaw, Seth; Shih, Ie-Ming; Wang, Tian-Li

    2014-01-01

    The Notch3 signaling pathway is thought to play a critical role in cancer development, as evidenced by the Notch3 amplification and rearrangement observed in human cancers. However, the molecular mechanism by which Notch3 signaling contributes to tumorigenesis is largely unknown. In an effort to identify the molecular modulators of the Notch3 signaling pathway, we screened for Notch3-intracellular domain (N3-ICD) interacting proteins using a human proteome microarray. Pathway analysis of the Notch3 interactome demonstrated that ubiquitin C was the molecular hub of the top functional network, suggesting the involvement of ubiquitination in modulating Notch3 signaling. Thereby, we focused on functional characterization of an E3 ubiquitin-protein ligase, WWP2, a top candidate in the Notch3 interactome list. Co-immunoprecipitation experiments showed that WWP2 interacted with N3-ICD but not with intracellular domains from other Notch receptors. Wild-type WWP2 but not ligase-deficient mutant WWP2 increases mono-ubiquitination of the membrane-tethered Notch3 fragment, therefore attenuating Notch3 pathway activity in cancer cells and leading to cell cycle arrest. The mono-ubiquitination by WWP2 may target an endosomal/lysosomal degradation fate for Notch3 as suggested by the fact that the process could be suppressed by the endosomal/lysosomal inhibitor. Analysis of The Cancer Genome Atlas dataset showed that the majority of ovarian carcinomas harbored homozygous or heterozygous deletions in WWP2 locus, and there was an inverse correlation in the expression levels between WWP2 and Notch3 in ovarian carcinomas. Furthermore, ectopic expression of WWP2 decreased tumor development in a mouse xenograft model and suppressed the Notch3-induced phenotypes including increase in cancer stem cell-like cell population and platinum resistance. Taken together, our results provide evidence that WWP2 serves as a tumor suppressor by negatively regulating Notch3 signaling in ovarian cancer

  7. The Histone Deacetylase Inhibitor Vaproic Acid Induces Cell Growth Arrest in Hepatocellular Carcinoma Cells via Suppressing Notch Signaling

    PubMed Central

    Sun, Guangchun; Mackey, Lily V.; Coy, David H.; Yu, Cui-Yun; Sun, Lichun

    2015-01-01

    Hepatocellular carcinoma (HCC) is a type of malignant cancer. Notch signaling is aberrantly expressed in HCC tissues with more evidence showing that this signaling plays a critical role in HCCs. In the present study, we investigate the effects of the anti-convulsant drug valproic acid (VPA) in HCC cells and its involvement in modulating Notch signaling. We found that VPA, acting as a histone deacetylase (HDAC) inhibitor, induced a decrease in HDAC4 and an increase in acetylated histone 4 (AcH4) and suppressed HCC cell growth. VPA also induced down-regulation of Notch signaling via suppressing the expression of Notch1 and its target gene HES1, with an increase of tumor suppressor p21 and p63. Furthermore, Notch1 activation via overexpressing Notch1 active form ICN1 induced HCC cell proliferation and anti-apoptosis, indicating Notch signaling played an oncogenic role in HCC cells. Meanwhile, VPA could reverse Notch1-induced increase of cell proliferation. Interestingly, VPA was also observed to stimulate the expression of G protein-coupled somatostatin receptor type 2 (SSTR2) that has been used in receptor-targeting therapies. This discovery supports a combination therapy of VPA with the SSTR2-targeting agents. Our in vitro assay did show that the combination of VPA and the peptide-drug conjugate camptothecin-somatostatin (CPT-SST) displayed more potent anti-proliferative effects on HCC cells than did each alone. VPA may be a potential drug candidate in the development of anti-HCC drugs via targeting Notch signaling, especially in combination with receptor-targeting cytotoxic agents. PMID:26366213

  8. Notch-regulated ankyrin-repeat protein inhibits Notch1 signaling: multiple Notch1 signaling pathways involved in T cell development.

    PubMed

    Yun, Theodore J; Bevan, Michael J

    2003-06-15

    We have characterized the function of Notch-regulated ankyrin-repeat protein (Nrarp) in mouse cell lines and in hematopoietic stem cells (HSCs). Nrarp overexpression is able to block Notch-induced activation of CBF-1. In AKR1010 thymoma cells, Nrarp overexpression blocks CBF-1-dependent transcriptional activation of Notch-responsive genes and inhibits phenotypic changes associated with Notch activation. Enforced expression of Nrarp in mouse HSCs results in a profound block in T lineage commitment and progression through early stages of thymocyte maturation. In contrast, Deltex-1 overexpression in HSCs can also block T lineage commitment but not progression through the early double negative stages of thymocyte maturation. The different effects of Deltex-1 and Nrarp overexpression suggest that alternate Notch signaling pathways mediate T vs B lineage commitment and thymocyte maturation.

  9. Dynamic binding of RBPJ is determined by Notch signaling status

    PubMed Central

    Castel, David; Mourikis, Philippos; Bartels, Stefanie J.J.; Brinkman, Arie B.; Tajbakhsh, Shahragim; Stunnenberg, Hendrik G.

    2013-01-01

    Notch signaling plays crucial roles in mediating cell fate choices in all metazoans largely by specifying the transcriptional output of one cell in response to a neighboring cell. The DNA-binding protein RBPJ is the principle effector of this pathway in mammals and, together with the transcription factor moiety of Notch (NICD), regulates the expression of target genes. The prevalent view presumes that RBPJ statically occupies consensus binding sites while exchanging repressors for activators in response to NICD. We present the first specific RBPJ chromatin immunoprecipitation and high-throughput sequencing study in mammalian cells. To dissect the mode of transcriptional regulation by RBPJ and identify its direct targets, whole-genome binding profiles were generated for RBPJ; its coactivator, p300; NICD; and the histone H3 modifications H3 Lys 4 trimethylation (H3K4me3), H3 Lys 4 monomethylation (H3K4me1), and histone H3 Lys 27 acetylation (H3K27ac) in myogenic cells under active or inhibitory Notch signaling conditions. Our results demonstrate dynamic binding of RBPJ in response to Notch activation at essentially all sites co-occupied by NICD. Additionally, we identify a distinct set of sites where RBPJ recruits neither NICD nor p300 and binds DNA statically, irrespective of Notch activity. These findings significantly modify our views on how RBPJ and Notch signaling mediate their activities and consequently impact on cell fate decisions. PMID:23651858

  10. Lateral inhibition of Notch signaling in neoplastic cells

    PubMed Central

    Heth, Jason A.; Muraszko, Karin M.; Fan, Xing; Bar, Eli E.; Eberhart, Charles G.

    2015-01-01

    During normal development, heterogeneous expression of Notch ligands can result in pathway suppression in the signal-sending cell, a process known as lateral inhibition. It is unclear if an analogous phenomenon occurs in malignant cells. We observed significant induction of Notch ligands in glioblastoma neurospheres and pancreatic carcinoma cells cultured in low oxygen, suggesting that this phenomenon could occur around hypoxic regions. To model lateral inhibition in these tumors, the ligand Jagged1 was overexpressed in glioblastoma and pancreatic carcinoma cells, resulting in overall induction of pathway targets. However, when ligand high and ligand low cells from a single line were co-cultured and then separated, we noted suppression of Notch pathway targets in the former and induction in the latter, suggesting that neoplastic lateral inhibition can occur. We also found that repression of Notch pathway targets in signal-sending cells may occur through the activity of a Notch ligand intracellular domain, which translocates into the nucleus. Understanding how this neoplastic lateral inhibition process functions in cancer cells may be important in targeting ligand driven Notch signaling in solid tumors. PMID:25557173

  11. Notch signal strength controls cell fate in the haemogenic endothelium

    PubMed Central

    Gama-Norton, Leonor; Ferrando, Eva; Ruiz-Herguido, Cristina; Liu, Zenhy; Guiu, Jordi; Islam, Abul B. M. M. K.; Lee, Sung-Uk; Yan, Minhong; Guidos, Cynthia J.; López-Bigas, Nuria; Maeda, Takahiro; Espinosa, Lluis; Kopan, Raphael; Bigas, Anna

    2015-01-01

    Acquisition of the arterial and haemogenic endothelium fates concurrently occur in the aorta–gonad–mesonephros (AGM) region prior to haematopoietic stem cell (HSC) generation. The arterial programme depends on Dll4 and the haemogenic endothelium/HSC on Jag1-mediated Notch1 signalling. How Notch1 distinguishes and executes these different programmes in response to particular ligands is poorly understood. By using two Notch1 activation trap mouse models with different sensitivity, here we show that arterial endothelial cells and HSCs originate from distinct precursors, characterized by different Notch1 signal strengths. Microarray analysis on AGM subpopulations demonstrates that the Jag1 ligand stimulates low Notch strength, inhibits the endothelial programme and is permissive for HSC specification. In the absence of Jag1, endothelial cells experience high Dll4-induced Notch activity and select the endothelial programme, thus precluding HSC formation. Interference with the Dll4 signal by ligand-specific blocking antibodies is sufficient to inhibit the endothelial programme and favour specification of the haematopoietic lineage. PMID:26465397

  12. Notch1 signaling stimulates proliferation of immature cardiomyocytes

    PubMed Central

    Collesi, Chiara; Zentilin, Lorena; Sinagra, Gianfranco; Giacca, Mauro

    2008-01-01

    The identification of the molecular mechanisms controlling cardiomyocyte proliferation during the embryonic, fetal, and early neonatal life appears of paramount interest in regard to exploiting this information to promote cardiac regeneration. Here, we show that the proliferative potential of neonatal rat cardiomyocytes is powerfully stimulated by the sustained activation of the Notch pathway. We found that Notch1 is expressed in proliferating ventricular immature cardiac myocytes (ICMs) both in vitro and in vivo, and that the number of Notch1-positive cells in the heart declines with age. Notch1 expression in ICMs paralleled the expression of its Jagged1 ligand on non-myocyte supporting cells. The inhibition of Notch signaling in ICMs blocked their proliferation and induced apoptosis; in contrast, its activation by Jagged1 or by the constitutive expression of its activated form using an adeno-associated virus markedly stimulated proliferative signaling and promoted ICM expansion. Maintenance or reactivation of Notch signaling in cardiac myocytes might represent an interesting target for innovative regenerative therapy. PMID:18824567

  13. Maintenance of Bone Homeostasis by DLL1-Mediated Notch Signaling.

    PubMed

    Muguruma, Yukari; Hozumi, Katsuto; Warita, Hiroyuki; Yahata, Takashi; Uno, Tomoko; Ito, Mamoru; Ando, Kiyoshi

    2016-10-13

    Adult bone mass is maintained through a balance of the activities of osteoblasts and osteoclasts. Although Notch signaling has been shown to maintain bone homeostasis by controlling the commitment, differentiation, and function of cells in both the osteoblast and osteoclast lineages, the precise mechanisms by which Notch performs such diverse and complex roles in bone physiology remain unclear. By using a transgenic approach that modified the expression of delta-like 1 (DLL1) or Jagged1 (JAG1) in an osteoblast-specific manner, we investigated the ligand-specific effects of Notch signaling in bone homeostasis. This study demonstrated for the first time that the proper regulation of DLL1 expression, but not JAG1 expression, in osteoblasts is essential for the maintenance of bone remodeling. DLL1-induced Notch signaling was responsible for the expansion of the bone-forming cell pool by promoting the proliferation of committed but immature osteoblasts. However, DLL1-Notch signaling inhibited further differentiation of the expanded osteoblasts to become fully matured functional osteoblasts, thereby substantially decreasing bone formation. Osteoblast-specific expression of DLL1 did not alter the intrinsic differentiation ability of cells of the osteoclast lineage. However, maturational arrest of osteoblasts caused by the DLL1 transgene impaired the maturation and function of osteoclasts due to a failed osteoblast-osteoclast coupling, resulting in severe suppression of bone metabolic turnover. Taken together, DLL1-mediated Notch signaling is critical for proper bone remodeling as it regulates the differentiation and function of both osteoblasts and osteoclasts. Our study elucidates the importance of ligand-specific activation of Notch signaling in the maintenance of bone homeostasis. This article is protected by copyright. All rights reserved.

  14. Cell-Cell Contact Area Affects Notch Signaling and Notch-Dependent Patterning.

    PubMed

    Shaya, Oren; Binshtok, Udi; Hersch, Micha; Rivkin, Dmitri; Weinreb, Sheila; Amir-Zilberstein, Liat; Khamaisi, Bassma; Oppenheim, Olya; Desai, Ravi A; Goodyear, Richard J; Richardson, Guy P; Chen, Christopher S; Sprinzak, David

    2017-03-13

    During development, cells undergo dramatic changes in their morphology. By affecting contact geometry, these morphological changes could influence cellular communication. However, it has remained unclear whether and how signaling depends on contact geometry. This question is particularly relevant for Notch signaling, which coordinates neighboring cell fates through direct cell-cell signaling. Using micropatterning with a receptor trans-endocytosis assay, we show that signaling between pairs of cells correlates with their contact area. This relationship extends across contact diameters ranging from micrometers to tens of micrometers. Mathematical modeling predicts that dependence of signaling on contact area can bias cellular differentiation in Notch-mediated lateral inhibition processes, such that smaller cells are more likely to differentiate into signal-producing cells. Consistent with this prediction, analysis of developing chick inner ear revealed that ligand-producing hair cell precursors have smaller apical footprints than non-hair cells. Together, these results highlight the influence of cell morphology on fate determination processes.

  15. Zinc-induced downregulation of Notch signaling is associated with cytoplasmic retention of Notch1-IC and RBP-Jk via PI3k-Akt signaling pathway.

    PubMed

    Baek, Sang-Hyun; Kim, Mi-Yeon; Mo, Jung-Soon; Ann, Eun-Jung; Lee, Kyu Shik; Park, Ji-Hye; Kim, Jin-Young; Seo, Mi-Sun; Choi, Eui-Ju; Park, Hee-Sae

    2007-09-18

    The Notch signaling pathway appears to perform an important function in the determination of cell fate and in differentiation, in a wide variety of organisms and cell types. In this study, we provide evidence that the inactivation of Notch signaling by zinc is achieved via a PI3K-Akt-dependent, cytoplasmic retention of Notch1-IC and RBP-Jk. Extracellular zinc has been determined to inhibit constitutive active mutants of both Notch1 (DeltaEN1) and Notch1-IC-mediated transcription. However, in such cases, neither the cleavage pattern of Notch nor the protein stability of Notch1-IC and RBP-Jk was found to have significantly changed. With regard to the modulation of Notch signaling, zinc appears to exert a significant negative influence on the binding occurring between Notch1 and RBP-Jk, both in vivo and in vitro. The zinc-induced inhibition of Notch signaling can be rescued via pretreatment with wortmannin or LY294002, both of which are specific PI3K signaling pathway inhibitors. Furthermore, we ascertained that zinc triggers the cytoplasmic retention of Notch1-IC and RBP-Jk, and that cytoplasmic retention could be rescued via treatment with wortmannin. Overall, we have determined that an important relationship exists between zinc and the Notch1 signaling pathway, and that this relationship is intimately involved with the cytoplasmic retention of Notch and RBP-Jk.

  16. NOTCH signaling in skeletal progenitors is critical for fracture repair

    PubMed Central

    Wang, Cuicui; Inzana, Jason A.; Mirando, Anthony J.; Liu, Zhaoyang; Shen, Jie; O’Keefe, Regis J.; Awad, Hani A.; Hilton, Matthew J.

    2016-01-01

    Fracture nonunions develop in 10%–20% of patients with fractures, resulting in prolonged disability. Current data suggest that bone union during fracture repair is achieved via proliferation and differentiation of skeletal progenitors within periosteal and soft tissues surrounding bone, while bone marrow stromal/stem cells (BMSCs) and other skeletal progenitors may also contribute. The NOTCH signaling pathway is a critical maintenance factor for BMSCs during skeletal development, although the precise role for NOTCH and the requisite nature of BMSCs following fracture is unknown. Here, we evaluated whether NOTCH and/or BMSCs are required for fracture repair by performing nonstabilized and stabilized fractures on NOTCH-deficient mice with targeted deletion of RBPjk in skeletal progenitors, maturing osteoblasts, and committed chondrocytes. We determined that removal of NOTCH signaling in BMSCs and subsequent depletion of this population result in fracture nonunion, as the fracture repair process was normal in animals harboring either osteoblast- or chondrocyte-specific deletion of RBPjk. Together, this work provides a genetic model of a fracture nonunion and demonstrates the requirement for NOTCH and BMSCs in fracture repair, irrespective of fracture stability and vascularity. PMID:26950423

  17. Protein O-fucosyltransferase 1 is an essential component of Notch signaling pathways

    PubMed Central

    Shi, Shaolin; Stanley, Pamela

    2003-01-01

    Notch receptor signaling regulates cell growth and differentiation, and core components of Notch signaling pathways are conserved from Drosophila to humans. Fringe glycosyltransferases are crucial modulators of Notch signaling that act on epidermal growth factor (EGF)-like repeats in the Notch receptor extracellular domain. The substrate of Fringe is EGF-O-fucose and the transfer of fucose to Notch by protein O-fucosyltransferase 1 is necessary for Fringe to function. O-fucose also occurs on Cripto and on Notch ligands. Here we show that mouse embryos lacking protein O-fucosyltransferase 1 die at midgestation with severe defects in somitogenesis, vasculogenesis, cardiogenesis, and neurogenesis. The phenotype is similar to that of embryos lacking downstream effectors of all Notch signaling pathways such as presenilins or RBP-Jκ, and is different from Cripto, Notch receptor, Notch ligand, or Fringe null phenotypes. Protein O-fucosyltransferase 1 is therefore an essential core member of Notch signaling pathways in mammals. PMID:12697902

  18. Notch signaling in response to excitotoxicity induces neurodegeneration via erroneous cell cycle reentry

    PubMed Central

    Marathe, S; Liu, S; Brai, E; Kaczarowski, M; Alberi, L

    2015-01-01

    Neurological disorders such as Alzheimer's disease, stroke and epilepsy are currently marred by the lack of effective treatments to prevent neuronal death. Erroneous cell cycle reentry (CCR) is hypothesized to have a causative role in neurodegeneration. We show that forcing S-phase reentry in cultured hippocampal neurons is sufficient to induce neurodegeneration. We found that kainic-acid treatment in vivo induces erroneous CCR and neuronal death through a Notch-dependent mechanism. Ablating Notch signaling in neurons provides neuroprotection against kainic acid-induced neuronal death. We further show that kainic-acid treatment activates Notch signaling, which increases the bioavailability of CyclinD1 through Akt/GSK3β pathway, leading to aberrant CCR via activation of CyclinD1-Rb-E2F1 axis. In addition, pharmacological blockade of this pathway at critical steps is sufficient to confer resistance to kainic acid-induced neurotoxicity in mice. Taken together, our results demonstrate that excitotoxicity leads to neuronal death in a Notch-dependent manner through erroneous CCR. PMID:25822340

  19. Role of Notch signaling pathway in pancreatic cancer

    PubMed Central

    Gao, Jiankun; Long, Bo; Wang, Zhiwei

    2017-01-01

    Pancreatic cancer (PC) is one of the highly aggressive malignancies in the United States. It has been shown that multiple signaling pathways are involved in the pathogenesis of PC, such as JNK, PI3K/AKT, Rho GTPase, Hedgehog (Hh) and Skp2. In recent years, accumulated evidence has demonstrated that Notch signaling pathway plays critical roles in the development and progression of PC. Therefore, in this review we discuss the recent literature regarding the function and regulation of Notch in the pathogenesis of PC. Moreover, we describe that Notch signaling pathway could be down-regulated by its inhibitors or natural compounds, which could be a novel approach for the treatment of PC patients. PMID:28337369

  20. A dual role for NOTCH signaling in joint cartilage maintenance and osteoarthritis.

    PubMed

    Liu, Zhaoyang; Chen, Jianquan; Mirando, Anthony J; Wang, Cuicui; Zuscik, Michael J; O'Keefe, Regis J; Hilton, Matthew J

    2015-07-21

    Loss of NOTCH signaling in postnatal murine joints results in osteoarthritis, indicating a requirement for NOTCH during maintenance of joint cartilage. However, NOTCH signaling components are substantially increased in abundance in posttraumatic osteoarthritis in humans and mice, suggesting either a reparative or a pathological role for NOTCH activation in osteoarthritis. We investigated a potential dual role for NOTCH in joint maintenance and osteoarthritis by generating two mouse models overexpressing the NOTCH1 intracellular domain (NICD) within postnatal joint cartilage. The first mouse model exhibited sustained NOTCH activation to resemble pathological NOTCH signaling, whereas the second model had transient NOTCH activation, which more closely reflected physiological NOTCH signaling. Sustained NOTCH signaling in joint cartilage led to an early and progressive osteoarthritic-like pathology, whereas transient NOTCH activation enhanced the synthesis of cartilage matrix and promoted joint maintenance under normal physiological conditions. Through RNA-sequencing, immunohistochemical, and biochemical approaches, we identified several targets that could be responsible for NOTCH-mediated cartilage degradation, fibrosis, and osteoarthritis progression. These targets included components of the interleukin-6 (IL-6)-signal transducer and activator of transcription 3 (STAT3) and mitogen-activated protein kinase signaling pathways, which may also contribute to the posttraumatic development of osteoarthritis. Together, these data suggest a dual role for the NOTCH pathway in joint cartilage, and they identify downstream effectors of NOTCH signaling as potential targets for disease-modifying osteoarthritis drugs.

  1. FGF and Notch signaling in sensory neuron formation: a multifactorial approach to understanding signaling pathway hierarchy.

    PubMed

    Voelkel, Jacob E; Harvey, Jamison A; Adams, Jason S; Lassiter, Rhonda N; Stark, Michael R

    2014-11-01

    The ophthalmic trigeminal (opV) placode exclusively gives rise to sensory neurons, making it a good model to study the molecular regulation of sensory neurogenesis. A number of signaling pathways including Wnt, PDGF, FGF, and Notch have been shown to be involved in the process of opV placode cell development. However, the regulatory relationships between these signaling pathways in placode cells are still unknown and have been difficult to study experimentally. Using a novel multifactorial approach in chick embryos that allows for inhibition of FGF throughout the tissue or in individual cells, with simultaneous inactivation of Notch signaling, we investigated the potential interaction between the FGF and Notch signaling pathways in trigeminal sensory neurogenesis. This study builds on prior research describing the individual role of FGF signaling or Notch signaling in opV placode development, where blocking FGF signaling resulted in neurogenesis failure, while blocking Notch signaling resulted in enhanced neurogenesis. Reported here, blocking both pathways simultaneously resulted in a reduction in the number of cells delaminating from the opV placode and undergoing sensory neuron differentiation. Further, Notch inhibition alone did not lead to an increase in the number of cells expressing FGFR4 or in the FGFR4 expression domain, but did result in a highly fragmented basal lamina, which was reversed when blocking FGF signaling. Cumulatively, the results presented here do not support a model of Notch/FGF interdependence, rather that FGF and Notch act in parallel to promote sensory neurogenesis.

  2. Notchless encodes a novel WD40-repeat-containing protein that modulates Notch signaling activity.

    PubMed Central

    Royet, J; Bouwmeester, T; Cohen, S M

    1998-01-01

    Signaling by Notch family receptors is involved in many cell-fate decisions during development. Several modifiers of Notch activity have been identified, suggesting that regulation of Notch signaling is complex. In a genetic screen for modifiers of Notch activity, we identified a gene encoding a novel WD40-repeat protein. The gene is called Notchless, because loss-of-function mutant alleles dominantly suppress the wing notching caused by certain Notch alleles. Reducing Notchless activity increases Notch activity. Overexpression of Notchless in Xenopus or Drosophila appears to have a dominant-negative effect in that it also increases Notch activity. Biochemical studies show that Notchless binds to the cytoplasmic domain of Notch, suggesting that it serves as a direct regulator of Notch signaling activity. PMID:9857191

  3. Targeting stem cell signaling pathways for drug discovery: advances in the Notch and Wnt pathways.

    PubMed

    An, Songzhu Michael; Ding, Qiang; Zhang, Jie; Xie, JingYi; Li, LingSong

    2014-06-01

    Signaling pathways transduce extracellular stimuli into cells through molecular cascades to regulate cellular functions. In stem cells, a small number of pathways, notably those of TGF-β/BMP, Hedgehog, Notch, and Wnt, are responsible for the regulation of pluripotency and differentiation. During embryonic development, these pathways govern cell fate specifications as well as the formation of tissues and organs. In adulthood, their normal functions are important for tissue homeostasis and regeneration, whereas aberrations result in diseases, such as cancer and degenerative disorders. In complex biological systems, stem cell signaling pathways work in concert as a network and exhibit crosstalk, such as the negative crosstalk between Wnt and Notch. Over the past decade, genetic and genomic studies have identified a number of potential drug targets that are involved in stem cell signaling pathways. Indeed, discovery of new targets and drugs for these pathways has become one of the most active areas in both the research community and pharmaceutical industry. Remarkable progress has been made and several promising drug candidates have entered into clinical trials. This review focuses on recent advances in the discovery of novel drugs which target the Notch and Wnt pathways.

  4. HIV Tat Impairs Neurogenesis through Functioning As a Notch Ligand and Activation of Notch Signaling Pathway.

    PubMed

    Fan, Yan; Gao, Xiang; Chen, Jinhui; Liu, Ying; He, Johnny J

    2016-11-02

    Alterations in adult neurogenesis have been noted in the brain of HIV-infected individuals and are likely linked to HIV-associated neurocognitive deficits, including those in learning and memory. But the underlying molecular mechanisms are not fully understood. In the study, we took advantage of doxycycline-inducible and astrocyte-specific HIV-1 Tat transgenic mice (iTat) and determined the relationship between Tat expression and neurogenesis. Tat expression in astrocytes was associated with fewer neuron progenitor cells (NPCs), fewer immature neurons, and fewer mature neurons in the dentate gyrus of the hippocampus of the mouse brain. In vitro NPC-derived neurosphere assays showed that Tat-containing conditioned media from astrocytes or recombinant Tat protein inhibited NPC proliferation and migration and altered NPC differentiation, while immunodepletion of Tat from Tat-containing conditioned media or heat inactivation of recombinant Tat abrogated those effects. Notch signaling downstream gene Hes1 promoter-driven luciferase reporter gene assay and Western blotting showed that recombinant Tat or Tat-containing conditioned media activated Hes1 transcription and protein expression, which were abrogated by Tat heat inactivation, immunodepletion, and cysteine mutation at position 30. Last, Notch signaling inhibitor N-[N-(3,5-difluorophenacetyl)-l-alanyl]-S-phenylglycine t-butyl ester (DAPT) significantly rescued Tat-impaired NPC differentiation in vitro and neurogenesis in vivo Together, these results show that Tat adversely affects NPCs and neurogenesis through Notch signaling and point to the potential of developing Notch signaling inhibitors as HIV/neuroAIDS therapeutics.

  5. Notch Signaling is Essential for Ventricular Chamber Development

    PubMed Central

    Grego-Bessa, Joaquín; Luna-Zurita, Luis; Monte, Gonzalo del; Bolós, Victoria; Melgar, Pedro; Arandilla, Alejandro; Garratt, Alistair N.; Zang, Heesuk; Mukouyama, Yoh-suke; Chen, Hanying; Shou, Weinian; Ballestar, Esteban; Esteller, Manel; Rojas, Ana; Pérez-Pomares, José María; de la Pompa, José Luis

    2009-01-01

    Summary Ventricular chamber morphogenesis, first manifested by trabeculae formation, is crucial for cardiac function and embryonic viability and depends on cellular interactions between endocardium and myocardium. We show that ventricular Notch1 activity is highest at presumptive trabecular endocardium. RBPJk and Notch1 mutants show impaired trabeculation and marker expression, attenuated EphrinB2, NRG1 and BMP10 expression and signaling and decreased myocardial proliferation. Functional and molecular analyses show that Notch inhibition prevents EphrinB2 expression and that EphrinB2 is a direct Notch target acting upstream of NRG1 in the ventricles. However, BMP10 levels are found to be independent of both EphrinB2 and NRG1 during trabeculation. Accordingly, exogenous BMP10 rescues the myocardial proliferative defect of in vitro cultured RBPJk mutants, while exogenous NRG1 rescues differentiation in parallel. We suggest that during trabeculation Notch independently regulates cardiomyocyte proliferation and differentiation, two exquisitely balanced processes whose perturbation may result in congenital heart disease. PMID:17336907

  6. Notch signaling is essential for ventricular chamber development.

    PubMed

    Grego-Bessa, Joaquín; Luna-Zurita, Luis; del Monte, Gonzalo; Bolós, Victoria; Melgar, Pedro; Arandilla, Alejandro; Garratt, Alistair N; Zang, Heesuk; Mukouyama, Yoh-Suke; Chen, Hanying; Shou, Weinian; Ballestar, Esteban; Esteller, Manel; Rojas, Ana; Pérez-Pomares, José María; de la Pompa, José Luis

    2007-03-01

    Ventricular chamber morphogenesis, first manifested by trabeculae formation, is crucial for cardiac function and embryonic viability and depends on cellular interactions between the endocardium and myocardium. We show that ventricular Notch1 activity is highest at presumptive trabecular endocardium. RBPJk and Notch1 mutants show impaired trabeculation and marker expression, attenuated EphrinB2, NRG1, and BMP10 expression and signaling, and decreased myocardial proliferation. Functional and molecular analyses show that Notch inhibition prevents EphrinB2 expression, and that EphrinB2 is a direct Notch target acting upstream of NRG1 in the ventricles. However, BMP10 levels are found to be independent of both EphrinB2 and NRG1 during trabeculation. Accordingly, exogenous BMP10 rescues the myocardial proliferative defect of in vitro-cultured RBPJk mutants, while exogenous NRG1 rescues differentiation in parallel. We suggest that during trabeculation Notch independently regulates cardiomyocyte proliferation and differentiation, two exquisitely balanced processes whose perturbation may result in congenital heart disease.

  7. Honokiol inhibits melanoma stem cells by targeting notch signaling.

    PubMed

    Kaushik, Gaurav; Venugopal, Anand; Ramamoorthy, Prabhu; Standing, David; Subramaniam, Dharmalingam; Umar, Shahid; Jensen, Roy A; Anant, Shrikant; Mammen, Joshua M V

    2015-12-01

    Melanoma is an aggressive disease with limited therapeutic options. Here, we determined the effects of honokiol (HNK), a biphenolic natural compound on melanoma cells and stemness. HNK significantly inhibited melanoma cell proliferation, viability, clonogenicity and induced autophagy. In addition, HNK significantly inhibited melanosphere formation in a dose dependent manner. Western blot analyses also demonstrated reduction in stem cell markers CD271, CD166, Jarid1b, and ABCB5. We next examined the effect of HNK on Notch signaling, a pathway involved in stem cell self-renewal. Four different Notch receptors exist in cells, which when cleaved by a series of enzymatic reactions catalyzed by Tumor Necrosis Factor-α-Converting Enzyme (TACE) and γ-secretase protein complex, results in the release of the Notch intracellular domain (NICD), which then translocates to the nucleus and induces target gene expression. Western blot analyses demonstrated that in HNK treated cells there is a significant reduction in the expression of cleaved Notch-2. In addition, there was a reduction in the expression of downstream target proteins, Hes-1 and cyclin D1. Moreover, HNK treatment suppressed the expression of TACE and γ-secretase complex proteins in melanoma cells. To confirm that suppression of Notch-2 activation is critical for HNK activity, we overexpressed NICD1, NICD2, and performed HNK treatment. NICD2, but not NICD1, partially restored the expression of Hes-1 and cyclin D1, and increased melanosphere formation. Taken together, these data suggest that HNK is a potent inhibitor of melanoma cells, in part, through the targeting of melanoma stem cells by suppressing Notch-2 signaling.

  8. The matricellular protein CCN3 regulates NOTCH1 signalling in chronic myeloid leukaemia.

    PubMed

    Suresh, Sukanya; McCallum, Lynn; Crawford, Lisa J; Lu, Wan Hua; Sharpe, Daniel J; Irvine, Alexandra E

    2013-11-01

    Deregulated NOTCH1 has been reported in lymphoid leukaemia, although its role in chronic myeloid leukaemia (CML) is not well established. We previously reported BCR-ABL down-regulation of a novel haematopoietic regulator, CCN3, in CML; CCN3 is a non-canonical NOTCH1 ligand. This study characterizes the NOTCH1–CCN3 signalling axis in CML. In K562 cells, BCR-ABL silencing reduced full-length NOTCH1 (NOTCH1-FL) and inhibited the cleavage of NOTCH1 intracellular domain (NOTCH1-ICD), resulting in decreased expression of the NOTCH1 targets c-MYC and HES1. K562 cells stably overexpressing CCN3 (K562/CCN3) or treated with recombinant CCN3(rCCN3) showed a significant reduction in NOTCH1 signalling (> 50% reduction in NOTCH1-ICD, p < 0.05).Gamma secretase inhibitor (GSI), which blocks NOTCH1 signalling, reduced K562/CCN3 colony formation but increased that of K562/control cells. GSI combined with either rCCN3 or imatinib reduced K562 colony formation with enhanced reduction of NOTCH1 signalling observed with combination treatments. We demonstrate an oncogenic role for NOTCH1 in CML and suggest that BCR-ABL disruption of NOTCH1–CCN3 signalling contributes to the pathogenesis of CML.

  9. Effects of S1 Cleavage on the Structure, Surface Export, and Signaling Activity of Human Notch1 and Notch2

    SciTech Connect

    Gordon, Wendy R.; Vardar-Ulu, Didem; L'Heureux, Sarah; Ashworth, Todd; Malecki, Michael J.; Sanchez-Irizarry, Cheryll; McArthur, Debbie G.; Histen, Gavin; Mitchell, Jennifer L.; Aster, Jon C.; Blacklow, Stephen C.

    2009-09-25

    Notch receptors are normally cleaved during maturation by a furin-like protease at an extracellular site termed S1, creating a heterodimer of non-covalently associated subunits. The S1 site lies within a key negative regulatory region (NRR) of the receptor, which contains three highly conserved Lin12/Notch repeats and a heterodimerization domain (HD) that interact to prevent premature signaling in the absence of ligands. Because the role of S1 cleavage in Notch signaling remains unresolved, we investigated the effect of S1 cleavage on the structure, surface trafficking and ligand-mediated activation of human Notch1 and Notch2, as well as on ligand-independent activation of Notch1 by mutations found in human leukemia. The X-ray structure of the Notch1 NRR after furin cleavage shows little change when compared with that of an engineered Notch1 NRR lacking the S1-cleavage loop. Likewise, NMR studies of the Notch2 HD domain show that the loop containing the S1 site can be removed or cleaved without causing a substantial change in its structure. However, Notch1 and Notch2 receptors engineered to resist S1 cleavage exhibit unexpected differences in surface delivery and signaling competence: S1-resistant Notch1 receptors exhibit decreased, but detectable, surface expression and ligand-mediated receptor activation, whereas S1-resistant Notch2 receptors are fully competent for cell surface delivery and for activation by ligands. Variable dependence on S1 cleavage also extends to T-ALL-associated NRR mutations, as common class 1 mutations display variable decrements in ligand-independent activation when introduced into furin-resistant receptors, whereas a class 2 mutation exhibits increased signaling activity. S1 cleavage has distinct effects on the surface expression of Notch1 and Notch2, but is not generally required for physiologic or pathophysiologic activation of Notch proteins. These findings are consistent with models for receptor activation in which ligand-binding or

  10. Effects of S1 Cleavage on the Structure, Surface Export, and Signaling Activity of Human Notch1 and Notch2

    PubMed Central

    Gordon, Wendy R.; Vardar-Ulu, Didem; L'Heureux, Sarah; Ashworth, Todd; Malecki, Michael J.; Sanchez-Irizarry, Cheryll; McArthur, Debbie G.; Histen, Gavin; Mitchell, Jennifer L.; Aster, Jon C.; Blacklow, Stephen C.

    2009-01-01

    Background Notch receptors are normally cleaved during maturation by a furin-like protease at an extracellular site termed S1, creating a heterodimer of non-covalently associated subunits. The S1 site lies within a key negative regulatory region (NRR) of the receptor, which contains three highly conserved Lin12/Notch repeats and a heterodimerization domain (HD) that interact to prevent premature signaling in the absence of ligands. Because the role of S1 cleavage in Notch signaling remains unresolved, we investigated the effect of S1 cleavage on the structure, surface trafficking and ligand-mediated activation of human Notch1 and Notch2, as well as on ligand-independent activation of Notch1 by mutations found in human leukemia. Principal Findings The X-ray structure of the Notch1 NRR after furin cleavage shows little change when compared with that of an engineered Notch1 NRR lacking the S1-cleavage loop. Likewise, NMR studies of the Notch2 HD domain show that the loop containing the S1 site can be removed or cleaved without causing a substantial change in its structure. However, Notch1 and Notch2 receptors engineered to resist S1 cleavage exhibit unexpected differences in surface delivery and signaling competence: S1-resistant Notch1 receptors exhibit decreased, but detectable, surface expression and ligand-mediated receptor activation, whereas S1-resistant Notch2 receptors are fully competent for cell surface delivery and for activation by ligands. Variable dependence on S1 cleavage also extends to T-ALL-associated NRR mutations, as common class 1 mutations display variable decrements in ligand-independent activation when introduced into furin-resistant receptors, whereas a class 2 mutation exhibits increased signaling activity. Conclusions/Significance S1 cleavage has distinct effects on the surface expression of Notch1 and Notch2, but is not generally required for physiologic or pathophysiologic activation of Notch proteins. These findings are consistent with

  11. Homozygous NOTCH3 null mutation and impaired NOTCH3 signaling in recessive early-onset arteriopathy and cavitating leukoencephalopathy

    PubMed Central

    Pippucci, Tommaso; Maresca, Alessandra; Magini, Pamela; Cenacchi, Giovanna; Donadio, Vincenzo; Palombo, Flavia; Papa, Valentina; Incensi, Alex; Gasparre, Giuseppe; Valentino, Maria Lucia; Preziuso, Carmela; Pisano, Annalinda; Ragno, Michele; Liguori, Rocco; Giordano, Carla; Tonon, Caterina; Lodi, Raffaele; Parmeggiani, Antonia; Carelli, Valerio; Seri, Marco

    2015-01-01

    Notch signaling is essential for vascular physiology. Neomorphic heterozygous mutations in NOTCH3, one of the four human NOTCH receptors, cause cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL). Hypomorphic heterozygous alleles have been occasionally described in association with a spectrum of cerebrovascular phenotypes overlapping CADASIL, but their pathogenic potential is unclear. We describe a patient with childhood-onset arteriopathy, cavitating leukoencephalopathy with cerebral white matter abnormalities presented as diffuse cavitations, multiple lacunar infarctions and disseminated microbleeds. We identified a novel homozygous c.C2898A (p.C966*) null mutation in NOTCH3 abolishing NOTCH3 expression and causing NOTCH3 signaling impairment. NOTCH3 targets acting in the regulation of arterial tone (KCNA5) or expressed in the vasculature (CDH6) were downregulated. Patient's vessels were characterized by smooth muscle degeneration as in CADASIL, but without deposition of granular osmiophilic material (GOM), the CADASIL hallmark. The heterozygous parents displayed similar but less dramatic trends in decrease in the expression of NOTCH3 and its targets, as well as in vessel degeneration. This study suggests a functional link between NOTCH3 deficiency and pathogenesis of vascular leukoencephalopathies. PMID:25870235

  12. NOTCH, a new signaling pathway implicated in holoprosencephaly

    PubMed Central

    Dupé, Valérie; Rochard, Lucie; Mercier, Sandra; Le Pétillon, Yann; Gicquel, Isabelle; Bendavid, Claude; Bourrouillou, Georges; Kini, Usha; Thauvin-Robinet, Christel; Bohan, Timothy P.; Odent, Sylvie; Dubourg, Christèle; David, Véronique

    2011-01-01

    Genetics of Holoprosencephaly (HPE), a congenital malformation of the developing human forebrain, is due to multiple genetic defects. Most genes that have been implicated in HPE belong to the Sonic Hedgehog (SHH) signaling pathway. Here we describe a new candidate gene isolated from array CGH redundant 6qter deletions, DELTA Like 1 (DLL1), which is a ligand of NOTCH. We show that DLL1 is co-expressed in the developing chick forebrain with Fgf8. By treating chick embryos with a pharmacological inhibitor, we demonstrate that DLL1 interacts with FGF signaling pathway. Moreover, a mutation analysis of DLL1 in HPE patients, revealed a three-nucleotide deletion. These various findings implicate DLL1 in early patterning of the forebrain and identify NOTCH as a new signaling pathway involved in HPE. PMID:21196490

  13. Faster embryonic segmentation through elevated Delta-Notch signalling

    PubMed Central

    Liao, Bo-Kai; Jörg, David J.; Oates, Andrew C.

    2016-01-01

    An important step in understanding biological rhythms is the control of period. A multicellular, rhythmic patterning system termed the segmentation clock is thought to govern the sequential production of the vertebrate embryo's body segments, the somites. Several genetic loss-of-function conditions, including the Delta-Notch intercellular signalling mutants, result in slower segmentation. Here, we generate DeltaD transgenic zebrafish lines with a range of copy numbers and correspondingly increased signalling levels, and observe faster segmentation. The highest-expressing line shows an altered oscillating gene expression wave pattern and shortened segmentation period, producing embryos with more, shorter body segments. Our results reveal surprising differences in how Notch signalling strength is quantitatively interpreted in different organ systems, and suggest a role for intercellular communication in regulating the output period of the segmentation clock by altering its spatial pattern. PMID:27302627

  14. Constitutive activation of NF-κB signaling by NOTCH1 mutations in chronic lymphocytic leukemia.

    PubMed

    Xu, Zhen-Shu; Zhang, Ju-Shun; Zhang, Jing-Yan; Wu, Shun-Quan; Xiong, Dong-Lian; Chen, Hui-Jun; Chen, Zhi-Zhe; Zhan, Rong

    2015-04-01

    NOTCH1 mutations occur in approximately 10% of patients with chronic lymphocytic leukemia (CLL). However, the relationship between the genetic aberrations and tumor cell drug resistance or disease progression remains unclear. Frameshift deletions were detected by gene sequencing in the NOTCH1 PEST domain in three naive CLL patients. These mutations were associated with chromosomal abnormalities including trisomy 12 or 13q deletion. Of note, one of the patients developed Richter's transformation during FCR treatment. Immunofluorescent and western blot analyses revealed a markedly higher intracellular domain of NOTCH (ICN) expression in the mutated cells compared with their unmutated counterparts and normal CD19+ B lymphocytes (P<0.01 and P<0.001, respectively). In addition, strong DNA-κB binding activities were observed in the mutant cells by gel shift assays. RT-PCR analysis revealed elevated RelA mRNA expression in the mutant cells, while RelB levels were variable. Reduced levels of RelA and RelB mRNA were observed in unmutated CLL and normal B cells. Compared to unmutated CLL and normal B cells, increased apoptosis occurred in the mutant cells in the presence of GSI (ICN inhibitor) and PDTC (NF-κB inhibitor), particularly under the synergistic effects of the two drugs (P=0.03). Moreover, IKKα and IKKβ, the active components in the NF-κB pathway, were markedly inhibited following prolonged treatment with GSI and PDTC. These results suggested that NOTCH1 mutations constitutively activate the NF-κB signaling pathway in CLL, which is likely related to ICN overexpression, indicating NOTCH1 and NF-κB as potential therapeutic targets in the treatment of CLL.

  15. [Research Progress on Notch Signal Pathway in Acute Graft-Versus-Host Disease -Review].

    PubMed

    Guo, Dong-Mei; Li, Ban-Ban; Li, Chun-Pu; Teng, Qing-Liang

    2017-02-01

    The Notch signaling pathway is a highly conserved cell signaling system that plays an essential role in many biological processes. Notch signaling regulates multiple aspects of hematopoiesis, especially during T cell develop-ment. Recent data suggest that Notch also regulates mature T cell differentiation and function. The latest data show that Notch also plays an essential role in alloreactive T cells mediating acute graft-versus-host disease (aGVHD), the most severe complication of allogeneic hematopoietic stem cell transplantation (allo-HSCT). Notch inhibition in donor-derived T cells or blockade of individual Notch ligands and receptors after transplantation can reduce GVHD severity and mortality in mouse models of allo-HSCT, without causing global immunosuppression. These findings indicate Notch in T cells as an attractive therapeutic target to control aGVHD. In this article, the pathophysiology of aGVHD, the Notch signal pathway and aGVHD are reviewed.

  16. Significance of Notch1-signaling pathway in human pancreatic development and carcinogenesis.

    PubMed

    Hu, Huankai; Zhou, Lan; Awadallah, Amad; Xin, Wei

    2013-05-01

    In animal studies, Notch1-signaling pathway plays an important role in the pancreatic embryogenesis by promoting pancreatic progenitor cells self-renewal and exocrine linage development. The persistent activation of Notch pathway could arrest the organ development and keep cells at an undifferentiated stage. Studies have shown that Notch1-signaling pathway is upregulated in invasive pancreatic ductal adenocarcinoma (PDAC). Here we examined the expression pattern of Notch1 and Hes1 in human fetal pancreatic tissues to elucidate the role of Notch1 in human pancreatic embryonic development. We also compared Notch1 expression in tissues from PDAC, chronic pancreatitis and pancreatic intraepithelial neoplasm. Our data show that Notch1/Hes1-signaling pathway is activated during early pancreatic embryogenesis and reaches the highest at birth. After pancreas is fully developed, Notch1/Hes1 pathway is inactivated even though Notch1 protein cell-surface expression is upregulated. We also showed that the expression of both Notch1 and Hes1 are present in 50% (33/66) of PDACs, but not in pancreatic intraepithelial neoplasms. These findings indicate that Notch1 activation is only apparent in late stage of pancreatic carcinogenesis, suggesting that treatment with Notch-signaling inhibitors including γ-secretase should be selectively used for PDACs with confirmed Notch1-signaling activation.

  17. NOTCH2 signaling confers immature morphology and aggressiveness in human hepatocellular carcinoma cells.

    PubMed

    Hayashi, Yoshihiro; Osanai, Makoto; Lee, Gang-Hong

    2015-10-01

    The NOTCH family of membranous receptors plays key roles during development and carcinogenesis. Since NOTCH2, yet not NOTCH1 has been shown essential for murine hepatogenesis, NOTCH2 rather than NOTCH1 may be more relevant to human hepatocarcinogenesis; however, no previous studies have supported this hypothesis. We therefore assessed the role of NOTCH2 in human hepatocellular carcinoma (HCC) by immunohistochemistry and cell culture. Immunohistochemically, 19% of primary HCCs showed nuclear staining for NOTCH2, indicating activated NOTCH2 signaling. NOTCH2-positive HCCs were on average in more advanced clinical stages, and exhibited more immature cellular morphology, i.e. higher nuclear-cytoplasmic ratios and nuclear densities. Such features were not evident in NOTCH1‑positive HCCs. In human HCC cell lines, abundant NOTCH2 expression was associated with anaplasia, represented by loss of E-cadherin. When NOTCH2 signaling was stably downregulated in HLF cells, an anaplastic HCC cell line, the cells were attenuated in potential for in vitro invasiveness and migration, as well as in vivo tumorigenicity accompanied by histological maturation. Generally, inverse results were obtained for a differentiated HCC cell line, Huh7, manipulated to overexpress activated NOTCH2. These findings suggested that the NOTCH2 signaling may confer aggressive behavior and immature morphology in human HCC cells.

  18. Loss of CCM3 impairs DLL4-Notch signalling: implication in endothelial angiogenesis and in inherited cerebral cavernous malformations.

    PubMed

    You, Chao; Sandalcioglu, Ibrahim Erol; Dammann, Philipp; Felbor, Ute; Sure, Ulrich; Zhu, Yuan

    2013-03-01

    CCM3, a product of the cerebral cavernous malformation 3 or programmed cell death 10 gene (CCM3/PDCD10), is broadly expressed throughout development in both vertebrates and invertebrates. Increasing evidence indicates a crucial role of CCM3 in vascular development and in regulation of angiogenesis and apoptosis. Furthermore, loss of CCM3 causes inherited (familial) cerebral cavernous malformation (CCM), a common brain vascular anomaly involving aberrant angiogenesis. This study focused on signalling pathways underlying the angiogenic functions of CCM3. Silencing CCM3 by siRNA stimulated endothelial proliferation, migration and sprouting accompanied by significant downregulation of the core components of Notch signalling including DLL4, Notch4, HEY2 and HES1 and by activation of VEGF and Erk pathways. Treatment with recombinant DLL4 (rhDLL4) restored DLL4 expression and reversed CCM3-silence-mediated impairment of Notch signalling and reduced the ratio of VEGF-R2 to VEGF-R1 expression. Importantly, restoration of DLL4-Notch signalling entirely rescued the hyper-angiogenic phenotype induced by CCM3 silence. A concomitant loss of CCM3 and the core components of DLL4-Notch signalling were also demonstrated in CCM3-deficient endothelial cells derived from human CCM lesions (CCMEC) and in a CCM3 germline mutation carrier. This study defined DLL4 as a key downstream target of CCM3 in endothelial cells. CCM3/DLL4-Notch pathway serves as an important signalling for endothelial angiogenesis and is potentially implicated in the pathomechanism of human CCMs.

  19. Blockade of Notch signaling promotes acetaminophen-induced liver injury.

    PubMed

    Jiang, Longfeng; Ke, Michael; Yue, Shi; Xiao, Wen; Yan, Youde; Deng, Xiaozhao; Ying, Qi-Long; Li, Jun; Ke, Bibo

    2017-03-13

    Liver injury after experimental acetaminophen treatment is mediated both by direct hepatocyte injury through a P450-generated toxic metabolite and indirectly by activated liver Kupffer cells and neutrophils. This study was designed to investigate the role of Notch signaling in the regulation of innate immune responses in acetaminophen (APAP)-induced liver injury. Using a mouse model of APAP-induced liver injury, wild-type (WT) and toll-like receptor 4 knockout (TLR4 KO) mice were injected intraperitoneally with APAP or PBS. Some animals were injected with γ-secretase inhibitor DAPT or DMSO vehicle. For the in vitro study, bone marrow-derived macrophages (BMMs) were transfected with Notch1 siRNA, TLR4 siRNA, and non-specific (NS) siRNA and stimulated with LPS. Indeed, paracetamol/acetaminophen-induced liver damage was worse after Notch blockade with DAPT in wild-type mice, which was accompanied by significantly increased ALT levels, diminished hairy and enhancer of split-1 (Hes1), and phosphorylated Stat3 and Akt but enhanced high mobility group box 1 (HMGB1), TLR4, NF-κB, and NLRP3 activation after APAP challenge. Mice receiving DAPT increased macrophage and neutrophil accumulation and hepatocellular apoptosis. However, TLR4 KO mice that received DAPT reduced APAP-induced liver damage and NF-κB, NLRP3, and cleaved caspase-1 activation. BMMs transfected with Notch1 siRNA reduced Hes1 and phosphorylated Stat3 and Akt but augmented HMGB1, TLR4, NF-κB, and NLRP3. Furthermore, TLR4 siRNA knockdown resulted in decreased NF-κB and NLRP3 and cleaved caspase-1 and IL-1β levels following LPS stimulation. These results demonstrate that Notch signaling regulates innate NLRP3 inflammasome activation through regulation of HMGB1/TLR4/NF-κB activation in APAP-induced liver injury. Our novel findings underscore the critical role of the Notch1-Hes1 signaling cascade in the regulation of innate immunity in APAP-triggered liver inflammation. This might imply a novel therapeutic

  20. Autophagy is involved in mouse kidney development and podocyte differentiation regulated by Notch signalling.

    PubMed

    Zhang, Chuyue; Li, Wen; Wen, Junkai; Yang, Zhuo

    2017-02-03

    Podocyte dysfunction results in glomerular diseases accounted for 90% of end-stage kidney disease. The evolutionarily conserved Notch signalling makes a crucial contribution in podocyte development and function. However, the underlying mechanism of Notch pathway modulating podocyte differentiation remains less obvious. Autophagy, reported to be related with Notch signalling pathways in different animal models, is regarded as a possible participant during podocyte differentiation. Here, we found the dynamic changes of Notch1 were coincided with autophagy: they both increased during kidney development and podocyte differentiation. Intriguingly, when Notch signalling was down-regulated by DAPT, autophagy was greatly diminished, and differentiation was also impaired. Further, to better understand the relationship between Notch signalling and autophagy in podocyte differentiation, rapamycin was added to enhance autophagy levels in DAPT-treated cells, and as a result, nephrin was recovered and DAPT-induced injury was ameliorated. Therefore, we put forward that autophagy is involved in kidney development and podocyte differentiation regulated by Notch signalling.

  1. TSG attenuates LPC-induced endothelial cells inflammatory damage through notch signaling inhibition.

    PubMed

    Zhao, Jing; Liang, Yuan; Song, Fan; Xu, Shouzhu; Nian, Lun; Zhou, Xuanxuan; Wang, Siwang

    2016-01-01

    Lysophosphatidylcholine (LPC) induces inflammation in endothelial cells (ECs) but the mechanism is not fully understood. The Notch signaling pathway is involved in chronic EC inflammation, but its functions in LPC-induced endothelial inflammatory damage and 2,3,5,4'-tetrahydroxystilbene-2-O-β-d-glucoside's (TSG) protective effect during LPC-induced inflammatory damage in human umbilical vein endothelial cells (HUVECs) is largely unknown. We report that Notch signaling activation contributed to LPC-induced injury in HUVECs, and that TSG protected HUVECs from LPC-induced injury by antagonizing Notch signaling activation by LPC. γ-secretase inhibitor (DAPT), a specific inhibitor of the Notch signaling pathway, and Notch1 siRNA were used to inhibit Notch activity. HUVECs were exposed to LPC in the presence or absence of TSG, DAPT, and Notch1 siRNA. LPC treatment of HUVECs resulted in reduced cell viability, and Notch1 and Hes1 upregulation. Either silencing of Notch1 by siRNA or pharmacological inhibition of Notch signaling by DAPT prevented the loss of cell viability, and induction of apoptosis, and enhanced expression Notch1, Hes1 and MCP-1 by LPC in HUVECs. Similarly, TSG reduced LPC stimulation of Notch1, Hes1, and MCP-1 expression, prevented the release of IL-6 and CRP and rescued HUVECs from LPC-induced cell damage. Our data indicate that the Notch signaling pathway is a crucial mediator of endothelial inflammatory damage and that TSG protects against endothelial inflammatory damage by inhibiting the Notch signaling pathway. Our findings suggest that targeting Notch signaling by natural products such as TSG is a promising strategy for the prevention and treatment of chronic inflammation associated diseases, including atherosclerosis. © 2015 IUBMB Life, 68(1):37-50, 2016.

  2. Nandrolone reduces activation of Notch signaling in denervated muscle associated with increased Numb expression

    SciTech Connect

    Liu, Xin-Hua; Yao, Shen; Qiao, Rui-Fang; Levine, Alice C.; Kirschenbaum, Alexander; Pan, Jiangping; Wu, Yong; Qin, Weiping; Bauman, William A.; Cardozo, Christopher P.

    2011-10-14

    Highlights: {yields} Nerve transection increased Notch signaling in paralyzed muscle. {yields} Nandrolone prevented denervation-induced Notch signaling. {yields} Nandrolone induced the expression of an inhibitor of the Notch signaling, Numb. {yields} Reduction of denervation-induced Notch signaling by nandrolone is likely through upregulation of Numb. -- Abstract: Nandrolone, an anabolic steroid, slows denervation-atrophy in rat muscle. The molecular mechanisms responsible for this effect are not well understood. Androgens and anabolic steroids activate Notch signaling in animal models of aging and thereby mitigate sarcopenia. To explore the molecular mechanisms by which nandrolone prevents denervation-atrophy, we investigated the effects of nandrolone on Notch signaling in denervated rat gastrocnemius muscle. Denervation significantly increased Notch activity reflected by elevated levels of nuclear Notch intracellular domain (NICD) and expression of Hey1 (a Notch target gene). Activation was greatest at 7 and 35 days after denervation but remained present at 56 days after denervation. Activation of Notch in denervated muscle was prevented by nandrolone associated with upregulated expression of Numb mRNA and protein. These data demonstrate that denervation activates Notch signaling, and that nandrolone abrogates this response associated with increased expression of Numb, suggesting a potential mechanism by which nandrolone reduces denervation-atrophy.

  3. Regulation of Notch signaling during T- and B-cell development by O-fucose glycans.

    PubMed

    Stanley, Pamela; Guidos, Cynthia J

    2009-07-01

    Notch signaling is required for the development of all T cells and marginal zone (MZ) B cells. Specific roles in T- and B-cell differentiation have been identified for different Notch receptors, the canonical Delta-like (Dll) and Jagged (Jag) Notch ligands, and downstream effectors of Notch signaling. Notch receptors and ligands are post-translationally modified by the addition of glycans to extracellular domain epidermal growth factor-like (EGF) repeats. The O-fucose glycans of Notch cell-autonomously modulate Notch-ligand interactions and the strength of Notch signaling. These glycans are initiated by protein O-fucosyltransferase 1 (Pofut1), and elongated by the transfer of N-acetylglucosamine (GlcNAc) to the fucose by beta1,3GlcNAc-transferases termed lunatic, manic, or radical fringe. This review discusses T- and B-cell development from progenitors deficient in O-fucose glycans. The combined data show that Lfng and Mfng regulate T-cell development by enhancing the interactions of Notch1 in T-cell progenitors with Dll4 on thymic epithelial cells. In the spleen, Lfng and Mfng cooperate to modify Notch2 in MZ B progenitors, enhancing their interaction with Dll1 on endothelial cells and regulating MZ B-cell production. Removal of O-fucose affects Notch signaling in myelopoiesis and lymphopoiesis, and the O-fucose glycan in the Notch1 ligand-binding domain is required for optimal T-cell development.

  4. Xanthohumol inhibits Notch signaling and induces apoptosis in hepatocellular carcinoma.

    PubMed

    Kunnimalaiyaan, Selvi; Sokolowski, Kevin M; Balamurugan, Mariappan; Gamblin, T Clark; Kunnimalaiyaan, Muthusamy

    2015-01-01

    Despite improvement in therapeutic strategies, median survival in advanced hepatocellular carcinoma (HCC) remains less than one year. Therefore, molecularly targeted compounds with less toxic profiles are needed. Xanthohumol (XN), a prenylated chalcone has been shown to have anti-proliferative effects in various cancers types in vitro. XN treatment in healthy mice and humans yielded favorable pharmacokinetics and bioavailability. Therefore, we determined to study the effects of XN and understand the mechanism of its action in HCC. The effects of XN on a panel of HCC cell lines were assessed for cell viability, colony forming ability, and cellular proliferation. Cell lysates were analyzed for pro-apoptotic (c-PARP and cleaved caspase-3) and anti-apoptotic markers (survivin, cyclin D1, and Mcl-1). XN concentrations of 5 μM and above significantly reduced the cell viability, colony forming ability and also confluency of all four HCC cell lines studied. Furthermore, growth suppression due to apoptosis was evidenced by increased expression of pro-apoptotic and reduced expression of anti-apoptotic proteins. Importantly, XN treatment inhibited the Notch signaling pathway as evidenced by the decrease in the expression of Notch1 and HES-1 proteins. Ectopic expression of Notch1 in HCC cells reverses the anti-proliferative effect of XN as evidenced by reduced growth suppression compared to control. Taken together these results suggested that XN mediated growth suppression is appeared to be mediated by the inhibition of the Notch signaling pathway. Therefore, our findings warrants further studies on XN as a potential agent for the treatment for HCC.

  5. Xanthohumol Inhibits Notch Signaling and Induces Apoptosis in Hepatocellular Carcinoma

    PubMed Central

    Kunnimalaiyaan, Selvi; Gamblin, T. Clark; Kunnimalaiyaan, Muthusamy

    2015-01-01

    Despite improvement in therapeutic strategies, median survival in advanced hepatocellular carcinoma (HCC) remains less than one year. Therefore, molecularly targeted compounds with less toxic profiles are needed. Xanthohumol (XN), a prenylated chalcone has been shown to have anti-proliferative effects in various cancers types in vitro. XN treatment in healthy mice and humans yielded favorable pharmacokinetics and bioavailability. Therefore, we determined to study the effects of XN and understand the mechanism of its action in HCC. The effects of XN on a panel of HCC cell lines were assessed for cell viability, colony forming ability, and cellular proliferation. Cell lysates were analyzed for pro-apoptotic (c-PARP and cleaved caspase-3) and anti-apoptotic markers (survivin, cyclin D1, and Mcl-1). XN concentrations of 5μM and above significantly reduced the cell viability, colony forming ability and also confluency of all four HCC cell lines studied. Furthermore, growth suppression due to apoptosis was evidenced by increased expression of pro-apoptotic and reduced expression of anti-apoptotic proteins. Importantly, XN treatment inhibited the Notch signaling pathway as evidenced by the decrease in the expression of Notch1 and HES-1 proteins. Ectopic expression of Notch1 in HCC cells reverses the anti-proliferative effect of XN as evidenced by reduced growth suppression compared to control. Taken together these results suggested that XN mediated growth suppression is appeared to be mediated by the inhibition of the Notch signaling pathway. Therefore, our findings warrants further studies on XN as a potential agent for the treatment for HCC. PMID:26011160

  6. Modeling notch signaling in normal and neoplastic hematopoiesis: global gene expression profiling in response to activated notch expression.

    PubMed

    Ganapati, Uma; Tan, Hongying Tina; Lynch, Maureen; Dolezal, Milana; de Vos, Sven; Gasson, Judith C

    2007-08-01

    In normal hematopoiesis, proliferation is tightly linked to differentiation in ways that involve cell-cell interaction with stromal elements in the bone marrow stem cell niche. Numerous in vitro and in vivo studies strongly support a role for Notch signaling in the regulation of stem cell renewal and hematopoiesis. Not surprisingly, mutations in the Notch gene have been linked to a number of types of malignancies. To better define the function of Notch in both normal and neoplastic hematopoiesis, a tetracycline-inducible system regulating expression of a ligand-independent, constitutively active form of Notch1 was introduced into murine E14Tg2a embryonic stem cells. During coculture, OP9 stromal cells induce the embryonic stem cells to differentiate first to hemangioblasts and subsequently to hematopoietic stem cells. Our studies indicate that activation of Notch signaling in flk+ hemangioblasts dramatically reduces their survival and proliferative capacity and lowers the levels of hematopoietic stem cell markers CD34 and c-Kit and the myeloid marker CD11b. Global gene expression profiling of day 8 hematopoietic progenitors in the absence and presence of activated Notch yield candidate genes required for normal hematopoietic differentiation, as well as putative downstream targets of oncogenic forms of Notch including the noncanonical Wnts Wnt4 and 5A. Disclosure of potential conflicts of interest is found at the end of this article.

  7. FGF signaling specifies hematopoietic stem cells through its regulation of somitic Notch signaling

    PubMed Central

    Lee, Yoonsung; Manegold, Jennifer E; Kim, Albert D; Pouget, Claire; Stachura, David L; Clements, Wilson K; Traver, David

    2014-01-01

    Hematopoietic stem cells (HSCs) derive from hemogenic endothelial cells of the primitive dorsal aorta (DA) during vertebrate embryogenesis. The molecular mechanisms governing this unique endothelial to hematopoietic transition remain unclear. Here, we demonstrate a novel requirement for fibroblast growth factor (FGF) signaling in HSC emergence. This requirement is non-cell-autonomous, and acts within the somite to bridge the Wnt and Notch signaling pathways. We previously demonstrated that Wnt16 regulates the somitic expression of two Notch ligands, deltaC (dlc) and deltaD (dld), whose combined function is required for HSC fate. How Wnt16 connects to Notch function has remained an open question. Our current studies demonstrate that FGF signaling, via FGF receptor 4 (Fgfr4), mediates a signal transduction pathway between Wnt16 and Dlc, but not Dld, to regulate HSC specification. Our findings demonstrate that FGF signaling acts as a key molecular relay within the developmental HSC niche to instruct HSC fate. PMID:25428693

  8. Compensatory Flux Changes within an Endocytic Trafficking Network Maintain Thermal Robustness of Notch Signaling

    PubMed Central

    Shimizu, Hideyuki; Woodcock, Simon A.; Wilkin, Marian B.; Trubenová, Barbora; Monk, Nicholas A.M.; Baron, Martin

    2014-01-01

    Summary Developmental signaling is remarkably robust to environmental variation, including temperature. For example, in ectothermic animals such as Drosophila, Notch signaling is maintained within functional limits across a wide temperature range. We combine experimental and computational approaches to show that temperature compensation of Notch signaling is achieved by an unexpected variety of endocytic-dependent routes to Notch activation which, when superimposed on ligand-induced activation, act as a robustness module. Thermal compensation arises through an altered balance of fluxes within competing trafficking routes, coupled with temperature-dependent ubiquitination of Notch. This flexible ensemble of trafficking routes supports Notch signaling at low temperature but can be switched to restrain Notch signaling at high temperature and thus compensates for the inherent temperature sensitivity of ligand-induced activation. The outcome is to extend the physiological range over which normal development can occur. Similar mechanisms may provide thermal robustness for other developmental signals. PMID:24855951

  9. Nandrolone reduces activation of Notch signaling in denervated muscle associated with increased Numb expression.

    PubMed

    Liu, Xin-Hua; Yao, Shen; Qiao, Rui-Fang; Levine, Alice C; Kirschenbaum, Alexander; Pan, Jiangping; Wu, Yong; Qin, Weiping; Bauman, William A; Cardozo, Christopher P

    2011-10-14

    Nandrolone, an anabolic steroid, slows denervation-atrophy in rat muscle. The molecular mechanisms responsible for this effect are not well understood. Androgens and anabolic steroids activate Notch signaling in animal models of aging and thereby mitigate sarcopenia. To explore the molecular mechanisms by which nandrolone prevents denervation-atrophy, we investigated the effects of nandrolone on Notch signaling in denervated rat gastrocnemius muscle. Denervation significantly increased Notch activity reflected by elevated levels of nuclear Notch intracellular domain (NICD) and expression of Hey1 (a Notch target gene). Activation was greatest at 7 and 35 days after denervation but remained present at 56 days after denervation. Activation of Notch in denervated muscle was prevented by nandrolone associated with upregulated expression of Numb mRNA and protein. These data demonstrate that denervation activates Notch signaling, and that nandrolone abrogates this response associated with increased expression of Numb, suggesting a potential mechanism by which nandrolone reduces denervation-atrophy.

  10. Heterogeneity of Notch signaling in astrocytes and the effects of GFAP and vimentin deficiency.

    PubMed

    Lebkuechner, Isabell; Wilhelmsson, Ulrika; Möllerström, Elin; Pekna, Marcela; Pekny, Milos

    2015-10-01

    Astrocytes have multiple roles in the CNS including control of adult neurogenesis. We recently showed that astrocyte inhibition of neurogenesis through Notch signaling depends on the intermediate filament proteins glial fibrillary acidic protein (GFAP) and vimentin. Here, we used real-time quantitative PCR to analyze gene expression in individual mouse astrocytes in primary cultures and in GFAP(POS) or Aldh1L1(POS) astrocytes freshly isolated from uninjured, contralesional and lesioned hippocampus 4 days after entorhinal cortex lesion. To determine the Notch signaling competence of individual astrocytes, we measured the mRNA levels of Notch ligands and Notch1 receptor. We found that whereas most cultured and freshly isolated astrocytes were competent to receive Notch signals, only a minority of astrocytes were competent to send Notch signals. Injury increased the fraction of astrocyte subpopulation unable to send and receive Notch signals, thus resembling primary astrocytes in vitro. Astrocytes deficient of GFAP and vimentin showed decreased Notch signal sending competence and altered expression of Notch signaling pathway-related genes Dlk2, Notch1, and Sox2. Furthermore, we identified astrocyte subpopulations based on their mRNA and protein expression of nestin and HB-EGF. This study improves our understanding of astrocyte heterogeneity, and points to astrocyte cytoplasmic intermediate filaments as targets for neural cell replacement strategies.

  11. Aging neural progenitors lose competence to respond to mitogenic Notch signaling

    PubMed Central

    Farnsworth, Dylan R.; Bayraktar, Omer Ali; Doe, Chris Q.

    2015-01-01

    Drosophila neural stem cells (neuroblasts) are a powerful model system for investigating stem cell self-renewal, specification of temporal identity, and progressive restriction in competence. Notch signaling is a conserved cue that is an important determinant of cell fate in many contexts across animal development; for example mammalian T cell differentiation in the thymus and neuroblast specification in Drosophila are both regulated by Notch signaling. However, Notch also functions as a mitogen, and constitutive Notch signaling potentiates T cell leukemia as well as Drosophila neuroblast tumors. While the role of Notch signaling has been studied in these and other cell types, it remains unclear how stem cells and progenitors change competence to respond to Notch over time. Notch is required in type II neuroblasts for normal development of their transit amplifying progeny, intermediate neural progenitors (INPs). Here we find that aging INPs lose competence to respond to constitutively active Notch signaling. Moreover, we show that reducing the levels of the old INP temporal transcription factor Eyeless/Pax6 allows Notch signaling to promote the de-differentiation of INP progeny into ectopic INPs, thereby creating a proliferative mass of ectopic progenitors in the brain. These findings provide a new system for studying progenitor competence, and identify a novel role for the conserved transcription factor Eyeless/Pax6 in blocking Notch signaling during development. PMID:26585279

  12. Genetic deletion of Rnd3 results in aqueductal stenosis leading to hydrocephalus through up-regulation of Notch signaling.

    PubMed

    Lin, Xi; Liu, Baohui; Yang, Xiangsheng; Yue, Xiaojing; Diao, Lixia; Wang, Jing; Chang, Jiang

    2013-05-14

    Rho family guanosine triphosphatase (GTPase) 3 (Rnd3), a member of the small Rho GTPase family, is involved in the regulation of cell actin cytoskeleton dynamics, cell migration, and proliferation through the Rho kinase-dependent signaling pathway. We report a role of Rnd3 in the pathogenesis of hydrocephalus disorder. Mice with Rnd3 genetic deletion developed severe obstructive hydrocephalus with enlargement of the lateral and third ventricles, but not of the fourth ventricles. The cerebral aqueducts in Rnd3-null mice were partially or completely blocked by the overgrowth of ependymal epithelia. We examined the molecular mechanism contributing to this Rnd3-deficiency-mediated hydrocephalus and found that Rnd3 is a regulator of Notch signaling. Rnd3 deficiency, through either gene deletion or siRNA knockdown, resulted in a decrease in Notch intracellular domain (NICD) protein degradation. However, there was no correlated change in mRNA change, which in turn led to an increase in NICD protein levels. Immunoprecipitation analysis demonstrated that Rnd3 and NICD physically interacted, and that down-regulation of Rnd3 attenuated NICD protein ubiquitination. This eventually enhanced Notch signaling activity and promoted aberrant growth of aqueduct ependymal cells, resulting in aqueduct stenosis and the development of congenital hydrocephalus. Inhibition of Notch activity rescued the hydrocephalus disorder in the mutant animals.

  13. Notch1-mediated signaling regulates proliferation of porcine satellite cells (PSCs).

    PubMed

    Qin, Lili; Xu, Jian; Wu, Zhenfang; Zhang, Zhe; Li, Jiaqi; Wang, Chong; Long, Qiaoming

    2013-02-01

    Notch signaling is an evolutionarily conserved cell-cell communication mechanism involved in the regulation of cell proliferation, differentiation and fate decisions of mammalian cells. In the present study, we investigated the possible requirement for Notch signaling in the proliferation and differentiation of porcine satellite cells. We show that Notch1, 2 and 3 are expressed in cultured porcine satellite cells. Knock-down of NOTCH1, but not NOTCH2 and NOTCH3, decreases the proliferation of porcine satellite cells. In contrast, enhancement of NOTCH1 expression via treatment of porcine satellite cells with recombinant NF-κB increases the proliferation of porcine satellite cells. The alteration of porcine satellite cell proliferation is associated with significant changes in the expression of cell cycle related genes (cyclin B1, D1, D2, E1 and p21), myogenic regulatory factors (MyoD and myogenin) and the Notch effector Hes5. In addition, alteration of Notch1 expression in porcine satellite cells causes changes in the expression of GSK3β-3. Taken together, these findings suggest that of the four notch-related genes, Notch1is likely to be required for regulating the proliferation and therefore the maintenance of porcine satellite cells in vivo, and do so through activation of the Notch effector gene Hes5.

  14. The Role of Notch Signaling Pathway in Breast Cancer Pathogenesis

    DTIC Science & Technology

    2004-07-01

    coexpression of a constitutively active form of Notch1 in immortalized breast epithelial HMLE cells expressing low levels of oncogenic Ras rendered them...the Notch-Ras pathway interaction revealed that nuclear localization of Notch1 is essential for this cooperation. Dissection of Ras-pathways using the...activates Raf/MAPK pathway, formed efficient colonies with activated Notch1 . Interestingly, I found that expression of activated Notch1 rendered the

  15. Enhancement of Notch receptor maturation and signaling sensitivity by Cripto-1.

    PubMed

    Watanabe, Kazuhide; Nagaoka, Tadahiro; Lee, Joseph M; Bianco, Caterina; Gonzales, Monica; Castro, Nadia P; Rangel, Maria Cristina; Sakamoto, Kei; Sun, Youping; Callahan, Robert; Salomon, David S

    2009-11-02

    Nodal and Notch signaling pathways play essential roles in vertebrate development. Through a yeast two-hybrid screening, we identified Notch3 as a candidate binding partner of the Nodal coreceptor Cripto-1. Coimmunoprecipitation analysis confirmed the binding of Cripto-1 with all four mammalian Notch receptors. Deletion analyses revealed that the binding of Cripto-1 and Notch1 is mediated by the Cripto-1/FRL-1/Cryptic domain of Cripto-1 and the C-terminal region of epidermal growth factor-like repeats of Notch1. Binding of Cripto-1 to Notch1 occurred mainly in the endoplasmic reticulum-Golgi network. Cripto-1 expression resulted in the recruitment of Notch1 protein into lipid raft microdomains and enhancement of the furin-like protein convertase-mediated proteolytic maturation of Notch1 (S1 cleavage). Enhanced S1 cleavage resulted in the sensitization to ligand-induced activation of Notch signaling. In addition, knockdown of Cripto-1 expression in human and mouse embryonal carcinoma cells desensitized the ligand-induced Notch signaling activation. These results suggest a novel role of Cripto-1 in facilitating the posttranslational maturation of Notch receptors.

  16. Regulation of notch endosomal sorting and signaling by Drosophila Nedd4 family proteins.

    PubMed

    Wilkin, Marian B; Carbery, Ann-Marie; Fostier, Maggy; Aslam, Hanna; Mazaleyrat, Sabine L; Higgs, Jenny; Myat, Anna; Evans, Dana A P; Cornell, Michael; Baron, Martin

    2004-12-29

    The Notch receptor mediates a short-range signal that regulates many cell fate decisions. The misregulation of Notch has been linked to cancer and to developmental disorders. Upon binding to its ligands, Delta (Dl) or Serrate (Ser), the Notch ectodomain is shed by the action of an ADAM protease. The Notch intracellular domain is subsequently released proteolytically from the membrane by Presenilin and translocates to the nucleus to activate the transcription factor, Suppressor of Hairless. We show in Drosophila that Notch signaling is limited by the activity of two Nedd4 family HECT domain proteins, Suppressor of deltex [Su(dx)] and DNedd4. We rule out models by which Su(dx) downregulates Notch through modulating Deltex or by limiting the adherens junction accumulation of Notch. Instead, we show that Su(dx) regulates the postendocytic sorting of Notch within the early endosome to an Hrs- and ubiquitin-enriched subdomain en route to the late endosome. We propose a model in which endocytic sorting of Notch mediates a decision between its activation and downregulation. Such intersections between trafficking routes may provide key points at which other signals can modulate Notch activity in both normal development and in the pathological misactivation of Notch.

  17. Notch signalling suppresses regulatory T-cell function in murine experimental autoimmune uveitis.

    PubMed

    Rong, Hua; Shen, Hongjie; Xu, Yueli; Yang, Hai

    2016-12-01

    Autoimmune uveitis is an intraocular inflammatory disorder in developed countries. Understanding the mechanisms underlying the development and modulation of immune reaction in uveitic eyes is critical for designing therapeutic interventions. Here we investigated the role of Notch signalling in regulatory T-cell (Treg cell) function during experimental autoimmune uveitis (EAU). Using the Foxp3-GFP reporter mouse strain, the significance of Notch signalling for the function of infiltrating Treg cells was characterized in an EAU model. We found that infiltrating Treg cells substantially expressed Notch-1, Notch-2, JAG1 and DLL1 in uveitic eyes. Activation of Notch signalling, represented by expression of HES1 and HES5, was enhanced in infiltrating Treg cells. Treatment with JAG1 and DLL1 down-regulated Foxp3 expression and immunosuppressive activity of isolated infiltrating Treg cells in vitro, whereas neutralizing antibodies against JAG1 and DLL1 diminished Notch ligand-mediated negative effects on Treg cells. To investigate the significance of Notch signalling for Treg cell function in vivo, lentivirus-derived Notch short hairpin RNAs were transduced into in vitro expanded Treg cells before adoptive transfer of Treg cells into EAU mice. Transfer of Notch-1-deficient Treg cells remarkably reduced pro-inflammatory cytokine production and inflammatory cell infiltration in uveitic eyes. Taken together, Notch signalling negatively modulates the immunosuppressive function of infiltrating Treg cells in mouse EAU.

  18. Notch Signaling Mediates Skeletal Muscle Atrophy in Cancer Cachexia Caused by Osteosarcoma

    PubMed Central

    Agarwal, Rashmi; March, Daniel; Voigt, Clifford

    2016-01-01

    Skeletal muscle atrophy in cancer cachexia is mediated by the interaction between muscle stem cells and various tumor factors. Although Notch signaling has been known as a key regulator of both cancer development and muscle stem cell activity, the potential involvement of Notch signaling in cancer cachexia and concomitant muscle atrophy has yet to be elucidated. The murine K7M2 osteosarcoma cell line was used to generate an orthotopic model of sarcoma-associated cachexia, and the role of Notch signaling was evaluated. Skeletal muscle atrophy was observed in the sarcoma-bearing mice, and Notch signaling was highly active in both tumor tissues and the atrophic skeletal muscles. Systemic inhibition of Notch signaling reduced muscle atrophy. In vitro coculture of osteosarcoma cells with muscle-derived stem cells (MDSCs) isolated from normal mice resulted in decreased myogenic potential of MDSCs, while the application of Notch inhibitor was able to rescue this repressed myogenic potential. We further observed that Notch-activating factors reside in the exosomes of osteosarcoma cells, which activate Notch signaling in MDSCs and subsequently repress myogenesis. Our results revealed that signaling between tumor and muscle via the Notch pathway may play an important role in mediating the skeletal muscle atrophy seen in cancer cachexia. PMID:27378829

  19. Curcumin attenuates endothelial cell oxidative stress injury through Notch signaling inhibition.

    PubMed

    Yang, Yang; Duan, Weixun; Liang, Zhenxin; Yi, Wei; Yan, Juanjuan; Wang, Ning; Li, Yue; Chen, Wensheng; Yu, Shiqiang; Jin, Zhenxiao; Yi, Dinghua

    2013-03-01

    Previous studies have demonstrated that Notch signaling pathway plays a regulatory role in cellular oxidative stress injury (OSI). In this study, our aim was to explore the role of the Notch signaling pathway in hydrogen peroxide (H(2)O(2))-induced OSI and the protective effect of curcumin during (H(2)O(2))-induced injury in human umbilical vein endothelial cells (HUVECs). DAPT, a specific inhibitor of the Notch signaling pathway, and Notch1 siRNA were used to study Notch activity. Further, HUVECs were exposed to H(2)O(2) in the absence or presence of curcumin. DAPT and Notch1 siRNA significantly inhibited OSI and the expression of Notch1 and Hes1. Curcumin conferred a protective effect on the HUVECs against H(2)O(2), which was evidenced by improved cell viability, adhesive ability and migratory ability and a decreased apoptotic index, decreased production of reactive oxygen species (ROS) and a reduction in several biochemical parameters. Immunofluorescence and Western blotting analyses demonstrated that H(2)O(2) treatment upregulated the expression of Notch1, Hes1, Caspase3, Bax and cytochrome c downregulated the expression of Bcl2, and treatment with curcumin reversed these effects. We demonstrated for the first time that the inhibition of Notch signaling pathway imparts a protective effect against endothelial OSI. The protective effects of curcumin against OSI are at least in part dependent on Notch1 inhibition.

  20. Notch Signaling Coordinates Progenitor Cell-Mediated Biliary Regeneration Following Partial Hepatectomy

    PubMed Central

    Lu, Jie; Zhou, Yingqun; Hu, Tianyuan; Zhang, Hui; Shen, Miao; Cheng, Ping; Dai, Weiqi; Wang, Fan; Chen, Kan; Zhang, Yan; Wang, Chengfeng; Li, Jingjing; Zheng, Yuanyuan; Yang, Jing; Zhu, Rong; Wang, Jianrong; Lu, Wenxia; Zhang, Huawei; Wang, Junshan; Xia, Yujing; De Assuncao, Thiago M.; Jalan-Sakrikar, Nidhi; Huebert, Robert C.; Bin Zhou; Guo, Chuanyong

    2016-01-01

    Aberrant transcriptional regulation contributes to the pathogenesis of both congenital and adult forms of liver disease. Although the transcription factor RBPJ is essential for liver morphogenesis and biliary development, its specific function in the differentiation of hepatic progenitor cells (HPC) has not been investigated, and little is known about its role in adult liver regeneration. HPCs are bipotent liver stem cells that can self-replicate and differentiate into hepatocytes or cholangiocytes in vitro. HPCs are thought to play an important role in liver regeneration and repair responses. While the coordinated repopulation of both hepatocyte and cholangiocyte compartment is pivotal to the structure and function of the liver after regeneration, the mechanisms coordinating biliary regeneration remain vastly understudied. Here, we utilized complex genetic manipulations to drive liver-specific deletion of the Rbpj gene in conjunction with lineage tracing techniques to delineate the precise functions of RBPJ during biliary development and HPC-associated biliary regeneration after hepatectomy. Furthermore, we demonstrate that RBPJ promotes HPC differentiation toward cholangiocytes in vitro and blocks hepatocyte differentiation through mechanisms involving Hippo-Notch crosstalk. Overall, this study demonstrates that the Notch-RBPJ signaling axis critically regulates biliary regeneration by coordinating the fate decision of HPC and clarifies the molecular mechanisms involved. PMID:26951801

  1. A role for the primary cilium in Notch signaling and epidermal differentiation during skin development.

    PubMed

    Ezratty, Ellen J; Stokes, Nicole; Chai, Sophia; Shah, Alok S; Williams, Scott E; Fuchs, Elaine

    2011-06-24

    Ciliogenesis precedes lineage-determining signaling in skin development. To understand why, we performed shRNA-mediated knockdown of seven intraflagellar transport proteins (IFTs) and conditional ablation of Ift-88 and Kif3a during embryogenesis. In both cultured keratinocytes and embryonic epidermis, all of these eliminated cilia, and many (not Kif3a) caused hyperproliferation. Surprisingly and independent of proliferation, ciliary mutants displayed defects in Notch signaling and commitment of progenitors to differentiate. Notch receptors and Notch-processing enzymes colocalized with cilia in wild-type epidermal cells. Moreover, differentiation defects in ciliary mutants were cell autonomous and rescued by activated Notch (NICD). By contrast, Shh signaling was neither operative nor required for epidermal ciliogenesis, Notch signaling, or differentiation. Rather, Shh signaling defects in ciliary mutants occurred later, arresting hair follicle morphogenesis in the skin. These findings unveil temporally and spatially distinct functions for primary cilia at the nexus of signaling, proliferation, and differentiation.

  2. Targeting aberrant expression of Notch-1 in ALDH(+) cancer stem cells in breast cancer.

    PubMed

    Pal, Deeksha; Kolluru, Venkatesh; Chandrasekaran, Balaji; Baby, Becca V; Aman, Masarath; Suman, Suman; Sirimulla, Suman; Sanders, Mary Ann; Alatassi, Houda; Ankem, Murali K; Damodaran, Chendil

    2017-03-01

    We have previously reported that high aldehyde dehydrogenase (ALDH) enzyme activity in breast cancer cells results in breast cancer stem cell (BCSC) properties by upregualting Notch-1 and epithelial mesenchymal markers. This results in chemoresistance in breast cancer. Here, we examined the functional and clinical significance of ALDH expression by measuring the ALDH levels in breast cancer tissues by immunohistochemistry. There was a significantly higher ALDH expression in higher grade breast cancer tumor tissues (Grade- II and III) versus normal breast tissues. Injection of BCSC (ALDH(+) and CD44(+) /CD22(-) ) cells resulted in aggressive tumor growth in athymic mice versus ALDH(-) cells. The ALDH(+) and CD44(+) /CD22(-) tumors grow rapidly and are larger than ALDH(-) tumors which were slow growing and smaller. Molecularly, ALDH(+) tumors expressed higher expression of Notch-1 and EMT markers than ALDH(-) tumors. Oral administration of the naturally occurring Psoralidin (Pso, 25 mg/kg of body weight) significantly inhibited the growth in ALDH(+) and ALDH(-) tumors as well. Psoralidin inhibited Notch-1 mediated EMT activation in ALDH(+) and ALDH(-) tumors-this confirms our in vitro findings. Our results suggest that Notch-1 could be an attractive target and inhibition of Notch-1 by Psoralidin may prevent pathogenesis of breast cancer as well as metastasis. © 2016 Wiley Periodicals, Inc.

  3. Notch signalling drives bone marrow stromal cell-mediated chemoresistance in acute myeloid leukemia.

    PubMed

    Takam Kamga, Paul; Bassi, Giulio; Cassaro, Adriana; Midolo, Martina; Di Trapani, Mariano; Gatti, Alessandro; Carusone, Roberta; Resci, Federica; Perbellini, Omar; Gottardi, Michele; Bonifacio, Massimiliano; Nwabo Kamdje, Armel Hervé; Ambrosetti, Achille; Krampera, Mauro

    2016-04-19

    Both preclinical and clinical investigations suggest that Notch signalling is critical for the development of many cancers and for their response to chemotherapy. We previously showed that Notch inhibition abrogates stromal-induced chemoresistance in lymphoid neoplasms. However, the role of Notch in acute myeloid leukemia (AML) and its contribution to the crosstalk between leukemia cells and bone marrow stromal cells remain controversial. Thus, we evaluated the role of the Notch pathway in the proliferation, survival and chemoresistance of AML cells in co-culture with bone marrow mesenchymal stromal cells expanded from both healthy donors (hBM-MSCs) and AML patients (hBM-MSCs*). As compared to hBM-MSCs, hBM-MSCs* showed higher level of Notch1, Jagged1 as well as the main Notch target gene HES1. Notably, hBM-MSCs* induced expression and activation of Notch signalling in AML cells, supporting AML proliferation and being more efficientin inducing AML chemoresistance than hBM-MSCs*. Pharmacological inhibition of Notch using combinations of Notch receptor-blocking antibodies or gamma-secretase inhibitors (GSIs), in presence of chemotherapeutic agents, significant lowered the supportive effect of hBM-MSCs and hBM-MSCs* towards AML cells, by activating apoptotic cascade and reducing protein level of STAT3, AKT and NF-κB.These results suggest that Notch signalling inhibition, by overcoming the stromal-mediated promotion of chemoresistance,may represent a potential therapeutic targetnot only for lymphoid neoplasms, but also for AML.

  4. SHARP is a novel component of the Notch/RBP-Jκ signalling pathway

    PubMed Central

    Oswald, Franz; Kostezka, Ulrike; Astrahantseff, Kathy; Bourteele, Soizic; Dillinger, Karin; Zechner, Ulrich; Ludwig, Leopold; Wilda, Monika; Hameister, Horst; Knöchel, Walter; Liptay, Susanne; Schmid, Roland M.

    2002-01-01

    Notch proteins are the receptors for an evolutionarily highly conserved signalling pathway that regulates numerous cell fate decisions during development. Signal transduction involves the presenilin-dependent intracellular processing of Notch and nuclear translocation of the intracellular domain of Notch, Notch-IC. Notch-IC associates with the DNA-binding protein RBP-Jκ/CBF-1 to activate transcription of Notch target genes. In the absence of Notch signalling, RBP-Jκ/CBF-1 acts as a transcriptional repressor through the recruitment of histone deacetylase (HDAC) corepressor complexes. We identified SHARP as an RBP-Jκ/CBF-1-interacting corepressor in a yeast two-hybrid screen. In cotransfection experiments, SHARP-mediated repression was sensitive to the HDAC inhibitor TSA and facilitated by SKIP, a highly conserved SMRT and RBP-Jκ-interacting protein. SHARP repressed Hairy/Enhancer of split (HES)-1 promoter activity, inhibited Notch-1-mediated transactivation and rescued Notch-1-induced inhibition of primary neurogenesis in Xenopus laevis embryos. Based on our data, we propose a model in which SHARP is a novel component of the HDAC corepressor complex, recruited by RBP-Jκ to repress transcription of target genes in the absence of activated Notch. PMID:12374742

  5. Cellular Notch responsiveness is defined by phosphoinositide 3-kinase-dependent signals

    PubMed Central

    Mckenzie, Grahame; Ward, George; Stallwood, Yvette; Briend, Emmanuel; Papadia, Sofia; Lennard, Andrew; Turner, Martin; Champion, Brian; Hardingham, Giles E

    2006-01-01

    Background Notch plays a wide-ranging role in controlling cell fate, differentiation and development. The PI3K-Akt pathway is a similarly conserved signalling pathway which regulates processes such as differentiation, proliferation and survival. Mice with disrupted Notch and PI3K signalling show phenotypic similarities during haematopoietic cell development, suggesting functional interaction between these pathways. Results We show that cellular responsiveness to Notch signals depends on the activity of the PI3K-Akt pathway in cells as diverse as CHO cells, primary T-cells and hippocampal neurons. Induction of the endogenous PI3K-Akt pathway in CHO cells (by the insulin pathway), in T-cells (via TCR activation) or in neurons (via TrKB activation) potentiates Notch-dependent responses. We propose that the PI3K-Akt pathway exerts its influence on Notch primarily via inhibition of GSK3-beta, a kinase known to phosphorylate and regulate Notch signals. Conclusion The PI3K-Akt pathway acts as a "gain control" for Notch signal responses. Since physiological levels of intracellular Notch are often low, coincidence with PI3K-activation may be crucial for induction of Notch-dependent responses. PMID:16507111

  6. Notch-1 Signalling Is Activated in Brain Arteriovenous Malformations in Humans

    ERIC Educational Resources Information Center

    ZhuGe, Qichuan; Zhong, Ming; Zheng, WeiMing; Yang, Guo-Yuan; Mao, XiaoOu; Xie, Lin; Chen, Gourong; Chen, Yongmei; Lawton, Michael T.; Young, William L.; Greenberg, David A.; Jin, Kunlin

    2009-01-01

    A role for the Notch signalling pathway in the formation of arteriovenous malformations during development has been suggested. However, whether Notch signalling is involved in brain arteriovenous malformations in humans remains unclear. Here, we performed immunohistochemistry on surgically resected brain arteriovenous malformations and found that,…

  7. Deadly crosstalk: Notch signaling at the intersection of EMT and cancer stem cells.

    PubMed

    Espinoza, Ingrid; Miele, Lucio

    2013-11-28

    Notch signaling is an evolutionarily conserved pathway involved in cell fate control during development, stem cell self-renewal and postnatal tissue differentiation. Roles for Notch in carcinogenesis, in the biology of cancer stem cells, tumor angiogenesis and epithelial-to-mesenchymal transition (EMT) have been reported. This mini-review describes the role of Notch signaling deregulation in EMT and tumor aggressiveness. We describe how accumulated evidence suggests that Notch inhibition is an attractive strategy for the treatment of several cancers, at least in part because of its potential to reverse or prevent EMT.

  8. The Notch intracellular domain integrates signals from Wnt, Hedgehog, TGFβ/BMP and hypoxia pathways.

    PubMed

    Borggrefe, Tilman; Lauth, Matthias; Zwijsen, An; Huylebroeck, Danny; Oswald, Franz; Giaimo, Benedetto Daniele

    2016-02-01

    Notch signaling is a highly conserved signal transduction pathway that regulates stem cell maintenance and differentiation in several organ systems. Upon activation, the Notch receptor is proteolytically processed, its intracellular domain (NICD) translocates into the nucleus and activates expression of target genes. Output, strength and duration of the signal are tightly regulated by post-translational modifications. Here we review the intracellular post-translational regulation of Notch that fine-tunes the outcome of the Notch response. We also describe how crosstalk with other conserved signaling pathways like the Wnt, Hedgehog, hypoxia and TGFβ/BMP pathways can affect Notch signaling output. This regulation can happen by regulation of ligand, receptor or transcription factor expression, regulation of protein stability of intracellular key components, usage of the same cofactors or coregulation of the same key target genes. Since carcinogenesis is often dependent on at least two of these pathways, a better understanding of their molecular crosstalk is pivotal.

  9. Notch1 Regulates Hippocampal Plasticity Through Interaction with the Reelin Pathway, Glutamatergic Transmission and CREB Signaling

    PubMed Central

    Brai, Emanuele; Marathe, Swananda; Astori, Simone; Fredj, Naila Ben; Perry, Elisabeth; Lamy, Christophe; Scotti, Alessandra; Alberi, Lavinia

    2015-01-01

    Notch signaling plays a crucial role in adult brain function such as synaptic plasticity, memory and olfaction. Several reports suggest an involvement of this pathway in neurodegenerative dementia. Yet, to date, the mechanism underlying Notch activity in mature neurons remains unresolved. In this work, we investigate how Notch regulates synaptic potentiation and contributes to the establishment of memory in mice. We observe that Notch1 is a postsynaptic receptor with functional interactions with the Reelin receptor, apolipoprotein E receptor 2 (ApoER2) and the ionotropic receptor, N-methyl-D-aspartate receptor (NMDAR). Targeted loss of Notch1 in the hippocampal CA fields affects Reelin signaling by influencing Dab1 expression and impairs the synaptic potentiation achieved through Reelin stimulation. Further analysis indicates that loss of Notch1 affects the expression and composition of the NMDAR but not AMPAR. Glutamatergic signaling is further compromised through downregulation of CamKII and its secondary and tertiary messengers resulting in reduced cAMP response element-binding (CREB) signaling. Our results identify Notch1 as an important regulator of mechanisms involved in synaptic plasticity and memory formation. These findings emphasize the possible involvement of this signaling receptor in dementia. Highlights In this paper, we propose a mechanism for Notch1-dependent plasticity that likely underlies the function of Notch1 in memory formation: Notch1 interacts with another important developmental pathway, the Reelin cascade. Notch1 regulates both NMDAR expression and composition. Notch1 influences a cascade of cellular events culminating in CREB activation. PMID:26635527

  10. Characterization of Notch Signalling Pathway Members in Normal Prostate, Prostatic Intraepithelial Neoplasia (PIN) and Prostatic Adenocarcinoma.

    PubMed

    Soylu, Hakan; Acar, Nuray; Ozbey, Ozlem; Unal, Betul; Koksal, Ismail Turker; Bassorgun, Ibrahim; Ciftcioglu, Akif; Ustunel, Ismail

    2016-01-01

    Prostate Cancer (PCa) holds the second place in terms of cancer-related mortality rate among men. The Notch signalling pathway regulates the proliferation and differentiation in embryonic and adult tissues and determines the cell fate. The body of knowledge in the present literature is currently controversial about the effect of the Notch pathway on prostatic cancer. Therefore, the present study aimed to examine the immunolocalization and expression levels of Notch1-4, Jagged1-2, Delta, HES1 and HES5 from among the members of the Notch signalling pathway in tissues of normal, prostatic intraepithelial neoplasia (PIN) and malignant prostate. The current study included a sample of 20 patients with localised prostatic adenocarcinoma, 18 patients with high grade PIN (H-PIN) and 18 normal prostatic tissue. Immunolocalisations of Notch1, 2, 3, 4, Jagged1, 2, Delta, HES1 and HES5 were identified through the immunohistochemical method. The findings of the present study showed that all in-scope members of the Notch signalling pathway were localised in PIN structures to a greater extent than in other tissues and from amongst these members, specifically Notch1, Notch4, Jagged1 and HES1 were at more significant levels. Consequently, the findings of the present study may indicate that the Notch signalling pathway can play a role especially in the formation of PIN structures.

  11. A novel non-canonical Notch signaling regulates expression of synaptic vesicle proteins in excitatory neurons

    PubMed Central

    Hayashi, Yukari; Nishimune, Hiroshi; Hozumi, Katsuto; Saga, Yumiko; Harada, Akihiro; Yuzaki, Michisuke; Iwatsubo, Takeshi; Kopan, Raphael; Tomita, Taisuke

    2016-01-01

    Notch signaling plays crucial roles for cellular differentiation during development through γ-secretase-dependent intramembrane proteolysis followed by transcription of target genes. Although recent studies implicate that Notch regulates synaptic plasticity or cognitive performance, the molecular mechanism how Notch works in mature neurons remains uncertain. Here we demonstrate that a novel Notch signaling is involved in expression of synaptic proteins in postmitotic neurons. Levels of several synaptic vesicle proteins including synaptophysin 1 and VGLUT1 were increased when neurons were cocultured with Notch ligands-expressing NIH3T3 cells. Neuron-specific deletion of Notch genes decreased these proteins, suggesting that Notch signaling maintains the expression of synaptic vesicle proteins in a cell-autonomous manner. Unexpectedly, cGMP-dependent protein kinase (PKG) inhibitor, but not γ-secretase inhibitor, abolished the elevation of synaptic vesicle proteins, suggesting that generation of Notch intracellular domain is dispensable for this function. These data uncover a ligand-dependent, but γ-secretase-independent, non-canonical Notch signaling involved in presynaptic protein expression in postmitotic neurons. PMID:27040987

  12. Epithelial Notch signaling regulates lung alveolar morphogenesis and airway epithelial integrity

    PubMed Central

    Tsao, Po-Nien; Matsuoka, Chisa; Wei, Shu-Chen; Sato, Atsuyasu; Sato, Susumu; Hasegawa, Koichi; Chen, Hung-kuan; Ling, Thai-Yen; Mori, Munemasa; Cardoso, Wellington V.; Morimoto, Mitsuru

    2016-01-01

    Abnormal enlargement of the alveolar spaces is a hallmark of conditions such as chronic obstructive pulmonary disease and bronchopulmonary dysplasia. Notch signaling is crucial for differentiation and regeneration and repair of the airway epithelium. However, how Notch influences the alveolar compartment and integrates this process with airway development remains little understood. Here we report a prominent role of Notch signaling in the epithelial–mesenchymal interactions that lead to alveolar formation in the developing lung. We found that alveolar type II cells are major sites of Notch2 activation and show by Notch2-specific epithelial deletion (Notch2cNull) a unique contribution of this receptor to alveologenesis. Epithelial Notch2 was required for type II cell induction of the PDGF-A ligand and subsequent paracrine activation of PDGF receptor-α signaling in alveolar myofibroblast progenitors. Moreover, Notch2 was crucial in maintaining the integrity of the epithelial and smooth muscle layers of the distal conducting airways. Our data suggest that epithelial Notch signaling regulates multiple aspects of postnatal development in the distal lung and may represent a potential target for intervention in pulmonary diseases. PMID:27364009

  13. Interferon regulatory factor 4 attenuates Notch signaling to suppress the development of chronic lymphocytic leukemia.

    PubMed

    Shukla, Vipul; Shukla, Ashima; Joshi, Shantaram S; Lu, Runqing

    2016-07-05

    Molecular pathogenesis of Chronic Lymphocytic Leukemia (CLL) is not fully elucidated. Genome wide association studies have linked Interferon Regulatory Factor 4 (IRF4) to the development of CLL. We recently established a causal relationship between low levels of IRF4 and development of CLL. However, the molecular mechanism through which IRF4 suppresses CLL development remains unclear. Deregulation of Notch signaling pathway has been identified as one of the most recurrent molecular anomalies in the pathogenesis of CLL. Yet, the role of Notch signaling as well as its regulation during CLL development remains poorly understood. Previously, we demonstrated that IRF4 deficient mice expressing immunoglobulin heavy chain Vh11 (IRF4-/-Vh11) developed spontaneous CLL with complete penetrance. In this study, we show that elevated Notch2 expression and the resulting hyperactivation of Notch signaling are common features of IRF4-/-Vh11 CLL cells. Our studies further reveal that Notch signaling is indispensable for CLL development in the IRF4-/-Vh11 mice. Moreover, we identify E3 ubiquitin ligase Nedd4, which targets Notch for degradation, as a direct target of IRF4 in CLL cells and their precursors. Collectively, our studies provide the first in vivo evidence for an essential role of Notch signaling in the development of CLL and establish IRF4 as a critical regulator of Notch signaling during CLL development.

  14. Fringe proteins modulate Notch-ligand cis and trans interactions to specify signaling states

    PubMed Central

    LeBon, Lauren; Lee, Tom V; Sprinzak, David; Jafar-Nejad, Hamed; Elowitz, Michael B

    2014-01-01

    The Notch signaling pathway consists of multiple types of receptors and ligands, whose interactions can be tuned by Fringe glycosyltransferases. A major challenge is to determine how these components control the specificity and directionality of Notch signaling in developmental contexts. Here, we analyzed same-cell (cis) Notch-ligand interactions for Notch1, Dll1, and Jag1, and their dependence on Fringe protein expression in mammalian cells. We found that Dll1 and Jag1 can cis-inhibit Notch1, and Fringe proteins modulate these interactions in a way that parallels their effects on trans interactions. Fringe similarly modulated Notch-ligand cis interactions during Drosophila development. Based on these and previously identified interactions, we show how the design of the Notch signaling pathway leads to a restricted repertoire of signaling states that promote heterotypic signaling between distinct cell types, providing insight into the design principles of the Notch signaling system, and the specific developmental process of Drosophila dorsal-ventral boundary formation. DOI: http://dx.doi.org/10.7554/eLife.02950.001 PMID:25255098

  15. Role of CSL-dependent and independent Notch signaling pathways in cell apoptosis.

    PubMed

    Zeng, Chong; Xing, Rui; Liu, Jing; Xing, Feiyue

    2016-01-01

    Apoptosis is a normally biological phenomenon in various organisms, involving complexly molecular mechanisms with a series of signaling processes. Notch signaling is found evolutionarily conserved in many species, playing a critical role in embryonic development, normal tissue homeostasis, angiogenesis and immunoregulation. The focus of this review is on currently novel advances about roles of CSL-dependent and independent Notch signaling pathways in cell apoptosis. The CSL can bind Notch intracellular domain (NIC) to act as a switch in mediating transcriptional activation or inactivation of the Notch signaling pathway downstream genes in the nucleus. It shows that CSL-dependent signaling regulates the cell apoptosis through Hes-1-PTEN-AKT-mTOR signaling, but rather the CSL-independent signaling mediates the cell apoptosis possibly via NIC-mTORC2-AKT-mTOR signaling, providing a new insight into apoptotic mechanisms.

  16. Notch signaling and EMT in non-small cell lung cancer: biological significance and therapeutic application.

    PubMed

    Yuan, Xun; Wu, Hua; Han, Na; Xu, Hanxiao; Chu, Qian; Yu, Shiying; Chen, Yuan; Wu, Kongming

    2014-12-05

    Through epithelial-mesenchymal transition (EMT), cancer cells acquire enhanced ability of migration and invasion, stem cell like characteristics and therapeutic resistance. Notch signaling regulates cell-cell connection, cell polarity and motility during organ development. Recent studies demonstrate that Notch signaling plays an important role in lung cancer initiation and cross-talks with several transcriptional factors to enhance EMT, contributing to the progression of non-small cell lung cancer (NSCLC). Correspondingly, blocking of Notch signaling inhibits NSCLC migration and tumor growth by reversing EMT. Clinical trials have showed promising effect in some cancer patients received treatment with Notch1 inhibitor. This review attempts to provide an overview of the Notch signal in NSCLC: its biological significance and therapeutic application.

  17. Conservation of the Notch1 signaling pathway in gastrointestinal carcinoid cells.

    PubMed

    Kunnimalaiyaan, Muthusamy; Traeger, Kelly; Chen, Herbert

    2005-10-01

    Gastrointestinal (GI) carcinoid cells secrete multiple neuroendocrine (NE) markers and hormones including 5-hydroxytryptamine and chromogranin A. We were interested in determining whether activation of the Notch1 signal transduction pathway in carcinoid cells could modulate production of NE markers and hormones. Human pancreatic carcinoid cells (BON cells) were stably transduced with an estrogen-inducible Notch1 construct, creating BON-NIER cells. In the present study, we found that Notch1 is not detectable in human GI carcinoid tumor cells. The induction of Notch1 in human BON carcinoid cells led to high levels of functional Notch1, as measured by CBF-1 binding studies, resulting in activation of the Notch1 pathway. Similar to its developmental role in the GI tract, Notch1 pathway activation led to an increase in hairy enhancer of split 1 (HES-1) protein and a concomitant silencing of human Notch1/HES-1/achaete-scute homolog 1. Furthermore, Notch1 activation led to a significant reduction in NE markers. Most interestingly, activation of the Notch1 pathway caused a significant reduction in 5-hydroxytryptamine, an important bioactive hormone in carcinoid syndrome. In addition, persistent activation of the Notch1 pathway in BON cells led to a notable reduction in cellular proliferation. These results demonstrate that the Notch1 pathway, which plays a critical role in the differentiation of enteroendocrine cells, is highly conserved in the gut. Therefore, manipulation of the Notch1 signaling pathway may be useful for expanding the targets for therapeutic and palliative treatment of patients with carcinoid tumors.

  18. Notch signaling is required for normal prostatic epithelial cell proliferation and differentiation.

    PubMed

    Wang, Xi-De; Leow, Ching Ching; Zha, Jiping; Tang, Zhijun; Modrusan, Zora; Radtke, Freddy; Aguet, Michel; de Sauvage, Frederic J; Gao, Wei-Qiang

    2006-02-01

    Notch pathway is crucial for stem/progenitor cell maintenance, growth and differentiation in a variety of tissues. Using a transgenic cell ablation approach, we found in our previous study that cells expressing Notch1 are crucial for prostate early development and re-growth. Here, we further define the role of Notch signaling in regulating prostatic epithelial cell growth and differentiation using biochemical and genetic approaches in ex vivo or in vivo systems. Treatment of developing prostate grown in culture with inhibitors of gamma-secretase/presenilin, which is required for Notch cleavage and activation, caused a robust increase in proliferation of epithelial cells co-expressing cytokeratin 8 and 14, lack of luminal/basal layer segregation and dramatically reduced branching morphogenesis. Using conditional Notch1 gene deletion mouse models, we found that inactivation of Notch1 signaling resulted in profound prostatic alterations, including increased tufting, bridging and enhanced epithelial proliferation. Cells within these lesions co-expressed both luminal and basal cell markers, a feature of prostatic epithelial cells in predifferentiation developmental stages. Microarray analysis revealed that the gene expression in a number of genetic networks was altered following Notch1 gene deletion in prostate. Furthermore, expression of Notch1 and its effector Hey-1 gene in human prostate adenocarcinomas were found significantly down-regulated compared to normal control tissues. Taken together, these data suggest that Notch signaling is critical for normal cell proliferation and differentiation in the prostate, and deregulation of this pathway may facilitate prostatic tumorigenesis.

  19. hCLP46 regulates U937 cell proliferation via Notch signaling pathway

    SciTech Connect

    Ma, Wenzhan; Du, Jie; Chu, Qiaoyun; Wang, Youxin; Liu, Lixin; Song, Manshu; Wang, Wei

    2011-04-29

    Highlights: {yields} Knock down of hCLP46 by RNAi impairs mammalian Notch signaling. {yields} hCLP46 affects neither cell surface Notch1 expression nor ligand-receptor binding. {yields} Knock down of hCLP46 inhibits U937 cell-growth by up-regulation of CDKN1B. -- Abstract: Human CAP10-like protein 46 kDa (hCLP46) is the homolog of Rumi, which is the first identified protein O-glucosyltransferase that modifies Notch receptor in Drosophila. Dysregulation of hCLP46 occurs in many hematologic diseases, but the role of hCLP46 remains unclear. Knockdown of hCLP46 by RNA interference resulted in decreased protein levels of endogenous Notch1, Notch intracellular domain (NICD) and Notch target gene Hes-1, suggesting the impairment of the Notch signaling. However, neither cell surface Notch expression nor ligand binding activities were affected. In addition, down-regulated expression of hCLP46 inhibited the proliferation of U937 cells, which was correlated with increased cyclin-dependent kinase inhibitor (CDKI) CDKN1B (p27) and decreased phosphorylation of retinoblastoma (RB) protein. We showed that lack of hCLP46 results in impaired ligand induced Notch activation in mammalian cell, and hCLP46 regulates the proliferation of U937 cell through CDKI-RB signaling pathway, which may be important for the pathogenesis of leukemia.

  20. Metalloprotease-disintegrin ADAM12 expression is regulated by Notch signaling via microRNA-29.

    PubMed

    Li, Hui; Solomon, Emilia; Duhachek Muggy, Sara; Sun, Danqiong; Zolkiewska, Anna

    2011-06-17

    Metalloprotease-disintegrin ADAM12 is overexpressed and frequently mutated in breast cancer. We report here that ADAM12 expression in cultured mammalian cells is up-regulated by Notch signals. Expression of a constitutively active form of Notch1 in murine fibroblasts, myoblasts, or mammary epithelial cells or activation of the endogenous Notch signaling by co-culture with ligand-expressing cells increases ADAM12 protein and mRNA levels. Up-regulation of ADAM12 expression by Notch requires new transcription, is activated in a CSL-dependent manner, and is abolished upon inhibition of IκB kinase. Expression of a constitutively active Notch1 in NIH3T3 cells increases the stability of Adam12 mRNA. We further show that the microRNA-29 family, which has a predicted conserved site in the 3'-untranslated region of mouse Adam12, plays a critical role in mediating the stimulatory effect of Notch on ADAM12 expression. In human cells, Notch up-regulates the expression of the long form, but not the short form, of ADAM12 containing a divergent 3'-untranslated mRNA region. These studies uncover a novel paradigm in Notch signaling and establish Adam12 as a Notch-related gene.

  1. Dicer is Required for the Maintenance of Notch Signaling and Gliogenic Competence During Mouse Retinal Development

    PubMed Central

    Georgi, Sean A.; Reh, Thomas A.

    2017-01-01

    MicroRNAs (miRNAs) are 19–25 nucleotide RNAs that regulate messenger RNA translation and stability. Recently, we performed a conditional knockout (CKO) of the miRNA-processing enzyme Dicer during mouse retinal development and showed an essential role for miRNAs in the transition of retinal progenitors from an early to a late competence state (Georgi and Reh [2010]: J Neurosci 30:4048–4061). Notably, Dicer CKO progenitors failed to express Ascl1 and generated ganglion cells beyond their normal competence window. Because Ascl1 regulates multiple Notch signaling components, we hypothesized that Notch signaling is downregulated in Dicer CKO retinas. We show here that Notch signaling is severely reduced in Dicer CKO retinas, but that retinal progenitors still retain a low level of Notch signaling. By increasing Notch signaling in Dicer CKO progenitors through constitutive expression of the Notch intra-cellular domain (NICD), we show that transgenic rescue of Notch signaling has little effect on the competence of retinal progenitors or the enhanced generation of ganglion cells, suggesting that loss of Notch signaling is not a major determinant of these phenotypes. Nevertheless, transgenic NICD expression restored horizontal cells, suggesting an interaction between miRNAs and Notch signaling in the development of this cell type. Furthermore, while NICD overexpression leads to robust glial induction in control retinas, NICD overexpression was insufficient to drive Dicer-null retinal progenitors to a glial fate. Surprisingly, the presence of transgenic NICD expression did not prevent the differentiation of some types of retinal neurons, suggesting that Notch inactivation is not an absolute requirement for the initial stages of neuronal differentiation. PMID:21542136

  2. Effects of notch signaling on regulation of myeloid cell differentiation in cancer.

    PubMed

    Cheng, Pingyan; Kumar, Vinit; Liu, Hao; Youn, Je-In; Fishman, Mayer; Sherman, Simon; Gabrilovich, Dmitry

    2014-01-01

    Functionally altered myeloid cells play an important role in immune suppression in cancer, in angiogenesis, and in tumor cells' invasion and metastases. Here, we report that inhibition of Notch signaling in hematopoietic progenitor cells (HPC), myeloid-derived suppressor cells (MDSC), and dendritic cells is directly involved in abnormal myeloid cell differentiation in cancer. Inhibition of Notch signaling was caused by the disruption of the interaction between Notch receptor and transcriptional repressor CSL, which is normally required for efficient transcription of target genes. This disruption was the result of serine phosphorylation of Notch. We demonstrated that increased activity of casein kinase 2 (CK2) observed in HPC and in MDSC could be responsible for the phosphorylation of Notch and downregulation of Notch signaling. Inhibition of CK2 by siRNA or by pharmacological inhibitor restored Notch signaling in myeloid cells and substantially improved their differentiation, both in vitro and in vivo. This study demonstrates a novel mechanism regulation of Notch signaling in cancer. This may suggest a new perspective for pharmacological regulation of differentiation of myeloid cells in cancer.

  3. Evidence of non-canonical NOTCH signaling: Delta-like 1 homolog (DLK1) directly interacts with the NOTCH1 receptor in mammals.

    PubMed

    Traustadóttir, Gunnhildur Ásta; Jensen, Charlotte H; Thomassen, Mads; Beck, Hans Christian; Mortensen, Sussi B; Laborda, Jorge; Baladrón, Victoriano; Sheikh, Søren P; Andersen, Ditte C

    2016-04-01

    Canonical NOTCH signaling, known to be essential for tissue development, requires the Delta-Serrate-LAG2 (DSL) domain for NOTCH to interact with its ligand. However, despite lacking DSL, Delta-like 1 homolog (DLK1), a protein that plays a significant role in mammalian development, has been suggested to interact with NOTCH1 and act as an antagonist. This non-canonical interaction is, however controversial, and evidence for a direct interaction, still lacking in mammals. In this study, we elucidated the putative DLK1-NOTCH1 interaction in a mammalian context. Taking a global approach and using Dlk1(+/+) and Dlk1(-/-) mouse tissues at E16.5, we demonstrated that several NOTCH signaling pathways indeed are affected by DLK1 during tissue development, and this was supported by a lower activation of NOTCH1 protein in Dlk1(+/+) embryos. Likewise, but using a distinct Dlk1-manipulated (siRNA) setup in a mammalian cell line, NOTCH signaling was substantially inhibited by DLK1. Using a mammalian two-hybrid system, we firmly established that the effect of DLK1 on NOTCH signaling was due to a direct interaction between DLK1 and NOTCH1. By careful dissection of this mechanism, we found this interaction to occur between EGF domains 5 and 6 of DLK1 and EGF domains 10-15 of NOTCH1. Thus, our data provide the first evidence for a direct interaction between DLK1 and NOTCH1 in mammals, and substantiate that non-canonical NOTCH ligands exist, adding to the complexity of NOTCH signaling.

  4. The Notch-2 Gene Is Regulated by Wnt Signaling in Cultured Colorectal Cancer Cells

    PubMed Central

    Ungerbäck, Jonas; Elander, Nils; Grünberg, John; Sigvardsson, Mikael; Söderkvist, Peter

    2011-01-01

    Background Notch and Wnt pathways are key regulators of intestinal homeostasis and alterations in these pathways may lead to the development of colorectal cancer (CRC). In CRC the Apc/β-catenin genes in the Wnt signaling pathway are frequently mutated and active Notch signaling contributes to tumorigenesis by keeping the epithelial cells in a proliferative state. These pathways are simultaneously active in proliferative adenoma cells and a crosstalk between them has previously been suggested in normal development as well as in cancer. Principal Findings In this study, in silico analysis of putative promoters involved in transcriptional regulation of genes coding for proteins in the Notch signaling pathway revealed several putative LEF-1/TCF sites as potential targets for β-catenin and canonical Wnt signaling. Further results from competitive electrophoretic mobility-shift assay (EMSA) studies suggest binding of several putative sites in Notch pathway gene promoters to in vitro translated β-catenin/Lef-1. Wild type (wt)-Apc negatively regulates β-catenin. By induction of wt-Apc or β-catenin silencing in HT29 cells, we observed that several genes in the Notch pathway, including Notch-2, were downregulated. Finally, active Notch signaling was verified in the ApcMin/+ mouse model where Hes-1 mRNA levels were found significantly upregulated in intestinal tumors compared to normal intestinal mucosa. Luciferase assays showed an increased activity for the core and proximal Notch-2 promoter upon co-transfection of HCT116 cells with high expression recombinant Tcf-4, Lef-1 or β-catenin. Conclusions In this paper, we identified Notch-2 as a novel target for β-catenin-dependent Wnt signaling. Furthermore our data supports the notion that additional genes in the Notch pathway might be transcriptionally regulated by Wnt signaling in colorectal cancer. PMID:21437251

  5. Notch signaling: targeting cancer stem cells and epithelial-to-mesenchymal transition.

    PubMed

    Espinoza, Ingrid; Pochampally, Radhika; Xing, Fei; Watabe, Kounosuke; Miele, Lucio

    2013-09-06

    Notch signaling is an evolutionarily conserved pathway involved in cell fate control during development, stem cell self-renewal, and postnatal tissue differentiation. Roles for Notch in carcinogenesis, the biology of cancer stem cells, tumor angiogenesis, and epithelial-to-mesenchymal transition (EMT) have been reported. This review describes the role of Notch in the "stemness" program in cancer cells and in metastases, together with a brief update on the Notch inhibitors currently under investigation in oncology. These agents may be useful in targeting cancer stem cells and to reverse the EMT process.

  6. Notch signaling: targeting cancer stem cells and epithelial-to-mesenchymal transition

    PubMed Central

    Espinoza, Ingrid; Pochampally, Radhika; Xing, Fei; Watabe, Kounosuke; Miele, Lucio

    2013-01-01

    Notch signaling is an evolutionarily conserved pathway involved in cell fate control during development, stem cell self-renewal, and postnatal tissue differentiation. Roles for Notch in carcinogenesis, the biology of cancer stem cells, tumor angiogenesis, and epithelial-to-mesenchymal transition (EMT) have been reported. This review describes the role of Notch in the “stemness” program in cancer cells and in metastases, together with a brief update on the Notch inhibitors currently under investigation in oncology. These agents may be useful in targeting cancer stem cells and to reverse the EMT process. PMID:24043949

  7. The V-ATPase a2 isoform controls mammary gland development through Notch and TGF-β signaling

    PubMed Central

    Pamarthy, Sahithi; Mao, Liquin; Katara, Gajendra K; Fleetwood, Sara; Kulshreshta, Arpita; Gilman-Sachs, Alice; Beaman, Kenneth D

    2016-01-01

    Among all tissues and organs, the mammary gland is unique because most of its development occurs in adulthood. Notch signaling has a major role in mammary gland development and has been implicated in breast cancer. The vacuolar-ATPase (V-ATPase) is a proton pump responsible for the regulation and control of pH in intracellular vesicles and the extracellular milieu. We have previously reported that a2V-ATPase (a2V), an isoform of ‘a' subunit of V-ATPase, regulates processing of Notch receptor and alters Notch signaling in breast cancer. To study the role of a2V in mammary gland development, we generated an a2V-KO model (conditional mammary knockout a2V mouse strain). During normal mammary gland development, the basal level expression of a2V increased from puberty, virginity, and pregnancy through the lactation stage and then decreased during involution. Litters of a2V-KO mice weighed significantly less when compared with litters from wild-type mice and showed reduced expression of the lactation marker β-casein. Whole-mount analysis of mammary glands demonstrated impaired ductal elongation and bifurcation in a2V-KO mice. Consequently, we found disintegrated mammary epithelium as seen by basal and luminal epithelial staining, although the rate of proliferation remained unchanged. Delayed mammary morphogenesis in a2V-KO mice was associated with aberrant activation of Notch and TGF-β (transforming growth factor-β) pathways. Notably, Hey1 (hairy/enhancer-of-split related with YRPW motif) and Smad2, the key downstream mediators of Notch and TGF-β pathways, respectively, were upregulated in a2V-KO mice and also in human mammary epithelial cells treated with a2V siRNA. Taken together, our results show that a2V deficiency disrupts the endolysosomal route in Notch and TGF signaling, thereby impairing mammary gland development. Our findings have broader implications in developmental and oncogenic cellular environments where V-ATPase, Notch and TGF-β are crucial for cell

  8. The functional role of Notch signaling in triple-negative breast cancer.

    PubMed

    Speiser, Jodi J; Erşahin, Cağatay; Osipo, Clodia

    2013-01-01

    The term "triple-negative breast cancer" (TNBC) is a heterogeneous subtype of breast cancer. Unfortunately, due to the lack of expression of hormone receptors and human epidermal growth factor receptor-2, therefore the lack of US Food and Drug Administration-approved targeted therapies, TNBC has the worst prognosis of all subtypes of breast cancer. Notch signaling has emerged as a pro-oncogene in several human malignancies and has particularly been associated with the triple-negative subtype of breast cancer. This chapter explores the role of Notch signaling in triple negative and other subtypes of breast cancer, the relationship of Notch with other breast cancer biomarkers, prognostic indicators associated with Notch, and potential therapeutic strategies targeting Notch inhibition. Hopefully, better understanding of this signaling pathway in the future will lead to optimal molecular therapeutic treatments for TNBC patients, improving their quality of life and outcome.

  9. [Novel signal transduction pathways: the molecular basis for targeted cancer therapies in Hedgehog/Notch/Wnt pathway].

    PubMed

    Shimizu, Toshio; Nakagawa, Kazuhiko

    2015-08-01

    Aberrant activation of the Wnt, Notch and Hedgehog pathways via mutations or ligand overexpression has been implicated in a large number of cancer types where they are involved in functions ranging from tumor initiation to cancer stem cell (CSC) maintenance and angiogenesis. Agents targeting each one of these three pathways have now reached clinical trials, and the first one of these, Vismodegib, a hedgehog pathway inhibitor, was approved in 2012 by US FDA for the treatment of advanced basal cell carcinoma. Development of agents that target critical steps in these pathways as novel signal transduction pathways will be complicated by signaling cross-talk. The role that embryonic signaling pathways play in the function of CSCs, the development of new anti-CSC therapeutic agents, and the complexity of potential CSC signaling cross-talk are being explored coupled with early phase I clinical studies.

  10. Phosphorylation of Notch1 by Pim kinases promotes oncogenic signaling in breast and prostate cancer cells

    PubMed Central

    Vahtera, Laura; Ylä-Pelto, Jani; Paloniemi, Elina; Imanishi, Susumu Y.; Corthals, Garry; Varjosalo, Markku; Manoharan, Ganesh Babu; Uri, Asko; Lendahl, Urban; Sahlgren, Cecilia; Koskinen, Päivi J.

    2016-01-01

    Tumorigenesis is a multistep process involving co-operation between several deregulated oncoproteins. In this study, we unravel previously unrecognized interactions and crosstalk between Pim kinases and the Notch signaling pathway, with implications for both breast and prostate cancer. We identify Notch1 and Notch3, but not Notch2, as novel Pim substrates and demonstrate that for Notch1, the serine residue 2152 is phosphorylated by all three Pim family kinases. This target site is located in the second nuclear localization sequence (NLS) of the Notch1 intracellular domain (N1ICD), and is shown to be important for both nuclear localization and transcriptional activity of N1ICD. Phosphorylation-dependent stimulation of Notch1 signaling promotes migration of prostate cancer cells, balances glucose metabolism in breast cancer cells, and supports in vivo growth of both types of cancer cells on chick embryo chorioallantoic membranes. Furthermore, Pim-induced growth of orthotopic prostate xenografts in mice is associated with enhanced nuclear Notch1 activity. Finally, simultaneous inhibition of Pim and Notch abrogates the cellular responses more efficiently than individual treatments, opening up new vistas for combinatorial cancer therapy. PMID:27281612

  11. Dll1- and Dll4-mediated Notch signaling is required for homeostasis of intestinal stem cells

    PubMed Central

    Pellegrinet, Luca; Rodilla, Veronica; Liu, Zhenyi; Chen, Shuang; Koch, Ute; Espinosa, Lluis; Kaestner, Klaus H.; Kopan, Raphael; Lewis, Julian; Radtke, Freddy

    2011-01-01

    Background & Aims Ablation of Notch signaling within the intestinal epithelium results in loss of proliferating crypt progenitors, due to their conversion into post-mitotic secretory cells. We aimed to confirm that Notch was active in stem cells (SC), investigate consequences of loss of Notch signaling within the intestinal SC compartment, and identify the physiological ligands of Notch in mouse intestine. Furthermore, we investigated whether the induction of goblet cell differentiation that results from loss of Notch requires the transcription factor Krüppel-like factor 4 (Klf4). Methods Trasgenic mice that carried a reporter of Notch1 activation were used for lineage tracing experiments. The in vivo functions of the Notch ligands Jagged1 (Jag1), Delta-like1 (Dll1), Delta-like4 (Dll4), and the transcription factor Klf4 were assessed in mice with inducible, gut-specific gene targeting (Vil-Cre-ERT2). Results Notch1 signaling was found to be activated in intestinal SC. Although deletion of Jag1 or Dll4 did not perturb the intestinal epithelium, inactivation of Dll1 resulted in a moderate increase in number of goblet cells without noticeable effects of progenitor proliferation. However, simultaneous inactivation of Dll1 and Dll4 resulted in the complete conversion of proliferating progenitors into post-mitotic goblet cells, concomitant with loss of SC (Olfm4+, Lgr5+ and Ascl2+). Klf4 inactivation did not interfere with goblet cell differentiation in adult wild-type or in Notch pathway-deficient gut. Conclusions Notch signaling in SC and progenitors is activated by Dll1 and Dll4 ligands and is required for maintenance of intestinal progenitor and SC. Klf4 is dispensable for goblet cell differentiation in intestines of adult Notch-deficient mice. PMID:21238454

  12. ASIC1 promotes differentiation of neuroblastoma by negatively regulating Notch signaling pathway.

    PubMed

    Liu, Mingli; Inoue, Koichi; Leng, Tiandong; Zhou, An; Guo, Shanchun; Xiong, Zhi-Gang

    2017-01-31

    In neurons, up-regulation of Notch activity either inhibits neurite extension or causes retraction of neurites. Conversely, inhibition of Notch1 facilitates neurite extension. Acid-sensing ion channels (ASICs) are a family of proton-gated cation channels, which play critical roles in synaptic plasticity, learning and memory and spine morphogenesis. Our pilot proteomics data from ASIC1a knock out mice implicated that ASIC1a may play a role in regulating Notch signaling, therefore, we explored whether or not ASIC1a regulates neurite growth during neuronal development through Notch signaling. In this study, we determined the effects of ASIC1a on neurite growth in a mouse neuroblastoma cell line, NS20Y cells, by modulating ASIC1a expression. We also determined the relationship between ASIC1a and Notch signaling on neuronal differentiation. Our results showed that down-regulation of ASIC1a in NS20Y cells inhibits CPT-cAMP induced neurite growth, while over expression of ASIC1a promotes its growth. In addition, down-regulation of ASIC1a increased the expression of Notch1 and its target gene Survivin while inhibitor of Notch significantly prevented the neurite extension induced by ASIC1a in NS20Y cells. These data indicate that Notch1 signaling may be required for ASIC1a-mediated neurite growth and neuronal differentiation.

  13. Sanpodo: a context-dependent activator and inhibitor of Notch signaling during asymmetric divisions

    PubMed Central

    Babaoglan, A. Burcu; O'Connor-Giles, Kate M.; Mistry, Hemlata; Schickedanz, Adam; Wilson, Beth A.; Skeath, James B.

    2009-01-01

    Asymmetric cell divisions generate sibling cells of distinct fates (‘A’, ‘B’) and constitute a fundamental mechanism that creates cell-type diversity in multicellular organisms. Antagonistic interactions between the Notch pathway and the intrinsic cell-fate determinant Numb appear to regulate asymmetric divisions in flies and vertebrates. During these divisions, productive Notch signaling requires sanpodo, which encodes a novel transmembrane protein. Here, we demonstrate that Drosophila sanpodo plays a dual role to regulate Notch signaling during asymmetric divisions — amplifying Notch signaling in the absence of Numb in the ‘A’ daughter cell and inhibiting Notch signaling in the presence of Numb in the ‘B’ daughter cell. In so doing, sanpodo ensures the asymmetry in Notch signaling levels necessary for the acquisition of distinct fates by the two daughter cells. These findings answer long-standing questions about the restricted ability of Numb and Sanpodo to inhibit and to promote, respectively, Notch signaling during asymmetric divisions. PMID:19906847

  14. Pleiotropic roles of Notch signaling in normal, malignant, and developmental hematopoiesis in the human

    PubMed Central

    Kushwah, Rahul; Guezguez, Borhane; Lee, Jung Bok; Hopkins, Claudia I; Bhatia, Mickie

    2014-01-01

    The Notch signaling pathway is evolutionarily conserved across species and plays an important role in regulating cell differentiation, proliferation, and survival. It has been implicated in several different hematopoietic processes including early hematopoietic development as well as adult hematological malignancies in humans. This review focuses on recent developments in understanding the role of Notch signaling in the human hematopoietic system with an emphasis on hematopoietic initiation from human pluripotent stem cells and regulation within the bone marrow. Based on recent insights, we summarize potential strategies for treatment of human hematological malignancies toward the concept of targeting Notch signaling for fate regulation. PMID:25252682

  15. Regulation of Notch Signaling by an Evolutionary Conserved DEAD Box RNA Helicase, Maheshvara in Drosophila melanogaster.

    PubMed

    Surabhi, Satya; Tripathi, Bipin K; Maurya, Bhawana; Bhaskar, Pradeep K; Mukherjee, Ashim; Mutsuddi, Mousumi

    2015-11-01

    Notch signaling is an evolutionary conserved process that influences cell fate determination, cell proliferation, and cell death in a context-dependent manner. Notch signaling is fine-tuned at multiple levels and misregulation of Notch has been implicated in a variety of human diseases. We have characterized maheshvara (mahe), a novel gene in Drosophila melanogaster that encodes a putative DEAD box protein that is highly conserved across taxa and belongs to the largest group of RNA helicase. A dynamic pattern of mahe expression along with the maternal accumulation of its transcripts is seen during early stages of embryogenesis. In addition, a strong expression is also seen in the developing nervous system. Ectopic expression of mahe in a wide range of tissues during development results in a variety of defects, many of which resemble a typical Notch loss-of-function phenotype. We illustrate that ectopic expression of mahe in the wing imaginal discs leads to loss of Notch targets, Cut and Wingless. Interestingly, Notch protein levels are also lowered, whereas no obvious change is seen in the levels of Notch transcripts. In addition, mahe overexpression can significantly rescue ectopic Notch-mediated proliferation of eye tissue. Further, we illustrate that mahe genetically interacts with Notch and its cytoplasmic regulator deltex in trans-heterozygous combination. Coexpression of Deltex and Mahe at the dorso-ventral boundary results in a wing-nicking phenotype and a more pronounced loss of Notch target Cut. Taken together we report identification of a novel evolutionary conserved RNA helicase mahe, which plays a vital role in regulation of Notch signaling.

  16. Cisplatin selects for stem-like cells in osteosarcoma by activating Notch signaling.

    PubMed

    Yu, Ling; Fan, Zhengfu; Fang, Shuo; Yang, Jian; Gao, Tian; Simões, Bruno M; Eyre, Rachel; Guo, Weichun; Clarke, Robert B

    2016-05-31

    Notch signaling regulates normal stem cells and is also thought to regulate cancer stem cells (CSCs). Recent data indicate that Notch signaling plays a role in the development and progression of osteosarcoma, however the regulation of Notch in chemo-resistant stem-like cells has not yet been fully elucidated. In this study we generated cisplatin-resistant osteosarcoma cells by treating them with sub-lethal dose of cisplatin, sufficient to induce DNA damage responses. Cisplatin-resistant osteosarcoma cells exhibited lower proliferation, enhanced spheroid formation and more mesenchymal characteristics than cisplatin-sensitive cells, were enriched for Stro-1+/CD117+ cells and showed increased expression of stem cell-related genes. A similar effect was observed in vivo, and in addition in vivo tumorigenicity was enhanced during serial transplantation. Using several publicly available datasets, we identified that Notch expression was closely associated with osteosarcoma stem cells and chemotherapy resistance. We confirmed that cisplatin-induced enrichment of osteosarcoma stem cells was mediated through Notch signaling in vitro, and immunohistochemistry showed that cleaved Notch1 (NICD1) positive cells were significantly increased in a relapsed xenograft which had received cisplatin treatment. Furthermore, pretreatment with a γ-secretase inhibitor (GSI) to prevent Notch signalling inhibited cisplatin-enriched osteosarcoma stem cell activity in vitro, including Stro-1+/CD117+ double positive cells and spheroid formation capacity. The Notch inhibitor DAPT also prevented tumor recurrence in resistant xenograft tumors. Overall, our results show that cisplatin induces the enrichment of osteosarcoma stem-like cells through Notch signaling, and targeted inactivation of Notch may be useful for the elimination of CSCs and overcoming drug resistance.

  17. Lunatic Fringe-mediated Notch signaling is required for lung alveogenesis.

    PubMed

    Xu, Keli; Nieuwenhuis, Erica; Cohen, Brenda L; Wang, Wei; Canty, Angelo J; Danska, Jayne S; Coultas, Leigh; Rossant, Janet; Wu, Megan Y J; Piscione, Tino D; Nagy, Andras; Gossler, Achim; Hicks, Geoff G; Hui, Chi-Chung; Henkelman, R Mark; Yu, Lisa X; Sled, John G; Gridley, Thomas; Egan, Sean E

    2010-01-01

    Distal lung development occurs through coordinated induction of myofibroblasts, epithelial cells, and capillaries. Lunatic Fringe (Lfng) is a beta(1-3) N-acetylglucosamine transferase that modifies Notch receptors to facilitate their activation by Delta-like (Dll1/4) ligands. Lfng is expressed in the distal lung during saccular development, and deletion of this gene impairs myofibroblast differentiation and alveogenesis in this context. A similar defect was observed in Notch2(beta-geo/+)Notch3(beta-geo/beta-geo) compound mutant mice but not in Notch2(beta-geo/+) or Notch3(beta-geo/beta-geo) single mutants. Finally, to directly test for the role of Notch signaling in myofibroblast differentiation in vivo, we used ROSA26-rtTA(/+);tetO-CRE(/+);RBPJkappa(flox/flox) inducible mutant mice to show that disruption of canonical Notch signaling during late embryonic development prevents induction of smooth muscle actin in mesenchymal cells of the distal lung. In sum, these results demonstrate that Lfng functions to enhance Notch signaling in myofibroblast precursor cells and thereby to coordinate differentiation and mobilization of myofibroblasts required for alveolar septation.

  18. Notch1 functions as a tumor suppressor in mouse skin.

    PubMed

    Nicolas, Michael; Wolfer, Anita; Raj, Kenneth; Kummer, J Alain; Mill, Pleasantine; van Noort, Mascha; Hui, Chi-chung; Clevers, Hans; Dotto, G Paolo; Radtke, Freddy

    2003-03-01

    Notch proteins are important in binary cell-fate decisions and inhibiting differentiation in many developmental systems, and aberrant Notch signaling is associated with tumorigenesis. The role of Notch signaling in mammalian skin is less well characterized and is mainly based on in vitro studies, which suggest that Notch signaling induces differentiation in mammalian skin. Conventional gene targeting is not applicable to establishing the role of Notch receptors or ligands in the skin because Notch1-/- embryos die during gestation. Therefore, we used a tissue-specific inducible gene-targeting approach to study the physiological role of the Notch1 receptor in the mouse epidermis and the corneal epithelium of adult mice. Unexpectedly, ablation of Notch1 results in epidermal and corneal hyperplasia followed by the development of skin tumors and facilitated chemical-induced skin carcinogenesis. Notch1 deficiency in skin and in primary keratinocytes results in increased and sustained expression of Gli2, causing the development of basal-cell carcinoma-like tumors. Furthermore, Notch1 inactivation in the epidermis results in derepressed beta-catenin signaling in cells that should normally undergo differentiation. Enhanced beta-catenin signaling can be reversed by re-introduction of a dominant active form of the Notch1 receptor. This leads to a reduction in the signaling-competent pool of beta-catenin, indicating that Notch1 can inhibit beta-catenin-mediated signaling. Our results indicate that Notch1 functions as a tumor-suppressor gene in mammalian skin.

  19. Regulation of Notch signaling and endocytosis by the Lgl neoplastic tumor suppressor

    PubMed Central

    Portela, Marta; Parsons, Linda M; Grzeschik, Nicola A; Richardson, Helena E

    2015-01-01

    The evolutionarily conserved neoplastic tumor suppressor protein, Lethal (2) giant larvae (Lgl), plays roles in cell polarity and tissue growth via regulation of the Hippo pathway. In our recent study, we showed that in the developing Drosophila eye epithelium, depletion of Lgl leads to increased ligand-dependent Notch signaling. lgl mutant tissue also exhibits an accumulation of early endosomes, recycling endosomes, early-multivesicular body markers and acidic vesicles. We showed that elevated Notch signaling in lgl− tissue can be rescued by feeding larvae the vesicle de-acidifying drug chloroquine, revealing that Lgl attenuates Notch signaling by limiting vesicle acidification. Strikingly, chloroquine also rescued the lgl− overgrowth phenotype, suggesting that the Hippo pathway defects were also rescued. In this extraview, we provide additional data on the regulation of Notch signaling and endocytosis by Lgl, and discuss possible mechanisms by which Lgl depletion contributes to signaling pathway defects and tumorigenesis. PMID:25789785

  20. An Overview of Notch Signaling in Adult Tissue Renewal and Maintenance

    PubMed Central

    Sato, Chihiro; Zhao, Guojun; Ilagan, Ma. Xenia G.

    2015-01-01

    The Notch pathway is a critical mediator of short-range cell-cell communication that is reiteratively used to regulate a diverse array of cellular processes during embryonic development and the renewal and maintenance of adult tissues. Most Notch-dependent processes utilize a core signaling mechanism that is dependent on regulated intramembrane proteolysis: Upon ligand binding, Notch receptors undergo ectodomain shedding by ADAM metalloproteases, followed by γ-secretase-mediated intramembrane proteolysis. This releases the Notch intracellular domain, which translocates to the nucleus to activate transcription. In this review, we highlight the roles of Notch signaling particularly in self-renewing tissues in adults and several human diseases and raise some key considerations when targeting ADAMs and γ-secretase as disease-modifying strategies for Alzheimer's Disease. PMID:21605032

  1. Regulation of Notch1 signaling by the APP intracellular domain facilitates degradation of the Notch1 intracellular domain and RBP-Jk.

    PubMed

    Kim, Mi-Yeon; Mo, Jung-Soon; Ann, Eun-Jung; Yoon, Ji-Hye; Jung, Jane; Choi, Yun-Hee; Kim, Su-Man; Kim, Hwa-Young; Ahn, Ji-Seon; Kim, Hangun; Kim, Kwonseop; Hoe, Hyang-Sook; Park, Hee-Sae

    2011-06-01

    The Notch1 receptor is a crucial controller of cell fate decisions, and is also a key regulator of cell growth and differentiation in a variety of contexts. In this study, we have demonstrated that the APP intracellular domain (AICD) attenuates Notch1 signaling by accelerated degradation of the Notch1 intracellular domain (Notch1-IC) and RBP-Jk, through different degradation pathways. AICD suppresses Notch1 transcriptional activity by the dissociation of the Notch1-IC-RBP-Jk complex after processing by γ-secretase. Notch1-IC is capable of forming a trimeric complex with Fbw7 and AICD, and AICD enhances the protein degradation of Notch1-IC through an Fbw7-dependent proteasomal pathway. AICD downregulates the levels of RBP-Jk protein through the lysosomal pathway. AICD-mediated degradation is involved in the preferential degradation of non-phosphorylated RBP-Jk. Collectively, our results demonstrate that AICD functions as a negative regulator in Notch1 signaling through the promotion of Notch1-IC and RBP-Jk protein degradation.

  2. Hippo signaling is required for Notch-dependent smooth muscle differentiation of neural crest

    PubMed Central

    Manderfield, Lauren J.; Aghajanian, Haig; Engleka, Kurt A.; Lim, Lillian Y.; Liu, Feiyan; Jain, Rajan; Li, Li; Olson, Eric N.; Epstein, Jonathan A.

    2015-01-01

    Notch signaling has well-defined roles in the assembly of arterial walls and in the development of the endothelium and smooth muscle of the vasculature. Hippo signaling regulates cellular growth in many tissues, and contributes to regulation of organ size, in addition to other functions. Here, we show that the Notch and Hippo pathways converge to regulate smooth muscle differentiation of the neural crest, which is crucial for normal development of the aortic arch arteries and cranial vasculature during embryonic development. Neural crest-specific deletion of the Hippo effectors Yap and Taz produces neural crest precursors that migrate normally, but fail to produce vascular smooth muscle, and Notch target genes such as Jagged1 fail to activate normally. We show that Yap is normally recruited to a tissue-specific Jagged1 enhancer by directly interacting with the Notch intracellular domain (NICD). The Yap-NICD complex is recruited to chromatin by the DNA-binding protein Rbp-J in a Tead-independent fashion. Thus, Hippo signaling can modulate Notch signaling outputs, and components of the Hippo and Notch pathways physically interact. Convergence of Hippo and Notch pathways by the mechanisms described here might be relevant for the function of these signaling cascades in many tissues and in diseases such as cancer. PMID:26253400

  3. Novel transcript nort is a downstream target gene of the Notch signaling pathway in zebrafish.

    PubMed

    Tsutsumi, Makiko; Itoh, Motoyuki

    2007-01-01

    The Notch signaling pathway plays important roles in the regulation of diverse developmental processes. Although many Notch-signal target genes with different specificities have been identified, their regulation and functions are not fully understood. Here, we conducted a microarray screen to search for novel downstream target genes of the Notch pathway in zebrafish. From the screen, we isolated nort (Notch-regulated transcript) as a transcript whose expression was reduced by the inhibition of Notch signaling. The expression level of nort increased when Notch signaling was activated. nort was expressed in hypoblast cells and the developing nervous system. We found its expression pattern to be similar to that of her4, but it showed some differences, at least in the anterior and posterior neural plate at the 3-somite stage. The nort transcript did not contain any long open-reading frame (ORF) of more than 300 nt, and its ORF-encoded sequence showed no significant homology with the proteins in databases. However, nort has one SPS (suppressor of hairless paired binding site) in its 5'-flanking region. These data suggest that nort is a putative noncoding RNA regulated by Notch signaling.

  4. Dual tumor suppressing and promoting function of Notch1 signaling in human prostate cancer.

    PubMed

    Lefort, Karine; Ostano, Paola; Mello-Grand, Maurizia; Calpini, Valérie; Scatolini, Maria; Farsetti, Antonella; Dotto, G Paolo; Chiorino, Giovanna

    2016-07-26

    Adenocarcinomas of the prostate arise as multifocal heterogeneous lesions as the likely result of genetic and epigenetic alterations and deranged cell-cell communication. Notch signaling is an important form of intercellular communication with a role in growth/differentiation control and tumorigenesis. Contrasting reports exist in the literature on the role of this pathway in prostate cancer (PCa) development. We show here that i) compared to normal prostate tissue, Notch1 expression is significantly reduced in a substantial fraction of human PCas while it is unaffected or even increased in others; ii) acute Notch activation both inhibits and induces process networks associated with prostatic neoplasms; iii) down-modulation of Notch1 expression and activity in immortalized normal prostate epithelial cells increases their proliferation potential, while increased Notch1 activity in PCa cells suppresses growth and tumorigenicity through a Smad3-dependent mechanism involving p21WAF1/CIP1; iv) prostate cancer cells resistant to Notch growth inhibitory effects retain Notch1-induced upregulation of pro-oncogenic genes, like EPAS1 and CXCL6, also overexpressed in human PCas with high Notch1 levels. Taken together, these results reconcile conflicting data on the role of Notch1 in prostate cancer.

  5. Dual tumor suppressing and promoting function of Notch1 signaling in human prostate cancer

    PubMed Central

    Lefort, Karine; Ostano, Gian Paola; Mello-Grand, Maurizia; Calpini, Valérie; Scatolini, Maria; Farsetti, Antonella; Dotto, Gian Paolo; Chiorino, Giovanna

    2016-01-01

    Adenocarcinomas of the prostate arise as multifocal heterogeneous lesions as the likely result of genetic and epigenetic alterations and deranged cell-cell communication. Notch signaling is an important form of intercellular communication with a role in growth/differentiation control and tumorigenesis. Contrasting reports exist in the literature on the role of this pathway in prostate cancer (PCa) development. We show here that i) compared to normal prostate tissue, Notch1 expression is significantly reduced in a substantial fraction of human PCas while it is unaffected or even increased in others; ii) acute Notch activation both inhibits and induces process networks associated with prostatic neoplasms; iii) down-modulation of Notch1 expression and activity in immortalized normal prostate epithelial cells increases their proliferation potential, while increased Notch1 activity in PCa cells suppresses growth and tumorigenicity through a Smad3-dependent mechanism involving p21WAF1/CIP1; iv) prostate cancer cells resistant to Notch growth inhibitory effects retain Notch1-induced upregulation of pro-oncogenic genes, like EPAS1 and CXCL6, also overexpressed in human PCas with high Notch1 levels. Taken together, these results reconcile conflicting data on the role of Notch1 in prostate cancer. PMID:27384993

  6. Conservation of the biochemical mechanisms of signal transduction among mammalian Notch family members

    PubMed Central

    Mizutani, Tomohiko; Taniguchi, Yoshihito; Aoki, Tomokazu; Hashimoto, Nobuo; Honjo, Tasuku

    2001-01-01

    Mouse Notch1, which plays an important role in cell fate determination in development, is proteolytically processed within its transmembrane domain by unidentified γ-secretase-like activity that depends on presenilin. To study this proteolytic event, we established a cell-free Notch cleavage assay system using the membrane fraction of fibroblast transfectants of various Notch constructs with deletion of the extracellular portion (Notch ΔE). The cytoplasmic portion of Notch1 ΔE was released from the membrane upon incubation at 37°C, which was inhibited by the specific γ-secretase inhibitor, MW167, or by overexpression of dominant negative presenilin1. Likewise, other members of mouse Notch family were proteolytically cleaved in a presenilin-dependent, MW167-sensitive manner in vivo as well as in the cell-free Notch ΔE cleavage assay system. All four members of the mouse Notch family migrated to the nucleus and activated the transcription from the promoter carrying the RBP-J consensus sequences after they were released from the membrane. These results demonstrate the conserved biochemical mechanism of signal transduction among mammalian Notch family members. PMID:11459941

  7. Direct regulation of interleukin-6 expression by Notch signaling in macrophages

    PubMed Central

    Wongchana, Wipawee; Palaga, Tanapat

    2012-01-01

    Interleukin-6 (IL-6) is a pleiotropic, pro-inflammatory cytokine produced by various types of cells, including macrophages. Within the IL-6 gene promoter region, the signature binding motif of CBF1/Su(H)/Lag-1 (CSL), a key DNA-binding protein in the Notch signaling pathway, was identified and found to overlap with a consensus nuclear factor (NF)-κB-binding site. Notch signaling is highly conserved and is involved in the regulation of biological functions in immune cells. In this study, we investigated the role of Notch signaling in the regulation of the IL-6 transcript in murine macrophages. The upregulation of Notch1 protein levels and the appearance of cleaved Notch1 (Val1744) correlated well with the increased IL-6 mRNA expression levels in murine primary bone marrow-derived macrophages (BMMφ) after activation by lipopolysaccharide (LPS) together with interferon-gamma (IFN-γ). Treatment of BMMφ with the γ-secretase inhibitor IL-CHO to suppress the transduction of Notch signaling resulted in a partial decrease in the level of IL-6 mRNA and the amount of IL-6 protein produced. In contrast, the overexpression of a constitutively activated intracellular Notch1 protein (NIC) in the RAW264.7 macrophage-like cell line resulted in significantly higher IL-6 transcript expression levels than in cells transfected with the empty vector control. The NF-κB inhibitor completely abrogated IL-6 mRNA expression induced by the overexpression of NIC. Chromatin immunoprecipitation (ChIP) using an anti-Notch1 antibody demonstrated that Notch1 is associated with the IL-6 promoter in RAW264.7 cells activated by LPS/IFN-γ but not in unstimulated cells. Taken together, these results strongly suggest that Notch1 positively regulates IL-6 expression via NF-κB in activated macrophages. PMID:21983868

  8. p53 Modulates Notch Signaling in MCF-7 Breast Cancer Cells by Associating with the Notch Transcriptional Complex via MAML1†

    PubMed Central

    Yun, Jieun; Espinoza, Ingrid; Pannuti, Antonio; Romero, Damian; Martinez, Luis; Caskey, Mary; Stanculescu, Adina; Bocchetta, Maurizio; Rizzo, Paola; Band, Vimla; Band, Hamid; Kim, Hwan Mook; Park, Song-Kyu; Kang, Keon Wook; Avantaggiati, Maria Laura; Gomez, Christian R.; Golde, Todd; Osborne, Barbara; Miele, Lucio

    2015-01-01

    p53 and Notch-1 play important roles in breast cancer biology. Notch-1 inhibits p53 activity in cervical and breast cancer cells. Conversely, p53 inhibits Notch activity in T-cells but stimulates it in human keratinocytes. Notch co-activator MAML1 binds p53 and functions as a p53 co-activator. We studied the regulation of Notch signaling by p53 in MCF-7 cells and normal human mammary epithelial cells (HMEC). Results show that overexpression of p53 or activation of endogenous p53 with Nutlin-3 inhibits Notch-dependent transcriptional activity and Notch target expression in a dose-dependent manner. This effect could be partially rescued by transfection of MAML1 but not p300. Standard and quantitative co-immunoprecipitation experiments readily detected a complex containing p53 and Notch-1 in MCF-7 cells. Formation of this complex was inhibited by dominant negative MAML1 (DN-MAML1) and stimulated by wild-type MAML1. Standard and quantitative far-Western experiments showed a complex including p53, Notch-1 and MAML1. Chromatin immunoprecipitation (ChIP) experiments showed that p53 can associate with Notch-dependent HEY1 promoter and this association is inhibited by DN-MAML1 and stimulated by wild-type MAML1. Our data support a model in which p53 associates with the Notch transcriptional complex (NTC) in a MAML1-dependent fashion, most likely through a p53-MAML1 interaction. In our cellular models, the effect of this association is to inhibit Notch-dependent transcription. Our data suggest that p53-null breast cancers may lack this Notch-modulatory mechanism, and that therapeutic strategies that activate wild-type p53 can indirectly cause inhibition of Notch transcriptional activity. PMID:26033683

  9. p53 Modulates Notch Signaling in MCF-7 Breast Cancer Cells by Associating With the Notch Transcriptional Complex Via MAML1.

    PubMed

    Yun, Jieun; Espinoza, Ingrid; Pannuti, Antonio; Romero, Damian; Martinez, Luis; Caskey, Mary; Stanculescu, Adina; Bocchetta, Maurizio; Rizzo, Paola; Band, Vimla; Band, Hamid; Kim, Hwan Mook; Park, Song-Kyu; Kang, Keon Wook; Avantaggiati, Maria Laura; Gomez, Christian R; Golde, Todd; Osborne, Barbara; Miele, Lucio

    2015-12-01

    p53 and Notch-1 play important roles in breast cancer biology. Notch-1 inhibits p53 activity in cervical and breast cancer cells. Conversely, p53 inhibits Notch activity in T-cells but stimulates it in human keratinocytes. Notch co-activator MAML1 binds p53 and functions as a p53 co-activator. We studied the regulation of Notch signaling by p53 in MCF-7 cells and normal human mammary epithelial cells (HMEC). Results show that overexpression of p53 or activation of endogenous p53 with Nutlin-3 inhibits Notch-dependent transcriptional activity and Notch target expression in a dose-dependent manner. This effect could be partially rescued by transfection of MAML1 but not p300. Standard and quantitative co-immunoprecipitation experiments readily detected a complex containing p53 and Notch-1 in MCF-7 cells. Formation of this complex was inhibited by dominant negative MAML1 (DN-MAML1) and stimulated by wild-type MAML1. Standard and quantitative far-Western experiments showed a complex including p53, Notch-1, and MAML1. Chromatin immunoprecipitation (ChIP) experiments showed that p53 can associate with Notch-dependent HEY1 promoter and this association is inhibited by DN-MAML1 and stimulated by wild-type MAML1. Our data support a model in which p53 associates with the Notch transcriptional complex (NTC) in a MAML1-dependent fashion, most likely through a p53-MAML1 interaction. In our cellular models, the effect of this association is to inhibit Notch-dependent transcription. Our data suggest that p53-null breast cancers may lack this Notch-modulatory mechanism, and that therapeutic strategies that activate wild-type p53 can indirectly cause inhibition of Notch transcriptional activity.

  10. Notch signaling modulates proliferative vitreoretinopathy via regulating retinal pigment epithelial-to-mesenchymal transition.

    PubMed

    Zhang, Jingjing; Yuan, Gongqiang; Dong, Muchen; Zhang, Ting; Hua, Gao; Zhou, Qingjun; Shi, Weiyun

    2016-09-07

    Elevated Notch signaling has been verified in a large range of fibrotic diseases developed in the kidney, liver, and lung, inducing the development of the epithelial-mesenchymal transition (EMT). The aim of this study was to observe the involvement of Notch signaling in the EMT of retinal pigment epithelial (RPE) cells and the pathogenesis of proliferative vitreoretinopathy (PVR). In vitro cultivated human RPE cells (ARPE-19) were treated with 10 ng/mL transforming growth factor (TGF)-β1 for 24, 48, and 72 h. The expression levels of ZO-1, α-SMA, vimentin, Notch1 intracellular domain (NICD1), and Hes-1 were evaluated with quantitative real-time polymerase chain reaction (qRT-PCR), immunofluorescence staining or Western blot. TGF-β1 induced EMT and the activation of Notch signaling in ARPE-19 cells. To examine the effect of Notch inhibition on TGF-β1-induced EMT and PVR formation, ARPE-19 cells were preincubated with γ-secretase inhibitor LY411575 before TGF-β1 treatment. Mouse PVR model was used for in vivo study. ARPE-19 cells were injected intravitreously with or without the LY411575 to examine the effect of Notch inhibition on PVR formation. LY411575 significantly attenuated EMT by inhibiting the Notch signaling activation in vitro. PVR was induced by intravitreal injections of ARPE-19 cells, while LY411575 inhibited mouse PVR formation in vivo. Notch signaling plays a critical role in TGF-β1-induced EMT in vitro and mice PVR model, which provides a novel insight into the pathogenesis of PVR. The specific inhibition of Notch signaling by γ-secretase inhibitor may provide a new approach for the prevention of PVR.

  11. Molecular pathways: translational and therapeutic implications of the Notch signaling pathway in cancer.

    PubMed

    Previs, Rebecca A; Coleman, Robert L; Harris, Adrian L; Sood, Anil K

    2015-03-01

    Over 100 years have passed since the first observation of the notched wing phenotype in Drosophila melanogaster, and significant progress has been made to characterize the role of the Notch receptor, its ligands, downstream targets, and cross-talk with other signaling pathways. The canonical Notch pathway with four Notch receptors (Notch1-4) and five ligands (DLL1, 3-4, Jagged 1-2) is an evolutionarily conserved cell signaling pathway that plays critical roles in cell-fate determination, differentiation, development, tissue patterning, cell proliferation, and death. In cancer, these roles have a critical impact on tumor behavior and response to therapy. Because the role of Notch remains tissue and context dependent, alterations within this pathway may lead to tumor suppressive or oncogenic phenotypes. Although no FDA-approved therapies currently exist for the Notch pathway, multiple therapeutics (e.g., demcizumab, tarextumab, GSI MK-0752, R04929097, and PF63084014) have been developed to target different aspects of this pathway for both hematologic and solid malignancies. Understanding the context-specific effects of the Notch pathway will be important for individualized therapies targeting this pathway.

  12. Molecular Profiling of the Developing Lacrimal Gland Reveals Putative Role of Notch Signaling in Branching Morphogenesis

    PubMed Central

    Dvoriantchikova, Galina; Tao, Wensi; Pappas, Steve; Gaidosh, Gabriel; Tse, David T.; Ivanov, Dmitry; Pelaez, Daniel

    2017-01-01

    Purpose Although normal function of the lacrimal gland is essential for vision (and thus for human well-being), the lacrimal gland remains rather poorly understood at a molecular level. The purpose of this study was to identify new genes and signaling cascades involved in lacrimal gland development. Methods To identify these genes, we used microarray analysis to compare the gene expression profiles of developing (embryonic) and adult lacrimal glands. Differential data were validated by quantitative RT-PCR, and several corresponding proteins were confirmed by immunohistochemistry and Western blot analysis. To evaluate the role of NOTCH signaling in lacrimal gland (LG) development, we used the NOTCH inhibitor DAPT and conditional Notch1 knockouts. Results Our microarray data and an in silico reconstruction of cellular networks revealed significant changes in the expression patterns of genes from the NOTCH, WNT, TGFβ, and Hedgehog pathways, all of which are involved in the regulation of epithelial-to-mesenchymal transition (EMT). Our study also revealed new putative lacrimal gland stem cell/progenitor markers. We found that inhibiting Notch signaling both increases the average number of lacrimal gland lobules and reduces the size of each lobule. Conclusions Our findings suggest that NOTCH-, WNT-, TGFβ-, and Hedgehog-regulated EMT transition are critical mechanisms in lacrimal gland development and morphogenesis. Our data also supports the hypothesis that NOTCH signaling regulates branching morphogenesis in the developing lacrimal gland by suppressing cleft formation. PMID:28192800

  13. Quinomycin A targets Notch signaling pathway in pancreatic cancer stem cells.

    PubMed

    Ponnurangam, Sivapriya; Dandawate, Prasad R; Dhar, Animesh; Tawfik, Ossama W; Parab, Rajashri R; Mishra, Prabhu Dutt; Ranadive, Prafull; Sharma, Rajiv; Mahajan, Girish; Umar, Shahid; Weir, Scott J; Sugumar, Aravind; Jensen, Roy A; Padhye, Subhash B; Balakrishnan, Arun; Anant, Shrikant; Subramaniam, Dharmalingam

    2016-01-19

    Cancer stem cells (CSCs) appear to explain many aspects of the neoplastic evolution of tumors and likely account for enhanced therapeutic resistance following treatment. Dysregulated Notch signaling, which affects CSCs plays an important role in pancreatic cancer progression. We have determined the ability of Quinomycin to inhibit CSCs and the Notch signaling pathway. Quinomycin treatment resulted in significant inhibition of proliferation and colony formation in pancreatic cancer cell lines, but not in normal pancreatic epithelial cells. Moreover, Quinomycin affected pancreatosphere formation. The compound also decreased the expression of CSC marker proteins DCLK1, CD44, CD24 and EPCAM. In addition, flow cytometry studies demonstrated that Quinomycin reduced the number of DCLK1+ cells. Furthermore, levels of Notch 1-4 receptors, their ligands Jagged1, Jagged2, DLL1, DLL3, DLL4 and the downstream target protein Hes-1 were reduced. The γ-secretase complex proteins, Presenilin 1, Nicastrin, Pen2, and APH-1, required for Notch activation also exhibited decreased expression. Ectopic expression of the Notch Intracellular Domain (NICD) partially rescued the cells from Quinomycin mediated growth suppression. To determine the effect of Quinomycin on tumor growth in vivo, nude mice carrying tumor xenografts were administered Quinomycin intraperitoneally every day for 21 days. Treatment with the compound significantly inhibited tumor xenograft growth, coupled with significant reduction in the expression of CSC markers and Notch signaling proteins. Together, these data suggest that Quinomycin is a potent inhibitor of pancreatic cancer that targets the stem cells by inhibiting Notch signaling proteins.

  14. Inhibition of Notch Signaling Attenuates Schistosomiasis Hepatic Fibrosis via Blocking Macrophage M2 Polarization

    PubMed Central

    Chen, Yixiong; Zheng, Shaojiang; Zheng, Liping; Weng, Zhihong

    2016-01-01

    Macrophages play a key role in the pathogenesis of liver granuloma and fibrosis in schistosomiasis. However, the underlying mechanisms have not been fully characterized. This study revealed that the macrophages infiltrating the liver tissues in a murine model of Schistosoma japonica infection exhibited M2 functional polarization, and Notch1/Jagged1 signaling was significantly upregulated in the M2 polarized macrophages in vivo and in vitro. Furthermore, the blockade of Notch signaling pathway by a γ–secretase inhibitor could reverse macrophage M2 polarization in vitro and alleviate liver granuloma and fibrosis in the murine model of schistosomiasis. These results implied that the Notch1/Jagged1 signaling-dependent M2 polarization of macrophages might play an important role in liver granuloma and fibrosis in schistosomiasis, and the inhibition of Notch1/Jagged1 signaling might provide a novel therapeutic approach to administrate patients with schistosomiasis. PMID:27875565

  15. Inflammation and Notch signaling: a crosstalk with opposite effects on tumorigenesis

    PubMed Central

    Fazio, Chiara; Ricciardiello, Luigi

    2016-01-01

    The Notch cascade is a fundamental and highly conserved pathway able to control cell-fate. The Notch pathway arises from the interaction of one of the Notch receptors (Notch1–4) with different types of ligands; in particular, the Notch pathway can be activated canonically (through the ligands Jagged1, Jagged2, DLL1, DLL3 or DLL4) or non-canonically (through various molecules shared by other pathways). In the context of tumor biology, the deregulation of Notch signaling is found to be crucial, but it is still not clear if the activation of this pathway exerts a tumor-promoting or a tumor suppressing function in different cancer settings. Untill now, it is well known that the inflammatory compartment is critically involved in tumor progression; however, inflammation, which occurs as a physiological response to damage, can also drive protective processes toward carcinogenesis. Therefore, the role of inflammation in cancer is still controversial and needs to be further clarified. Interestingly, recent literature reports that some of the signaling molecules modulated by the cells of the immune system also belong to or interact with the canonical and non-canonical Notch pathways, delineating a possible link between Notch activation and inflammatory environment. In this review we analyze the hypothesis that specific inflammatory conditions can control the activation of the Notch pathway in terms of biological effect, partially explaining the dichotomy of both phenomena. For this purpose, we detail the molecular links reported in the literature connecting inflammation and Notch signaling in different types of tumor, with a particular focus on colorectal carcinogenesis, which represents a perfect example of context-dependent interaction between malignant transformation and immune response. PMID:27929540

  16. Delta-1 Activation of Notch-1 Signaling Results in HES-1 Transactivation

    PubMed Central

    Jarriault, Sophie; Le Bail, Odile; Hirsinger, Estelle; Pourquié, Olivier; Logeat, Frédérique; Strong, Clare F.; Brou, Christel; Seidah, Nabil G.; Israël, Alain

    1998-01-01

    The Notch receptor is involved in many cell fate determination events in vertebrates and invertebrates. It has been shown in Drosophila melanogaster that Delta-dependent Notch signaling activates the transcription factor Suppressor of Hairless, leading to an increased expression of the Enhancer of Split genes. Genetic evidence has also implicated the kuzbanian gene, which encodes a disintegrin metalloprotease, in the Notch signaling pathway. By using a two-cell coculture assay, we show here that vertebrate Dl-1 activates the Notch-1 cascade. Consistent with previous data obtained with active forms of Notch-1 a HES-1-derived promoter construct is transactivated in cells expressing Notch-1 in response to Dl-1 stimulation. Impairing the proteolytic maturation of the full-length receptor leads to a decrease in HES-1 transactivation, further supporting the hypothesis that only mature processed Notch is expressed at the cell surface and activated by its ligand. Furthermore, we observed that Dl-1-induced HES-1 transactivation was dependent both on Kuzbanian and RBP-J activities, consistent with the involvement of these two proteins in Notch signaling in Drosophila. We also observed that exposure of Notch-1-expressing cells to Dl-1 results in an increased level of endogenous HES-1 mRNA. Finally, coculture of Dl-1-expressing cells with myogenic C2 cells suppresses differentiation of C2 cells into myotubes, as previously demonstrated for Jagged-1 and Jagged-2, and also leads to an increased level of endogenous HES-1 mRNA. Thus, Dl-1 behaves as a functional ligand for Notch-1 and has the same ability to suppress cell differentiation as the Jagged proteins do. PMID:9819428

  17. CaMK-II activation is essential for zebrafish inner ear development and acts through Delta-Notch signaling.

    PubMed

    Rothschild, Sarah C; Lahvic, Jamie; Francescatto, Ludmila; McLeod, Jamie J A; Burgess, Shawn M; Tombes, Robert M

    2013-09-01

    Zebrafish inner ear development is characterized by the crystallization of otoliths onto immotile kinocilia that protrude from sensory "hair" cells. The stereotypical formation of these sensory structures is dependent on the expression of key patterning genes and on Ca2+ signals. One potential target of Ca2+ signaling in the inner ear is the type II Ca2+/calmodulin-dependent protein kinase (CaMK-II), which is preferentially activated in hair cells, with intense activation at the base of kinocilia. In zebrafish, CaMK-II is encoded by seven genes; the expression of one of these genes (camk2g1) is enriched in hair cells. The suppression of camk2g1 expression by antisense morpholino oligonucleotides or inhibition of CaMK-II activation by the pharmacological antagonist, KN-93, results in aberrant otolith formation without preventing cilia formation. In fact, CaMK-II suppression results in additional ciliated hair cells and altered levels of Delta-Notch signaling members. DeltaA and deltaD transcripts are increased and DeltaD protein accumulates in hair cells of CaMK-II morphants, indicative of defective recycling and/or exocytosis. Our findings indicate that CaMK-II plays a critical role in the developing ear, influencing cell differentiation through extranuclear effects on Delta-Notch signaling. Continued expression and activation of CaMK-II in maculae and cristae in older embryos suggests continued roles in auditory sensory maturation and transduction.

  18. Reduced Notch signalling leads to postnatal skeletal muscle hypertrophy in Pofut1cax/cax mice.

    PubMed

    Al Jaam, Bilal; Heu, Katy; Pennarubia, Florian; Segelle, Alexandre; Magnol, Laetitia; Germot, Agnès; Legardinier, Sébastien; Blanquet, Véronique; Maftah, Abderrahman

    2016-09-01

    Postnatal skeletal muscle growth results from the activation of satellite cells and/or an increase in protein synthesis. The Notch signalling pathway maintains satellite cells in a quiescent state, and once activated, sustains their proliferation and commitment towards differentiation. In mammals, POFUT1-mediated O-fucosylation regulates the interactions between NOTCH receptors and ligands of the DELTA/JAGGED family, thus initiating the activation of canonical Notch signalling. Here, we analysed the consequences of downregulated expression of the Pofut1 gene on postnatal muscle growth in mutant Pofut1(cax/cax) (cax, compact axial skeleton) mice and differentiation of their satellite cell-derived myoblasts (SCDMs). Pofut1(cax/cax) mice exhibited muscle hypertrophy, no hyperplasia and a decrease in satellite cell numbers compared with wild-type C3H mice. In agreement with these observations, Pofut1(cax/cax) SCDMs differentiated earlier concomitant with reduced Pax7 expression and decrease in PAX7(+)/MYOD(-) progenitor cells. In vitro binding assays showed a reduced interaction of DELTA-LIKE 1 ligand (DLL1) with NOTCH receptors expressed at the cell surface of SCDMs, leading to a decreased Notch signalling as seen by the quantification of cleaved NICD and Notch target genes. These results demonstrated that POFUT1-mediated O-fucosylation of NOTCH receptors regulates myogenic cell differentiation and affects postnatal muscle growth in mice.

  19. Reduced Notch signalling leads to postnatal skeletal muscle hypertrophy in Pofut1cax/cax mice

    PubMed Central

    Al Jaam, Bilal; Heu, Katy; Pennarubia, Florian; Segelle, Alexandre; Magnol, Laetitia; Germot, Agnès; Blanquet, Véronique; Maftah, Abderrahman

    2016-01-01

    Postnatal skeletal muscle growth results from the activation of satellite cells and/or an increase in protein synthesis. The Notch signalling pathway maintains satellite cells in a quiescent state, and once activated, sustains their proliferation and commitment towards differentiation. In mammals, POFUT1-mediated O-fucosylation regulates the interactions between NOTCH receptors and ligands of the DELTA/JAGGED family, thus initiating the activation of canonical Notch signalling. Here, we analysed the consequences of downregulated expression of the Pofut1 gene on postnatal muscle growth in mutant Pofut1cax/cax (cax, compact axial skeleton) mice and differentiation of their satellite cell-derived myoblasts (SCDMs). Pofut1cax/cax mice exhibited muscle hypertrophy, no hyperplasia and a decrease in satellite cell numbers compared with wild-type C3H mice. In agreement with these observations, Pofut1cax/cax SCDMs differentiated earlier concomitant with reduced Pax7 expression and decrease in PAX7+/MYOD− progenitor cells. In vitro binding assays showed a reduced interaction of DELTA-LIKE 1 ligand (DLL1) with NOTCH receptors expressed at the cell surface of SCDMs, leading to a decreased Notch signalling as seen by the quantification of cleaved NICD and Notch target genes. These results demonstrated that POFUT1-mediated O-fucosylation of NOTCH receptors regulates myogenic cell differentiation and affects postnatal muscle growth in mice. PMID:27628322

  20. Radiation-Induced Notch Signaling in Breast Cancer Stem Cells

    SciTech Connect

    Lagadec, Chann; Vlashi, Erina; Alhiyari, Yazeed; Phillips, Tiffany M.; Bochkur Dratver, Milana; Pajonk, Frank

    2013-11-01

    Purpose: To explore patterns of Notch receptor and ligand expression in response to radiation that could be crucial in defining optimal dosing schemes for γ-secretase inhibitors if combined with radiation. Methods and Materials: Using MCF-7 and T47D breast cancer cell lines, we used real-time reverse transcription–polymerase chain reaction to study the Notch pathway in response to radiation. Results: We show that Notch receptor and ligand expression during the first 48 hours after irradiation followed a complex radiation dose–dependent pattern and was most pronounced in mammospheres, enriched for breast cancer stem cells. Additionally, radiation activated the Notch pathway. Treatment with a γ-secretase inhibitor prevented radiation-induced Notch family gene expression and led to a significant reduction in the size of the breast cancer stem cell pool. Conclusions: Our results indicate that, if combined with radiation, γ-secretase inhibitors may prevent up-regulation of Notch receptor and ligand family members and thus reduce the number of surviving breast cancer stem cells.

  1. Roles of Notch1 Signaling in Regulating Satellite Cell Fates Choices and Postnatal Skeletal Myogenesis.

    PubMed

    Shan, Tizhong; Xu, Ziye; Wu, Weiche; Liu, Jiaqi; Wang, Yizhen

    2016-12-12

    Adult skeletal muscle stem cells, also called satellite cells, are indispensable for the growth, maintenance, and regeneration of the postnatal skeletal muscle. Satellite cells, predominantly quiescent in mature resting muscles, are activated after skeletal muscle injury or degeneration. Notch1 signaling is an evolutionarily conserved pathway that plays crucial roles in satellite cells homeostasis and postnatal skeletal myogenesis and regeneration. Activation of Notch1 signaling promotes the muscle satellite cells quiescence and proliferation, but inhibits differentiation of muscle satellite cells. Notably, the new roles of Notch1 signaling during late-stage of skeletal myogenesis including in post-differentiation myocytes and post-fusion myotubes have been recently reported. Here, we mainly review and discuss the regulatory roles of Notch1 in regulating satellite cell fates choices and skeletal myogenesis. This article is protected by copyright. All rights reserved.

  2. Inhibition of Notch Signaling During Mouse Incisor Renewal Leads to Enamel Defects.

    PubMed

    Jheon, Andrew H; Prochazkova, Michaela; Meng, Bo; Wen, Timothy; Lim, Young-Jun; Naveau, Adrien; Espinoza, Ruben; Cox, Timothy C; Sone, Eli D; Ganss, Bernhard; Siebel, Christian W; Klein, Ophir D

    2016-01-01

    The continuously growing rodent incisor is an emerging model for the study of renewal of mineralized tissues by adult stem cells. Although the Bmp, Fgf, Shh, and Wnt pathways have been studied in this organ previously, relatively little is known about the role of Notch signaling during incisor renewal. Notch signaling components are expressed in enamel-forming ameloblasts and the underlying stratum intermedium (SI), which suggested distinct roles in incisor renewal and enamel mineralization. Here, we injected adult mice with inhibitory antibodies against several components of the Notch pathway. This blockade led to defects in the interaction between ameloblasts and the SI cells, which ultimately affected enamel formation. Furthermore, Notch signaling inhibition led to the downregulation of desmosome-specific proteins such as PERP and desmoplakin, consistent with the importance of desmosomes in the integrity of ameloblast-SI attachment and enamel formation. Together, our data demonstrate that Notch signaling is critical for proper enamel formation during incisor renewal, in part by regulating desmosome-specific components, and that the mouse incisor provides a model system to dissect Jag-Notch signaling mechanisms in the context of mineralized tissue renewal.

  3. Notch as a tumour suppressor.

    PubMed

    Nowell, Craig S; Radtke, Freddy

    2017-03-01

    The Notch signalling cascade is an evolutionarily conserved pathway that has a crucial role in regulating development and homeostasis in various tissues. The cellular processes and events that it controls are diverse, and continued investigation over recent decades has revealed how the role of Notch signalling is multifaceted and highly context dependent. Consistent with the far-reaching impact that Notch has on development and homeostasis, aberrant activity of the pathway is also linked to the initiation and progression of several malignancies, and Notch can in fact be either oncogenic or tumour suppressive depending on the tissue and cellular context. The Notch pathway therefore represents an important target for therapeutic agents designed to treat many types of cancer. In this Review, we focus on the latest developments relating specifically to the tumour-suppressor activity of Notch signalling and discuss the potential mechanisms by which Notch can inhibit carcinogenesis in various tissues. Potential therapeutic strategies aimed at restoring or augmenting Notch-mediated tumour suppression will also be highlighted.

  4. Constitutively active Notch1 induces growth arrest of HPV-positive cervical cancer cells via separate signaling pathways.

    PubMed

    Talora, Claudio; Cialfi, Samantha; Segatto, Oreste; Morrone, Stefania; Kim Choi, John; Frati, Luigi; Paolo Dotto, Gian; Gulino, Alberto; Screpanti, Isabella

    2005-05-01

    Notch signaling plays a key role in cell-fate determination and differentiation in different organisms and cell types. Several reports suggest that Notch signaling may be involved in neoplastic transformation. However, in primary keratinocytes, Notch1 can function as a tumor suppressor. Similarly, in HPV-positive cervical cancer cells, constitutively active Notch1 signaling was found to cause growth suppression. Activated Notch1 in these cells represses viral E6/E7 expression through AP-1 down-modulation, resulting in increased p53 expression and a block of pRb hyperphosphorylation. Here we show that in cervical cancer cell lines in which Notch1 ability to repress AP-1 activity is impaired, Notch1-enforced expression elicits an alternative pathway leading to growth arrest. Indeed, activated Notch1 signaling suppresses activity of the helix-loop-helix transcription factor E47, via ERK1/2 activation, resulting in inhibition of cell cycle progression. Moreover, we found that RBP-Jkappa-dependent Notch signaling is specifically repressed in cervical cancer cells and this repression could provide one such mechanism that needs to be activated for cervical carcinogenesis. Finally, we show that inhibition of endogenous Notch1 signaling, although results in a proliferative advantage, sensitizes cervical cancer cell lines to drug-induced apoptosis. Together, our results provide novel molecular insights into Notch1-dependent growth inhibitory effects, counteracting the transforming potential of HPV.

  5. Constitutively active Notch1 induces growth arrest of HPV-positive cervical cancer cells via separate signaling pathways

    SciTech Connect

    Talora, Claudio; Cialfi, Samantha; Segatto, Oreste; Morrone, Stefania; Kim Choi, John; Frati, Luigi; Paolo Dotto, Gian; Gulino, Alberto; Screpanti, Isabella . E-mail: isabella.screpanti@uniroma1.it

    2005-05-01

    Notch signaling plays a key role in cell-fate determination and differentiation in different organisms and cell types. Several reports suggest that Notch signaling may be involved in neoplastic transformation. However, in primary keratinocytes, Notch1 can function as a tumor suppressor. Similarly, in HPV-positive cervical cancer cells, constitutively active Notch1 signaling was found to cause growth suppression. Activated Notch1 in these cells represses viral E6/E7 expression through AP-1 down-modulation, resulting in increased p53 expression and a block of pRb hyperphosphorylation. Here we show that in cervical cancer cell lines in which Notch1 ability to repress AP-1 activity is impaired, Notch1-enforced expression elicits an alternative pathway leading to growth arrest. Indeed, activated Notch1 signaling suppresses activity of the helix-loop-helix transcription factor E47, via ERK1/2 activation, resulting in inhibition of cell cycle progression. Moreover, we found that RBP-J{kappa}-dependent Notch signaling is specifically repressed in cervical cancer cells and this repression could provide one such mechanism that needs to be activated for cervical carcinogenesis. Finally, we show that inhibition of endogenous Notch1 signaling, although results in a proliferative advantage, sensitizes cervical cancer cell lines to drug-induced apoptosis. Together, our results provide novel molecular insights into Notch1-dependent growth inhibitory effects, counteracting the transforming potential of HPV.

  6. Effects of Linker Length and Transient Secondary Structure Elements in the Intrinsically Disordered Notch RAM Region on Notch Signaling.

    PubMed

    Sherry, Kathryn P; Johnson, Scott E; Hatem, Christine L; Majumdar, Ananya; Barrick, Doug

    2015-11-06

    Formation of the bivalent interaction between the Notch intracellular domain (NICD) and the transcription factor CBF-1/RBP-j, Su(H), Lag-1 (CSL) is a key event in Notch signaling because it switches Notch-responsive genes from a repressed state to an activated state. Interaction of the intrinsically disordered RBP-j-associated molecule (RAM) region of NICD with CSL is thought to both disrupt binding of corepressor proteins to CSL and anchor NICD to CSL, promoting interaction of the ankyrin domain of NICD with CSL through an effective concentration mechanism. To quantify the role of disorder in the RAM linker region on the effective concentration enhancement of Notch transcriptional activation, we measured the effects of linker length variation on activation. The resulting activation profile has general features of a worm-like chain model for effective concentration. However, deviations from the model for short sequence deletions suggest that RAM contains sequence-specific structural elements that may be important for activation. Structural characterization of the RAM linker with sedimentation velocity analytical ultracentrifugation and NMR spectroscopy reveals that the linker is compact and contains three transient helices and two extended and dynamic regions. To test if these secondary structure elements are important for activation, we made sequence substitutions to change the secondary structure propensities of these elements and measured transcriptional activation of the resulting variants. Substitutions to two of these nonrandom elements (helix 2, extended region 1) have effects on activation, but these effects do not depend on the nature of the substituting residues. Thus, the primary sequences of these elements, but not their secondary structures, are influencing signaling.

  7. Renal tubular Notch signaling triggers a prosenescent state after acute kidney injury.

    PubMed

    Sörensen-Zender, Inga; Rong, Song; Susnik, Nathan; Zender, Steffen; Pennekamp, Petra; Melk, Anette; Haller, Hermann; Schmitt, Roland

    2014-04-15

    The aging kidney has a diminished regenerative potential and an increased tendency to develop tubular atrophy and fibrosis after acute injury. In this study, we found that activation of tubular epithelial Notch1 signaling was prolonged in the aging kidney after ischemia/reperfusion (IR) damage. To analyze the consequences of sustained Notch activation, we generated mice with conditional inducible expression of Notch1 intracellular domain (NICD) in proximal tubules. NICD kidneys were analyzed 1 and 4 wk after renal IR. Conditional NICD expression was associated with aggravated tubular damage, a fibrotic phenotype, and the expression of cellular senescence markers p21 and p16(INK4a). In wild-type mice pharmacological inhibition of Notch using the γ-secretase inhibitor N-[N-(3,5-difluorophenacetyl)-l-alanyl]-S-phenylglycine t-butyl ester (DAPT) improved tubulo-interstitial damage and antagonized the prosenescent pathway activation after IR. In vitro, activation of Notch signaling with delta-like-ligand-4 caused prosenescent changes in tubular cells while inhibition with DAPT attenuated these changes. In conclusion, our data suggest that sustained epithelial Notch activation after IR might contribute to the inferior outcome of old kidneys after injury. Sustained epithelial activation of Notch is associated with a prosenescent phenotype and maladaptive repair.

  8. Zebrafish nephrogenesis is regulated by interactions between retinoic acid, mecom, and Notch signaling.

    PubMed

    Li, Yue; Cheng, Christina N; Verdun, Valerie A; Wingert, Rebecca A

    2014-02-01

    The zebrafish pronephros provides a conserved model to study kidney development, in particular to delineate the poorly understood processes of how nephron segment pattern and cell type choice are established. Zebrafish nephrons are divided into distinct epithelial regions that include a series of proximal and distal tubule segments, which are comprised of intercalated transporting epithelial cells and multiciliated cells (MCC). Previous studies have shown that retinoic acid (RA) regionalizes the renal progenitor field into proximal and distal domains and that Notch signaling later represses MCC differentiation, but further understanding of these pathways has remained unknown. The transcription factor mecom (mds1/evi1 complex) is broadly expressed in renal progenitors, and then subsequently marks the distal tubule. Here, we show that mecom is necessary to form the distal tubule and to restrict both proximal tubule formation and MCC fate choice. We found that mecom and RA have opposing roles in patterning discrete proximal and distal segments. Further, we discovered that RA is required for MCC formation, and that one mechanism by which RA promotes MCC fate choice is to inhibit mecom. Next, we determined the epistatic relationship between mecom and Notch signaling, which limits MCC fate choice by lateral inhibition. Abrogation of Notch signaling with the γ-secretase inhibitor DAPT revealed that Notch and mecom did not have additive effects in blocking MCC formation, suggesting that they function in the same pathway. Ectopic expression of the Notch signaling effector, Notch intracellular domain (NICD), rescued the expansion of MCCs in mecom morphants, indicating that mecom acts upstream to induce Notch signaling. These findings suggest a model in which mecom and RA arbitrate proximodistal segment domains, while MCC fate is modulated by a complex interplay in which RA inhibition of mecom, and mecom promotion of Notch, titrates MCC number. Taken together, our studies

  9. Zebrafish nephrogenesis is regulated by interactions between retinoic acid, mecom, and Notch signaling

    PubMed Central

    Li, Yue; Cheng, Christina N.; Verdun, Valerie A.; Wingert, Rebecca A.

    2014-01-01

    The zebrafish pronephros provides a conserved model to study kidney development, in particular to delineate the poorly understood processes of how nephron segment pattern and cell type choice are established. Zebrafish nephrons are divided into distinct epithelial regions that include a series of proximal and distal tubule segments, which are comprised of intercalated transporting epithelial cells and multiciliated cells (MCC). Previous studies have shown that retinoic acid (RA) regionalizes the renal progenitor field into proximal and distal domains and that Notch signaling later represses MCC differentiation, but further understanding of these pathways has remained unknown. The transcription factor mecom (mds1/evi1 complex) is broadly expressed in renal progenitors, and then subsequently marks the distal tubule. Here, we show that mecom is necessary to form the distal tubule and to restrict both proximal tubule formation and MCC fate choice. We found that mecom and RA have opposing roles in patterning discrete proximal and distal segments. Further, we discovered that RA is required for MCC formation, and that one mechanism by which RA promotes MCC fate choice is to inhibit mecom. Next, we determined the epistatic relationship between mecom and Notch signaling, which limits MCC fate choice by lateral inhibition. Abrogation of Notch signaling with the y-secretase inhibitor DAPT revealed that Notch and mecom did not have additive effects in blocking MCC formation, suggesting that they function in the same pathway. Ectopic expression of the Notch signaling effector, Notch intracellular domain (NICD), rescued the expansion of MCCs in mecom morphants, indicating that mecom acts upstream to induce Notch signaling. These findings suggest a model in which mecom and RA arbitrate proximodistal segment domains, while MCC fate is modulated by a complex interplay in which RA inhibition of mecom, and mecom promotion of Notch, titrates MCC number. Taken together, our studies

  10. A Disintegrin and Metalloproteinase10 (ADAM10) Regulates NOTCH Signaling during Early Retinal Development.

    PubMed

    Toonen, Joseph A; Ronchetti, Adam; Sidjanin, D J

    2016-01-01

    ADAM10 and ADAM17 are two closely related members of the ADAM (a disintegrin and metalloprotease) family of membrane-bound sheddases, which proteolytically cleave surface membrane proteins. Both ADAM10 and ADAM17 have been implicated in the proteolytic cleavage of NOTCH receptors and as such regulators of NOTCH signaling. During retinal development, NOTCH signaling facilitates retinal neurogenesis by maintaining progenitor cells in a proliferative state and by mediating retinal cell fates. However, the roles of ADAM10 and ADAM17 in the retina are not well defined. In this study, we set out to clarify the roles of ADAM10 and ADAM17 during early retinal development. The retinal phenotype of conditionally abated Adam17 retinae (Adam17 CKO) did not differ from the controls whereas conditionally ablated Adam10 retinae (Adam10 CKO) exhibited abnormal morphogenesis characterized by the formation of rosettes and a loss of retinal laminae phenotypically similar to morphological abnormalities identified in mice with retinal NOTCH signaling deficiency. Additionally, Adam10 CKO retinae exhibited abnormal neurogenesis characterized by fewer proliferating progenitor cells and greater differentiation of early photoreceptors and retinal ganglion cells. Moreover, constitutive activation of the NOTCH1-intracellular domain (N1-ICD) rescued Adam10 CKO abnormal neurogenesis, as well as abnormal retinal morphology by maintaining retinal cells in the progenitor state. Collectively these findings provide in vivo genetic evidence that ADAM10, and not ADAM17, is indispensable for proper retinal development as a regulator of NOTCH signaling.

  11. A Disintegrin and Metalloproteinase10 (ADAM10) Regulates NOTCH Signaling during Early Retinal Development

    PubMed Central

    Toonen, Joseph A.; Ronchetti, Adam; Sidjanin, D. J.

    2016-01-01

    ADAM10 and ADAM17 are two closely related members of the ADAM (a disintegrin and metalloprotease) family of membrane-bound sheddases, which proteolytically cleave surface membrane proteins. Both ADAM10 and ADAM17 have been implicated in the proteolytic cleavage of NOTCH receptors and as such regulators of NOTCH signaling. During retinal development, NOTCH signaling facilitates retinal neurogenesis by maintaining progenitor cells in a proliferative state and by mediating retinal cell fates. However, the roles of ADAM10 and ADAM17 in the retina are not well defined. In this study, we set out to clarify the roles of ADAM10 and ADAM17 during early retinal development. The retinal phenotype of conditionally abated Adam17 retinae (Adam17 CKO) did not differ from the controls whereas conditionally ablated Adam10 retinae (Adam10 CKO) exhibited abnormal morphogenesis characterized by the formation of rosettes and a loss of retinal laminae phenotypically similar to morphological abnormalities identified in mice with retinal NOTCH signaling deficiency. Additionally, Adam10 CKO retinae exhibited abnormal neurogenesis characterized by fewer proliferating progenitor cells and greater differentiation of early photoreceptors and retinal ganglion cells. Moreover, constitutive activation of the NOTCH1-intracellular domain (N1-ICD) rescued Adam10 CKO abnormal neurogenesis, as well as abnormal retinal morphology by maintaining retinal cells in the progenitor state. Collectively these findings provide in vivo genetic evidence that ADAM10, and not ADAM17, is indispensable for proper retinal development as a regulator of NOTCH signaling. PMID:27224017

  12. Origin and evolution of the Notch signalling pathway: an overview from eukaryotic genomes

    PubMed Central

    Gazave, Eve; Lapébie, Pascal; Richards, Gemma S; Brunet, Frédéric; Ereskovsky, Alexander V; Degnan, Bernard M; Borchiellini, Carole; Vervoort, Michel; Renard, Emmanuelle

    2009-01-01

    Background Of the 20 or so signal transduction pathways that orchestrate cell-cell interactions in metazoans, seven are involved during development. One of these is the Notch signalling pathway which regulates cellular identity, proliferation, differentiation and apoptosis via the developmental processes of lateral inhibition and boundary induction. In light of this essential role played in metazoan development, we surveyed a wide range of eukaryotic genomes to determine the origin and evolution of the components and auxiliary factors that compose and modulate this pathway. Results We searched for 22 components of the Notch pathway in 35 different species that represent 8 major clades of eukaryotes, performed phylogenetic analyses and compared the domain compositions of the two fundamental molecules: the receptor Notch and its ligands Delta/Jagged. We confirm that a Notch pathway, with true receptors and ligands is specific to the Metazoa. This study also sheds light on the deep ancestry of a number of genes involved in this pathway, while other members are revealed to have a more recent origin. The origin of several components can be accounted for by the shuffling of pre-existing protein domains, or via lateral gene transfer. In addition, certain domains have appeared de novo more recently, and can be considered metazoan synapomorphies. Conclusion The Notch signalling pathway emerged in Metazoa via a diversity of molecular mechanisms, incorporating both novel and ancient protein domains during eukaryote evolution. Thus, a functional Notch signalling pathway was probably present in Urmetazoa. PMID:19825158

  13. Radiation driven epithelial-mesenchymal transition is mediated by Notch signaling in breast cancer

    PubMed Central

    Suh, Yongjoon; Yoo, Ki-Chun; Cui, Yan-Hong; Kim, Min-Jung; Lee, Hae-June; Kim, In-Gyu; Lee, Su-Jae

    2016-01-01

    Epithelial to mesenchymal transition (EMT) is developmental process associated with cancer metastasis. Here, we found that breast carcinoma cells adopt epithelial-to-mesenchymal transition (EMT) in response to fractionated-radiation. Importantly, we show that Notch signaling is highly activated in fractionally-irradiated tumors as compared to non-irradiated tumors that are accompanied by an EMT. Moreover, we uncovered the mechanism of Notch-driven EMT, in which Notch enhanced EMT through IL-6/JAK/STAT3 signaling axis in mammary tumor cells. Collectively, we present converging evidence from our studies that Notch2 is a critical mediator of radiation-induced EMT and responsible for induced malignant tumor growth. PMID:27462787

  14. BMP signaling orchestrates photoreceptor specification in the zebrafish pineal gland in collaboration with Notch.

    PubMed

    Quillien, Aurélie; Blanco-Sanchez, Bernardo; Halluin, Caroline; Moore, John C; Lawson, Nathan D; Blader, Patrick; Cau, Elise

    2011-06-01

    A variety of signaling pathways have been shown to regulate specification of neuronal subtype identity. However, the mechanisms by which future neurons simultaneously process information from multiple pathways to establish their identity remain poorly understood. The zebrafish pineal gland offers a simple system with which to address questions concerning the integration of signaling pathways during neural specification as it contains only two types of neurons - photoreceptors and projection neurons. We have previously shown that Notch signaling inhibits the projection neuron fate. Here, we show that BMP signaling is both necessary and sufficient to promote the photoreceptor fate. We also demonstrate that crosstalk between BMP and Notch signaling is required for the inhibition of a projection neuron fate in future photoreceptors. In this case, BMP signaling is required as a competence factor for the efficient activation of Notch targets. Our results indicate that both the induction of a photoreceptor fate and the interaction with Notch relies on a canonical BMP/Smad5 pathway. However, the activation of Notch-dependent transcription does not require a canonical Smad5-DNA interaction. Our results provide new insights into how multiple signaling influences are integrated during cell fate specification in the vertebrate CNS.

  15. Proliferation-independent regulation of organ size by Fgf/Notch signaling

    PubMed Central

    Kozlovskaja-Gumbrienė, Agnė; Yi, Ren; Alexander, Richard; Aman, Andy; Jiskra, Ryan; Nagelberg, Danielle; Knaut, Holger; McClain, Melainia; Piotrowski, Tatjana

    2017-01-01

    Organ morphogenesis depends on the precise orchestration of cell migration, cell shape changes and cell adhesion. We demonstrate that Notch signaling is an integral part of the Wnt and Fgf signaling feedback loop coordinating cell migration and the self-organization of rosette-shaped sensory organs in the zebrafish lateral line system. We show that Notch signaling acts downstream of Fgf signaling to not only inhibit hair cell differentiation but also to induce and maintain stable epithelial rosettes. Ectopic Notch expression causes a significant increase in organ size independently of proliferation and the Hippo pathway. Transplantation and RNASeq analyses revealed that Notch signaling induces apical junctional complex genes that regulate cell adhesion and apical constriction. Our analysis also demonstrates that in the absence of patterning cues normally provided by a Wnt/Fgf signaling system, rosettes still self-organize in the presence of Notch signaling. DOI: http://dx.doi.org/10.7554/eLife.21049.001 PMID:28085667

  16. Synergistic association of Notch and NFκB signaling and role of Notch signaling in modulating epithelial to mesenchymal transition in colorectal adenocarcinoma.

    PubMed

    Gopalakrishnan, Natarajan; Sivasithamparam, Niranjali Devaraj; Devaraj, Halagowder

    2014-12-01

    Notch1 signaling plays a key role in normal developmental processes and in cancer. The association between Notch activation and development of cancer has been well documented. Notch activation and outcome of the disease depend upon the crosstalk with other regulatory pathways including Nuclear Factor kappa B (NFκB) pathway. In this study, we have investigated the interaction of Notch intracellular domain (NICD) with NFκBp65 in colorectal cancer which resulted in the upregulation of Bcl-xL resulting in the inhibition of apoptosis. Mesenchymal marker Slug expression and down regulation of E-cadherin, an epithelial phenotypic marker were demonstrated in colon cancer tissues. The study was also illustrated by using the gamma secretase inhibitor, N-[N-(3,5-difluorophenacetyl)-L-alanyl]-S-phenylglycine t-butyl ester (DAPT) in HT29 cells. Immunohistochemistry (NICD, NFκBp65, and Slug) and double immunofluorescence analysis (NICD, NFκBp65) revealed that NICD and NFκBp65 were highly expressed in HT29 cells and in tumor tissue compared to normal tissue. Slug and Bcl-xL protein expressions were significantly reduced in DAPT treated HT 29 cells. Immunoprecipitation and dual staining emphasized the strong interaction of NICD with NFκBp65 in adenocarcinoma than in normal tissue. It appeared that Notch1 and NFκB could independently contribute to tumor progression. However, their interaction and synergism might be the determinants that would affect the outcome of the disease and therapeutic interventions.

  17. Split ends antagonizes the Notch and potentiates the EGFR signaling pathways during Drosophila eye development

    PubMed Central

    Doroquez, David B.; Orr-Weaver, Terry L.; Rebay, Ilaria

    2007-01-01

    The Notch and Epidermal Growth Factor Receptor (EGFR) signaling pathways interact cooperatively and antagonistically to regulate many aspects of Drosophila development, including the eye. How output from these two signaling networks is fine-tuned to achieve the precise balance needed for specific inductive interactions and patterning events remains an open and important question. Previously, we reported that the gene split ends (spen) functions within or parallel to the EGFR pathway during midline glial cell development in the embryonic central nervous system. Here, we report that the cellular defects caused by loss of spen function in the developing eye imaginal disc place spen as both an antagonist of the Notch pathway and a positive contributor to EGFR signaling during retinal cell differentiation. Specifically, loss of spen results in broadened expression of Scabrous, ectopic activation of Notch signaling, and a corresponding reduction in Atonal expression at the morphogenetic furrow. Consistent with Spen’s role in antagonizing Notch signaling, reduction of spen levels is sufficient to suppress Notch-dependent phenotypes. At least in part due to loss of Spen-dependent down-regulation of Notch signaling, loss of spen also dampens EGFR signaling as evidenced by reduced activity of MAP kinase (MAPK). This reduced MAPK activity in turn leads to a failure to limit expression of the EGFR pathway antagonist and the ETS-domain transcriptional repressor Yan and to a corresponding loss of cell fate specification in spen mutant ommatidia. We propose that Spen plays a role in modulating output from the Notch and EGFR pathways to ensure appropriate patterning during eye development. PMID:17588724

  18. Activation of the Notch1/STAT3/Twist signaling axis promotes gastric cancer progression.

    PubMed

    Hsu, Kai-Wen; Hsieh, Rong-Hong; Huang, Kuo-Hung; Fen-Yau Li, Anna; Chi, Chin-Wen; Wang, Tzu-Yin; Tseng, Min-Jen; Wu, Kou-Juey; Yeh, Tien-Shun

    2012-08-01

    Gastric carcinoma is one of the most common malignancies and a lethal cancer in the world. Notch signaling and transcription factors STAT3 (signal transducer and activator of transcription 3) and Twist regulate tumor development and are critical regulators of gastric cancer progression. Herein, the relationship among Notch, STAT3 and Twist pathways in the control of gastric cancer progression was studied. We found that Twist and phosphorylated STAT3 levels were promoted by the activated Notch1 receptor in human stomach adenocarcinoma SC-M1, embryonic kidney HEK293 and erythroleukemia K562 cells. Notch1 signaling dramatically induced Twist promoter activity through a C promoter binding factor-1-independent manner and STAT3 phosphorylation. Overexpression of Notch1 receptor intracellular domain (N1IC) enhanced the interaction between nuclear STAT3 and Twist promoter in cells. Gastric cancer progression of SC-M1 cells was promoted by N1IC through STAT3 phosphorylation and Twist expression including colony formation, migration and invasion. STAT3 regulated gastric cancer progression of SC-M1 cells via Twist. N1IC also elevated the progression of other gastric cancer cells such as AGS and KATO III cells through STAT3 and Twist. The N1IC-promoted tumor growth and lung metastasis of SC-M1 cells in mice were suppressed by the STAT3 inhibitor JSI-124 and Twist knockdown. Furthermore, Notch1 and Notch ligand Jagged1 expressions were significantly associated with phosphorylated STAT3 and Twist levels in gastric cancer tissues of patients. Taken together, these results suggest that Notch1/STAT3/Twist signaling axis is involved in progression of human gastric cancer and modulation of this cascade has potential for the targeted combination therapy.

  19. Notch Signaling and Schwann Cell Transformation: Development of a Model System and Application to Human MPNSTs

    DTIC Science & Technology

    2008-09-01

    TITLE: Notch Signaling and Schwann Cell Transformation: Development of a Model System and Application to Human MPNSTs PRINCIPAL INVESTIGATOR...Schwann cell transformation: Development of a model system and 5a. CONTRACT NUMBER application to human MPNSTs . 5b. GRANT NUMBER W81XWH-04-1-0209...of neurofibromas to MPNSTs in patients with NF1. Our previous work has shown that constitutive expression of Notch can transform rat Schwann cells

  20. Investigation of deregulated genes of Notch signaling pathway in human T cell acute lymphoblastic leukemia cell lines and clinical samples.

    PubMed

    Paryan, Mahdi; Mohammadi-Yeganeh, Samira; Samiee, Siamak Mirab; Soleimani, Masoud; Arefian, Ehsan; Azadmanesh, Keyhan; Poopak, Behzad; Mostafavi, Ehsan; Karimipoor, Morteza; Mahdian, Reza

    2013-10-01

    In diagnostic research challenges, quantitative real-time PCR (QPCR) has been widely utilized in gene expression analysis because of its sensitivity, accuracy, reproducibility, and most importantly, quantitativeness. Real-time PCR base kits are wildly applicable in cancer signaling pathways, especially in cancer investigations. T-cell acute lymphoblastic leukemia (T-ALL) is a type of leukemia that is more common in older children and teenagers. Deregulation of the Notch signaling pathway promotes proliferation and inhibits apoptosis of the lymphoblastic T cells. The aim of this study was to investigate the effect of Notch signaling activation on the expression of target genes using real-time QPCR and further use this method in clinical examination after validation. Two T-ALL cell lines, Jurkat and Molt-4, were used as models for activation of the Notch signaling via over-expression of the Notch1 intracellular domain. Expression analysis was performed for six downstream target genes (NCSTN, APH1, PSEN1, ADAM17, NOTCH1 and C-MYC) which play critical roles in the Notch signaling pathway. The results showed significant difference in the expression of target genes in the deregulated Notch signaling pathway. These results were also verified in 12 clinical samples bearing over-expression of the Notch signaling pathway. Identification of such downstream Notch target genes, which have not been studied inclusively, provides insights into the mechanisms of the Notch function in T cell leukemia, and may help identify novel diagnoses and therapeutic targets in acute lymphoblastic leukemia.

  1. The matricellular protein CCN1 controls retinal angiogenesis by targeting VEGF, Src homology 2 domain phosphatase-1 and Notch signaling.

    PubMed

    Chintala, Hemabindu; Krupska, Izabela; Yan, Lulu; Lau, Lester; Grant, Maria; Chaqour, Brahim

    2015-07-01

    Physiological angiogenesis depends on the highly coordinated actions of multiple angiogenic regulators. CCN1 is a secreted cysteine-rich and integrin-binding matricellular protein required for proper cardiovascular development. However, our understanding of the cellular origins and activities of this molecule is incomplete. Here, we show that CCN1 is predominantly expressed in angiogenic endothelial cells (ECs) at the leading front of actively growing vessels in the mouse retina. Endothelial deletion of CCN1 in mice using a Cre-Lox system is associated with EC hyperplasia, loss of pericyte coverage and formation of dense retinal vascular networks lacking the normal hierarchical arrangement of arterioles, capillaries and venules. CCN1 is a product of an immediate-early gene that is transcriptionally induced in ECs in response to stimulation by vascular endothelial growth factor (VEGF). We found that CCN1 activity is integrated with VEGF receptor 2 (VEGF-R2) activation and downstream signaling pathways required for tubular network formation. CCN1-integrin binding increased the expression of and association between Src homology 2 domain-containing protein tyrosine phosphatase-1 (SHP-1) and VEGF-R2, which leads to rapid dephosphorylation of VEGF-R2 tyrosine, thus preventing EC hyperproliferation. Predictably, CCN1 further brings receptors/signaling molecules into proximity that are otherwise spatially separated. Furthermore, CCN1 induces integrin-dependent Notch activation in cultured ECs, and its targeted gene inactivation in vivo alters Notch-dependent vascular specification and remodeling, suggesting that functional levels of Notch signaling requires CCN1 activity. These data highlight novel functions of CCN1 as a naturally optimized molecule, fine-controlling key processes in physiological angiogenesis and safeguarding against aberrant angiogenic responses.

  2. Jagged-Notch signaling ensures dorsal skeletal identity in the vertebrate face

    PubMed Central

    Zuniga, Elizabeth; Stellabotte, Frank; Crump, J. Gage

    2010-01-01

    The development of the vertebrate face relies on the regionalization of neural crest-derived skeletal precursors along the dorsoventral (DV) axis. Here we show that Jagged-Notch signaling ensures dorsal identity within the hyoid and mandibular components of the facial skeleton by repressing ventral fates. In a genetic screen in zebrafish, we identified a loss-of-function mutation in jagged 1b (jag1b) that results in dorsal expansion of ventral gene expression and partial transformation of the dorsal hyoid skeleton to a ventral morphology. Conversely, misexpression of human jagged 1 (JAG1) represses ventral gene expression and dorsalizes the ventral hyoid and mandibular skeletons. We further show that jag1b is expressed specifically in dorsal skeletal precursors, where it acts through the Notch2 receptor to activate hey1 expression. Whereas Jagged-Notch positive feedback propagates jag1b expression throughout the dorsal domain, Endothelin 1 (Edn1) inhibits jag1b and hey1 expression in the ventral domain. Strikingly, reduction of Jag1b or Notch2 function partially rescues the ventral defects of edn1 mutants, indicating that Edn1 promotes facial skeleton development in part by inhibiting Jagged-Notch signaling in ventral skeletal precursors. Together, these results indicate a novel function of Jagged-Notch signaling in ensuring dorsal identity within broad fields of facial skeletal precursors. PMID:20431122

  3. PKCθ links proximal T cell and Notch signaling through localized regulation of the actin cytoskeleton

    PubMed Central

    Britton, Graham J; Ambler, Rachel; Clark, Danielle J; Hill, Elaine V; Tunbridge, Helen M; McNally, Kerrie E; Burton, Bronwen R; Butterweck, Philomena; Sabatos-Peyton, Catherine; Hampton-O’Neil, Lea A; Verkade, Paul; Wuelfing, Christoph; Wraith, David Cameron

    2017-01-01

    Notch is a critical regulator of T cell differentiation and is activated through proteolytic cleavage in response to ligand engagement. Using murine myelin-reactive CD4 T cells, we demonstrate that proximal T cell signaling modulates Notch activation by a spatiotemporally constrained mechanism. The protein kinase PKCθ is a critical mediator of signaling by the T cell antigen receptor and the principal costimulatory receptor CD28. PKCθ selectively inactivates the negative regulator of F-actin generation, Coronin 1A, at the center of the T cell interface with the antigen presenting cell (APC). This allows for effective generation of the large actin-based lamellum required for recruitment of the Notch-processing membrane metalloproteinase ADAM10. Such enhancement of Notch activation is critical for efficient T cell proliferation and Th17 differentiation. We reveal a novel mechanism that, through modulation of the cytoskeleton, controls Notch activation at the T cell:APC interface thereby linking T cell receptor and Notch signaling pathways. DOI: http://dx.doi.org/10.7554/eLife.20003.001 PMID:28112644

  4. Notch signaling maintains bone marrow mesenchymal progenitors by suppressing osteoblast differentiation

    PubMed Central

    Hilton, Matthew J.; Tu, Xiaolin; Wu, Ximei; Bai, Shuting; Zhao, Haibo; Kobayashi, Tatsuya; Kronenberg, Henry M.; Teitelbaum, Steven L.; Ross, F. Patrick; Kopan, Raphael; Long, Fanxin

    2009-01-01

    Postnatal bone marrow houses mesenchymal progenitor cells that are osteoblast precursors. These cells have established therapeutic potential 1 but they are difficult to maintain and expand in vitro, presumably because little is known about the mechanisms controlling their fate decisions. To investigate the potential role of Notch signaling in osteoblastogenesis, we used conditional alleles to genetically remove components of the Notch signaling system during skeletal development. We find that Notch disruption in the limb skeletogenic mesenchyme markedly enhanced trabecular bone mass in adolescent mice. Notably, mesenchymal progenitors were virtually depleted in the bone marrow of the high-bone-mass animals. As a result, these animals developed severe osteopenia as they aged. Moreover, Notch appeared to inhibit osteoblast differentiation through Hes/Hey proteins that diminished Runx2 transcriptional activity via physical interaction. These results support a model wherein Notch signaling in bone marrow normally acts to maintain a pool of mesenchymal progenitors by suppressing osteoblast differentiation. Thus, mesechymal progenitors may be expanded in vitro by activating Notch, whereas bone formation in vivo may be enhanced by transiently suppressing this pathway. PMID:18297083

  5. Melatonin Inhibits Glioblastoma Stem-like cells through Suppression of EZH2-NOTCH1 Signaling Axis

    PubMed Central

    Zheng, Xiangrong; Pang, Bo; Gu, Guangyan; Gao, Taihong; Zhang, Rui; Pang, Qi; Liu, Qian

    2017-01-01

    Glioblastoma stem-like cells (GSCs) play essential roles in glioma growth, radio- and chemo-resistance, and recurrence. Elimination of GSCs has therefore become a key strategy and challenge in glioblastoma therapy. Here, we show that melatonin, an indolamine derived from I-tryptophan, significantly inhibited viability and self-renewal ability of GSCs accompanied by a decrease of stem cell markers. We have identified EZH2-NOTCH1 signaling as the key signal pathway that regulated the effects of melatonin in the GSCs. Instead of transcriptionally silencing gene expression by generating a methylated epigenetic mark at histone 3 at lysine 27 (H3K27), EZH2 regulates NOTCH1 expression by directly binding to the NOTCH1 promoter. Moreover, correlation between the expressions of EZH2 and NOTCH intracellular domain 1 (NICD1) was observed in the clinical tumor samples, evidently supporting the existence of EZH2-NOTCH1 interaction in the gliomas and GSCs. Collectively, we demonstrated that melatonin, a potential tumor inhibitor, performs its function partly by suppressing GSC properties through EZH2-NOTCH1 signaling axis. PMID:28255276

  6. Interacting network of Hippo, Wnt/β-catenin and Notch signaling represses liver tumor formation

    PubMed Central

    Kim, Wantae; Khan, Sanjoy Kumar; Yang, Yingzi

    2017-01-01

    Acquiring a selective growth advantage by breaking the proliferation barrier established by gatekeeper genes is a centrally important event in tumor formation. Removal of the mammalian Hippo kinase Mst1 and Mst2 in hepatocytes leads to rapid hepatocellular carcinoma (HCC) formation, indicating that the Hippo signaling pathway is a critical gatekeeper that restrains abnormal growth in hepatocytes. By rigorous genetic approaches, we identified an interacting network of the Hippo, Wnt/β-catenin and Notch signaling pathways that control organ size and HCC development. We found that in hepatocytes, the loss of Mst1/2 leads to the activation of Notch signaling, which forms a positive feedback loop with Yap/Taz (transcription factors controlled by Mst1/2). This positive feedback loop results in severe liver enlargement and rapid HCC formation. Blocking the Yap/Taz-Notch positive feedback loop by Notch inhibition in vivo significantly reduced the Yap/Taz activities, hepatocyte proliferation and tumor formation. Furthermore, we uncovered a surprising inhibitory role of Wnt/β-catenin signaling to Yap/Taz activities, which are important in tumor initiation. Genetic removal of β-catenin in the liver of the Mst1/2 mutants significantly accelerates tumoriogenesis. Therefore, Wnt/β-catenin signaling, known for its oncogenic property, exerts an unexpected function in restricting Yap/Taz and Notch activities in HCC initiation. The molecular interplay between the three signaling pathways identified in our study provides new insights in developing novel therapeutic strategies to treat liver tumors. PMID:27881216

  7. Notch signaling in the epididymal epithelium regulates sperm motility and is transferred at a distance within epididymosomes.

    PubMed

    Murta, D; Batista, M; Silva, E; Trindade, A; Henrique, D; Duarte, A; Lopes-da-Costa, L

    2016-03-01

    Spermatozoa undergo sequential maturation changes during their transit along the epididymis. These changes are modulated by the epididymal epithelium and require a finely tuned gene expression. The Notch cell signaling pathway is a major regulator of cell fate decisions in several tissues, including the testis. Here, we evaluated the transcription and expression patterns of Notch components (Notch1-3, Dll1, Dll4, and Jagged1) and effectors (Hes1-2 and Hes5) in the adult mouse epididymis, and evaluated the role of Notch signaling in the epididymis through its in vivo blockade following administration of an inhibitor (DAPT). Notch components and effectors were dynamically transcribed and expressed in the epididymis and vas deferens, each segment exhibiting a specific combination of epithelial receptor/ligand/effector expression patterns. Nuclear detection of Notch effectors indicates that Notch signaling was active. Notch components (but not effectors) were identified in the cytoplasmic droplet of spermatozoa, in a dynamic and specific pattern along the epididymis. In addition, Notch components were identified within large and small vesicles in the epididymal lumen. A purified population of these membranous vesicles from different epididymal segments was obtained, and through dot blot analysis, it was confirmed that Notch components were carried within these vesicles in a dynamic pattern along the epididymal lumen. We hypothesize that these vesicles (epididymosomes) allow Notch signaling at distance from epididymal epithelial cells to spermatozoa. DAPT-induced in vivo Notch signaling blockade, although showing a low efficiency, disrupted the expression patterns of Notch components and effectors in the epididymal epithelium and in spermatozoa, and significantly decreased sperm motility, although not affecting male fertility. These results prompt for a regulatory role of Notch signaling in epididymal epithelial function and sperm maturation.

  8. Activity-dependent Notch signalling in the hypothalamic-neurohypophysial system of adult mouse brains.

    PubMed

    Mannari, T; Miyata, S

    2014-08-01

    Notch signalling has a key role in cell fate specification in developing brains; however, recent studies have shown that Notch signalling also participates in the regulation of synaptic plasticity in adult brains. In the present study, we examined the expression of Notch3 and Delta-like ligand 4 (DLL4) in the hypothalamic-neurohypophysial system (HNS) of the adult mouse. The expression of DLL4 was higher in the supraoptic nucleus (SON) and paraventricular nucleus (PVN) compared to adjacent hypothalamic regions. Double-labelling immunohistochemistry using vesicular GABA transporter and glutamate transporter revealed that DLL4 was localised at a subpopulation of excitatory and inhibitory axonal boutons against somatodendrites of arginine vasopressin (AVP)- and oxytocin (OXT)-containing magnocellular neurones. In the neurohypophysis (NH), the expression of DLL4 was seen at OXT- but not AVP-containing axonal terminals. The expression of Notch3 was seen at somatodendrites of AVP- and OXT-containing magnocellular neurones in the SON and PVN and at pituicytes in the NH. Chronic physiological stimulation by salt loading, which remarkably enhances the release of AVP and OXT, decreased the number of DLL4-immunoreactive axonal boutons in the SON and PVN. Moreover, chronic and acute osmotic stimulation promoted proteolytic cleavage of Notch3 to yield the intracellular fragments of Notch3 in the HNS. Thus, the present study demonstrates activity-dependent reduction of DLL4 expression and proteolytic cleavage of Notch3 in the HNS, suggesting that Notch signalling possibly participates in synaptic interaction in the hypothalamic nuclei and neuroglial interaction in the NH.

  9. Ehrlichia chaffeensis TRP120 Activates Canonical Notch Signaling To Downregulate TLR2/4 Expression and Promote Intracellular Survival

    PubMed Central

    Lina, Taslima T.; Dunphy, Paige S.; Luo, Tian

    2016-01-01

    ABSTRACT Ehrlichia chaffeensis preferentially targets mononuclear phagocytes and survives through a strategy of subverting innate immune defenses, but the mechanisms are unknown. We have shown E. chaffeensis type 1 secreted tandem repeat protein (TRP) effectors are involved in diverse molecular pathogen-host interactions, such as the TRP120 interaction with the Notch receptor-cleaving metalloprotease ADAM17. In the present study, we demonstrate E. chaffeensis, via the TRP120 effector, activates the canonical Notch signaling pathway to promote intracellular survival. We found that nuclear translocation of the transcriptionally active Notch intracellular domain (NICD) occurs in response to E. chaffeensis or recombinant TRP120, resulting in upregulation of Notch signaling pathway components and target genes notch1, adam17, hes, and hey. Significant differences in canonical Notch signaling gene expression levels (>40%) were observed during early and late stages of infection, indicating activation of the Notch pathway. We linked Notch pathway activation specifically to the TRP120 effector, which directly interacts with the Notch metalloprotease ADAM17. Using pharmacological inhibitors and small interfering RNAs (siRNAs) against γ-secretase enzyme, Notch transcription factor complex, Notch1, and ADAM17, we demonstrated that Notch signaling is required for ehrlichial survival. We studied the downstream effects and found that E. chaffeensis TRP120-mediated activation of the Notch pathway causes inhibition of the extracellular signal-regulated kinase 1/2 (ERK1/2) and p38 mitogen-activated protein kinase (MAPK) pathways required for PU.1 and subsequent Toll-like receptor 2/4 (TLR2/4) expression. This investigation reveals a novel mechanism whereby E. chaffeensis exploits the Notch pathway to evade the host innate immune response for intracellular survival. PMID:27381289

  10. Notch1-STAT3-ETBR signaling axis controls reactive astrocyte proliferation after brain injury.

    PubMed

    LeComte, Matthew D; Shimada, Issei S; Sherwin, Casey; Spees, Jeffrey L

    2015-07-14

    Defining the signaling network that controls reactive astrogliosis may provide novel treatment targets for patients with diverse CNS injuries and pathologies. We report that the radial glial cell antigen RC2 identifies the majority of proliferating glial fibrillary acidic protein-positive (GFAP(+)) reactive astrocytes after stroke. These cells highly expressed endothelin receptor type B (ETB(R)) and Jagged1, a Notch1 receptor ligand. To study signaling in adult reactive astrocytes, we developed a model based on reactive astrocyte-derived neural stem cells isolated from GFAP-CreER-Notch1 conditional knockout (cKO) mice. By loss- and gain-of-function studies and promoter activity assays, we found that Jagged1/Notch1 signaling increased ETB(R) expression indirectly by raising the level of phosphorylated signal transducer and activator of transcription 3 (STAT3), a previously unidentified EDNRB transcriptional activator. Similar to inducible transgenic GFAP-CreER-Notch1-cKO mice, GFAP-CreER-ETB(R)-cKO mice exhibited a defect in reactive astrocyte proliferation after cerebral ischemia. Our results indicate that the Notch1-STAT3-ETB(R) axis connects a signaling network that promotes reactive astrocyte proliferation after brain injury.

  11. Leptin-Notch signaling axis is involved in pancreatic cancer progression.

    PubMed

    Harbuzariu, Adriana; Rampoldi, Antonio; Daley-Brown, Danielle S; Candelaria, Pierre; Harmon, Tia L; Lipsey, Crystal C; Beech, Derrick J; Quarshie, Alexander; Ilies, Gabriela Oprea; Gonzalez-Perez, Ruben R

    2017-01-31

    Pancreatic cancer (PC) shows a high death rate. PC incidence and prognosis are affected by obesity, a pandemic characterized by high levels of leptin. Notch is upregulated by leptin in breast cancer. Thus, leptin and Notch crosstalk could influence PC progression. Here we investigated in PC cell lines (BxPC-3, MiaPaCa-2, Panc-1, AsPC-1), derived tumorspheres and xenografts whether a functional leptin-Notch axis affects PC progression and expansion of pancreatic cancer stem cells (PCSC). PC cells and tumorspheres were treated with leptin and inhibitors of Notch (gamma-secretase inhibitor, DAPT) and leptin (iron oxide nanoparticle-leptin peptide receptor antagonist 2, IONP-LPrA2). Leptin treatment increased cell cycle progression and proliferation, and the expression of Notch receptors, ligands and targeted molecules (Notch1-4, DLL4, JAG1, Survivin and Hey2), PCSC markers (CD24/CD44/ESA, ALDH, CD133, Oct-4), ABCB1 protein, as well as tumorsphere formation. Leptin-induced effects on PC and tumorspheres were decreased by IONP-LPrA2 and DAPT. PC cells secreted leptin and expressed the leptin receptor, OB-R, which indicates a leptin autocrine/paracrine signaling loop could also affect tumor progression. IONP-LPrA2 treatment delayed the onset of MiaPaCa-2 xenografts, and decreased tumor growth and the expression of proliferation and PCSC markers. Present data suggest that leptin-Notch axis is involved in PC. PC has no targeted therapy and is mainly treated with chemotherapy, whose efficiency could be decreased by leptin and Notch activities. Thus, the leptin-Notch axis could be a novel therapeutic target, particularly for obese PC patients.

  12. Notch signaling mediates the age-associated decrease in adhesion of germline stem cells to the niche.

    PubMed

    Tseng, Chen-Yuan; Kao, Shih-Han; Wan, Chih-Ling; Cho, Yueh; Tung, Shu-Yun; Hsu, Hwei-Jan

    2014-12-01

    Stem cells have an innate ability to occupy their stem cell niche, which in turn, is optimized to house stem cells. Organ aging is associated with reduced stem cell occupancy in the niche, but the mechanisms involved are poorly understood. Here, we report that Notch signaling is increased with age in Drosophila female germline stem cells (GSCs), and this results in their removal from the niche. Clonal analysis revealed that GSCs with low levels of Notch signaling exhibit increased adhesiveness to the niche, thereby out-competing their neighbors with higher levels of Notch; adhesiveness is altered through regulation of E-cadherin expression. Experimental enhancement of Notch signaling in GSCs hastens their age-dependent loss from the niche, and such loss is at least partially mediated by Sex lethal. However, disruption of Notch signaling in GSCs does not delay GSC loss during aging, and nor does it affect BMP signaling, which promotes self-renewal of GSCs. Finally, we show that in contrast to GSCs, Notch activation in the niche (which maintains niche integrity, and thus mediates GSC retention) is reduced with age, indicating that Notch signaling regulates GSC niche occupancy both intrinsically and extrinsically. Our findings expose a novel role of Notch signaling in controlling GSC-niche adhesion in response to aging, and are also of relevance to metastatic cancer cells, in which Notch signaling suppresses cell adhesion.

  13. Divergent effects of supraphysiologic Notch signals on leukemia stem cells and hematopoietic stem cells.

    PubMed

    Chiang, Mark Y; Shestova, Olga; Xu, Lanwei; Aster, Jon C; Pear, Warren S

    2013-02-07

    The leukemia stem cell (LSC) hypothesis proposes that a subset of cells in the bulk leukemia population propagates the leukemia.We tested the LSC hypothesis in a mouse model of Notch-induced T-cell acute lymphoblastic leukemia (T-ALL) in which the tumor cells were largely CD4+ CD8+ T cells. LSC activity was enriched but rare in the CD8+ CD4 HSA(hi) immature single-positive T-cell subset. Although our murine T-ALL model relies on transduction of HSCs, we were unable to isolate Notch-activated HSCs to test for LSC activity. Further analysis showed that Notch activation in HSCs caused an initial expansion of hematopoietic and T-cell progenitors and loss of stem cell quiescence, which was followed by progressive loss of long-term HSCs and T-cell production over several weeks. Similar results were obtained in a conditional transgenic model in which Notch activation is induced in HSCs by Cre recombinase. We conclude that although supraphysiologic Notch signaling in HSCs promotes LSC activity in T-cell progenitors, it extinguishes self-renewal of LT-HSCs. These results provide further evidence for therapeutically targeting T-cell progenitors in T-ALL while also underscoring the need to tightly regulate Notch signaling to expand normal HSC populations for clinical applications.

  14. Family Based Whole Exome Sequencing Reveals the Multifaceted Role of Notch Signaling in Congenital Heart Disease

    PubMed Central

    Chetaille, Philippe; Prince, Andrea; Godard, Beatrice; Leclerc, Severine; Sobreira, Nara; Ling, Hua; Awadalla, Philip; Thibeault, Maryse; Khairy, Paul; Samuels, Mark E.; Andelfinger, Gregor

    2016-01-01

    Left-ventricular outflow tract obstructions (LVOTO) encompass a wide spectrum of phenotypically heterogeneous heart malformations which frequently cluster in families. We performed family based whole-exome and targeted re-sequencing on 182 individuals from 51 families with multiple affected members. Central to our approach is the family unit which serves as a reference to identify causal genotype-phenotype correlations. Screening a multitude of 10 overlapping phenotypes revealed disease associated and co-segregating variants in 12 families. These rare or novel protein altering mutations cluster predominantly in genes (NOTCH1, ARHGAP31, MAML1, SMARCA4, JARID2, JAG1) along the Notch signaling cascade. This is in line with a significant enrichment (Wilcoxon, p< 0.05) of variants with a higher pathogenicity in the Notch signaling pathway in patients compared to controls. The significant enrichment of novel protein truncating and missense mutations in NOTCH1 highlights the allelic and phenotypic heterogeneity in our pediatric cohort. We identified novel co-segregating pathogenic mutations in NOTCH1 associated with left and right-sided cardiac malformations in three independent families with a total of 15 affected individuals. In summary, our results suggest that a small but highly pathogenic fraction of family specific mutations along the Notch cascade are a common cause of LVOTO. PMID:27760138

  15. Corilagin suppresses cholangiocarcinoma progression through Notch signaling pathway in vitro and in vivo

    PubMed Central

    GU, YUE; XIAO, LINFENG; MING, YANLIN; ZHENG, ZHIZHONG; LI, WENGANG

    2016-01-01

    Corilagin is a natural plant polyphenol tannic acid with antitumor, anti-inflammatory, and anti-oxidative properties. However, the mechanisms of its actions are largely unknown. Our group reported that corilagin could induce cell inhibition in human breast cancer cell line MCF-7 and human liver hepatocellular carcinoma cell lines HepG2. We report here that corilagin inhibits cholangiocarcinoma (CCA) development through regulating Notch signaling pathway. We found that, in vitro, corilagin inhibited CCA cell proliferation, migration and invasion, promoted CCA cell apoptosis, and inhibited Notch1 and Notch signaling pathway protein expression. Co-immunoprecipitation was used to establish Notch intracellular domain (NICD) interaction with MAML1 and P300 in CCA. Importantly, corilagin reduced Hes1 mRNA level through inhibiting Hes1 promoter activity. In nude mice, corilagin inhibited CCA growth and repressed the expression of Notch1 and mTOR. These results indicate that corilagin may control CCA cell growth by downregulating the expression of Notch1. Therefore, our findings suggest that corilagin may have the potential to become a new therapeutic drug for human CCA. PMID:26935808

  16. Presenilin-mediated transmembrane cleavage is required for Notch signal transduction in Drosophila

    PubMed Central

    Struhl, Gary; Greenwald, Iva

    2001-01-01

    The cleavage model for signal transduction by receptors of the LIN-12/Notch family posits that ligand binding leads to cleavage within the transmembrane domain, so that the intracellular domain is released to translocate to the nucleus and activate target gene expression. The familial Alzheimer's disease-associated protein Presenilin is required for LIN-12/Notch signaling, and several lines of evidence suggest that Presenilin mediates the transmembrane cleavage event that releases the LIN-12/Notch intracellular domain. However, doubt was cast on this possibility by a report that Presenilin is not required for the transducing activity of NECN, a constitutively active transmembrane form of Notch, in Drosophila. Here, we have reassessed this finding and show instead that Presenilin is required for activity of NECN for all cell fate decisions examined. Our results indicate that transmembrane cleavage and signal transduction are strictly correlated, supporting the cleavage model for signal transduction by LIN-12/Notch and a role for Presenilin in mediating the ligand-induced transmembrane cleavage. PMID:11134525

  17. Protein Tyrosine Phosphatase PRL2 Mediates Notch and Kit Signals in Early T Cell Progenitors.

    PubMed

    Kobayashi, Michihiro; Nabinger, Sarah C; Bai, Yunpeng; Yoshimoto, Momoko; Gao, Rui; Chen, Sisi; Yao, Chonghua; Dong, Yuanshu; Zhang, Lujuan; Rodriguez, Sonia; Yashiro-Ohtani, Yumi; Pear, Warren S; Carlesso, Nadia; Yoder, Mervin C; Kapur, Reuben; Kaplan, Mark H; Daniel Lacorazza, Hugo; Zhang, Zhong-Yin; Liu, Yan

    2017-04-01

    The molecular pathways regulating lymphoid priming, fate, and development of multipotent bone marrow hematopoietic stem and progenitor cells (HSPCs) that continuously feed thymic progenitors remain largely unknown. While Notch signal is indispensable for T cell specification and differentiation, the downstream effectors are not well understood. PRL2, a protein tyrosine phosphatase that regulates hematopoietic stem cell proliferation and self-renewal, is highly expressed in murine thymocyte progenitors. Here we demonstrate that protein tyrosine phosphatase PRL2 and receptor tyrosine kinase c-Kit are critical downstream targets and effectors of the canonical Notch/RBPJ pathway in early T cell progenitors. While PRL2 deficiency resulted in moderate defects of thymopoiesis in the steady state, de novo generation of T cells from Prl2 null hematopoietic stem cells was significantly reduced following transplantation. Prl2 null HSPCs also showed impaired T cell differentiation in vitro. We found that Notch/RBPJ signaling upregulated PRL2 as well as c-Kit expression in T cell progenitors. Further, PRL2 sustains Notch-mediated c-Kit expression and enhances stem cell factor/c-Kit signaling in T cell progenitors, promoting effective DN1-DN2 transition. Thus, we have identified a critical role for PRL2 phosphatase in mediating Notch and c-Kit signals in early T cell progenitors. Stem Cells 2017;35:1053-1064.

  18. Notch and the awesome power of genetics.

    PubMed

    Greenwald, Iva

    2012-07-01

    Notch is a receptor that mediates cell-cell interactions in animal development, and aberrations in Notch signal transduction can cause cancer and other human diseases. Here, I describe the major advances in the Notch field from the identification of the first mutant in Drosophila almost a century ago through the elucidation of the unusual mechanism of signal transduction a little over a decade ago. As an essay for the GENETICS Perspectives series, it is my personal and critical commentary as well as an historical account of discovery.

  19. Protective effect of curcumin on acute airway inflammation of allergic asthma in mice through Notch1-GATA3 signaling pathway.

    PubMed

    Chong, Lei; Zhang, Weixi; Nie, Ying; Yu, Gang; Liu, Liu; Lin, Li; Wen, Shunhang; Zhu, Lili; Li, Changchong

    2014-10-01

    Curcumin, a natural product derived from the plant Curcuma longa, has been found to have anti-inflammatory, antineoplastic and antifibrosis effects. It has been reported that curcumin attenuates allergic airway inflammation in mice through inhibiting NF-κB and its downstream transcription factor GATA3. It also has been proved the antineoplastic effect of curcumin through down-regulating Notch1 receptor and its downstream nuclear transcription factor NF-κB levels. In this study, we aimed to investigate the anti-inflammatory effect of curcumin on acute allergic asthma and its underlying mechanisms. 36 male BALB/c mice were randomly divided into four groups (normal, asthma, asthma+budesonide and asthma+curcumin groups). BALF (bronchoalveolar lavage fluid) and lung tissues were analyzed for airway inflammation and the expression of Notch1, Notch2, Notch3, Notch4 and the downstream transcription factor GATA3. Our findings showed that the levels of Notch1 and Notch2 receptors were up-regulated in asthma group, accompanied by the increased expression of GATA3. But the expression of Notch2 receptor was lower than Notch1 receptor. Curcumin pretreatment improved the airway inflammatory cells infiltration and reversed the increasing levels of Notch1/2 receptors and GATA3. Notch3 receptor was not expressed in all of the four groups. Notch4 receptor protein and mRNA expression level in the four groups had no significant differences. The results of the present study suggested that Notch1 and Notch2 receptor, major Notch1 receptor, played an important role in the development of allergic airway inflammation and the inhibition of Notch1-GATA3 signaling pathway by curcumin can prevent the development and deterioration of the allergic airway inflammation. This may be a possible therapeutic option of allergic asthma.

  20. Notch-HES1 signaling axis controls hemato-endothelial fate decisions of human embryonic and induced pluripotent stem cells.

    PubMed

    Lee, Jung Bok; Werbowetski-Ogilvie, Tamra E; Lee, Jong-Hee; McIntyre, Brendan A S; Schnerch, Angelique; Hong, Seok-Ho; Park, In-Hyun; Daley, George Q; Bernstein, Irwin D; Bhatia, Mickie

    2013-08-15

    Notch signaling regulates several cellular processes including cell fate decisions and proliferation in both invertebrates and mice. However, comparatively less is known about the role of Notch during early human development. Here, we examined the function of Notch signaling during hematopoietic lineage specification from human pluripotent stem cells of both embryonic and adult fibroblast origin. Using immobilized Notch ligands and small interfering RNA to Notch receptors we have demonstrated that Notch1, but not Notch2, activation induced hairy and enhancer of split 1 (HES1) expression and generation of committed hematopoietic progenitors. Using gain- and loss-of-function approaches, this was shown to be attributed to Notch-signaling regulation through HES1, which dictated cell fate decisions from bipotent precursors either to the endothelial or hematopoietic lineages at the clonal level. Our study reveals a previously unappreciated role for the Notch pathway during early human hematopoiesis, whereby Notch signaling via HES1 represents a toggle switch of hematopoietic vs endothelial fate specification.

  1. Rbpj-κ mediated Notch signaling plays a critical role in development of hypothalamic Kisspeptin neurons

    PubMed Central

    Biehl, Matthew J; Raetzman, Lori T

    2015-01-01

    The mammalian arcuate nucleus (ARC) houses neurons critical for energy homeostasis and sexual maturation. Proopiomelanocortin (POMC) and Neuropeptide Y (NPY) neurons function to balance energy intake and Kisspeptin neurons are critical for the onset of puberty and reproductive function. While the physiological roles of these neurons have been well established, their development remains unclear. We have previously shown that Notch signaling plays an important role in cell fate within the ARC of mice. Active Notch signaling prevented neural progenitors from differentiating into feeding circuit neurons, whereas conditional loss of Notch signaling lead to a premature differentiation of these neurons. Presently, we hypothesized that Kisspeptin neurons would similarly be affected by Notch manipulation. To address this, we utilized mice with a conditional deletion of the Notch signaling co-factor Rbpj-κ (Rbpj cKO), or mice persistently expressing the Notch1 intracellular domain (NICD tg) within Nkx2.1 expressing cells of the developing hypothalamus. Interestingly, we found that in both models, a lack of Kisspeptin neurons are observed. This suggests that Notch signaling must be properly titrated for formation of Kisspeptin neurons. These results led us to hypothesize that Kisspeptin neurons of the ARC may arise from a different lineage of intermediate progenitors than NPY neurons and that Notch was responsible for the fate choice between these neurons. To determine if Kisspeptin neurons of the ARC differentiate similarly through a Pomc intermediate, we utilized a genetic model expressing the tdTomato fluorescent protein in all cells that have ever expressed Pomc. We observed some Kisspeptin expressing neurons labeled with the Pomc reporter similar to NPY neurons, suggesting that these distinct neurons can arise from a common progenitor. Finally, we hypothesized that temporal differences leading to premature depletion of progenitors in cKO mice lead to our observed

  2. Notch signaling deficiency underlies age-dependent depletion of satellite cells in muscular dystrophy.

    PubMed

    Jiang, Chunhui; Wen, Yefei; Kuroda, Kazuki; Hannon, Kevin; Rudnicki, Michael A; Kuang, Shihuan

    2014-08-01

    Duchenne muscular dystrophy (DMD) is a devastating disease characterized by muscle wasting, loss of mobility and death in early adulthood. Satellite cells are muscle-resident stem cells responsible for the repair and regeneration of damaged muscles. One pathological feature of DMD is the progressive depletion of satellite cells, leading to the failure of muscle repair. Here, we attempted to explore the molecular mechanisms underlying satellite cell ablation in the dystrophin mutant mdx mouse, a well-established model for DMD. Initial muscle degeneration activates satellite cells, resulting in increased satellite cell number in young mdx mice. This is followed by rapid loss of satellite cells with age due to the reduced self-renewal ability of mdx satellite cells. In addition, satellite cell composition is altered even in young mdx mice, with significant reductions in the abundance of non-committed (Pax7+ and Myf5-) satellite cells. Using a Notch-reporter mouse, we found that the mdx satellite cells have reduced activation of Notch signaling, which has been shown to be necessary to maintain satellite cell quiescence and self-renewal. Concomitantly, the expression of Notch1, Notch3, Jag1, Hey1 and HeyL are reduced in the mdx primary myoblast. Finally, we established a mouse model to constitutively activate Notch signaling in satellite cells, and show that Notch activation is sufficient to rescue the self-renewal deficiencies of mdx satellite cells. These results demonstrate that Notch signaling is essential for maintaining the satellite cell pool and that its deficiency leads to depletion of satellite cells in DMD.

  3. Anticancer effect of eupatilin on glioma cells through inhibition of the Notch-1 signaling pathway

    PubMed Central

    WANG, YAWEI; HOU, HONGWEI; LI, MING; YANG, YANG; SUN, LAN

    2016-01-01

    Eupatilin, one of the major flavonoids in Artemisia asiatica Nakai (Asteraceae), has been reported to possess antitumor properties. However, thus far there have been no reports regarding the effects of eupatilin on glioma. Therefore, in the current study the effects of eupatilin on glioma and the underlying molecular mechanism were explored. The effect of eupatilin on cell viability was detected by the MTT assay. Cell invasion and migration were performed with Transwell assays and cell apoptosis was determined by flow cytometric analysis. Notch-1 knockdown cells were established by transfection with Notch-1 small interfering RNA (siRNA). The expression levels of Notch-1 were detected by quantitative reverse transcription-polymerase chain reaction and western blotting. The results of the present study indicated that eupatilin exhibits an anticancer effect on glioma cells. Eupatilin inhibited proliferation, reduced cell invasion and migration, and promoted the apoptosis of glioma cells. Additionally, it suppressed Notch-1 expression. Knockdown of Notch-1 by siRNA contributed to the inhibitory effect of eupatilin on proliferation and invasion of glioma cells. In conclusion, eupatilin had an inhibitory effect on proliferation, invasion and migration, and promoted apoptosis of glioma cells through suppression of the Notch-1 signaling pathway. Therefore, eupatilin may have potential as an effective agent for the treatment of glioma. PMID:26676446

  4. MyoD stimulates delta-1 transcription and triggers notch signaling in the Xenopus gastrula.

    PubMed Central

    Wittenberger, T; Steinbach, O C; Authaler, A; Kopan, R; Rupp, R A

    1999-01-01

    The Notch signaling cascade is involved in many developmental decisions, a paradigm of which has been the selection between epidermal and neural cell fates in both invertebrates and vertebrates. Notch has also been implicated as a regulator of myogenesis, although its precise function there has remained controversial. Here we show that the muscle-determining factor MyoD is a direct, positive regulator of the Notch ligand Delta-1 in prospective myoblasts of the pre-involuted mesoderm in Xenopus gastrulae. Injection of a dominant MyoD repressor variant ablates mesodermal Delta-1 expression in vivo. Furthermore, MyoD-dependent Delta-1 induction is sufficient to activate transcription from promoters of E(spl)-related genes in a Notch-dependent manner. These results indicate that a hallmark of neural cell fate determination, i.e. the feedback loop between differentiation promoting basic helix-loop-helix proteins and the Notch regulatory circuitry, is conserved in myogenesis, supporting a direct involvement of Notch in muscle determination. PMID:10202155

  5. Mutations in TSPEAR, Encoding a Regulator of Notch Signaling, Affect Tooth and Hair Follicle Morphogenesis

    PubMed Central

    Samuelov, Liat; Bertolini, Marta; Weissglas-Volkov, Daphna; Eskin-Schwartz, Marina; Malchin, Natalia; Bochner, Ron; Fainberg, Gilad; Goldberg, Ilan; Sugawara, Koji; Tsuruta, Daisuke; Morasso, Maria; Shalev, Stavit; Gallo, Richard L.; Shomron, Noam; Paus, Ralf; Sprecher, Eli

    2016-01-01

    Despite recent advances in our understanding of the pathogenesis of ectodermal dysplasias (EDs), the molecular basis of many of these disorders remains unknown. In the present study, we aimed at elucidating the genetic basis of a new form of ED featuring facial dysmorphism, scalp hypotrichosis and hypodontia. Using whole exome sequencing, we identified 2 frameshift and 2 missense mutations in TSPEAR segregating with the disease phenotype in 3 families. TSPEAR encodes the thrombospondin-type laminin G domain and EAR repeats (TSPEAR) protein, whose function is poorly understood. TSPEAR knock-down resulted in altered expression of genes known to be regulated by NOTCH and to be involved in murine hair and tooth development. Pathway analysis confirmed that down-regulation of TSPEAR in keratinocytes is likely to affect Notch signaling. Accordingly, using a luciferase-based reporter assay, we showed that TSPEAR knock-down is associated with decreased Notch signaling. In addition, NOTCH1 protein expression was reduced in patient scalp skin. Moreover, TSPEAR silencing in mouse hair follicle organ cultures was found to induce apoptosis in follicular epithelial cells, resulting in decreased hair bulb diameter. Collectively, these observations indicate that TSPEAR plays a critical, previously unrecognized role in human tooth and hair follicle morphogenesis through regulation of the Notch signaling pathway. PMID:27736875

  6. Endothelial nitric oxide signaling regulates Notch1 in aortic valve disease

    PubMed Central

    Bosse, Authors: Kevin; Hans, Chetan P.; Zhao, Ning; Koenig, Sara N.; Huang, Nianyuan; Guggilam, Anuradha; LaHaye, Stephanie; Tao, Ge; Lucchesi, Pamela A.; Lincoln, Joy; Lilly, Brenda; Garg, Vidu

    2013-01-01

    The mature aortic valve is composed of a structured trilaminar extracellular matrix that is interspersed with aortic valve interstitial cells (AVICs) and covered by endothelium. Dysfunction of the valvular endothelium initiates calcification of neighboring AVICs leading to calcific aortic valve disease (CAVD). The molecular mechanism by which endothelial cells communicate with AVICs and cause disease is not well understood. Using a co-culture assay, we show that endothelial cells secrete a signal to inhibit calcification of AVICs. Gain or loss of nitric oxide (NO) prevents or accelerates calcification of AVICs, respectively, suggesting that the endothelial cell-derived signal is NO. Overexpression of Notch1, which is genetically linked to human CAVD, retards the calcification of AVICs that occurs with NO inhibition. In AVICs, NO regulates the expression of Hey1, a downstream target of Notch1, and alters nuclear localization of Notch1 intracellular domain. Finally, Notch1 and NOS3 (endothelial NO synthase) display an in vivo genetic interaction critical for proper valve morphogenesis and the development of aortic valve disease. Our data suggests that endothelial cell-derived NO is a regulator of Notch1 signaling in AVICs in the development of the aortic valve and adult aortic valve disease. PMID:23583836

  7. Notch receptors and Smad3 signaling cooperate in the induction of interleukin-9-producing T cells.

    PubMed

    Elyaman, Wassim; Bassil, Ribal; Bradshaw, Elizabeth M; Orent, William; Lahoud, Youmna; Zhu, Bing; Radtke, Freddy; Yagita, Hideo; Khoury, Samia J

    2012-04-20

    Interleukin 9 (IL-9) is a pleiotropic cytokine that can regulate autoimmune responses by enhancing regulatory CD4(+)FoxP3(+) T regulatory (Treg) cell survival and T helper 17 (Th17) cell proliferation. Here, we analyzed the costimulatory requirements for the induction of Th9 cells, and demonstrated that Notch pathway cooperated with TGF-β signaling to induce IL-9. Conditional ablation of Notch1 and Notch2 receptors inhibited the development of Th9 cells. Notch1 intracellular domain (NICD1) recruited Smad3, downstream of TGF-β cytokine signaling, and together with recombining binding protein (RBP)-Jκ bound the Il9 promoter and induced its transactivation. In experimental autoimmune encephalomyelitis (EAE), Jagged2 ligation regulated clinical disease in an IL-9-dependent fashion. Signaling through Jagged2 expanded Treg cells and suppressed EAE when administered before antigen immunization, but worsened EAE when administered concurrently with immunization by favoring Th17 cell expansion. We propose that Notch and Smad3 cooperate to induce IL-9 and participate in regulating the immune response.

  8. Activation of Notch1 signaling in cardiogenic mesoderm induces abnormal heart morphogenesis in mouse.

    PubMed

    Watanabe, Yusuke; Kokubo, Hiroki; Miyagawa-Tomita, Sachiko; Endo, Maho; Igarashi, Katsuhide; Aisaki, Ken ichi; Kanno, Jun; Saga, Yumiko

    2006-05-01

    Notch signaling is implicated in many developmental processes. In our current study, we have employed a transgenic strategy to investigate the role of Notch signaling during cardiac development in the mouse. Cre recombinase-mediated Notch1 (NICD1) activation in the mesodermal cell lineage leads to abnormal heart morphogenesis, which is characterized by deformities of the ventricles and atrioventricular (AV) canal. The major defects observed include impaired ventricular myocardial differentiation, the ectopic appearance of cell masses in the AV cushion, the right-shifted interventricular septum (IVS) and impaired myocardium of the AV canal. However, the fates of the endocardium and myocardium were not disrupted in NICD1-activated hearts. One of the Notch target genes, Hesr1, was found to be strongly induced in both the ventricle and the AV canal of NICD1-activated hearts. However, a knockout of the Hesr1 gene from NICD-activated hearts rescues only the abnormality of the AV myocardium. We searched for additional possible targets of NICD1 activation by GeneChip analysis and found that Wnt2, Bmp6, jagged 1 and Tnni2 are strongly upregulated in NICD1-activated hearts, and that the activation of these genes was also observed in the absence of Hesr1. Our present study thus indicates that the Notch1 signaling pathway plays a suppressive role both in AV myocardial differentiation and the maturation of the ventricular myocardium.

  9. BLOS2 negatively regulates Notch signaling during neural and hematopoietic stem and progenitor cell development

    PubMed Central

    Zhou, Wenwen; He, Qiuping; Zhang, Chunxia; He, Xin; Cui, Zongbin; Liu, Feng; Li, Wei

    2016-01-01

    Notch signaling plays a crucial role in controling the proliferation and differentiation of stem and progenitor cells during embryogenesis or organogenesis, but its regulation is incompletely understood. BLOS2, encoded by the Bloc1s2 gene, is a shared subunit of two lysosomal trafficking complexes, biogenesis of lysosome-related organelles complex-1 (BLOC-1) and BLOC-1-related complex (BORC). Bloc1s2−/− mice were embryonic lethal and exhibited defects in cortical development and hematopoiesis. Loss of BLOS2 resulted in elevated Notch signaling, which consequently increased the proliferation of neural progenitor cells and inhibited neuronal differentiation in cortices. Likewise, ablation of bloc1s2 in zebrafish or mice led to increased hematopoietic stem and progenitor cell production in the aorta-gonad-mesonephros region. BLOS2 physically interacted with Notch1 in endo-lysosomal trafficking of Notch1. Our findings suggest that BLOS2 is a novel negative player in regulating Notch signaling through lysosomal trafficking to control multiple stem and progenitor cell homeostasis in vertebrates. DOI: http://dx.doi.org/10.7554/eLife.18108.001 PMID:27719760

  10. Demonstration of optical steganography transmission using temporal phase coded optical signals with spectral notch filtering.

    PubMed

    Hong, Xuezhi; Wang, Dawei; Xu, Lei; He, Sailing

    2010-06-07

    A novel approach is proposed and experimentally demonstrated for optical steganography transmission in WDM networks using temporal phase coded optical signals with spectral notch filtering. A temporal phase coded stealth channel is temporally and spectrally overlaid onto a public WDM channel. Direct detection of the public channel is achieved in the presence of the stealth channel. The interference from the public channel is suppressed by spectral notching before the detection of the optical stealth signal. The approach is shown to have good compatibility and robustness to the existing WDM network for optical steganography transmission.

  11. Activation of Notch Signaling Is Required for Cholangiocarcinoma Progression and Is Enhanced by Inactivation of p53 In Vivo

    PubMed Central

    Palagani, Vindhya; Malek, Nisar P.; Wilkens, Ludwig; Plentz, Ruben R.

    2013-01-01

    Cholangiocacinoma (CC) is a cancer disease with rising incidence. Notch signaling has been shown to be deregulated in many cancers. However, the role of this signaling pathway in the carcinogenesis of CC is still not fully explored. In this study, we investigated the effects of Notch inhibition by γ-secretase inhibitor IX (GSI IX) in cultured human CC cell lines and we established a transgenic mouse model with liver specific expression of the intracellular domain of Notch (Notch-ICD) and inactivation of tumor suppressor p53. GSI IX treatment effectively impaired cell proliferation, migration, invasion, epithelial to mesenchymal transition and growth of softagar colonies. In vivo overexpression of Notch-ICD together with an inactivation of p53 significantly increased tumor burden and showed CC characteristics. Conclusion: Our study highlights the importance of Notch signaling in the tumorigenesis of CC and demonstrates that additional inactivation of p53 in vivo. PMID:24204826

  12. Notch signaling regulates expression of Mcl-1 and apoptosis in PPD-treated macrophages.

    PubMed

    Palaga, Tanapat; Ratanabunyong, Siriluk; Pattarakankul, Thitiporn; Sangphech, Naunpun; Wongchana, Wipawee; Hadae, Yukihiro; Kueanjinda, Patipark

    2013-09-01

    Macrophages are cellular targets for infection by bacteria and viruses. The fate of infected macrophages plays a key role in determining the outcome of the host immune response. Apoptotic cell death of macrophages is considered to be a protective host defense that eliminates pathogens and infected cells. In this study, we investigated the involvement of Notch signaling in regulating apoptosis in macrophages treated with tuberculin purified protein derivative (PPD). Murine bone marrow-derived macrophages (BMMs) treated with PPD or infected with Mycobacterium bovis Bacillus Calmette-Guérin (BCG) induced upregulation of Notch1. This upregulation correlated well with the upregulation of the anti-apoptotic gene mcl-1 both at the transcriptional and translational levels. Decreased levels of Notch1 and Mcl-1 were observed in BMM treated with PPD when a gamma secretase inhibitor (GSI), which inhibits the processing of Notch receptors, was used. Moreover, silencing Notch1 in the macrophage-like cell line RAW264.7 decreased Mcl-1 protein expression, suggesting that Notch1 is critical for Mcl-1 expression in macrophages. A significant increase in apoptotic cells was observed upon treatment of BMM with PPD in the presence of GSI compared to the vehicle-control treated cells. Finally, analysis of the mcl-1 promoter in humans and mice revealed a conserved potential CSL/RBP-Jκ binding site. The association of Notch1 with the mcl-1 promoter was confirmed by chromatin immunoprecipitation. Taken together, these results indicate that Notch1 inhibits apoptosis of macrophages stimulated with PPD by directly controlling the mcl-1 promoter.

  13. NUMB negatively regulates the epithelial-mesenchymal transition of triple-negative breast cancer by antagonizing Notch signaling

    PubMed Central

    Zhang, Jianchao; Shao, Ximing; Sun, Haiyan; Liu, Ke; Ding, Zhihao; Chen, Juntao; Fang, Lijing; Su, Wu; Hong, Yang; Li, Huashun; Li, Hongchang

    2016-01-01

    Triple-negative breast cancer (TNBC), an aggressive subtype of breast cancer with higher rates of early relapse and metastasis, is frequently associated with aberrant activation of epithelial-mesenchymal transition (EMT). Nonetheless, how EMT is initiated and regulated during TNBC progression is not well understood. Here, we report that NUMB is a negative regulator of EMT in both human mammary epithelial cells and breast cancer cells. Reduced NUMB expression was significantly associated with elevated EMT in TNBC. Conversely, overexpression of NUMB strongly attenuated the EMT program and metastasis of TNBC cell lines. Interestingly, we showed that NUMB employs different molecular mechanisms to regulate EMT. In normal mammary epithelial cells and breast cancer cells expressing wild-type p53, NUMB suppressed EMT by stabilizing p53. However, in TNBC cells, loss of NUMB facilitated the EMT program by activating Notch signaling. Consistent with these findings, low NUMB expression and high Notch activity were significantly correlated with the TNBC subtype in patients. Collectively, these findings reveal novel molecular mechanisms of NUMB in the regulation of breast tumor EMT, especially in TNBC. PMID:27506933

  14. Notch Signaling Limits Supporting Cell Plasticity in the Hair Cell-Damaged Early Postnatal Murine Cochlea

    PubMed Central

    Korrapati, Soumya; Roux, Isabelle; Glowatzki, Elisabeth; Doetzlhofer, Angelika

    2013-01-01

    In mammals, auditory hair cells are generated only during embryonic development and loss or damage to hair cells is permanent. However, in non-mammalian vertebrate species, such as birds, neighboring glia-like supporting cells regenerate auditory hair cells by both mitotic and non-mitotic mechanisms. Based on work in intact cochlear tissue, it is thought that Notch signaling might restrict supporting cell plasticity in the mammalian cochlea. However, it is unresolved how Notch signaling functions in the hair cell-damaged cochlea and the molecular and cellular changes induced in supporting cells in response to hair cell trauma are poorly understood. Here we show that gentamicin-induced hair cell loss in early postnatal mouse cochlear tissue induces rapid morphological changes in supporting cells, which facilitate the sealing of gaps left by dying hair cells. Moreover, we provide evidence that Notch signaling is active in the hair cell damaged cochlea and identify Hes1, Hey1, Hey2, HeyL, and Sox2 as targets and potential Notch effectors of this hair cell-independent mechanism of Notch signaling. Using Cre/loxP based labeling system we demonstrate that inhibition of Notch signaling with a γ- secretase inhibitor (GSI) results in the trans-differentiation of supporting cells into hair cell-like cells. Moreover, we show that these hair cell-like cells, generated by supporting cells have molecular, cellular, and basic electrophysiological properties similar to immature hair cells rather than supporting cells. Lastly, we show that the vast majority of these newly generated hair cell-like cells express the outer hair cell specific motor protein prestin. PMID:24023676

  15. The effects of methylmercury on Notch signaling during embryonic neural development in Drosophila melanogaster

    PubMed Central

    Engel, GL; Delwig, A; Rand, MD

    2012-01-01

    Methylmercury (MeHg) is a ubiquitous toxicant that targets the developing fetal nervous system. MeHg interacts with the Notch signaling pathway, a highly-conserved intercellular signaling mechanism required for normal development. Notch signaling is conveyed by activation of the genes in the Enhancer of Split (E(spl)) locus in Drosophila. We have previously shown that acute high doses of MeHg upregulate several E(spl) genes in Drosophila neural-derived C6 cells. Furthermore, MeHg induction of E(spl) can occur independent of the Notch receptor itself. We now show that MeHg, unlike inorganic mercury (HgCl2), preferentially upregulates E(spl)mδ and E(spl)mγ in Drosophila C6 cells. This is distinct from Delta ligand-induced Notch signaling in which no induction of E(spl)mδ is seen. MeHg is also seen to specifically upregulate E(spl)mδ in Drosophila embryos where HgCl2 showed no such effect. Additionally, treatment of embryos with MeHg caused a consistent failure in axonal outgrowth of the intersegmental nerve (ISN). This ISN phenotype was partially replicated by genetic activation of the Notch pathway, but was not replicated by increasing expression of E(spl)mδ. These data suggest a role for Notch signaling and the E(spl)mδ target gene in MeHg toxicity, however, the site of action for E(spl)mδ in this system remains to be elucidated. PMID:22230562

  16. Reciprocal regulatory interactions between the Notch and Ras signaling pathways in the Drosophila embryonic mesoderm.

    PubMed

    Carmena, Ana; Buff, Eugene; Halfon, Marc S; Gisselbrecht, Stephen; Jiménez, Fernando; Baylies, Mary K; Michelson, Alan M

    2002-04-15

    Convergent intercellular signals must be precisely integrated in order to elicit specific biological responses. During specification of muscle and cardiac progenitors from clusters of equivalent cells in the Drosophila embryonic mesoderm, the Ras/MAPK pathway--activated by both epidermal and fibroblast growth factor receptors--functions as an inductive cellular determination signal, while lateral inhibition mediated by Notch antagonizes this activity. A critical balance between these signals must be achieved to enable one cell of an equivalence group to segregate as a progenitor while its neighbors assume a nonprogenitor identity. We have investigated whether these opposing signals directly interact with each other, and we have examined how they are integrated by the responding cells to specify their unique fates. Our findings reveal that Ras and Notch do not function independently; rather, we have uncovered several modes of cross-talk between these pathways. Ras induces Notch, its ligand Delta, and the epidermal growth factor receptor antagonist, Argos. We show that Delta and Argos then synergize to nonautonomously block a positive autoregulatory feedback loop that amplifies a fate-inducing Ras signal. This feedback loop is characterized by Ras-mediated upregulation of proximal components of both the epidermal and fibroblast growth factor receptor pathways. In turn, Notch activation in nonprogenitors induces its own expression and simultaneously suppresses both Delta and Argos levels, thereby reinforcing a unidirectional inhibitory response. These reciprocal interactions combine to generate the signal thresholds that are essential for proper specification of progenitors and nonprogenitors from groups of initially equivalent cells.

  17. Essential role of Notch4/STAT3 signaling in epithelial-mesenchymal transition of tamoxifen-resistant human breast cancer.

    PubMed

    Bui, Quyen Thu; Im, Ji Hye; Jeong, Sung Baek; Kim, Young-Mi; Lim, Sung Chul; Kim, Bumseok; Kang, Keon Wook

    2017-04-01

    We previously demonstrated that tamoxifen (TAM)-resistant human breast cancer (TAMR-MCF-7) cells showed increased expression of mesenchymal marker proteins compared to the parent MCF-7 cells. Notch is functionally important in the promotion of epithelial-mesenchymal transition (EMT) during both development and tumor progression. Notch1 and Notch4 have been reported as prognostic markers in human breast cancer. Here, we indicated that Notch4, but not Notch1, plays a critical role in the regulation of EMT signaling in TAMR-MCF-7 cells. Notch4 suppression by either Notch inhibitors or Notch4 siRNA attenuated EMT signaling. Tyrosine-phosphorylated STAT3 protein is known as a crucial signaling molecule in the regulation of tumorigenesis and metastasis. We found that TAMR-MCF-7 cells exhibited constitutive STAT3 phosphorylation, and Notch inhibition reduced the level of activated STAT3 in TAMR-MCF-7 cells. An intrasplenic injection model of liver metastases was performed using TAMR-MCF-7 cells. Mice injected with N-[N-(3,5-Difluorophenacetyl)-l-alanyl]-S-phenylglycine t-butyl ester (DAPT, 10 mg/kg) formed smaller splenic tumors and showed a reduced micrometastatic tumor burden in their livers compared with the control group treated with vehicle. To conclude, Notch4 could be a potential target to prevent metastasis in TAM-resistant breast cancer.

  18. Estrogen improves the proliferation and differentiation of hBMSCs derived from postmenopausal osteoporosis through notch signaling pathway.

    PubMed

    Fan, Jin-Zhu; Yang, Liu; Meng, Guo-Lin; Lin, Yan-shui; Wei, Bo-Yuan; Fan, Jing; Hu, Hui-Min; Liu, Yan-Wu; Chen, Shi; Zhang, Jin-Kang; He, Qi-Zhen; Luo, Zhuo-Jing; Liu, Jian

    2014-07-01

    Estrogen deficiency is the main reason of bone loss, leading to postmenopausal osteoporosis, and estrogen replacement therapy (ERT) has been demonstrated to protect bone loss efficiently. Notch signaling controls proliferation and differentiation of bone marrow-derived mesenchymal stem cells (BMSCs). Moreover, imperfect estrogen-responsive elements (EREs) were found in the 5'-untranslated region of Notch1 and Jagged1. Thus, we examined the molecular and biological links between estrogen and the Notch signaling in postmenopausal osteoporosis in vitro. hBMSCs were obtained from healthy women and patients with postmenopausal osteoporosis. Notch signaling molecules were quantified using real-time polymerase chain reaction (real-time PCR) and Western Blot. Luciferase reporter constructs with putative EREs were transfected into hBMSCs and analyzed. hBMSCs were transduced with lentiviral vectors containing human Notch1 intracellular domain (NICD1). We also used N-[N-(3, 5-diflurophenylacetate)-l-alanyl]-(S)-phenylglycine t-butyl ester, a γ-secretase inhibitor, to suppress the Notch signaling. We found that estrogen enhanced the Notch signaling in hBMSCs by promoting the expression of Jagged1. hBMSCs cultured with estrogen resulted in the up-regulation of Notch signaling and increased proliferation and differentiation. Enhanced Notch signaling could enhance the proliferation and differentiation of hBMSCs from patients with postmenopausal osteoporosis (OP-hBMSCs). Our results demonstrated that estrogen preserved bone mass partly by activating the Notch signaling. Because long-term ERT has been associated with several side effects, the Notch signaling could be a potential target for treating postmenopausal osteoporosis.

  19. Identification of Novel Targets of CSL-Dependent Notch Signaling in Hematopoiesis

    PubMed Central

    Hamidi, Habib; Gustafason, Derek; Pellegrini, Matteo; Gasson, Judith

    2011-01-01

    Somatic activating mutations in the Notch1 receptor result in the overexpression of activated Notch1, which can be tumorigenic. The goal of this study is to understand the molecular mechanisms underlying the phenotypic changes caused by the overexpression of ligand independent Notch 1 by using a tetracycline inducible promoter in an in vitro embryonic stem (ES) cells/OP9 stromal cells coculture system, recapitulating normal hematopoiesis. First, an in silico analysis of the promoters of Notch regulated genes (previously determined by microarray analysis) revealed that the motifs recognized by regulatory proteins known to mediate hematopoiesis were overrepresented. Notch 1 does not bind DNA but instead binds the CSL transcription factor to regulate gene expression. The in silico analysis also showed that there were putative CSL binding sites observed in the promoters of 28 out of 148 genes. A custom ChIP-chip array was used to assess the occupancy of CSL in the promoter regions of the Notch1 regulated genes in vivo and showed that 61 genes were bound by activated Notch responsive CSL. Then, comprehensive mapping of the CSL binding sites genome-wide using ChIP-seq analysis revealed that over 10,000 genes were bound within 10 kb of the TSS (transcription start site). The majority of the targets discovered by ChIP-seq belong to pathways that have been shown by others to crosstalk with Notch signaling. Finally, 83 miRNAs were significantly differentially expressed by greater than 1.5-fold during the course of in vitro hematopoiesis. Thirty one miRNA were up-regulated and fifty two were down-regulated. Overexpression of Notch1 altered this pattern of expression of microRNA: six miRNAs were up-regulated and four were down regulated as a result of activated Notch1 overexpression during the course of hematopoiesis. Time course analysis of hematopoietic development revealed that cells with Notch 1 overexpression mimic miRNA expression of cells in a less mature stage, which

  20. Identification of novel targets of CSL-dependent Notch signaling in hematopoiesis.

    PubMed

    Hamidi, Habib; Gustafason, Derek; Pellegrini, Matteo; Gasson, Judith

    2011-01-01

    Somatic activating mutations in the Notch1 receptor result in the overexpression of activated Notch1, which can be tumorigenic. The goal of this study is to understand the molecular mechanisms underlying the phenotypic changes caused by the overexpression of ligand independent Notch 1 by using a tetracycline inducible promoter in an in vitro embryonic stem (ES) cells/OP9 stromal cells coculture system, recapitulating normal hematopoiesis. First, an in silico analysis of the promoters of Notch regulated genes (previously determined by microarray analysis) revealed that the motifs recognized by regulatory proteins known to mediate hematopoiesis were overrepresented. Notch 1 does not bind DNA but instead binds the CSL transcription factor to regulate gene expression. The in silico analysis also showed that there were putative CSL binding sites observed in the promoters of 28 out of 148 genes. A custom ChIP-chip array was used to assess the occupancy of CSL in the promoter regions of the Notch1 regulated genes in vivo and showed that 61 genes were bound by activated Notch responsive CSL. Then, comprehensive mapping of the CSL binding sites genome-wide using ChIP-seq analysis revealed that over 10,000 genes were bound within 10 kb of the TSS (transcription start site). The majority of the targets discovered by ChIP-seq belong to pathways that have been shown by others to crosstalk with Notch signaling. Finally, 83 miRNAs were significantly differentially expressed by greater than 1.5-fold during the course of in vitro hematopoiesis. Thirty one miRNA were up-regulated and fifty two were down-regulated. Overexpression of Notch1 altered this pattern of expression of microRNA: six miRNAs were up-regulated and four were down regulated as a result of activated Notch1 overexpression during the course of hematopoiesis. Time course analysis of hematopoietic development revealed that cells with Notch 1 overexpression mimic miRNA expression of cells in a less mature stage, which

  1. Roles of PINK1, mTORC2, and mitochondria in preserving brain tumor-forming stem cells in a noncanonical Notch signaling pathway.

    PubMed

    Lee, Kyu-Sun; Wu, Zhihao; Song, Yan; Mitra, Siddhartha S; Feroze, Abdullah H; Cheshier, Samuel H; Lu, Bingwei

    2013-12-15

    The self-renewal versus differentiation choice of Drosophila and mammalian neural stem cells (NSCs) requires Notch (N) signaling. How N regulates NSC behavior is not well understood. Here we show that canonical N signaling cooperates with a noncanonical N signaling pathway to mediate N-directed NSC regulation. In the noncanonical pathway, N interacts with PTEN-induced kinase 1 (PINK1) to influence mitochondrial function, activating mechanistic target of rapamycin complex 2 (mTORC2)/AKT signaling. Importantly, attenuating noncanonical N signaling preferentially impaired the maintenance of Drosophila and human cancer stem cell-like tumor-forming cells. Our results emphasize the importance of mitochondria to N and NSC biology, with important implications for diseases associated with aberrant N signaling.

  2. Notch and Wnt/β-catenin signaling pathway play important roles in activating liver cancer stem cells.

    PubMed

    Wang, Ronghua; Sun, Qian; Wang, Peng; Liu, Man; Xiong, Si; Luo, Jing; Huang, Hai; Du, Qiang; Geller, David A; Cheng, Bin

    2016-02-02

    Human hepatocellular carcinoma (HCC) is driven and maintained by liver cancer stem cells (LCSCs) that display stem cell properties. These LCSCs are promoted by the intersecting of Notch and Wnt/β-Catenin signaling pathways. In this study, we demonstrate that LCSCs with markers CD90, CD24, CD13, and CD133 possess stem properties of self-renewal and tumorigenicity in NOD/SCID mice. The increased expression of these markers was correlated with advanced disease stage, larger tumors, and worse overall survival in 61 HCC cases. We also found that both Notch and Wnt/β-catenin signaling pathways played important roles in increasing the stem-ness characteristics of LCSCs. Our data suggested that Notch1 was downstream of Wnt/β-catenin. The active form of Notch1 intracellular domain (NICD) expression depended on Wnt/β-catenin pathway activation. Moreover, Notch1 negatively contributed to Wnt/β-catenin signaling modulation. Knock down of Notch1 with lentivirus N1ShRNA up-regulated the active form of β-catenin. Ectopic expression of NICD with LV-Notch1 in LCSCs attenuated β-catenin/TCF dependent luciferase activity significantly. In addition, there was a non-proteasome mediated feedback loop between Notch1 and Wnt/β-catenin signaling in LCSCs. The central role of Notch and the Wnt/β-catenin signaling pathway in LCSCs may provide an attractive therapeutic strategy against HCC.

  3. Crucial role of the Rap G protein signal in Notch activation and leukemogenicity of T-cell acute lymphoblastic leukemia.

    PubMed

    Doi, Keiko; Imai, Takahiko; Kressler, Christopher; Yagita, Hideo; Agata, Yasutoshi; Vooijs, Marc; Hamazaki, Yoko; Inoue, Joe; Minato, Nagahiro

    2015-01-23

    The Rap G protein signal regulates Notch activation in early thymic progenitor cells, and deregulated Rap activation (Rap(high)) results in the development of Notch-dependent T-cell acute lymphoblastic leukemia (T-ALL). We demonstrate that the Rap signal is required for the proliferation and leukemogenesis of established Notch-dependent T-ALL cell lines. Attenuation of the Rap signal by the expression of a dominant-negative Rap1A17 or Rap1GAP, Sipa1, in a T-ALL cell line resulted in the reduced Notch processing at site 2 due to impaired maturation of Adam10. Inhibition of the Rap1 prenylation with a geranylgeranyl transferase inhibitor abrogated its membrane-anchoring to Golgi-network and caused reduced proprotein convertase activity required for Adam10 maturation. Exogenous expression of a mature form of Adam10 overcame the Sipa1-induced inhibition of T-ALL cell proliferation. T-ALL cell lines expressed Notch ligands in a Notch-signal dependent manner, which contributed to the cell-autonomous Notch activation. Although the initial thymic blast cells barely expressed Notch ligands during the T-ALL development from Rap(high) hematopoietic progenitors in vivo, the ligands were clearly expressed in the T-ALL cells invading extrathymic vital organs. These results reveal a crucial role of the Rap signal in the Notch-dependent T-ALL development and the progression.

  4. Involvement of Notch1 signaling in neurogenesis in the subventricular zone of normal and ischemic rat brain in vivo.

    PubMed

    Wang, Xiaomei; Mao, Xiaoou; Xie, Lin; Greenberg, David A; Jin, Kunlin

    2009-10-01

    The Notch1 signaling pathway is regarded as one of the main regulators of neural stem cell behavior during development, but its role in the adult brain is less well understood. We found that Notch1 was mainly expressed in doublecortin (DCX)-positive cells corresponding to newborn neurons, whereas the Notch1 ligand, Jagged1, was predominantly expressed in glial fibrillary acidic protein (GFAP)-positive astrocytic cells in the subventricular zone (SVZ) of the normal adult brain. These findings were confirmed by conditional depletion of DCX-positive cells in transgenic mice carrying herpes simplex virus thymidine kinase (HSV-TK) under the control of the DCX promoter. In addition, the activated form of Notch1 (Notch intracellular domain, NICD) and its downstream transcriptional targets, Hes1 and sonic hedgehog (Shh), were also expressed in SVZ cells. Increased activation of Notch1 signaling increased SVZ cell proliferation, whereas inhibiting Notch1 signaling resulted in a reduction of proliferating cells in the SVZ. Levels of NICD, Hes1, and Shh were increased in the SVZ at 4 and 24 h after focal cerebral ischemia. Finally, ischemia-induced cell proliferation in the SVZ was blocked by inhibition of the Notch1 signaling pathway, suggesting that Notch1 signaling may have a key role in normal adult and ischemia-induced neurogenesis.

  5. NOTCH SIGNALLING MODULATES HYPOXIA-INDUCED NEUROENDOCRINE DIFFERENTIATION OF HUMAN PROSTATE CANCER CELLS

    PubMed Central

    Danza, Giovanna; Di Serio, Claudia; Rosati, Fabiana; Lonetto, Giuseppe; Sturli, Niccolò; Kacer, Doreen; Pennella, Antonio; Ventimiglia, Giuseppina; Barucci, Riccardo; Piscazzi, Annamaria; Prudovsky, Igor; Landriscina, Matteo; Marchionni, Niccolò; Tarantini, Francesca

    2012-01-01

    Prostate carcinoma is among the most common causes of cancer-related death in men, representing 15% of all male malignancies in developed countries. Neuroendocrine differentiation has been associated with tumor progression, poor prognosis and with the androgen-independent status. Currently, no successful therapy exists for advanced, castration-resistant disease. Because hypoxia has been linked to prostate cancer progression and unfavourable outcome, we sought to determine whether hypoxia would impact the degree of neuroendocrine differentiation of prostate cancer cells, in vitro. Results exposure of LNCaP cells to low oxygen tension induced a neuroendocrine phenotype, associated with an increased expression of the transcription factor neurogenin3 and neuroendocrine markers, such as neuron-specific enolase, chromogranin A and β3-tubulin. Moreover, hypoxia triggered a significant decrease of Notch 1 and Notch 2 mRNA and protein expression, with subsequent down regulation of Notch-mediated signalling, as demonstrated by reduced levels of the Notch target genes, Hes1 and Hey1. Neuroendocrine differentiation was promoted by attenuation of Hes1 transcription, as cells expressing a dominant negative form of Hes1 displayed increased levels of neuroendocrine markers under normoxic conditions. Although hypoxia down regulated Notch 1 and Notch 2 mRNA transcription and receptor activation also in the androgen independent cell lines, PC3 and Du145, it did not change the extent of NE differentiation in these cultures, suggesting that androgen sensitivity may be required for transdifferentiation to occur. Conclusions hypoxia induces neuroendocrine differentiation of LNCaP cells in vitro, which appears to be driven by the inhibition of Notch signalling with subsequent down-regulation of Hes1 transcription. PMID:22172337

  6. CAF-1 promotes Notch signaling through epigenetic control of target gene expression during Drosophila development.

    PubMed

    Yu, Zhongsheng; Wu, Honggang; Chen, Hanqing; Wang, Ruoqi; Liang, Xuehong; Liu, Jiyong; Li, Changqing; Deng, Wu-Min; Jiao, Renjie

    2013-09-01

    The histone chaperone CAF-1 is known for its role in DNA replication-coupled histone deposition. However, loss of function causes lethality only in higher multicellular organisms such as mice and flies, but not in unicellular organisms such as yeasts, suggesting that CAF-1 has other important functions than histone deposition during animal development. Emerging evidence indicates that CAF-1 also has a role in higher order chromatin organization and heterochromatin-mediated gene expression; it remains unclear whether CAF-1 has a role in specific signaling cascades to promote gene expression during development. Here, we report that knockdown of one of the subunits of Drosophila CAF-1, dCAF-1-p105 (Caf1-105), results in phenotypes that resemble those of, and are augmented synergistically by, mutations of Notch positive regulatory pathway components. Depletion of dCAF-1-p105 leads to abrogation of cut expression and to downregulation of other Notch target genes in wing imaginal discs. dCAF-1-p105 is associated with Suppressor of Hairless [Su(H)] and regulates its binding to the enhancer region of E(spl)mβ. The association of dCAF-1-p105 with Su(H) on chromatin establishes an active local chromatin status for transcription by maintaining a high level of histone H4 acetylation. In response to induced Notch activation, dCAF-1 associates with the Notch intracellular domain to activate the expression of Notch target genes in cultured S2 cells, manifesting the role of dCAF-1 in Notch signaling. Together, our results reveal a novel epigenetic function of dCAF-1 in promoting Notch pathway activity that regulates normal Drosophila development.

  7. The hepatitis B virus X protein downregulates NF-κB signaling pathways through decreasing the Notch signaling pathway in HBx-transformed L02 cells.

    PubMed

    Luo, Jing; Zhou, Haiyan; Wang, Fan; Xia, Xiumei; Sun, Qian; Wang, Ronghua; Cheng, Bin

    2013-05-01

    Hepatitis B virus X protein (HBx) is implicated in the pathogenesis of hepatocellular carcinoma, which has been found to be associated with Notch and NF-κB signaling. This study aimed to investigate the crosstalk between Notch and NF-κB pathways in HBx-related hepatocellular carcinoma. An HBx-transformed non-tumor hepatic cell line L02 (L02/HBx) was previously established. Immunofluorescence assays were performed to visualize HBx and the Notch intracellular domain (NICD) in cell nuclei. Co-immunoprecipitation assays were used to investigate physical interactions between HBx and components of the Notch signaling pathway (NICD and JAG1), NF-κB signaling pathway (p65 and p50) or IκBα. L02/HBx cells were treated with the Notch signal inhibitor DAPT or Notch1 siRNA to inhibit the Notch1 pathway. qRT-PCR was used to quantify the expression of the p65, p50 and IκBα genes. Protein expression changes in cytoplasm and nuclei after treatment with DAPT or Notch1 siRNA were analyzed by western blotting and EMSA assays. We found that HBx directly regulated Notch1 signaling, which cross-talked with the NF-κB pathway. Downregulation of Notch1 decreased the binding of NF-κB p65 to its target gene promoter, reduced NF-κB expression and enhanced IκBα expression. The results suggest that HBx functions through the Notch signaling pathway; Notch contributes to hepatocarcinogenesis partially by regulating the NF-κB pathway. Our findings provide new insights into the role of Notch and NF-κB signaling in the progression of hepatocellular carcinoma related to HBx.

  8. Notch signaling alters sensory or neuronal cell fate specification of inner ear stem cells.

    PubMed

    Jeon, Sang-Jun; Fujioka, Masato; Kim, Shi-Chan; Edge, Albert S B

    2011-06-08

    Multipotent progenitor cells in the otic placode give rise to the specialized cell types of the inner ear, including neurons, supporting cells, and hair cells. The mechanisms governing acquisition of specific fates by the cells that form the cochleovestibular organs remain poorly characterized. Here we show that whereas blocking Notch signaling with a γ-secretase inhibitor increased the conversion of inner ear stem cells to hair cells by a mechanism that involved the upregulation of bHLH transcription factor, Math1 (mouse Atoh1), differentiation to a neuronal lineage was increased by expression of the Notch intracellular domain. The shift to a neuronal lineage could be attributed in part to continued cell proliferation in cells that did not undergo sensory cell differentiation due to the high Notch signaling, but also involved upregulation of Ngn1. The Notch intracellular domain influenced Ngn1 indirectly by upregulation of Sox2, a transcription factor expressed in many neural progenitor cells, and directly by an interaction with an RBP-J binding site in the Ngn1 promoter/enhancer. The induction of Ngn1 was blocked partially by mutation of the RBP-J site and nearly completely when the mutation was combined with inhibition of Sox2 expression. Thus, Notch signaling had a significant role in the fate specification of neurons and hair cells from inner ear stem cells, and decisions about cell fate were mediated in part by a differential effect of combinatorial signaling by Notch and Sox2 on the expression of bHLH transcription factors.

  9. Enhancement of early cardiac differentiation of dedifferentiated fat cells by dimethyloxalylglycine via notch signaling pathway

    PubMed Central

    Li, Fuhai; Li, Zongzhuang; Jiang, Zhi; Tian, Ye; Wang, Zhi; Yi, Wei; Zhang, Chenyun

    2016-01-01

    Background: Hypoxia has been reported to possess the ability to induce mature lipid-filled adipocytes to differentiate into fibroblast-like multipotent dedifferentiated fat (DFAT) cells and stem cells such as iPSCs (interstitial pluripotent stem cells) and ESCs (embryonic stem cells) and then to differentiate into cardiomyocytes. However, the effect of hypoxia on cardiac differentiation of DFAT cells and its underlying molecular mechanism remains to be investigated. Objective: To investigate the role of hypoxia in early cardiac differentiation of DFAT cells and the underlying molecular mechanism. Methods: DFAT cells were prepared from 4 to 6 week-age mice and cultured under hypoxic conditions by adding Prolyl hydroxylase inhibitor and dimethyloxalylglycine (DMOG) into the culture media. To inhibit or block Notch signaling, γ-secretase inhibitor-II (GSI-II) and Notch1 siRNA (si-Notch1) were used. DFAT cell viability was detected using MTT assay. qRT-PCR, immunofluorescence microscopy and western blotting were used to evaluate the cardiac differentiation of DFAT cells and co-immunoprecipitation was used to study the interaction between HIF-1α and Notch signaling. Results: 0.6-mM DMOG failed to affect the viability of DFAT cells, but stimulated the cells to express early cardiac transcription factors including Islet1, Nkx2.5 and Gata4 in a time-dependent manner and increase the number of cTnT+ cardiomyocytes (detected at the 28th day after stimulation). It was also demonstrated that DMOG was involved in HIF-1α and Notch signaling as well as HIF-1α-NICD complex formation. Conclusion: Hypoxia enhanced early cardiac differentiation of DFAT cells through HIF-1α and Notch signaling pathway. PMID:27904680

  10. Enhancement of early cardiac differentiation of dedifferentiated fat cells by dimethyloxalylglycine via notch signaling pathway.

    PubMed

    Li, Fuhai; Li, Zongzhuang; Jiang, Zhi; Tian, Ye; Wang, Zhi; Yi, Wei; Zhang, Chenyun

    2016-01-01

    Background: Hypoxia has been reported to possess the ability to induce mature lipid-filled adipocytes to differentiate into fibroblast-like multipotent dedifferentiated fat (DFAT) cells and stem cells such as iPSCs (interstitial pluripotent stem cells) and ESCs (embryonic stem cells) and then to differentiate into cardiomyocytes. However, the effect of hypoxia on cardiac differentiation of DFAT cells and its underlying molecular mechanism remains to be investigated. Objective: To investigate the role of hypoxia in early cardiac differentiation of DFAT cells and the underlying molecular mechanism. Methods: DFAT cells were prepared from 4 to 6 week-age mice and cultured under hypoxic conditions by adding Prolyl hydroxylase inhibitor and dimethyloxalylglycine (DMOG) into the culture media. To inhibit or block Notch signaling, γ-secretase inhibitor-II (GSI-II) and Notch1 siRNA (si-Notch1) were used. DFAT cell viability was detected using MTT assay. qRT-PCR, immunofluorescence microscopy and western blotting were used to evaluate the cardiac differentiation of DFAT cells and co-immunoprecipitation was used to study the interaction between HIF-1α and Notch signaling. Results: 0.6-mM DMOG failed to affect the viability of DFAT cells, but stimulated the cells to express early cardiac transcription factors including Islet1, Nkx2.5 and Gata4 in a time-dependent manner and increase the number of cTnT(+) cardiomyocytes (detected at the 28(th) day after stimulation). It was also demonstrated that DMOG was involved in HIF-1α and Notch signaling as well as HIF-1α-NICD complex formation. Conclusion: Hypoxia enhanced early cardiac differentiation of DFAT cells through HIF-1α and Notch signaling pathway.

  11. Huang Qi Decoction Prevents BDL-Induced Liver Fibrosis Through Inhibition of Notch Signaling Activation.

    PubMed

    Zhang, Xiao; Xu, Ying; Chen, Jia-Mei; Liu, Cheng; Du, Guang-Li; Zhang, Hua; Chen, Gao-Feng; Jiang, Shi-Li; Liu, Cheng-Hai; Mu, Yong-Ping; Liu, Ping

    2017-01-01

    Notch signaling has been demonstrated to be involved in ductular reactions and fibrosis. Previous studies have shown that Huang Qi Decoction (HQD) can prevent the progression of cholestatic liver fibrosis (CLF). However, whether HQD affects the Notch signaling pathway is unclear. In this study, CLF was established by common bile duct ligation (BDL) in rats. At the end of the first week, the rats were randomly divided into a model group (i.e., BDL), an HQD group, and a sorafenib positive control group (SORA) and were treated for 3 weeks. Bile duct proliferation and liver fibrosis were determined by tissue staining. Activation of the Notch signaling pathway was evaluated by analyzing expressions of Notch-1, -2, -3, and -4, Jagged (JAG) 1, and Delta like (DLL)-1, -3, and -4. The results showed that HQD significantly reduced the deposition of collagen and the Hyp content of liver tissue and inhibited the activation of HSCs compared with the BDL group. In addition, HQD significantly decreased the protein and mRNA expressions of TGF-[Formula: see text]1 and [Formula: see text]-SMA. In contrast, HQD significantly enhanced expression of the Smad 7 protein. HQD also reduced biliary epithelial cell proliferation, and reduced the mRNA levels of CK7, CK8, CK18, SRY-related high mobility group-box gene (SOX) 9, epithelial cell adhesion molecule (EpCAM) and the positive areas of CK19 and OV6. In addition, the mRNA and protein expressions of Notch-3, -4, JAG1, and DLL-1, -3 were significantly reduced in the HQD compared to the BDL group. These results demonstrated that HQD may prevent biliary liver fibrosis through inhibition of the Notch signaling pathway, and it may be a potential treatment for cholestatic liver disease.

  12. Regulation of striatal dopamine responsiveness by Notch/RBP-J signaling.

    PubMed

    Toritsuka, M; Kimoto, S; Muraki, K; Kitagawa, M; Kishimoto, T; Sawa, A; Tanigaki, K

    2017-03-07

    Dopamine signaling is essential for reward learning and fear-related learning, and thought to be involved in neuropsychiatric diseases. However, the molecular mechanisms underlying the regulation of dopamine responsiveness is unclear. Here we show the critical roles of Notch/RBP-J signaling in the regulation of dopamine responsiveness in the striatum. Notch/RBP-J signaling regulates various neural cell fate specification, and neuronal functions in the adult central nervous system. Conditional deletion of RBP-J specifically in neuronal cells causes enhanced response to apomorphine, a non-selective dopamine agonist, and SKF38393, a D1 agonist, and impaired dopamine-dependent instrumental avoidance learning, which is corrected by SCH23390, a D1 antagonist. RBP-J deficiency drastically reduced dopamine release in the striatum and caused a subtle decrease in the number of dopaminergic neurons. Lentivirus-mediated gene transfer experiments showed that RBP-J deficiency in the striatum was sufficient for these deficits. These findings demonstrated that Notch/RBP-J signaling regulates dopamine responsiveness in the striatum, which may explain the mechanism whereby Notch/RBP-J signaling affects an individual's susceptibility to neuropsychiatric disease.

  13. Critical roles of NOTCH1 in acute T-cell lymphoblastic leukemia.

    PubMed

    Liu, Hudan; Chiang, Mark Y; Pear, Warren S

    2011-08-01

    NOTCH1 plays a central role in T-cell development and, when aberrantly activated, in acute T-cell lymphoblastic leukemia (T-ALL). As a transmembrane receptor that is ultimately converted into a transcription factor, NOTCH1 directly activates a spectrum of target genes, which function to mediate NOTCH1 signaling in normal or transformed T cells. During physiologic T-cell development, NOTCH1 has important functions in cell fate determination, proliferation, survival and metabolism. Activating NOTCH1 mutations occur in more than half of human patients with T-ALL, suggesting an important role for aberrant NOTCH1 signaling in the pathogenesis of this disease. Inhibiting NOTCH1 signaling in patient-derived cell lines and murine T-ALLs leads to growth arrest and/or apoptosis suggesting that NOTCH1 inhibitors can improve T-ALL treatment. However, there are challenges to translate NOTCH1 inhibitors to the clinic because of toxicity and resistance. This review focuses on molecular mechanisms of oncogenic NOTCH1 signaling, molecular and functional analysis of NOTCH1 transcriptional targets in T-ALL, and recent advances in therapeutic targeting of NOTCH1.

  14. Targeting notch signaling pathway in cancer: clinical development advances and challenges.

    PubMed

    Takebe, Naoko; Nguyen, Dat; Yang, Sherry X

    2014-02-01

    Notch signaling plays an important role in development and cell fate determination, and it is deregulated in human hematologic malignancies and solid tumors. This review includes a brief introduction of the relevant pathophysiology of Notch signaling pathway and primarily focuses on the clinical development of promising agents that either obstruct Notch receptor cleavages such as γ-secretase inhibitors (GSIs) or interfere with the Notch ligand-receptor interaction by monoclonal antibodies (mAbs). Antitumor activity by GSIs and mAbs administered as single agent in early phases of clinical trials has been observed in advanced or metastatic thyroid cancer, non-small cell lung cancer, intracranial tumors, sarcoma or desmoid tumors, colorectal cancer with neuroendocrine features, melanoma and ovarian cancer. A number of mechanism-based adverse events particularly gastrointestinal toxicities emerged and mitigation strategies are developed after testing multiple GSIs and Notch targeting mAbs. We also discuss pharmacodynamic biomarkers in conjunction with methods of assessment of the molecular target inhibition validation. Biomarkers of efficacy or benefit may be of importance for a successful development of this class of drugs.

  15. Maternal and zygotic Zfp57 modulate NOTCH signaling in cardiac development

    PubMed Central

    Shamis, Yulia; Cullen, Dana E.; Liu, Lizhi; Yang, Guan; Ng, Sheau-Fang; Xiao, Lijuan; Bell, Fong T.; Ray, Chelsea; Takikawa, Sachiko; Moskowitz, Ivan P.; Cai, Chen-Leng; Yang, Xiao; Li, Xiajun

    2015-01-01

    Zfp57 is a maternal–zygotic effect gene that maintains genomic imprinting. Here we report that Zfp57 mutants exhibited a variety of cardiac defects including atrial septal defect (ASD), ventricular septal defect (VSD), thin myocardium, and reduced trabeculation. Zfp57 maternal-zygotic mutant embryos displayed more severe phenotypes with higher penetrance than the zygotic ones. Cardiac progenitor cells exhibited proliferation and differentiation defects in Zfp57 mutants. ZFP57 is a master regulator of genomic imprinting, so the DNA methylation imprint was lost in embryonic heart without ZFP57. Interestingly, the presence of imprinted DLK1, a target of ZFP57, correlated with NOTCH1 activation in cardiac cells. These results suggest that ZFP57 may modulate NOTCH signaling during cardiac development. Indeed, loss of ZFP57 caused loss of NOTCH1 activation in embryonic heart with more severe loss observed in the maternal-zygotic mutant. Maternal and zygotic functions of Zfp57 appear to play redundant roles in NOTCH1 activation and cardiomyocyte differentiation. This serves as an example of a maternal effect that can influence mammalian organ development. It also links genomic imprinting to NOTCH signaling and particular developmental functions. PMID:25848000

  16. Increasing Notch signaling antagonizes PRC2-mediated silencing to promote reprograming of germ cells into neurons

    PubMed Central

    Seelk, Stefanie; Adrian-Kalchhauser, Irene; Hargitai, Balázs; Hajduskova, Martina; Gutnik, Silvia; Tursun, Baris; Ciosk, Rafal

    2016-01-01

    Cell-fate reprograming is at the heart of development, yet very little is known about the molecular mechanisms promoting or inhibiting reprograming in intact organisms. In the C. elegans germline, reprograming germ cells into somatic cells requires chromatin perturbation. Here, we describe that such reprograming is facilitated by GLP-1/Notch signaling pathway. This is surprising, since this pathway is best known for maintaining undifferentiated germline stem cells/progenitors. Through a combination of genetics, tissue-specific transcriptome analysis, and functional studies of candidate genes, we uncovered a possible explanation for this unexpected role of GLP-1/Notch. We propose that GLP-1/Notch promotes reprograming by activating specific genes, silenced by the Polycomb repressive complex 2 (PRC2), and identify the conserved histone demethylase UTX-1 as a crucial GLP-1/Notch target facilitating reprograming. These findings have wide implications, ranging from development to diseases associated with abnormal Notch signaling. DOI: http://dx.doi.org/10.7554/eLife.15477.001 PMID:27602485

  17. In silico evidence of signaling pathways of notch mediated networks in leukemia

    PubMed Central

    Jamil, Kaiser; Jayaraman, Archana; Rao, Raghunatha; Raju, Suryanarayana

    2012-01-01

    Notch signaling plays a critical role in cell fate determination and maintenance of progenitors in many developmental systems. Notch receptors have been shown to be expressed on hematopoietic progenitor cells as well as to various degrees in peripheral blood T and B lymphocytes, monocytes, and neutrophils. Our aim was to understand the protein interaction network, using Notch1 protein name as query in STRING database and we generated a model to assess the significance of Notch1 associated proteins in Acute Lymphoblastic Leukemia (ALL). We further analyzed the expression levels of the genes encoding hub proteins, using Oncomine database, to determine their significance in leukemogenesis. Of the forty two hub genes, we observed that sixteen genes were underexpressed and eleven genes were overexpressed in T-cell Acute Lymphoblastic samples in comparison to their expression levels in normal cells. Of these, we found three novel genes which have not been reported earlier- KAT2B, PSEN1 (underexpressed) and CDH2 (overexpressed).These three identified genes may provide new insights into the abnormal hematopoietic process observed in Leukemia as these genes are involved in Notch signaling and cell adhesion processes. It is evident that experimental validation of the protein interactors in leukemic cells could help in the identification of new diagnostic markers for leukemia. PMID:24688641

  18. The disintegrin/metalloproteinase Adam10 is essential for epidermal integrity and Notch-mediated signaling.

    PubMed

    Weber, Silvio; Niessen, Michaela T; Prox, Johannes; Lüllmann-Rauch, Renate; Schmitz, Annika; Schwanbeck, Ralf; Blobel, Carl P; Jorissen, Ellen; de Strooper, Bart; Niessen, Carien M; Saftig, Paul

    2011-02-01

    The disintegrin and metalloproteinase Adam10 has been implicated in the regulation of key signaling pathways that determine skin morphogenesis and homeostasis. To address the in vivo relevance of Adam10 in the epidermis, we have selectively disrupted Adam10 during skin morphogenesis and in adult skin. K14-Cre driven epidermal Adam10 deletion leads to perinatal lethality, barrier impairment and absence of sebaceous glands. A reduction of spinous layers, not associated with differences in either proliferation or apoptosis, indicates that loss of Adam10 triggers a premature differentiation of spinous keratinocytes. The few surviving K14-Adam10-deleted mice and mice in which Adam10 was deleted postnatally showed loss of hair, malformed vibrissae, epidermal hyperproliferation, cyst formation, thymic atrophy and upregulation of the cytokine thymic stromal lymphopoetin (TSLP), thus indicating non cell-autonomous multi-organ disease resulting from a compromised barrier. Together, these phenotypes closely resemble skin specific Notch pathway loss-of-function phenotypes. Notch processing is indeed strongly reduced resulting in decreased levels of Notch intracellular domain fragment and functional Notch signaling. The data identify Adam10 as the major Site-2 processing enzyme for Notch in the epidermis in vivo, and thus as a central regulator of skin development and maintenance.

  19. Numb/Notch signaling pathway modulation enhances human pancreatic cancer cell radiosensitivity.

    PubMed

    Bi, Yi-Liang; Min, Min; Shen, Wei; Liu, Yan

    2016-11-01

    The present study aims to evaluate whether repression of the Numb/Notch signaling pathway affects the radiosensitivity of human pancreatic cancer cell lines. Different doses of X-rays (0, 2, 3, 4, and 5 Gy) were applied to the PANC-1, SW1990, and MIA PaCa-2 human pancreatic cancer cell lines, and the Numb/Notch pathway inhibitor DAPT was added at different doses (0, 1, 3, and 5 μmol/l). MTT assay, colony formation assay, flow cytometry, scratch assay, and Transwell experiments were performed, and qRT-PCR and Western blot were conducted for the detection of Numb expression. Tumorigenicity assay in nude mice was carried out to verify the influence of blocker of the Numb/Notch signaling pathway on the radiosensitivity of xenograft tumors. The MTT assay, colony formation assay and flow cytometry experiments revealed that proliferation decreased as radiation dose increased. The viability of PANC-1 cells at 5 Gy, SW 1990 cells at 4 Gy and 5 Gy, and MIA PaCa-2 cells at 2-5 Gy was significantly lower than that of non-irradiated cells (all P < 0.05). The migration and invasion assays indicated that the PANC-1 cell line was least radiosensitive, while the MIA PaCa-2 cell line was the most radiosensitive. Numb expression significantly increased with increasing radiation dose, whereas the expression of Hes1, Notch1, and Hes5 significantly decreased compared to non-irradiated cells (P < 0.05). Compared to untreated control cells, DAPT dose dependently increased Numb expression and inhibited Notch1, Hes1, and Hes5 expressions at 2 Gy (P < 0.05). Subcutaneous tumorigenicity assay in nude mice demonstrated that DAPT increased the radiosensitivity of PANC-1, SW 1990, and MIA PaCa-2 cells. These findings suggest that Numb/Notch signaling in pancreatic cancer cells is associated with X-ray radiation and that inhibition of the Numb/Notch signaling pathway can enhance radiosensitivity, suggesting that inhibition of the Numb/Notch signaling pathway may serve as a potential

  20. Repression of Ccr9 transcription in mouse T lymphocyte progenitors by the Notch signaling pathway.

    PubMed

    Krishnamoorthy, Veena; Carr, Tiffany; de Pooter, Renee F; Emanuelle, Akinola Olumide; Akinola, Emanuelle Olumide; Gounari, Fotini; Kee, Barbara L

    2015-04-01

    The chemokine receptor CCR9 controls the immigration of multipotent hematopoietic progenitor cells into the thymus to sustain T cell development. Postimmigration, thymocytes downregulate CCR9 and migrate toward the subcapsular zone where they recombine their TCR β-chain and γ-chain gene loci. CCR9 is subsequently upregulated and participates in the localization of thymocytes during their selection for self-tolerant receptor specificities. Although the dynamic regulation of CCR9 is essential for early T cell development, the mechanisms controlling CCR9 expression have not been determined. In this article, we show that key regulators of T cell development, Notch1 and the E protein transcription factors E2A and HEB, coordinately control the expression of Ccr9. E2A and HEB bind at two putative enhancers upstream of Ccr9 and positively regulate CCR9 expression at multiple stages of T cell development. In contrast, the canonical Notch signaling pathway prevents the recruitment of p300 to the putative Ccr9 enhancers, resulting in decreased acetylation of histone H3 and a failure to recruit RNA polymerase II to the Ccr9 promoter. Although Notch signaling modestly modulates the binding of E proteins to one of the two Ccr9 enhancers, we found that Notch signaling represses Ccr9 in T cell lymphoma lines in which Ccr9 transcription is independent of E protein function. Our data support the hypothesis that activation of Notch1 has a dominant-negative effect on Ccr9 transcription and that Notch1 and E proteins control the dynamic expression of Ccr9 during T cell development.

  1. Curcumin inhibits the survival and metastasis of prostate cancer cells via the Notch-1 signaling pathway.

    PubMed

    Yang, Jingzhe; Wang, Chengli; Zhang, Zhijie; Chen, Xiaojun; Jia, Yusen; Wang, Bin; Kong, Tao

    2017-02-01

    Prostate cancer is one of the most common malignancies in men, and it urgently demands precise interventions that target the signaling pathways implicated in its initiation, progression, and metastasis. The Notch-1 signaling pathway is closely associated with the pathophysiology of prostate cancer. This study investigated the antitumor effects and mechanisms of curcumin, which is a well-known natural compound from curcuminoids, in prostate cancer cells. Viability, proliferation, and migration were analyzed in two prostate cancer cell lines, DU145 and PC3, after curcumin treatment. Whether the Notch-1 signaling pathway is involved in the antitumor effects of curcumin was examined. Curcumin inhibited the survival and proliferation of PC3 and DU145 cells in a dose- and time-dependent manner and inhibited DU145 migration. Curcumin did not affect the expression of Notch-1 or its active product NICD, but it did inhibit the expression of MT1-MMP and MMP2 proteins in DU145 cells. We found that curcumin inhibited the DNA-binding ability of NICD in DU145 cells. In conclusion, curcumin inhibited the survival and metastasis of prostate cancer cells via the Notch-1 signaling pathway.

  2. A Presenilin-2–ARF4 trafficking axis modulates Notch signaling during epidermal differentiation

    PubMed Central

    Ezratty, Ellen J.; Pasolli, H. Amalia

    2016-01-01

    How primary cilia impact epidermal growth and differentiation during embryogenesis is poorly understood. Here, we show that during skin development, Notch signaling occurs within the ciliated, differentiating cells of the first few suprabasal epidermal layers. Moreover, both Notch signaling and cilia disappear in the upper layers, where key ciliary proteins distribute to cell–cell borders. Extending this correlation, we find that Presenilin-2 localizes to basal bodies/cilia through a conserved VxPx motif. When this motif is mutated, a GFP-tagged Presenilin-2 still localizes to intercellular borders, but basal body localization is lost. Notably, in contrast to wild type, this mutant fails to rescue epidermal differentiation defects seen upon Psen1 and 2 knockdown. Screening components implicated in ciliary targeting and polarized exocytosis, we provide evidence that the small GTPase ARF4 is required for Presenilin basal body localization, Notch signaling, and subsequent epidermal differentiation. Collectively, our findings raise the possibility that ARF4-dependent polarized exocytosis acts through the basal body–ciliary complex to spatially regulate Notch signaling during epidermal differentiation. PMID:27354375

  3. Notch Signaling and Schwann Cell Transformation: Development of a Model System and Application to Human MPNSTs

    DTIC Science & Technology

    2006-03-01

    MPNSTs PRINCIPAL INVESTIGATOR: Tom R. Kadesch, Ph.D...Signaling and Schwann Cell Tranformation: Development of a Model System and Application to Human MPNSTs 6. AUTHOR(S) Tom R. Kadesch, Ph.D. W81XWH-04-1...the malignant transformation of neurofibromas to MPNSTs in patients with NF1. Our previous work has shown that constitutive expression of Notch can

  4. ROS, Notch, and Wnt signaling pathways: crosstalk between three major regulators of cardiovascular biology.

    PubMed

    Caliceti, C; Nigro, P; Rizzo, P; Ferrari, R

    2014-01-01

    Reactive oxygen species (ROS), traditionally viewed as toxic by-products that cause damage to biomolecules, now are clearly recognized as key modulators in a variety of biological processes and pathological states. The development and regulation of the cardiovascular system require orchestrated activities; Notch and Wnt/β -catenin signaling pathways are implicated in many aspects of them, including cardiomyocytes and smooth muscle cells survival, angiogenesis, progenitor cells recruitment and differentiation, arteriovenous specification, vascular cell migration, and cardiac remodelling. Several novel findings regarding the role of ROS in Notch and Wnt/β-catenin modulation prompted us to review their emerging function in the cardiovascular system during embryogenesis and postnatally.

  5. CADASIL-associated Notch3 mutations have differential effects both on ligand binding and ligand-induced Notch3 receptor signaling through RBP-Jk.

    PubMed

    Peters, Nils; Opherk, Christian; Zacherle, Simone; Capell, Anja; Gempel, Petra; Dichgans, Martin

    2004-10-01

    Mutations in the NOTCH3 gene are the cause of cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL), a hereditary angiopathy leading to strokes and dementia. Pathogenic mutations remove or insert cysteine residues within epidermal growth factor (EGF) repeats in the extracellular domain of the Notch3 receptor (N3ECD). Vascular smooth muscle cells (VSMC) are the predominant site of Notch3 expression in adults. In CADASIL patients, VSMC degenerate and N3ECD is deposited within the vasculature. However, the mechanisms underlying VSMC degeneration and N3ECD accumulation are still unknown. In this study, we investigated the consequences of three pathogenic Notch3 mutations on the biological activity of the receptor by analyzing ligand (Delta-/Jagged-)-induced signaling via RBP-Jk. Two mutations (R133C and C183R) that are located outside the putative ligand binding domain (LBD) of the receptor were found to result in normal Jagged1-induced signaling in A7r5 VSMC, whereas the third mutation (C455R located within the putative LBD) showed strongly reduced signaling activity. Ligand binding assays with soluble Delta1 and Jagged1 revealed that C455R interferes with ligand binding through disruption of the LBD which, as we show here, is located in EGF repeats 10/11 of Notch3. All mutant receptors including Notch3C455R were targeted to the cell surface but showed an elevated ratio between the unprocessed full-length 280-kDa receptor and S1-cleaved receptor fragments. Taken together, these data indicate that CADASIL-associated Notch3 mutations differ with respect to their consequences both on ligand binding and ligand-induced signaling through RBP-Jk, whereas they have similar effects on receptor maturation. Moreover, the data suggest that ligand-induced receptor shedding may not be required for N3ECD deposition in CADASIL.

  6. Notch3 activation modulates cell growth behaviour and cross-talk to Wnt/TCF signalling pathway.

    PubMed

    Wang, Tao; Holt, Cathy M; Xu, Chiheng; Ridley, Caroline; P O Jones, Richard; Baron, Martin; Trump, Dorothy

    2007-12-01

    Notch3 is one of the four Notch receptors identified in mammal and expressed mainly in the arterial smooth muscle cells of human adult. Signalling via Notch3 is thought to be important in maintaining the phenotypic stability of the cells, but the nature of the signalling and its regulation to other signalling pathways are largely unknown. To understand further of the cellular function of Notch3 signalling, we generated cell lines stably expressing a constitutively active form of human Notch3 comprising of its soluble intracellular domain (N3IC). The N3IC expressing cells showed accelerated proliferation, decreased migration, increased cell surface N-cadherin, and growth in a colonised fashion that was reversible by N-cadherin blockade. N3IC expressing cells were also protected significantly against staurosporine-induced apoptosis and exhibited lower caspase 3/7 activity, accompanied by up-regulation of pAKT compared to control cells. We also found a complex cross-talk between Notch3 signalling and the Wnt pathway. N3IC stimulated Wnt-independent T-cell factor (TCF, the target transcription factor in the Wnt pathway) activation which was associated with increased Tyr-142 phosphorylation of beta-catenin. In contrast N3IC suppressed TCF activation in response to LiCl, which mimics the Wnt-dependent TCF activation mechanism. We conclude that Notch3 promotes cell growth and survival by activating PI3-kinase/AKT pathway; N-cadherin participates in the change of cell growth caused by Notch3 activation; and Notch3 signalling has dual-effects on the Wnt/TCF pathway suggesting a buffering role that Notch3 signalling may play in balancing these two important signalling pathways in regulating cell function.

  7. Targeting Tumor Initiating Cells through Inhibition of Cancer Testis Antigens and Notch Signaling: A Hypothesis.

    PubMed

    Colombo, Michela; Mirandola, Leonardo; Reidy, Adair; Suvorava, Natallia; Konala, Venu; Chiaramonte, Raffaella; Grizzi, Fabio; Rahman, Rakhshanda Layeequr; Jenkins, Marjorie R; Nugyen, Diane D; Dalhbeck, Scott; Cobos, Everardo; Figueroa, Jose A; Chiriva-Internati, Maurizio

    2015-03-01

    Tumor initiating cells (TICs) differ from normal stem cells (SCs) in their ability to initiate tumorigenesis, invasive growth, metastasis and the acquisition of chemo and/or radio-resistance. Over the past years, several studies have indicated the potential role of the Notch system as a key regulator of cellular stemness and tumor development. Furthermore, the expression of cancer testis antigens (CTA) in TICs, and their role in SC differentiation and biology, has become an important area of investigation. Here, we propose a model in which CTA expression and Notch signaling interacts to maintain the sustainability of self-replicating tumor populations, ultimately leading to the development of metastasis, drug resistance and cancer progression. We hypothesize that Notch-CTA interactions in TICs offer a novel opportunity for meaningful therapeutic interventions in cancer.

  8. Regulation of monocyte cell fate by blood vessels mediated by Notch signalling

    PubMed Central

    Gamrekelashvili, Jaba; Giagnorio, Roberto; Jussofie, Jasmin; Soehnlein, Oliver; Duchene, Johan; Briseño, Carlos G.; Ramasamy, Saravana K.; Krishnasamy, Kashyap; Limbourg, Anne; Kapanadze, Tamar; Ishifune, Chieko; Hinkel, Rabea; Radtke, Freddy; Strobl, Lothar J.; Zimber-Strobl, Ursula; Napp, L. Christian; Bauersachs, Johann; Haller, Hermann; Yasutomo, Koji; Kupatt, Christian; Murphy, Kenneth M.; Adams, Ralf H.; Weber, Christian; Limbourg, Florian P.

    2016-01-01

    A population of monocytes, known as Ly6Clo monocytes, patrol blood vessels by crawling along the vascular endothelium. Here we show that endothelial cells control their origin through Notch signalling. Using combinations of conditional genetic deletion strategies and cell-fate tracking experiments we show that Notch2 regulates conversion of Ly6Chi monocytes into Ly6Clo monocytes in vivo and in vitro, thereby regulating monocyte cell fate under steady-state conditions. This process is controlled by Notch ligand delta-like 1 (Dll1) expressed by a population of endothelial cells that constitute distinct vascular niches in the bone marrow and spleen in vivo, while culture on recombinant DLL1 induces monocyte conversion in vitro. Thus, blood vessels regulate monocyte conversion, a form of committed myeloid cell fate regulation. PMID:27576369

  9. Notch Signaling Modulates the Balance of Regulatory T Cells and T Helper 17 Cells in Patients with Chronic Hepatitis C.

    PubMed

    Qin, Lei; Zhou, Yan-Cai; Wu, Hong-Jie; Zhuo, Ya; Wang, Yan-Ping; Si, Chang-Yun; Qin, Yong-Mei

    2017-04-01

    The imbalance of regulatory T cells (Tregs) and T helper 17 (Th17) cells contributes to the persistent hepatitis C virus (HCV) infection. However, modulatory factors associated with Tregs-Th17 balance were not fully elucidated. A recent study demonstrated an immunoregulatory strategy by inactivation of Notch signaling to reverse the disequilibrium of Tregs-Th17 cells in immune thrombocytopenia. Thus, the aim of this study was to assess the effect of Notch signaling in regulating the functions of Tregs and Th17 cells in chronic hepatitis C. A total of 46 patients with chronic hepatitis C and 17 normal controls (NCs) were enrolled. mRNA expressions of Notch1 and Notch2 were semiquantified by real-time reserve polymerase chain reaction. Percentages of Tregs-Th17, levels of key transcriptional factors, and cytokine productions were measured in response to treatment by DAPT, a γ-secretase inhibitor to suppress Notch signaling. We found that Notch1 and Notch2 mRNAs were significantly elevated in peripheral blood mononuclear cells from chronic hepatitis C patients compared with those from NCs. DAPT treatment reduced Th17 response by downregulation of RORγt expression and interleukin (IL)-17/IL-22 secretion. Tregs proportion, FoxP3 expression, and IL-10 production did not change significantly with DAPT treatment in chronic hepatitis C; however, blockage of Notch signaling inhibited the suppressive function of Tregs. Moreover, effective anti-HCV therapy not only reduced Notch1 and Notch2 expression but also decreased Tregs and Th17 proportions. The current data provided a novel mechanism underlying the modulation of Treg-Th17 balance. The link between Notch signaling and Th cells might lead to a new intervention for breaking immunotolerance of chronic HCV infection.

  10. Systemic Inhibition of Canonical Notch Signaling Results in Sustained Callus Inflammation and Alters Multiple Phases of Fracture Healing

    PubMed Central

    Dishowitz, Michael I.; Mutyaba, Patricia L.; Takacs, Joel D.; Barr, Andrew M.; Engiles, Julie B.; Ahn, Jaimo; Hankenson, Kurt D.

    2013-01-01

    The Notch signaling pathway is an important regulator of embryological bone development, and many aspects of development are recapitulated during bone repair. We have previously reported that Notch signaling components are upregulated during bone fracture healing. However, the significance of the Notch pathway in bone regeneration has not been described. Therefore, the objective of this study was to determine the importance of Notch signaling in regulating bone fracture healing by using a temporally controlled inducible transgenic mouse model (Mx1-Cre;dnMAMLf/-) to impair RBPjκ-mediated canonical Notch signaling. The Mx1 promoter was synthetically activated resulting in temporally regulated systemic dnMAML expression just prior to creation of bilateral tibial fractures. This allowed for mice to undergo unaltered embryological and post-natal skeletal development. Results showed that systemic Notch inhibition prolonged expression of inflammatory cytokines and neutrophil cell inflammation, and reduced the proportion of cartilage formation within the callus at 10 days-post-fracture (dpf) Notch inhibition did not affect early bone formation at 10dpf, but significantly altered bone maturation and remodeling at 20dpf. Increased bone volume fraction in dnMAML fractures, which was due to a moderate decrease in callus size with no change in bone mass, coincided with increased trabecular thickness but decreased connectivity density, indicating that patterning of bone was altered. Notch inhibition decreased total osteogenic cell density, which was comprised of more osteocytes rather than osteoblasts. dnMAML also decreased osteoclast density, suggesting that osteoclast activity may also be important for altered fracture healing. It is likely that systemic Notch inhibition had both direct effects within cell types as well as indirect effects initiated by temporally upstream events in the fracture healing cascade. Surprisingly, Notch inhibition did not alter cell proliferation

  11. Notch1, Notch2, and Epstein-Barr virus-encoded nuclear antigen 2 signaling differentially affects proliferation and survival of Epstein-Barr virus-infected B cells.

    PubMed

    Kohlhof, Hella; Hampel, Franziska; Hoffmann, Reinhard; Burtscher, Helmut; Weidle, Ulrich H; Hölzel, Michael; Eick, Dirk; Zimber-Strobl, Ursula; Strobl, Lothar J

    2009-05-28

    The canonical mode of transcriptional activation by both the Epstein-Barr viral protein, Epstein-Barr virus-encoded nuclear antigen 2 (EBNA2), and an activated Notch receptor (Notch-IC) requires their recruitment to RBPJ, suggesting that EBNA2 uses the Notch pathway to achieve B-cell immortalization. To gain further insight into the biologic equivalence between Notch-IC and EBNA2, we performed a genome-wide expression analysis, revealing that Notch-IC and EBNA2 exhibit profound differences in the regulation of target genes. Whereas Notch-IC is more potent in regulating genes associated with differentiation and development, EBNA2 is more potent in inducing viral and cellular genes involved in proliferation, survival, and chemotaxis. Because both EBNA2 and Notch-IC induced the expression of cell cycle-associated genes, we analyzed whether Notch1-IC or Notch2-IC can replace EBNA2 in B-cell immortalization. Although Notch-IC could drive quiescent B cells into the cell cycle, B-cell immortalization was not maintained, partially due to an increased apoptosis rate in Notch-IC-expressing cells. Expression analysis revealed that both EBNA2 and Notch-IC induced the expression of proapoptotic genes, but only in EBNA2-expressing cells were antiapoptotic genes strongly up-regulated. These findings suggest that Notch signaling in B cells and B-cell lymphomas is only compatible with proliferation if pathways leading to antiapototic signals are active.

  12. The segmentation clock in mice: interaction between the Wnt and Notch signalling pathways.

    PubMed

    Rodríguez-González, J G; Santillán, M; Fowler, A C; Mackey, Michael C

    2007-09-07

    In the last few years, the efforts to elucidate the mechanisms underlying the segmentation clock in various vertebrate species have multiplied. Early evidence suggested that oscillations are caused by one of the genes under the Notch signalling pathway (like those of the her or Hes families). Recently, Aulehla et al. [Wnt3a plays a major role in the segmentation clock controlling somitogenesis. Dev. Cell 4, 395-406] discovered that Axin2 (a gene under the Wnt3a signalling pathway) also oscillates in the presomitic mesoderm (PSM) of mice embryos and proposed some mechanisms through which the Notch and Wnt3a pathways may interact. They further suggested that a decreasing concentration of Wnt3a along the PSM may be the gradient the segmentation clock interacts with to form somites. These results were reviewed by Rida et al. [A notch feeling of somite segmentation and beyond. Dev. Biol. 265, 2-22], who introduced a complex clockwork comprising genes Hes1, Lfng (under the Notch pathway), and Axin2, as well as their multiple interactions. In the present work we develop a mathematical model based on the Rida et al. review and use it to tackle some of the questions raided by the Aulehla et al. paper: can the Axin2 feedback loop constitute a clock? Could a decreasing Wnt3a signaling constitute the wavefront, where phase is recorded and the spatial pattern laid down? What is the master oscillator?

  13. Expanding role of delta-like 4 mediated notch signaling in cardiovascular and metabolic diseases.

    PubMed

    Fukuda, Daiju; Aikawa, Masanori

    2013-01-01

    Cardiometabolic disease, a global health threat, has been linked to chronic inflammation, in which activated macrophages play a key role. Macrophages are highly heterogeneous hematopoietic cells found in nearly every tissue in the body. Various stimuli recruit monocytes into the cardiovascular system and metabolic organs, where they differentiate to macrophages, and activate these pro-inflammatory phagocytes, leading to the initiation and development of inflammation in these organs. Key regulators of macrophage activation therefore may serve as therapeutic targets for cardiometabolic disease. The Notch signaling pathway, involving 5 ligands and 4 receptors, regulates the differentiation of various cell types during development, and also contributes to the disease processes in adults. We found that the Notch ligand delta-like 4 (Dll4) activates macrophages in vitro as determined by the induction of genes and pathways associated with cardiovascular and metabolic disorders. Our recent study demonstrated in vivo that blockade of Dll4 by a neutralizing antibody attenuates key features typical of cardiovascular and metabolic diseases, such as accumulation of activated macrophages in arteries and fat; chronic atherosclerosis; arterial and valvular calcification; insulin resistance; and fatty liver. These results suggest that Dll4-mediated Notch signaling participates in the shared disease mechanisms for cardiovascular and metabolic disorders. This review summarizes the role of macrophages and Dll4/Notch signaling in the development of inflammation in both the cardiovascular system and metabolic organs. 

  14. Myoblast cytonemes mediate Wg signaling from the wing imaginal disc and Delta-Notch signaling to the air sac primordium

    PubMed Central

    Huang, Hai; Kornberg, Thomas B

    2015-01-01

    The flight muscles, dorsal air sacs, wing blades, and thoracic cuticle of the Drosophila adult function in concert, and their progenitor cells develop together in the wing imaginal disc. The wing disc orchestrates dorsal air sac development by producing decapentaplegic and fibroblast growth factor that travel via specific cytonemes in order to signal to the air sac primordium (ASP). Here, we report that cytonemes also link flight muscle progenitors (myoblasts) to disc cells and to the ASP, enabling myoblasts to relay signaling between the disc and the ASP. Frizzled (Fz)-containing myoblast cytonemes take up Wingless (Wg) from the disc, and Delta (Dl)-containing myoblast cytonemes contribute to Notch activation in the ASP. Wg signaling negatively regulates Dl expression in the myoblasts. These results reveal an essential role for cytonemes in Wg and Notch signaling and for a signal relay system in the myoblasts. DOI: http://dx.doi.org/10.7554/eLife.06114.001 PMID:25951303

  15. Alterations in Notch signalling in skeletal muscles from mdx and dko dystrophic mice and patients with Duchenne muscular dystrophy.

    PubMed

    Church, Jarrod E; Trieu, Jennifer; Chee, Annabel; Naim, Timur; Gehrig, Stefan M; Lamon, Séverine; Angelini, Corrado; Russell, Aaron P; Lynch, Gordon S

    2014-04-01

    New Findings What is the central question of this study? The Notch signalling pathway plays an important role in muscle regeneration, and activation of the pathway has been shown to enhance muscle regeneration in aged mice. It is unknown whether Notch activation will have a similarly beneficial effect on muscle regeneration in the context of Duchenne muscular dystrophy (DMD). What is the main finding and its importance? Although expression of Notch signalling components is altered in both mouse models of DMD and in human DMD patients, activation of the Notch signalling pathway does not confer any functional benefit on muscles from dystrophic mice, suggesting that other signalling pathways may be more fruitful targets for manipulation in treating DMD. Abstract In Duchenne muscular dystrophy (DMD), muscle damage and impaired regeneration lead to progressive muscle wasting, weakness and premature death. The Notch signalling pathway represents a central regulator of gene expression and is critical for cellular proliferation, differentiation and apoptotic signalling during all stages of embryonic muscle development. Notch activation improves muscle regeneration in aged mice, but its potential to restore regeneration and function in muscular dystrophy is unknown. We performed a comprehensive examination of several genes involved in Notch signalling in muscles from dystrophin-deficient mdx and dko (utrophin- and dystrophin-null) mice and DMD patients. A reduction of Notch1 and Hes1 mRNA in tibialis anterior muscles of dko mice and quadriceps muscles of DMD patients and a reduction of Hes1 mRNA in the diaphragm of the mdx mice were observed, with other targets being inconsistent across species. Activation and inhibition of Notch signalling, followed by measures of muscle regeneration and function, were performed in the mouse models of DMD. Notch activation had no effect on functional regeneration in C57BL/10, mdx or dko mice. Notch inhibition significantly depressed the

  16. Notch signaling acts before cell division to promote asymmetric cleavage and cell fate of neural precursor cells.

    PubMed

    Bhat, Krishna Moorthi

    2014-10-21

    Asymmetric cell divisions in the central nervous system generate neurons of diverse fates. In Drosophila melanogaster, the protein Numb localizes asymmetrically to dividing neural precursor cells such that only one daughter cell inherits Numb. Numb inhibits Notch signaling in this daughter cell, resulting in a different cell fate from the Notch-induced fate in the other-Numb-negative-daughter cell. Precursor cells undergo asymmetric cytokinesis generating daughter cells of different sizes. I found that inactivation of Notch in fly embryonic neural precursor cells disrupted the asymmetric positioning of the cleavage furrow and produced daughter cells of the same size and fate. Moreover, inactivation of Notch at different times altered the degree of asymmetric Numb localization, such that earlier inactivation of Notch caused symmetric distribution of Numb and later inactivation produced incomplete asymmetric localization of Numb. The extent of asymmetrically localized Numb positively correlated with the degree of asymmetric cytokinesis and the size disparity in daughter cells. Loss of Numb or expression of constitutively active Notch led to premature specification of the precursor cells into the fate of one of the daughter cells. Thus, in addition to its role in the specification of daughter cell fate after division, Notch controls Numb localization in the precursor cells to determine the size and fate of daughter cells. Numb also inhibits Notch signaling in precursor cells to prevent Notch-induced differentiation of the precursor cell, forming an autoregulatory loop.

  17. Tripartite interactions between Wnt signaling, Notch and Myb for stem/progenitor cell functions during intestinal tumorigenesis.

    PubMed

    Germann, Markus; Xu, Huiling; Malaterre, Jordane; Sampurno, Shienny; Huyghe, Mathilde; Cheasley, Dane; Fre, Silvia; Ramsay, Robert G

    2014-11-01

    Deletion studies confirm Wnt, Notch and Myb transcriptional pathway engagement in intestinal tumorigenesis. Nevertheless, their contrasting and combined roles when activated have not been elucidated. This is important as these pathways are not ablated but rather are aberrantly activated during carcinogenesis. Using ApcMin/+ mice as a source of organoids we documented their transition, on a clone-by-clone basis, to cyst-like spheres with constitutively activated Wnt pathway, increased self-renewal and growth and reduced differentiation. We then looked at this transition when Myb and/or Notch1 are activated. Activated Notch promoted cyst-like organoids. Conversely growth and propagation of cyst-like, but not normal organoids were Notch-independent. Activated Myb promoted normal, but not cyst-like organoids. Interestingly the Wnt, Notch and Myb pathways were all involved in regulating the expression of the intestinal stem cell (ISC) gene Lgr5 in organoids, while ISC gene and Notch target Olfm4 was dominantly repressed by Wnt. These findings parallel mouse intestinal adenoma formation where Notch promoted the initiation, but not growth, of Wnt-driven Olfm4-repressed colon tumors. Also Myb was essential for colon tumor initiation and collateral mouse pathologies. These data reveal the complex interplay and hierarchy of transcriptional networks that operate in ISCs and uncover a shift in pathway-dependencies during tumor initiation.

  18. Genome-wide expression profiling in the Drosophila eye reveals unexpected repression of Notch signaling by the JAK/STAT pathway

    PubMed Central

    Flaherty, Maria Sol; Zavadil, Jiri; Ekas, Laura A.; Bach, Erika A.

    2010-01-01

    Although the JAK/STAT pathway regulates numerous processes in vertebrates and invertebrates through modulating transcription, its functionally-relevant transcriptional targets remain largely unknown. With one jak and one stat (stat92E), Drosophila provides a powerful system for finding new JAK/STAT target genes. Genome-wide expression profiling on eye discs in which Stat92E is hyperactivated, revealed 584 differentially-regulated genes, including known targets domeless, socs36E and wingless. Other differentially-regulated genes (chinmo, lama, Mo25, Imp-L2, Serrate, Delta) were validated and may represent new Stat92E targets. Genetic experiments revealed that Stat92E cell-autonomously represses Serrate, which encodes a Notch ligand. Loss of Stat92E led to de-repression of Serrate in the dorsal eye, resulting in ectopic Notch signaling and aberrant eye growth there. Thus, our micro-array documents a new Stat92E target gene and a previously-unidentified inhibitory action of Stat92E on Notch signaling. These data suggest that this study will be a useful resource for the identification of additional Stat92E targets. PMID:19504457

  19. O-fucose monosaccharide of Drosophila Notch has a temperature-sensitive function and cooperates with O-glucose glycan in Notch transport and Notch signaling activation.

    PubMed

    Ishio, Akira; Sasamura, Takeshi; Ayukawa, Tomonori; Kuroda, Junpei; Ishikawa, Hiroyuki O; Aoyama, Naoki; Matsumoto, Kenjiroo; Gushiken, Takuma; Okajima, Tetsuya; Yamakawa, Tomoko; Matsuno, Kenji

    2015-01-02

    Notch (N) is a transmembrane receptor that mediates the cell-cell interactions necessary for many cell fate decisions. N has many epidermal growth factor-like repeats that are O-fucosylated by the protein O-fucosyltransferase 1 (O-Fut1), and the O-fut1 gene is essential for N signaling. However, the role of the monosaccharide O-fucose on N is unclear, because O-Fut1 also appears to have O-fucosyltransferase activity-independent functions, including as an N-specific chaperon. Such an enzymatic activity-independent function could account for the essential role of O-fut1 in N signaling. To evaluate the role of the monosaccharide O-fucose modification in N signaling, here we generated a knock-in mutant of O-fut1 (O-fut1(R245A knock-in)), which expresses a mutant protein that lacks O-fucosyltransferase activity but maintains the N-specific chaperon activity. Using O-fut1(R245A knock-in) and other gene mutations that abolish the O-fucosylation of N, we found that the monosaccharide O-fucose modification of N has a temperature-sensitive function that is essential for N signaling. The O-fucose monosaccharide and O-glucose glycan modification, catalyzed by Rumi, function redundantly in the activation of N signaling. We also showed that the redundant function of these two modifications is responsible for the presence of N at the cell surface. Our findings elucidate how different forms of glycosylation on a protein can influence the protein's functions.

  20. Sex steroid blockade enhances thymopoiesis by modulating Notch signaling

    PubMed Central

    Tsai, Jennifer J.; Holland, Amanda M.; Wertheimer, Tobias; Yu, Vionnie W.C.; Zakrzewski, Johannes L.; Tuckett, Andrea Z.; Singer, Natalie V.; West, Mallory L.; Smith, Odette M.; Young, Lauren F.; Kreines, Fabiana M.; Levy, Emily R.; Boyd, Richard L.; Scadden, David T.

    2014-01-01

    Paradoxical to its importance for generating a diverse T cell repertoire, thymic function progressively declines throughout life. This process has been at least partially attributed to the effects of sex steroids, and their removal promotes enhanced thymopoiesis and recovery from immune injury. We show that one mechanism by which sex steroids influence thymopoiesis is through direct inhibition in cortical thymic epithelial cells (cTECs) of Delta-like 4 (Dll4), a Notch ligand crucial for the commitment and differentiation of T cell progenitors in a dose-dependent manner. Consistent with this, sex steroid ablation (SSA) led to increased expression of Dll4 and its downstream targets. Importantly, SSA induced by luteinizing hormone-releasing hormone (LHRH) receptor antagonism bypassed the surge in sex steroids caused by LHRH agonists, the gold standard for clinical ablation of sex steroids, thereby facilitating increased Dll4 expression and more rapid promotion of thymopoiesis. Collectively, these findings not only reveal a novel mechanism underlying improved thymic regeneration upon SSA but also offer an improved clinical strategy for successfully boosting immune function. PMID:25332287

  1. KCTD10 is involved in the cardiovascular system and Notch signaling during early embryonic development.

    PubMed

    Ren, Kaiqun; Yuan, Jing; Yang, Manjun; Gao, Xiang; Ding, Xiaofeng; Zhou, Jianlin; Hu, Xingwang; Cao, Jianguo; Deng, Xiyun; Xiang, Shuanglin; Zhang, Jian

    2014-01-01

    As a member of the polymerase delta-interacting protein 1 (PDIP1) gene family, potassium channel tetramerisation domain-containing 10 (KCTD10) interacts with proliferating cell nuclear antigen (PCNA) and polymerase δ, participates in DNA repair, DNA replication and cell-cycle control. In order to further investigate the physiological functions of KCTD10, we generated the KCTD10 knockout mice. The heterozygous KCTD10(+/-) mice were viable and fertile, while the homozygous KCTD10(-/-) mice showed delayed growth from E9.0, and died at approximately E10.5, which displayed severe defects in angiogenesis and heart development. Further study showed that VEGF induced the expression of KCTD10 in a time- and dose-dependent manner. Quantitative real-time PCR and western blotting results revealed that several key members in Notch signaling were up-regulated either in KCTD10-deficient embryos or in KCTD10-silenced HUVECs. Meanwhile, the endogenous immunoprecipitation (IP) analysis showed that KCTD10 interacted with Cullin3 and Notch1 simultaneously, by which mediating Notch1 proteolytic degradation. Our studies suggest that KCTD10 plays crucial roles in embryonic angiogenesis and heart development in mammalians by negatively regulating the Notch signaling pathway.

  2. Genomics and expression profiles of the Hedgehog and Notch signaling pathways in sea urchin development.

    PubMed

    Walton, Katherine D; Croce, Jenifer C; Glenn, Thomas D; Wu, Shu-Yu; McClay, David R

    2006-12-01

    The Hedgehog (Hh) and Notch signal transduction pathways control a variety of developmental processes including cell fate choice, differentiation, proliferation, patterning and boundary formation. Because many components of these pathways are conserved, it was predicted and confirmed that pathway components are largely intact in the sea urchin genome. Spatial and temporal location of these pathways in the embryo, and their function in development offer added insight into their mechanistic contributions. Accordingly, all major components of both pathways were identified and annotated in the sea urchin Strongylocentrotus purpuratus genome and the embryonic expression of key components was explored. Relationships of the pathway components, and modifiers predicted from the annotation of S. purpuratus, were compared against cnidarians, arthropods, urochordates, and vertebrates. These analyses support the prediction that the pathways are highly conserved through metazoan evolution. Further, the location of these two pathways appears to be conserved among deuterostomes, and in the case of Notch at least, display similar capacities in endomesoderm gene regulatory networks. RNA expression profiles by quantitative PCR and RNA in situ hybridization reveal that Hedgehog is produced by the endoderm beginning just prior to invagination, and signals to the secondary mesenchyme-derived tissues at least until the pluteus larva stage. RNA in situ hybridization of Notch pathway members confirms that Notch functions sequentially in the vegetal-most secondary mesenchyme cells and later in the endoderm. Functional analyses in future studies will embed these pathways into the growing knowledge of gene regulatory networks that govern early specification and morphogenesis.

  3. TNF-alpha and Notch signaling regulates the expression of HOXB4 and GATA3 during early T lymphopoiesis.

    PubMed

    Dos Santos Schiavinato, Josiane Lilian; Oliveira, Lucila Habib Bourguignon; Araujo, Amélia Goes; Orellana, Maristela Delgado; de Palma, Patrícia Viana Bonini; Covas, Dimas Tadeu; Zago, Marco Antonio; Panepucci, Rodrigo Alexandre

    2016-10-01

    During the early thymus colonization, Notch signaling activation on hematopoietic progenitor cells (HPCs) drives proliferation and T cell commitment. Although these processes are driven by transcription factors such as HOXB4 and GATA3, there is no evidence that Notch directly regulates their transcription. To evaluate the role of NOTCH and TNF signaling in this process, human CD34(+) HPCs were cocultured with OP9-DL1 cells, in the presence or absence of TNF. The use of a Notch signaling inhibitor and a protein synthesis inhibitor allowed us to distinguish primary effects, mediated by direct signaling downstream Notch and TNF, from secondary effects, mediated by de novo synthesized proteins. A low and physiologically relevant concentration of TNF promoted T lymphopoiesis in OP9-DL1 cocultures. TNF positively modulated the expression of both transcripts in a Notch-dependent manner; however, GATA3 induction was mediated by a direct mechanism, while HOXB4 induction was indirect. Induction of both transcripts was repressed by a GSK3β inhibitor, indicating that activation of canonical Wnt signaling inhibits rather than induces their expression. Our study provides novel evidences of the mechanisms integrating Notch and TNF-alpha signaling in the transcriptional induction of GATA3 and HOXB4. This mechanism has direct implications in the control of self-renewal, proliferation, commitment, and T cell differentiation.

  4. Brief report: Blockade of Notch signaling in muscle stem cells causes muscular dystrophic phenotype and impaired muscle regeneration.

    PubMed

    Lin, Shuibin; Shen, Huangxuan; Jin, Baofeng; Gu, Yumei; Chen, Zirong; Cao, Chunxia; Hu, Chengbin; Keller, Charles; Pear, Warren S; Wu, Lizi

    2013-04-01

    Muscular dystrophies are a group of devastating diseases characterized by progressive muscle weakness and degeneration, with etiologies including muscle gene mutations and regenerative defects of muscle stem cells. Notch signaling is critical for skeletal myogenesis and has important roles in maintaining the muscle stem cell pool and preventing premature muscle differentiation. To investigate the functional impact of Notch signaling blockade in muscle stem cells, we developed a conditional knock-in mouse model in which endogenous Notch signaling is specifically blocked in muscle stem cell compartment. Mice with Notch signaling inhibition in muscle stem cells showed several muscular dystrophic features and impaired muscle regeneration. Analyses of satellite cells and isolated primary myoblasts revealed that Notch signaling blockade in muscle stem cells caused reduced activation and proliferation of satellite cells but enhanced differentiation of myoblasts. Our data thus indicate that Notch signaling controls processes that are critical to regeneration in muscular dystrophy, suggesting that Notch inhibitor therapies could have potential side effects on muscle functions.

  5. Sonic Hedgehog promotes proliferation of Notch-dependent monociliated choroid plexus tumour cells

    PubMed Central

    Li, Li; Grausam, Katie B.; Wang, Jun; Lun, Melody P.; Ohli, Jasmin; Lidov, Hart G. W.; Calicchio, Monica L.; Zeng, Erliang; Salisbury, Jeffrey L.; Wechsler-Reya, Robert J.; Lehtinen, Maria K.; Schüller, Ulrich; Zhao, Haotian

    2016-01-01

    Aberrant Notch signaling has been linked to many cancers including choroid plexus (CP) tumours, a group of rare and predominantly pediatric brain neoplasms. We developed animal models of CP tumours by inducing sustained expression of Notch1 that recapitulate properties of human CP tumours with aberrant NOTCH signaling. Whole transcriptome and functional analyses showed that tumour cell proliferation is associated with Sonic Hedgehog (Shh) in the tumour microenvironment. Unlike CP epithelial cells, which have multiple primary cilia, tumour cells possess a solitary primary cilium as a result of Notch-mediated suppression of multiciliate diffferentiation. A Shh-driven signaling cascade in the primary cilium occurs in tumour cells but not in epithelial cells. Lineage studies show that CP tumours arise from mono-ciliated progenitors in the roof plate characterized by elevated Notch signaling. Abnormal SHH signaling and distinct ciliogenesis are detected in human CP tumours, suggesting SHH pathway and cilia differentiation as potential therapeutic avenues. PMID:26999738

  6. Structure of the Notch1-negative regulatory region: implications for normal activation and pathogenic signaling in T-ALL

    SciTech Connect

    Gordon, Wendy R.; Roy, Monideepa; Vardar-Ulu, Didem; Garfinkel, Megan; Mansour, Marc R.; Aster, Jon C.; Blacklow, Stephen C.

    2009-09-02

    Proteolytic resistance of Notch prior to ligand binding depends on the structural integrity of a negative regulatory region (NRR) of the receptor that immediately precedes the transmembrane segment. The NRR includes the 3 Lin12/Notch repeats and the juxtamembrane heterodimerization domain, the region of Notch1 most frequently mutated in T-cell acute lymphoblastic leukemia lymphoma (T-ALL). Here, we report the x-ray structure of the Notch1 NRR in its autoinhibited conformation. A key feature of the Notch1 structure that maintains its closed conformation is a conserved hydrophobic plug that sterically occludes the metalloprotease cleavage site. Crystal packing interactions involving a highly conserved, exposed face on the third Lin12/Notch repeat suggest that this site may normally be engaged in intermolecular or intramolecular protein-protein interactions. The majority of known T-ALL-associated point mutations map to residues in the hydrophobic interior of the Notch1 NRR. A novel mutation (H1545P), which alters a residue at the crystal-packing interface, leads to ligand-independent increases in signaling in reporter gene assays despite only mild destabilization of the NRR, suggesting that it releases the autoinhibitory clamp on the heterodimerization domain imposed by the Lin12/Notch repeats. The Notch1 NRR structure should facilitate a search for antibodies or compounds that stabilize the autoinhibited conformation.

  7. Notch Signaling Contributes to Liver Inflammation by Regulation of Interleukin-22-Producing Cells in Hepatitis B Virus Infection.

    PubMed

    Wei, Xin; Wang, Jiu-Ping; Hao, Chun-Qiu; Yang, Xiao-Fei; Wang, Lin-Xu; Huang, Chang-Xing; Bai, Xue-Fan; Lian, Jian-Qi; Zhang, Ye

    2016-01-01

    The mechanism of hepatitis B virus (HBV) induced liver inflammation is not fully elucidated. Notch signaling augmented interleukin (IL)-22 secretion in CD4(+) T cells, and Notch-IL-22 axis fine-tuned inflammatory response. We previously demonstrated a proinflammatory role of IL-22 in HBV infection. Thus, in this study, we analyzed the role of Notch in development of IL-22-producing cells in HBV infection by inhibition of Notch signaling using γ-secretase inhibitor DAPT in both hydrodynamic induced HBV-infected mouse model and in peripheral blood cells isolated from patients with HBV infection. mRNA expressions of Notch1 and Notch2 were significantly increased in livers and CD4(+) T cells upon HBV infection. Inhibition of Notch signaling in vivo leaded to the reduction in NKp46(+) innate lymphoid cells 22 (ILC22) and lymphoid tissue inducer 4 (LTi4) cells in the liver. This process was accompanied by downregulating the expressions of IL-22 and related proinflammatory cytokines and chemokines in the liver, as well as blocking the recruitment of antigen-non-specific inflammatory cells into the liver and subsequent liver injury, but did not affect HBV antigens production and IL-22 secretion in the serum. Furthermore, IL-22 production in HBV non-specific cultured CD4(+) T cells, but not HBV-specific CD4(+) T cells, was reduced in response to in vitro inhibition of Notch signaling. In conclusion, Notch siganling appears to be an important mediator of the liver inflammation by modulating hepatic ILC22. The potential proinflammatory effect of Notch-mediated ILC22 may be significant for the development of new therapeutic approaches for treatment of hepatitis B.

  8. Notch Signaling Contributes to Liver Inflammation by Regulation of Interleukin-22-Producing Cells in Hepatitis B Virus Infection

    PubMed Central

    Wei, Xin; Wang, Jiu-Ping; Hao, Chun-Qiu; Yang, Xiao-Fei; Wang, Lin-Xu; Huang, Chang-Xing; Bai, Xue-Fan; Lian, Jian-Qi; Zhang, Ye

    2016-01-01

    The mechanism of hepatitis B virus (HBV) induced liver inflammation is not fully elucidated. Notch signaling augmented interleukin (IL)-22 secretion in CD4+ T cells, and Notch-IL-22 axis fine-tuned inflammatory response. We previously demonstrated a proinflammatory role of IL-22 in HBV infection. Thus, in this study, we analyzed the role of Notch in development of IL-22-producing cells in HBV infection by inhibition of Notch signaling using γ-secretase inhibitor DAPT in both hydrodynamic induced HBV-infected mouse model and in peripheral blood cells isolated from patients with HBV infection. mRNA expressions of Notch1 and Notch2 were significantly increased in livers and CD4+ T cells upon HBV infection. Inhibition of Notch signaling in vivo leaded to the reduction in NKp46+ innate lymphoid cells 22 (ILC22) and lymphoid tissue inducer 4 (LTi4) cells in the liver. This process was accompanied by downregulating the expressions of IL-22 and related proinflammatory cytokines and chemokines in the liver, as well as blocking the recruitment of antigen-non-specific inflammatory cells into the liver and subsequent liver injury, but did not affect HBV antigens production and IL-22 secretion in the serum. Furthermore, IL-22 production in HBV non-specific cultured CD4+ T cells, but not HBV-specific CD4+ T cells, was reduced in response to in vitro inhibition of Notch signaling. In conclusion, Notch siganling appears to be an important mediator of the liver inflammation by modulating hepatic ILC22. The potential proinflammatory effect of Notch-mediated ILC22 may be significant for the development of new therapeutic approaches for treatment of hepatitis B. PMID:27800305

  9. Pre-clinical studies of Notch signaling inhibitor RO4929097 in inflammatory breast cancer cells.

    PubMed

    Debeb, Bisrat G; Cohen, Evan N; Boley, Kimberly; Freiter, Erik M; Li, Li; Robertson, Fredika M; Reuben, James M; Cristofanilli, Massimo; Buchholz, Thomas A; Woodward, Wendy A

    2012-07-01

    Basal breast cancer, common among patients presenting with inflammatory breast cancer (IBC), has been shown to be resistant to radiation and enriched in cancer stem cells. The Notch pathway plays an important role in self-renewal of breast cancer stem cells and contributes to inflammatory signaling which promotes the breast cancer stem cell phenotype. Herein, we inhibited Notch signaling using a gamma secretase inhibitor, RO4929097, in an in vitro model that enriches for cancer initiating cells (3D clonogenic assay) and conventional 2D clonogenic assay to compare the effect on radiosensitization of the SUM149 and SUM190 IBC cell lines. RO4929097 downregulated the Notch target genes Hes1, Hey1, and HeyL, and showed a significant reduction in anchorage independent growth in SUM190 and SUM149. However, the putative self-renewal assay mammosphere formation efficiency was increased with the drug. To assess radiosensitization of putative cancer stem cells, cells were exposed to increasing doses of radiation with or without 1 μM RO4929097 in their standard (2D) and self-renewal enriching (3D) culture conditions. In the conventional 2D clonogenic assay, RO4929097 significantly sensitized SUM190 cells to ionizing radiation and has a modest radiosensitization effect in SUM149 cells. In the 3D clonogenic assays, however, a radioprotective effect was seen in both SUM149 and SUM190 cells at higher doses. Both cell lines express IL-6 and IL-8 cytokines known to mediate the efficacy of Notch inhibition and to promote self-renewal of stem cells. We further showed that RO429097 inhibits normal T-cell synthesis of some inflammatory cytokines, including TNF-α, a potential mediator of IL-6 and IL-8 production in the microenvironment. These data suggest that additional targeting agents may be required to selectively target IBC stem cells through Notch inhibition, and that evaluation of microenvironmental influences may shed further light on the potential effects of this inhibitor.

  10. Honokiol in combination with radiation targets notch signaling to inhibit colon cancer stem cells.

    PubMed

    Ponnurangam, Sivapriya; Mammen, Joshua M V; Ramalingam, Satish; He, Zhiyun; Zhang, Youcheng; Umar, Shahid; Subramaniam, Dharmalingam; Anant, Shrikant

    2012-04-01

    Cancer stem cells are implicated in resistance to ionizing radiation (IR) and chemotherapy. Honokiol, a biphenolic compound has been used in traditional Chinese medicine for treating various ailments. In this study, we determined the ability of honokiol to enhance the sensitivity of colon cancer stem cells to IR. The combination of honokiol and IR suppressed proliferation and colony formation while inducing apoptosis of colon cancer cells in culture. There were also reduced numbers and size of spheroids, which was coupled with reduced expression of cancer stem cell marker protein DCLK1. Flow cytometry studies confirmed that the honokiol-IR combination reduced the number of DCLK1+ cells. In addition, there were reduced levels of activated Notch-1, its ligand Jagged-1, and the downstream target gene Hes-1. Furthermore, expression of components of the Notch-1 activating γ-secretase complex, presenilin 1, nicastrin, Pen2, and APH-1 was also suppressed. On the other hand, the honokiol effects were mitigated when the Notch intracellular domain was expressed. To determine the effect of honokiol-IR combination on tumor growth in vivo, nude mice tumor xenografts were administered honokiol intraperitoneally and exposed to IR. The honokiol-IR combination significantly inhibited tumor xenograft growth. In addition, there were reduced levels of DCLK1 and the Notch signaling-related proteins in the xenograft tissues. Together, these data suggest that honokiol is a potent inhibitor of colon cancer growth that targets the stem cells by inhibiting the γ-secretase complex and the Notch signaling pathway. These studies warrant further clinical evaluation for the combination of honokiol and IR for treating colon cancers.

  11. Reciprocal interaction between TRAF6 and notch signaling regulates adult myofiber regeneration upon injury.

    PubMed

    Hindi, Sajedah M; Paul, Pradyut K; Dahiya, Saurabh; Mishra, Vivek; Bhatnagar, Shephali; Kuang, Shihuan; Choi, Yongwon; Kumar, Ashok

    2012-12-01

    Skeletal muscle is a postmitotic tissue that repairs and regenerates through activation of a population of stem-cell-like satellite cells. However, signaling mechanisms governing adult skeletal muscle regeneration remain less understood. In the present study, we have investigated the role of tumor necrosis factor (TNF) receptor-associated factor 6 (TRAF6), an adaptor protein involved in receptor-mediated activation of multiple signaling pathways in regeneration of adult myofibers. Skeletal muscle-specific depletion of TRAF6 in mice (TRAF6(mko)) improved regeneration of myofibers upon injury with a concomitant increase in the number of satellite cells and activation of the Notch signaling pathway. Ex vivo cultures of TRAF6(mko) myofiber explants demonstrated an increase in the proliferative capacity of myofiber-associated satellite cells accompanied by an upregulation of Notch ligands. Deletion of TRAF6 also inhibited the activity of transcription factor NF-κB and the expression of inflammatory cytokines and augmented the M2c macrophage phenotype in injured muscle tissues. Collectively, our study demonstrates that specific inhibition of TRAF6 improves satellite cell activation and skeletal muscle regeneration through upregulation of Notch signaling and reducing the inflammatory repertoire.

  12. Notch signalling in adult neurons: a potential target for microtubule stabilization.

    PubMed

    Bonini, Sara Anna; Ferrari-Toninelli, Giulia; Montinaro, Mery; Memo, Maurizio

    2013-11-01

    Cytoskeletal dysfunction has been proposed during the last decade as one of the main mechanisms involved in the aetiology of several neurodegenerative diseases. Microtubules are basic elements of the cytoskeleton and the dysregulation of microtubule stability has been demonstrated to be causative for axonal transport impairment, synaptic contact degeneration, impaired neuronal function leading finally to neuronal loss. Several pathways are implicated in the microtubule assembly/disassembly process. Emerging evidence is focusing on Notch as a microtubule dynamics regulator. We demonstrated that activation of Notch signalling results in increased microtubule stability and changes in axonal morphology and branching. By contrast, Notch inhibition leads to an increase in cytoskeleton plasticity with intense neurite remodelling. Until now, several microtubule-binding compounds have been tested and the results have provided proof of concept that microtubule-binding agents or compounds with the ability to stabilize microtubules may have therapeutic potential for the treatment of Alzheimer's disease and other neurodegenerative diseases. In this review, based on its key role in cytoskeletal dynamics modulation, we propose Notch as a new potential target for microtubule stabilization.

  13. Notch signaling: a mediator of beta-cell de-differentiation in diabetes?

    PubMed

    Darville, Martine I; Eizirik, Décio L

    2006-01-27

    Cytokines are mediators of pancreatic beta-cell dysfunction and death in type 1 diabetes mellitus. Microarray analyses of insulin-producing cells exposed to interleukin-1beta+interferon-gamma showed decreased expression of genes related to beta-cell-differentiated functions and increased expression of members of the Notch signaling pathway. Re-expression of this developmental pathway may contribute for loss-of-function of beta-cells exposed to an autoimmune attack. In this study, we show that rat primary beta-cells exposed to cytokines up-regulate several Notch receptors and ligands, and the target gene Hes1. Transfection of insulin-producing INS-1E cells and primary rat beta-cells with a constitutively active form of the Notch receptor down-regulated Pdx1 and insulin expression in INS-1E cells but not in primary beta-cells. Thus, activation of the Notch pathway inhibits differentiated functions in dividing but not in terminally differentiated beta-cells.

  14. The γ-secretase-generated intracellular domain of β-amyloid precursor protein binds Numb and inhibits Notch signaling

    PubMed Central

    Roncarati, Roberta; Šestan, Nenad; Scheinfeld, Meir H.; Berechid, Bridget E.; Lopez, Peter A.; Meucci, Olimpia; McGlade, Jane C.; Rakic, Pasko; D'Adamio, Luciano

    2002-01-01

    The β-amyloid precursor protein (APP) and the Notch receptor undergo intramembranous proteolysis by the Presenilin-dependent γ-secretase. The cleavage of APP by γ-secretase releases amyloid-β peptides, which have been implicated in the pathogenesis of Alzheimer's disease, and the APP intracellular domain (AID), for which the function is not yet well understood. A similar γ-secretase-mediated cleavage of the Notch receptor liberates the Notch intracellular domain (NICD). NICD translocates to the nucleus and activates the transcription of genes that regulate the generation, differentiation, and survival of neuronal cells. Hence, some of the effects of APP signaling and Alzheimer's disease pathology may be mediated by the interaction of APP and Notch. Here, we show that membrane-tethered APP binds to the cytosolic Notch inhibitors Numb and Numb-like in mouse brain lysates. AID also binds Numb and Numb-like, and represses Notch activity when released by APP. Thus, γ-secretase may have opposing effects on Notch signaling; positive by cleaving Notch and generating NICD, and negative by processing APP and generating AID, which inhibits the function of NICD. PMID:12011466

  15. Colonization of the satellite cell niche by skeletal muscle progenitor cells depends on Notch signals.

    PubMed

    Bröhl, Dominique; Vasyutina, Elena; Czajkowski, Maciej T; Griger, Joscha; Rassek, Claudia; Rahn, Hans-Peter; Purfürst, Bettina; Wende, Hagen; Birchmeier, Carmen

    2012-09-11

    Skeletal muscle growth and regeneration rely on myogenic progenitor and satellite cells, the stem cells of postnatal muscle. Elimination of Notch signals during mouse development results in premature differentiation of myogenic progenitors and formation of very small muscle groups. Here we show that this drastic effect is rescued by mutation of the muscle differentiation factor MyoD. However, rescued myogenic progenitors do not assume a satellite cell position and contribute poorly to myofiber growth. The disrupted homing is due to a deficit in basal lamina assembly around emerging satellite cells and to their impaired adhesion to myofibers. On a molecular level, emerging satellite cells deregulate the expression of basal lamina components and adhesion molecules like integrin α7, collagen XVIIIα1, Megf10, and Mcam. We conclude that Notch signals control homing of satellite cells, stimulating them to contribute to their own microenvironment and to adhere to myofibers.

  16. Notch signalling inhibits the adipogenic differentiation of single-cell-derived mesenchymal stem cell clones isolated from human adipose tissue.

    PubMed

    Osathanon, Thanaphum; Subbalekha, Keskanya; Sastravaha, Panunn; Pavasant, Prasit

    2012-01-01

    ADSCs (adipose-derived mesenchymal stem cells) are candidate adult stem cells for regenerative medicine. Notch signalling participates in the differentiation of a heterogeneous ADSC population. We have isolated, human adipose tissue-derived single-cell clones using a cloning ring technique and characterized for their stem cell characteristics. The role of Notch signalling in the differentiation capacity of these adipose-derived single-cell-clones has also been investigated. All 14 clones expressed embryonic and mesenchymal stem cell marker genes. These clones could differentiate into both osteogenic and adipogenic lineages. However, the differentiation potential of each clone was different. Low adipogenic clones had significantly higher mRNA expression levels of Notch 2, 3 and 4, Jagged1, as well as Delta1, compared with those of high adipogenic clones. In contrast, no changes in expression of Notch signalling component mRNA between low and high osteogenic clones was found. Notch receptor mRNA expression decreased with the adipogenic differentiation of both low and high adipogenic clones. The γ-secretase inhibitor, DAPT (N-[N-(3,5-difluorophenacetyl)-l-alanyl]-(S)-phenylglycine t-butyl ester), enhanced adipogenic differentiation. Correspondingly, cells seeded on a Notch ligand (Jagged1) bound surface showed lower intracellular lipid accumulation. These results were noted in both low and high adipogenic clones, indicating that Notch signalling inhibited the adipogenic differentiation of adipose ADSC clones, and could be used to identify an adipogenic susceptible subpopulation for soft-tissue augmentation application.

  17. Gamma-Secretase and Notch Signaling: Novel Therapeutic Targets In Breast Cancer

    DTIC Science & Technology

    2005-05-01

    inflammation characteristic of auto- receptor-STATl-independent pathway for Tbx2l initiation, as these I.immune diseases such as multiple sclerosis, Crohn ...may be S effects the blocking of Notch signaling in vivo with GSI has on disease essential for the adoption of a TH1 cell fate by CD4+ cells. CD4...Unlimited The views, opinions and/or findings contained in this report are those of the author( s ) and should not be construed as an official

  18. Integration of Orthogonal Signaling by the Notch and Dpp Pathways in Drosophila.

    PubMed

    Stroebele, Elizabeth; Erives, Albert

    2016-05-01

    The transcription factor Suppressor of Hairless and its coactivator, the Notch intracellular domain, are polyglutamine (pQ)-rich factors that target enhancer elements and interact with other locally bound pQ-rich factors. To understand the functional repertoire of such enhancers, we identify conserved regulatory belts with binding sites for the pQ-rich effectors of both Notch and BMP/Dpp signaling, and the pQ-deficient tissue selectors Apterous (Ap), Scalloped (Sd), and Vestigial (Vg). We find that the densest such binding site cluster in the genome is located in the BMP-inducible nab locus, a homolog of the vertebrate transcriptional cofactors NAB1/NAB2 We report three major findings. First, we find that this nab regulatory belt is a novel enhancer driving dorsal wing margin expression in regions of peak phosphorylated Mad in wing imaginal discs. Second, we show that Ap is developmentally required to license the nab dorsal wing margin enhancer (DWME) to read out Notch and Dpp signaling in the dorsal compartment. Third, we find that the nab DWME is embedded in a complex of intronic enhancers, including a wing quadrant enhancer, a proximal wing disc enhancer, and a larval brain enhancer. This enhancer complex coordinates global nab expression via both tissue-specific activation and interenhancer silencing. We suggest that DWME integration of BMP signaling maintains nab expression in proliferating margin descendants that have divided away from Notch-Delta boundary signaling. As such, uniform expression of genes like nab and vestigial in proliferating compartments would typically require both boundary and nonboundary lineage-specific enhancers.

  19. Homemade notch filter to suppress strong FM or DAB - T/DVB - T signals

    NASA Astrophysics Data System (ADS)

    Monstein, Christian

    2016-04-01

    Many of the current 116 solar radio spectrometer instruments in the e-Callisto network are suffering from strong interference from FM-radio, DAB-T or DVB-T broadcast stations. With simple surface mount device (SMD) components a cheap notch (trap)filter can be produced to suppress these strong signals that otherwise may saturate the low noise amplifier and/or the receiver.

  20. Notch Signaling and Schwann Cell Transformation: Development of a Model System and Application to Human MPNSTs

    DTIC Science & Technology

    2007-03-01

    Transformation: Development of a Model System and Application to Human MPNSTs PRINCIPAL INVESTIGATOR: Tom Kadesch, Ph.D... MPNSTs 5b. GRANT NUMBER W81XWH-04-1-0209 5c. PROGRAM ELEMENT NUMBER 6. AUTHOR(S) Tom Kadesch, Ph.D. 5d. PROJECT NUMBER 5e. TASK...potential role of Notch signaling in the malignant transformation of neurofibromas to MPNSTs in patients with NF1. Our previous work has shown that

  1. Notch Signaling and Schwann Cell Transformation: Development of a Model System and Application to Human MPNSTs

    DTIC Science & Technology

    2008-03-01

    Transformation: Development of a Model System and Application to Human MPNSTs PRINCIPAL INVESTIGATOR: Tom Kadesch, Ph.D...and Application to Human MPNSTs 5b. GRANT NUMBER W81XWH-04-1-0209 5c. PROGRAM ELEMENT NUMBER 6. AUTHOR(S) Tom Kadesch, Ph.D. 5d. PROJECT...grant addresses the potential role of Notch signaling in the malignant transformation of neurofibromas to MPNSTs in patients with NF1. Our previous

  2. Insulin signals control the competence of the Drosophila female germline stem cell niche to respond to Notch ligands.

    PubMed

    Hsu, Hwei-Jan; Drummond-Barbosa, Daniela

    2011-02-15

    Adult stem cells reside in specialized microenvironments, or niches, that are essential for their function in vivo. Stem cells are physically attached to the niche, which provides secreted factors that promote their self-renewal and proliferation. Despite intense research on the role of the niche in regulating stem cell function, much less is known about how the niche itself is controlled. We previously showed that insulin signals directly stimulate germline stem cell (GSC) division and indirectly promote GSC maintenance via the niche in Drosophila. Insulin-like peptides are required for maintenance of cap cells (a major component of the niche) via modulation of Notch signaling, and they also control attachment of GSCs to cap cells and E-cadherin levels at the cap cell-GSC junction. Here, we further dissect the molecular and cellular mechanisms underlying these processes. We show that insulin and Notch ligands directly stimulate cap cells to maintain their numbers and indirectly promote GSC maintenance. We also report that insulin signaling, via phosphoinositide 3-kinase and FOXO, intrinsically controls the competence of cap cells to respond to Notch ligands and thereby be maintained. Contrary to a previous report, we also find that Notch ligands originated in GSCs are not required either for Notch activation in the GSC niche, or for cap cell or GSC maintenance. Instead, the niche itself produces ligands that activate Notch signaling within cap cells, promoting stability of the GSC niche. Finally, insulin signals control cap cell-GSC attachment independently of their role in Notch signaling. These results are potentially relevant to many systems in which Notch signaling modulates stem cells and demonstrate that complex interactions between local and systemic signals are required for proper stem cell niche function.

  3. BMP signaling balances murine myeloid potential through SMAD-independent p38MAPK and NOTCH pathways.

    PubMed

    Cook, Brandoch D; Evans, Todd

    2014-07-17

    Bone morphogenetic protein (BMP) signaling regulates early hematopoietic development, proceeding from mesoderm patterning through the progressive commitment and differentiation of progenitor cells. The BMP pathway signals largely through receptor-mediated activation of Mothers Against Decapentaplegic homolog (SMAD) proteins, although alternate pathways are modulated through various components of mitogen-activated protein kinase (MAPK) signaling. Using a conditional, short hairpin RNA (shRNA)-based knockdown system in the context of differentiating embryonic stem cells (ESCs), we demonstrated previously that Smad1 promotes hemangioblast specification, but then subsequently restricts primitive progenitor potential. Here we show that co-knockdown of Smad5 restores normal progenitor potential of Smad1-depleted cells, suggesting opposing functions for Smad1 and Smad5. This balance was confirmed by cotargeting Smad1/5 with a specific chemical antagonist, LDN193189 (LDN). However, we discovered that LDN treatment after hemangioblast commitment enhanced primitive myeloid potential. Moreover, inhibition with LDN (but not SMAD depletion) increased expression of Delta-like ligands Dll1 and Dll3 and NOTCH activity; abrogation of NOTCH activity restored LDN-enhanced myeloid potential back to normal, corresponding with expression levels of the myeloid master regulator, C/EBPα. LDN but not SMAD activity was also associated with activation of the p38MAPK pathway, and blocking this pathway was sufficient to enhance myelopoiesis. Therefore, NOTCH and p38MAPK pathways balance primitive myeloid progenitor output downstream of the BMP pathway.

  4. Gene expression analysis reveals that Delta/Notch signalling is not involved in onychophoran segmentation.

    PubMed

    Janssen, Ralf; Budd, Graham E

    2016-03-01

    Delta/Notch (Dl/N) signalling is involved in the gene regulatory network underlying the segmentation process in vertebrates and possibly also in annelids and arthropods, leading to the hypothesis that segmentation may have evolved in the last common ancestor of bilaterian animals. Because of seemingly contradicting results within the well-studied arthropods, however, the role and origin of Dl/N signalling in segmentation generally is still unclear. In this study, we investigate core components of Dl/N signalling by means of gene expression analysis in the onychophoran Euperipatoides kanangrensis, a close relative to the arthropods. We find that neither Delta or Notch nor any other investigated components of its signalling pathway are likely to be involved in segment addition in onychophorans. We instead suggest that Dl/N signalling may be involved in posterior elongation, another conserved function of these genes. We suggest further that the posterior elongation network, rather than classic Dl/N signalling, may be in the control of the highly conserved segment polarity gene network and the lower-level pair-rule gene network in onychophorans. Consequently, we believe that the pair-rule gene network and its interaction with Dl/N signalling may have evolved within the arthropod lineage and that Dl/N signalling has thus likely been recruited independently for segment addition in different phyla.

  5. The lack of autophagy triggers precocious activation of Notch signaling during Drosophila oogenesis

    PubMed Central

    2012-01-01

    Background The proper balance of autophagy, a lysosome-mediated degradation process, is indispensable for oogenesis in Drosophila. We recently demonstrated that egg development depends on autophagy in the somatic follicle cells (FC), but not in the germline cells (GCs). However, the lack of autophagy only affects oogenesis when FCs are autophagy-deficient but GCs are wild type, indicating that a dysfunctional signaling between soma and germline may be responsible for the oogenesis defects. Thus, autophagy could play an essential role in modulating signal transduction pathways during egg development. Results Here, we provide further evidence for the necessity of autophagy during oogenesis and demonstrate that autophagy is especially required in subsets of FCs. Generation of autophagy-deficient FCs leads to a wide range of phenotypes that are similar to mutants with defects in the classical cell-cell signaling pathways in the ovary. Interestingly, we observe that loss of autophagy leads to a precocious activation of the Notch pathway in the FCs as monitored by the expression of Cut and Hindsight, two downstream effectors of Notch signaling. Conclusion Our findings point to an unexpected function for autophagy in the modulation of the Notch signaling pathway during Drosophila oogenesis and suggest a function for autophagy in proper receptor activation. Egg development is affected by an imbalance of autophagy between signal sending (germline) and signal receiving cell (FC), thus the lack of autophagy in the germline is likely to decrease the amount of active ligand and accordingly compensates for increased signaling in autophagy-defective follicle cells. PMID:23217079

  6. Inhibition on Numb/Notch signal pathway enhances radiosensitivity of lung cancer cell line H358.

    PubMed

    Song, Shi-Gang; Yu, Hong-Yang; Ma, Yan-Wei; Zhang, Feng; Xu, Xiang-Ying

    2016-10-01

    The objective of the study is to investigate the effects of the Numb/Notch signal pathway on the radiosensitivity of lung cancer cell line H358. MTT assay and colony forming assay were used to detect the effects of different doses of X-rays and MW167 on the in vitro proliferation of the lung cancer cell line H358. Flow cytometry was applied to evaluate the effects of X rays on the apoptosis of H358. Scratch assay and Transwell invasion assay were used to examine the effects of X-rays on the migration and invasion abilities of H358. The mRNA and protein expressions in the signal pathway were detected by real-time PCR and western blot. Assays in vitro confirmed the effects of the Numb/Notch pathway inhibitor on the radiosensitivity to lung cancer. MW167 enhanced the inhibiting effects of X-ray on the proliferation of H358 cell line. After the addition of MW167, the apoptosis rates significantly increased, but the invasion and migration abilities decreased significantly. Meanwhile, MW167 could dose-dependently promote the increase of expression of Numb, which is the upstream gene of the Numb/Notch signaling pathway, but inhibit the expression of and HES1. In vivo experiments revealed that cell proliferation was suppressed in the radiation, pathway inhibitor, and pathway inhibitor + radiation groups, and the pathway inhibitor + radiation group exhibited more active anti-tumor ability when compared with the blank group (all P < 0.05); Numb expression was up-regulated, but Notch1 and HES1 expressions were down-regulated in those three groups, and also, the pathway inhibitor + radiation group exhibited more significant alternation when compared with the blank group (all P < 0.05); cell apoptosis was promoted in those three groups, and the pathway inhibitor + radiation group showed more active apoptosis when compared with the blank group (all P < 0.05). Repression of the Numb/Notch pathway enhances the effects of radiotherapy on the radiosensitivity of the lung

  7. Aldehyde dehydrogenase activity selects for lung adenocarcinoma stem cells dependent on notch signaling.

    PubMed

    Sullivan, James P; Spinola, Monica; Dodge, Michael; Raso, Maria G; Behrens, Carmen; Gao, Boning; Schuster, Katja; Shao, Chunli; Larsen, Jill E; Sullivan, Laura A; Honorio, Sofia; Xie, Yang; Scaglioni, Pier P; DiMaio, J Michael; Gazdar, Adi F; Shay, Jerry W; Wistuba, Ignacio I; Minna, John D

    2010-12-01

    Aldehyde dehydrogenase (ALDH) is a candidate marker for lung cancer cells with stem cell-like properties. Immunohistochemical staining of a large panel of primary non-small cell lung cancer (NSCLC) samples for ALDH1A1, ALDH3A1, and CD133 revealed a significant correlation between ALDH1A1 (but not ALDH3A1 or CD133) expression and poor prognosis in patients including those with stage I and N0 disease. Flow cytometric analysis of a panel of lung cancer cell lines and patient tumors revealed that most NSCLCs contain a subpopulation of cells with elevated ALDH activity, and that this activity is associated with ALDH1A1 expression. Isolated ALDH(+) lung cancer cells were observed to be highly tumorigenic and clonogenic as well as capable of self-renewal compared with their ALDH(-) counterparts. Expression analysis of sorted cells revealed elevated Notch pathway transcript expression in ALDH(+) cells. Suppression of the Notch pathway by treatment with either a γ-secretase inhibitor or stable expression of shRNA against NOTCH3 resulted in a significant decrease in ALDH(+) lung cancer cells, commensurate with a reduction in tumor cell proliferation and clonogenicity. Taken together, these findings indicate that ALDH selects for a subpopulation of self-renewing NSCLC stem-like cells with increased tumorigenic potential, that NSCLCs harboring tumor cells with ALDH1A1 expression have inferior prognosis, and that ALDH1A1 and CD133 identify different tumor subpopulations. Therapeutic targeting of the Notch pathway reduces this ALDH(+) component, implicating Notch signaling in lung cancer stem cell maintenance.

  8. Protein O-fucosyltransferase 1 expression impacts myogenic C2C12 cell commitment via the Notch signaling pathway.

    PubMed

    Der Vartanian, Audrey; Audfray, Aymeric; Al Jaam, Bilal; Janot, Mathilde; Legardinier, Sébastien; Maftah, Abderrahman; Germot, Agnès

    2015-01-01

    The Notch signaling pathway plays a crucial role in skeletal muscle regeneration in mammals by controlling the transition of satellite cells from quiescence to an activated state, their proliferation, and their commitment toward myotubes or self-renewal. O-fucosylation on Notch receptor epidermal growth factor (EGF)-like repeats is catalyzed by the protein O-fucosyltransferase 1 (Pofut1) and primarily controls Notch interaction with its ligands. To approach the role of O-fucosylation in myogenesis, we analyzed a murine myoblastic C2C12 cell line downregulated for Pofut1 expression by short hairpin RNA (shRNA) inhibition during the time course of differentiation. Knockdown of Pofut1 affected the signaling pathway activation by a reduction of the amount of cleaved Notch intracellular domain and a decrease in downstream Notch target gene expression. Depletion in Pax7(+)/MyoD(-) cells and earlier myogenic program entrance were observed, leading to an increase in myotube quantity with a small number of nuclei, reflecting fusion defects. The rescue of Pofut1 expression in knockdown cells restored Notch signaling activation and a normal course in C2C12 differentiation. Our results establish the critical role of Pofut1 on Notch pathway activation during myogenic differentiation.

  9. miR-216a regulates snx5, a novel Notch signaling pathway component, during zebrafish retinal development

    PubMed Central

    Thatcher, Elizabeth J.; Wu, Shu-Yu; Patton, James G.

    2015-01-01

    Precise regulation of Notch signaling is essential for normal vertebrate development. Mind bomb (Mib) is a ubiquitin ligase that is required for activation of Notch by Notch's ligand, Delta. Sorting Nexin 5 (SNX5) co-localizes with Mib and Delta complexes and has been shown to directly bind to Mib. We show that microRNA-216a (miR-216a) is expressed in the retina during early development and regulates snx5 to precisely regulate Notch signaling. miR-216a and snx5 have complementary expression patterns. Knocking down miR-216a and/or overexpression of snx5 resulted in increased Notch activation. Conversely, knocking down snx5 and/or miR-216a overexpression caused a decrease in Notch activation. We propose a model in which SNX5, precisely controlled by miR-216a, is a vital partner of Mib in promoting endocytosis of Delta and subsequent activation of Notch signaling. PMID:25645681

  10. Control of Neural Daughter Cell Proliferation by Multi-level Notch/Su(H)/E(spl)-HLH Signaling

    PubMed Central

    Bivik, Caroline; MacDonald, Ryan B.; Gunnar, Erika; Mazouni, Khalil; Schweisguth, Francois; Thor, Stefan

    2016-01-01

    The Notch pathway controls proliferation during development and in adulthood, and is frequently affected in many disorders. However, the genetic sensitivity and multi-layered transcriptional properties of the Notch pathway has made its molecular decoding challenging. Here, we address the complexity of Notch signaling with respect to proliferation, using the developing Drosophila CNS as model. We find that a Notch/Su(H)/E(spl)-HLH cascade specifically controls daughter, but not progenitor proliferation. Additionally, we find that different E(spl)-HLH genes are required in different neuroblast lineages. The Notch/Su(H)/E(spl)-HLH cascade alters daughter proliferation by regulating four key cell cycle factors: Cyclin E, String/Cdc25, E2f and Dacapo (mammalian p21CIP1/p27KIP1/p57Kip2). ChIP and DamID analysis of Su(H) and E(spl)-HLH indicates direct transcriptional regulation of the cell cycle genes, and of the Notch pathway itself. These results point to a multi-level signaling model and may help shed light on the dichotomous proliferative role of Notch signaling in many other systems. PMID:27070787

  11. Notch1 signaling inhibits growth of human hepatocellular carcinoma through induction of cell cycle arrest and apoptosis.

    PubMed

    Qi, Runzi; An, Huazhang; Yu, Yizhi; Zhang, Minghui; Liu, Shuxun; Xu, Hongmei; Guo, Zhenghong; Cheng, Tao; Cao, Xuetao

    2003-12-01

    Notch signaling plays a critical role in maintaining the balance between cell proliferation, differentiation, and apoptosis; hence, perturbed Notch signaling may contribute to tumorigenesis. Hepatocellular carcinoma (HCC) is one of the most common malignant tumors in Africa and Asia. The mechanisms that orchestrate the multiple oncogenic insults required for initiation and progression of HCC are not clear. We constitutively overexpressed active Notch1 in human HCC to explore the effects of Notch1 signaling on HCC cell growth and to investigate the underlying molecular mechanisms. We show here that overexpression of Notch1 was able to inhibit the growth of HCC cells in vitro and in vivo. Biochemical analysis revealed the involvement of cell cycle regulated proteins in Notch1-mediated G(0)/G(1) arrest of HCC cells. Compared with green fluorescent protein (GFP) control, transient transfection of Notch1 ICN decreased expression of cyclin A (3.5-fold), cyclin D1 (2-fold), cyclin E (4.5-fold), CDK2 (2.8-fold), and the phosphorylated form of retinoblastoma protein (3-fold). Up-regulation of p21(waf/cip1) protein expression was observed in SMMC7721-ICN cells stably expressing active Notch1 but not in SMMC7721-GFP cells, which only express GFP. Furthermore, a 12-fold increase in p53 expression and an increase (4.8-fold) in Jun-NH(2)-terminal kinase activation were induced in SMMC7721-ICN cells compared with SMMC7721-GFP cells. In contrast, expression of the antiapoptotic Bcl-2 protein could not be detected in SMMC7721-ICN cells. These findings suggest that Notch1 signaling may participate in the development of HCC cells, affecting multiple pathways that control both cell proliferation and apoptosis.

  12. Epithelial-stromal interaction via Notch signaling is essential for the full maturation of gut-associated lymphoid tissues.

    PubMed

    Obata, Yuuki; Kimura, Shunsuke; Nakato, Gaku; Iizuka, Keito; Miyagawa, Yurika; Nakamura, Yutaka; Furusawa, Yukihiro; Sugiyama, Machiko; Suzuki, Keiichiro; Ebisawa, Masashi; Fujimura, Yumiko; Yoshida, Hisahiro; Iwanaga, Toshihiko; Hase, Koji; Ohno, Hiroshi

    2014-12-01

    Intrinsic Notch signaling in intestinal epithelial cells restricts secretory cell differentiation. In gut-associated lymphoid tissue (GALT), stromal cells located beneath the follicle-associated epithelium (FAE) abundantly express the Notch ligand delta-like 1 (Dll1). Here, we show that mice lacking Rbpj-a gene encoding a transcription factor implicated in Notch signaling-in intestinal epithelial cells have defective GALT maturation. This defect can be attributed to the expansion of goblet cells, which leads to the down-regulation of CCL20 in FAE. These data demonstrate that epithelial Notch signaling maintained by stromal cells contributes to the full maturation of GALT by restricting secretory cell differentiation in FAE.

  13. The Notch Signaling Pathway Controls the Size of the Ocular Lens by Directly Suppressing p57Kip2 Expression▿ †

    PubMed Central

    Jia, Junling; Lin, Min; Zhang, Lingna; York, J. Philippe; Zhang, Pumin

    2007-01-01

    The size of an organ must be tightly controlled so that it fits within an organism. The mammalian lens is a relatively simple organ composed of terminally differentiated, amitotic lens fiber cells capped on the anterior surface by a layer of immature, mitotic epithelial cells. The proliferation of lens epithelial cells fuels the growth of the lens, thus controling the size of the lens. We report that the Notch signaling pathway defines the boundary between proliferation and differentiation in the developing lens. The loss of Notch signaling results in the loss of epithelial cells to differentiation and a much smaller lens. We found that the Notch effector Herp2 is expressed in lens epithelium and directly suppresses p57Kip2 expression, providing a molecular link between Notch signaling and the cell cycle control machinery during lens development. PMID:17709399

  14. Maternal Undernourished Fetal Kidneys Exhibit Differential Regulation of Nephrogenic Genes Including Downregulation of the Notch Signaling Pathway

    PubMed Central

    Magee, Thomas R.; Tafti, Sanaz A.; Desai, Mina; Liu, Qinghai; Ross, Michael G.; Nast, Cynthia C

    2011-01-01

    Maternal undernutrition results in offspring nephron number reduction and hypertension that are hypothesized to begin as compensatory changes in fetal gene expression during gestation. To evaluate mechanisms of dysregulated nephrogenesis, pregnant Sprague Dawley rats were 50% food restricted from embryonic day (E) 10 to E20. At E20, fetal male kidneys were examined by microarray analysis. A total of 476 differentially expressed transcripts were detected including those regulating development and differentiation, mitosis and cell cycle, chromatin assembly, and steroid hormone regulation. Differentially regulated genes were detected in MAPK/ERK, Wnt, and Notch signaling pathways. Validation of the microarray results was performed for the Notch signaling pathway, an important pathway in nephron formation. Protein expression of Notch pathway factors by Western blotting showed significantly decreased Notch2 and downstream effector Hey1 protein expression, while Ctbp1 co-repressor was increased. These data together show that maternal undernutrition results in developmental disruption in fetal nephrogenesis gene expression signaling. PMID:21273641

  15. PKCα Attenuates Jagged-1-mediated Notch Signaling in ErbB-2 positive Breast Cancer to Reverse Trastuzumab Resistance

    PubMed Central

    Pandya, Kinnari; Wyatt, Debra; Gallagher, Brian; Shah, Deep; Baker, Andrew; Bloodworth, Jeffrey; Zlobin, Andrei; Pannuti, Antonio; Green, Andrew; Ellis, Ian O.; Filipovic, Aleksandra; Sagert, Jason; Rana, Ajay; Albain, Kathy S.; Miele, Lucio; Denning, Mitchell F.; Osipo, Clodia

    2015-01-01

    Purpose Breast cancer is the second leading cause of cancer mortality among women worldwide. The major problem with current treatments is tumor resistance, recurrence, and disease progression. ErbB-2 positive breast tumors are aggressive and frequently become resistant to trastuzumab or lapatinib. We showed previously that Notch-1 is required for trastuzumab resistance in ErbB-2 positive breast cancer. Experimental Design Here, we sought to elucidate mechanisms by which ErbB-2 attenuates Notch signaling and how this is reversed by trastuzumab or lapatinib. Results The current study elucidates a novel Notch inhibitory mechanism by which PKCα downstream of ErbB-2: 1. Restricts the availability of Jagged-1 at the cell surface to trans activate Notch; 2. Restricts the critical interaction between Jagged-1 and Mindbomb-1, an E3 ligase that is required for Jagged-1 ubiquitinylation and subsequent Notch activation; 3. Reverses trastuzumab resistance in vivo; and 4. Predicts better outcome in women with ErbB-2 positive breast cancer. Conclusions The clinical impact of these studies is PKCα is potentially a good prognostic marker for low Notch activity and increased trastuzumab sensitivity in ErbB-2 positive breast cancer. Moreover, women with ErbB-2 positive breast tumors expressing high Notch activation and low PKCα expression could be the best candidates for anti-Notch therapy. PMID:26350262

  16. Notch1-Dll4 signalling and mechanical force regulate leader cell formation during collective cell migration.

    PubMed

    Riahi, Reza; Sun, Jian; Wang, Shue; Long, Min; Zhang, Donna D; Wong, Pak Kin

    2015-03-13

    At the onset of collective cell migration, a subset of cells within an initially homogenous population acquires a distinct 'leader' phenotype with characteristic morphology and motility. However, the factors driving the leader cell formation as well as the mechanisms regulating leader cell density during the migration process remain to be determined. Here we use single-cell gene expression analysis and computational modelling to show that the leader cell identity is dynamically regulated by Dll4 signalling through both Notch1 and cellular stress in a migrating epithelium. Time-lapse microscopy reveals that Dll4 is induced in leader cells after the creation of the cell-free region and leader cells are regulated via Notch1-Dll4 lateral inhibition. Furthermore, mechanical stress inhibits Dll4 expression and leader cell formation in the monolayer. Collectively, our findings suggest that a reduction of mechanical force near the boundary promotes Notch1-Dll4 signalling to dynamically regulate the density of leader cells during collective cell migration.

  17. Notch1-Dll4 signaling and mechanical force regulate leader cell formation during collective cell migration

    PubMed Central

    Riahi, Reza; Sun, Jian; Wang, Shue; Long, Min; Zhang, Donna D.; Wong, Pak Kin

    2015-01-01

    At the onset of collective cell migration, a subset of cells within an initially homogenous population acquires a distinct “leader” phenotype with characteristic morphology and motility. However, the factors driving leader cell formation as well as the mechanisms regulating leader cell density during the migration process remain to be determined. Here, we use single cell gene expression analysis and computational modeling to show that leader cell identity is dynamically regulated by Dll4 signaling through both Notch1 and cellular stress in a migrating epithelium. Time-lapse microscopy reveals that Dll4 is induced in leader cells after the creation of the cell-free region and leader cells are regulated via Notch1-Dll4 lateral inhibition. Furthermore, mechanical stress inhibits Dll4 expression and leader cell formation in the monolayer. Collectively, our findings suggest that a reduction of mechanical force near the boundary promotes Notch1-Dll4 signaling to dynamically regulate the density of leader cells during collective cell migration. PMID:25766473

  18. mTORC1 Prevents Preosteoblast Differentiation through the Notch Signaling Pathway

    PubMed Central

    Huang, Bin; Wang, Yongkui; Wang, Wenhao; Chen, Juan; Lai, Pinglin; Liu, Zhongyu; Yan, Bo; Xu, Song; Zhang, Zhongmin; Zeng, Chun; Rong, Limin; Liu, Bin; Cai, Daozhang; Jin, Dadi; Bai, Xiaochun

    2015-01-01

    The mechanistic target of rapamycin (mTOR) integrates both intracellular and extracellular signals to regulate cell growth and metabolism. However, the role of mTOR signaling in osteoblast differentiation and bone formation is undefined, and the underlying mechanisms have not been elucidated. Here, we report that activation of mTOR complex 1 (mTORC1) is required for preosteoblast proliferation; however, inactivation of mTORC1 is essential for their differentiation and maturation. Inhibition of mTORC1 prevented preosteoblast proliferation, but enhanced their differentiation in vitro and in mice. Activation of mTORC1 by deletion of tuberous sclerosis 1 (Tsc1) in preosteoblasts produced immature woven bone in mice due to excess proliferation but impaired differentiation and maturation of the cells. The mTORC1-specific inhibitor, rapamycin, restored these in vitro and in vivo phenotypic changes. Mechanistically, mTORC1 prevented osteoblast maturation through activation of the STAT3/p63/Jagged/Notch pathway and downregulation of Runx2. Preosteoblasts with hyperactive mTORC1 reacquired the capacity to fully differentiate and maturate when subjected to inhibition of the Notch pathway. Together, these findings identified the role of mTORC1 in osteoblast formation and established that mTORC1 prevents preosteoblast differentiation and maturation through activation of the Notch pathway. PMID:26241748

  19. Inhibition of cell proliferation and tumor growth of colorectal cancer by inhibitors of Wnt and Notch signaling pathways

    PubMed Central

    Xiao, Yuliang; Yang, Xiaojing; Miao, Yinglei; He, Xikun; Wang, Ming; Sha, Weihong

    2016-01-01

    Understanding the role and mechanism of signaling pathways including Notch and Wnt in colorectal carcinogenesis is critical to the development of novel therapeutics. In the present study, we analyzed the cell proliferation, migration, G2/M percentage and the expression of molecules of signaling pathways in HCT-116 cells through the inhibition of Wnt and Notch pathways, and also investigated the effect of inhibitors of Wnt and Notch pathways on tumor growth in a transplantation tumor model. We observed that rDDK-1 (an inhibitor of the Wnt signaling pathway) and LY374973 (an inhibitor of the Notch signaling pathway) synergistically inhibited the proliferation, migration and G2/M percentage of HCT-116 cell lines, and could further synergistically inhibit the tumor volume and weight in the transplantation tumor model. In the cell line and the transplantation tumor model, rDDK-1 and LY374973 further synergistically inhibited the expression level of all detected Wnt and Notch pathway genes. Our results may pave the way for using inhibitors of Wnt and Notch signaling pathways together to treat colorectal cancer. PMID:27900056

  20. “Methods to promote Notch signaling at the biomaterial interface and evaluation in a rafted organ culture model”

    PubMed Central

    Beckstead, Benjamin L.; Tung, Jason C.; Liang, Katharine J.; Tavakkol, Zarry; Usui, Marcia L.; Olerud, John E.; Giachelli, Cecilia M.

    2013-01-01

    The Notch signaling pathway is a promising target for controlling cell fate choices at the biomaterial-tissue interface. Building on our previous work in developing Notch-signaling biomaterials, we evaluated various immobilization schemes for Notch ligands and their effect on human foreskin keratinocytes. A peptide sequence derived from the Jagged-1 DSL-region and immobilized to poly (2-hydroxyethyl methacrylate) (polyHEMA) showed no bioactivity in relation to the Notch-CSL pathway. The full-length Jagged-1 protein immobilized directly to the polyHEMA surface showed activity in signaling the Notch-CSL pathway. However, an indirect affinity immobilization approach yielded a stronger signal. Human keratinocytes plated on bound Jagged-1 showed upregulated involucrin, keratin 10, and loricrin protein expression, with this expression being cell density-dependent. Utilizing a human foreskin rafted organ culture model as a bridge between in vitro and in vivo studies, Jagged-1-modified or control polyHEMA rods were implanted in human foreskin and cultured at the air-medium interface. Keratinocyte proliferation was suppressed and intermediate-stage differentiation promoted in Jagged-1-modified rods compared to control rods. Thus, Notch-signaling biomaterials provide a robust approach to control keratinocyte differentiation and may find application to other progenitor and stem cells. PMID:18985776

  1. The effects of notch filters on the correlation properties of a PN signal

    NASA Technical Reports Server (NTRS)

    Sussman, S. M.; Ferrari, E. J.

    1974-01-01

    With wideband pseudo-noise (PN) communications systems, it is sometimes desirable to supplement the inherent interference rejection capabilities by adding notch filters to attenuate relatively narrowband interference. This correspondence presents an investigation of the effects of notch filters on the performance of PN correlation receivers. A theoretical analysis of the correlation drop due to filter distortion has been conducted and confirmed by experimentation. Additional measurements and analysis have established the trade-off between correlation drop and interference suppression as a function of interference bandwidth. A typical result is that by incurring a penalty of a 1-dB drop in correlation peak, interfering signals having bandwidths of 2 to 3% of the PN chip rate can be attenuated by 25 dB.

  2. Epithelial expression and chromosomal location of human TLE genes: Implications for notch signaling and neoplasia

    SciTech Connect

    Liu, Yanling; Dehni, Ghassan; Stifani, S.

    1996-01-01

    The TLE genes are the human homologues of Drosophila groucho, a member of the Notch signaling pathway. This pathway controls a number of different cell-fate choices in invertebrates and vertebrates. We are interested in investigating the functions of the TLE gene family during epithelial determination and carcinogenesis. We show that expression of individual TLE genes correlates with immature epithelial cells that are progressing toward their terminally differentiated state, suggesting a role during epithelial differentiation. In both normal tissues and conditions resulting from incorrect or incomplete maturation events, such as metaplastic and neoplastic transformations, TLE expression is elevated and coincides with Notch expression, implicating these molecules in the maintenance of the undifferentiated state in epithelial cells. We also show that TLE1 and TLE2 are organized in a tandem array at chromosomal location 19p13.3, while TLE3 maps to 15q22. 26 refs., 4 figs.

  3. Rel/NF-kappaB can trigger the Notch signaling pathway by inducing the expression of Jagged1, a ligand for Notch receptors.

    PubMed Central

    Bash, J; Zong, W X; Banga, S; Rivera, A; Ballard, D W; Ron, Y; Gélinas, C

    1999-01-01

    Jagged1 belongs to the DSL family of ligands for Notch receptors that control the proliferation and differentiation of various cell lineages. However, little is known about the transcription factors that regulate its expression. Here, we show that Jagged1 is a Rel/NF-kappaB-responsive gene. Both c-Rel and RelA induced jagged1 gene expression, whereas a mutant defective for transactivation did not. Importantly, jagged1 transcripts were also upregulated by endogenous NF-kappaB activation and this effect was inhibited by a dominant mutant of IkappaBalpha, a physiological inhibitor of NF-kappaB. Cell surface expression of Jagged1 in c-Rel-expressing cell monolayers led to a functional interaction with lymphocytes expressing the Notch1/TAN-1 receptor. This correlated with the initiation of signaling downstream of Notch, as evidenced by increased levels of HES-1 transcripts in co-cultivated T cells and of CD23 transcripts in co-cultivated B cells. Consistent with its Rel/NF-kappaB-dependent induction, Jagged1 was found to be highly expressed in splenic B cells where c-Rel is expressed constitutively. These results demonstrate that c-Rel can trigger the Notch signaling pathway in neighboring cells by inducing jagged1 gene expression, and suggest a role for Jagged1 in B-cell activation, differentiation or function. These findings also highlight the potential for an interplay between the Notch and NF-kappaB signaling pathways in the immune system. PMID:10329626

  4. Inhibition of myostatin signaling through Notch activation following acute resistance exercise.

    PubMed

    MacKenzie, Matthew G; Hamilton, David Lee; Pepin, Mark; Patton, Amy; Baar, Keith

    2013-01-01

    Myostatin is a TGFβ family member and negative regulator of muscle size. Due to the complexity of the molecular pathway between myostatin mRNA/protein and changes in transcription, it has been difficult to understand whether myostatin plays a role in resistance exercise-induced skeletal muscle hypertrophy. To circumvent this problem, we determined the expression of a unique myostatin target gene, Mighty, following resistance exercise. Mighty mRNA increased by 6 h (82.9 ± 24.21%) and remained high out to 48 h (56.5 ± 19.67%) after resistance exercise. Further examination of the soleus, plantaris and tibialis anterior muscles showed that the change in Mighty mRNA at 6 h correlated with the increase in muscle size associated with this protocol (R(2) = 0.9996). The increase in Mighty mRNA occurred both independent of Smad2 phosphorylation and in spite of an increase in myostatin mRNA (341.8 ± 147.14% at 3 h). The myostatin inhibitor SKI remained unchanged. However, activated Notch, another potential inhibitor of TGFβ signaling, increased immediately following resistance exercise (83 ± 11.2%) and stayed elevated out to 6 h (78 ± 16.6%). Electroportion of the Notch intracellular domain into the tibialis anterior resulted in an increase in Mighty mRNA (63 ± 13.4%) that was equivalent to the canonical Notch target HES-1 (94.4 ± 7.32%). These data suggest that acute resistance exercise decreases myostatin signaling through the activation of the TGFβ inhibitor Notch resulting in a decrease in myostatin transcriptional activity that correlates well with muscle hypertrophy.

  5. TRPM7 channels regulate glioma stem cell through STAT3 and Notch signaling pathways.

    PubMed

    Liu, Mingli; Inoue, Koichi; Leng, Tiandong; Guo, Shanchun; Xiong, Zhi-gang

    2014-12-01

    Glioblastoma multiforme (GBM) is the most common and aggressive primary brain tumor in adults with median survival time of 14.6 months. A small fraction of cancer stem cells (CSC) initiate and maintain tumors thus driving glioma tumorigenesis and being responsible for resistance to classical chemo- and radio-therapies. It is desirable to identify signaling pathways related to CSC to develop novel therapies to selectively target them. Transient receptor potential cation channel, subfamily M, member 7, also known as TRPM7 is a ubiquitous, Ca(2+) and Mg(2+) permeable ion channels that are special in being both an ion channel and a serine/threonine kinase. In studies of glioma cells silenced for TRPM7, we demonstrated that Notch (Notch1, JAG1, Hey2, and Survivin) and STAT3 pathways are down regulated in glioma cells grown in monolayer. Furthermore, phospho-STAT3, Notch target genes and CSC markers (ALDH1 and CD133) were significantly higher in spheroid glioma CSCs when compared with monolayer cultures. The results further show that tyrosine-phosphorylated STAT3 binds and activates the ALDH1 promoters in glioma cells. We found that TRMP7-induced upregulation of ALDH1 expression is associated with increases in ALDH1 activity and is detectable in stem-like cells when expanded as spheroid CSCs. Finally, TRPM7 promotes proliferation, migration and invasion of glioma cells. These demonstrate that TRPM7 activates JAK2/STAT3 and/or Notch signaling pathways and leads to increased cell proliferation and migration. These findings for the first time demonstrates that TRPM7 (1) activates a previously unrecognized STAT3→ALDH1 pathway, and (2) promotes the induction of ALDH1 activity in glioma cells.

  6. Notch4 Signaling Induces a Mesenchymal–Epithelial–like Transition in Melanoma Cells to Suppress Malignant Behaviors

    PubMed Central

    Rad, Ehsan Bonyadi; Hammerlindl, Heinz; Wels, Christian; Popper, Ulrich; Menon, Dinoop Ravindran; Breiteneder, Heimo; Kitzwoegerer, Melitta; Hafner, Christine; Herlyn, Meenhard; Bergler, Helmut; Schaider, Helmut

    2016-01-01

    The effects of Notch signaling are context-dependent and both oncogenic and tumor-suppressive functions have been described. Notch signaling in melanoma is considered oncogenic, but clinical trials testing Notch inhibition in this malignancy have not proved successful. Here, we report that expression of the constitutively active intracellular domain of Notch4 (N4ICD) in melanoma cells triggered a switch from a mesenchymal-like parental phenotype to an epithelial-like phenotype. The epithelial-like morphology was accompanied by strongly reduced invasive, migratory, and proliferative properties concomitant with the downregulation of epithelial–mesenchymal transition markers Snail2 (SNAI2), Twist1, vimentin (VIM), and MMP2 and the reexpression of E-cadherin (CDH1). The N4ICD-induced phenotypic switch also resulted in significantly reduced tumor growth in vivo. Immunohistochemical analysis of primary human melanomas and cutaneous metastases revealed a significant correlation between Notch4 and E-cadherin expression. Mechanistically, we demonstrate that N4ICD induced the expression of the transcription factors Hey1 and Hey2, which bound directly to the promoter regions of Snail2 and Twist1 and repressed gene transcription, as determined by EMSA and luciferase assays. Taken together, our findings indicate a role for Notch4 as a tumor suppressor in melanoma, uncovering a potential explanation for the poor clinical efficacy of Notch inhibitors observed in this setting. PMID:26801977

  7. Salinomycin Suppresses PDGFRβ, MYC, and Notch Signaling in Human Medulloblastoma.

    PubMed

    Zhou, Shuang; Wang, Fengfei; Zhang, Ying; Johnson, Max R; Qian, Steven; Wu, Min; Wu, Erxi

    2014-01-01

    Medulloblastoma (MB) is the most common childhood brain tumor. Despite improved therapy and management, approximately 30% of patients die of the disease. To search for a more effective therapeutic strategy, the effects of salinomycin were tested on cell proliferation, cell death, and cell cycle progression in human MB cell lines. The results demonstrated that salinomycin inhibits cell proliferation, induces cell death , and disrupts cell cycle progression in MB cells. Salinomycin was also tested on the expression levels of key genes involved in proliferation and survival signaling and revealed that salinomycin down-regulates the expression of PDGFRβ, MYC, p21 and Bcl-2 as well as up-regulates the expression of cyclin A. In addition, the results reveal that salinomycin suppresses the expression of Hes1 and Hes5 in MB cells. Our data shed light on the potential of using salinomycin as a novel therapeutic agent for patients with MB.

  8. Downregulation of Notch1 and its potential correlation with epidermal growth factor receptor signalling in tongue squamous cell carcinoma.

    PubMed

    Huang, Hong-jie; Ping, Fei-yun; Hu, Ji-an; Zhao, Shi-fang

    2010-01-01

    We investigated the expression of Notch1 in human oral squamous cell carcinoma (SCC) and explored its potential correlation with epidermal growth factor receptor (EGFR) signalling in oral SCC. Paraffin sections of primary SCC of the tongue and normal mucosa were screened immunohistochemically for Notch1 and EGFR proteins. Human SCC of the tongue Tca8113 cells were treated with AG1478 to block EGFR signalling, and were transfected with the vector that encodes the specific short hairpin RNA (shRNA) that targets EGFR. In SCC of the tongue expression of Notch1 was cancelled except in sites of squamous metaplasia where it was raised, while expression of EGFR was found in the peripheral cells of carcinomas, but not in sites of squamous metaplasia. In normal tongue mucosa, Notch1 was expressed mainly in the stratum corneum, but not in the stratum basale, while EGFR was expressed mainly in the stratum basale, but not in the stratum granulosum or stratum corneum. The blocking of EGFR signalling or the silencing of the EGFR gene resulted in upregulation of Notch1 at mRNA and protein levels in Tca8113 cells. These observations suggest that downregulation of Notch1 in oral SCC may be associated with upregulation of EGFR signalling.

  9. Modulation of Notch Signaling Elicits Signature Tumors and Inhibits Hras1-Induced Oncogenesis in the Mouse Mammary Epithelium

    PubMed Central

    Kiaris, Hippokratis; Politi, Katerina; Grimm, Lisa M.; Szabolcs, Matthias; Fisher, Peter; Efstratiadis, Argiris; Artavanis-Tsakonas, Spyros

    2004-01-01

    Deregulation of Notch signaling, which normally affects a broad spectrum of cell fates, has been implicated in various neoplastic conditions. Here we describe a transgenic mouse model, which demonstrates that expression of a constitutively active form of the Notch1 receptor in the mammary epithelium induces the rapid development of pregnancy/lactation-dependent neoplasms that consistently exhibit a characteristic histopathological pattern. These signature tumors retain the ability to respond to apoptotic stimuli and regress on initiation of mammary gland involution, but eventually appear to progress in subsequent pregnancies to nonregressing malignant adenocarcinomas. Additionally, we present evidence indicating that cyclin D1 is an in vivo target of Notch signals in the mammary glands and demonstrate that we can effectively inhibit Hras1-driven, cyclin D1-dependent mammary oncogenesis by transgenic expression of the Notch antagonist Deltex. PMID:15277242

  10. Mouse Nkd1, a Wnt antagonist, exhibits oscillatory gene expression in the PSM under the control of Notch signaling.

    PubMed

    Ishikawa, Aki; Kitajima, Satoshi; Takahashi, Yu; Kokubo, Hiroki; Kanno, Jun; Inoue, Tohru; Saga, Yumiko

    2004-12-01

    During vertebrate embryogenesis, the formation of reiterated structures along the body axis is dependent upon the generation of the somite by segmentation of the presomitic mesoderm (PSM). Notch signaling plays a crucial role in both the generation and regulation of the molecular clock that provides the spatial information for PSM cells to form somites. In a screen for novel genes involved in somitogenesis, we identified a gene encoding a Wnt antagonist, Nkd1, which is transcribed in an oscillatory manner, and may represent a new member of the molecular clock constituents. The transcription of nkd1 is extremely downregulated in the PSM of vestigial tail (vt/vt), a hypomorphic mutant of Wnt3a, whereas nkd1 oscillations have a similar phase to lunatic fringe (L-fng) transcription and they are arrested in Hes7 (a negative regulator of Notch signaling) deficient embryos. These results suggest that the transcription of nkd1 requires Wnt3a, and that its oscillation patterns depend upon the function of Hes7. Wnt signaling has been postulated to be upstream of Notch signaling but we demonstrate in this study that a Wnt-signal-related gene may also be regulated by Notch signaling. Collectively, our data suggest that the reciprocal interaction of Notch and Wnt signals, and of their respective negative feedback loops, function to organize the segmentation clock required for somitogenesis.

  11. The bHLH Transcription Factor NeuroD Governs Photoreceptor Genesis and Regeneration Through Delta-Notch Signaling

    PubMed Central

    Taylor, Scott M.; Alvarez-Delfin, Karen; Saade, Carole J.; Thomas, Jennifer L.; Thummel, Ryan; Fadool, James M.; Hitchcock, Peter F.

    2015-01-01

    Purpose Photoreceptor genesis in the retina requires precise regulation of progenitor cell competence, cell cycle exit, and differentiation, although information around the mechanisms that govern these events currently is lacking. In zebrafish, the basic helix-loop-helix (bHLH) transcription factor NeuroD governs photoreceptor genesis, but the signaling pathways through which NeuroD functions are unknown. The purpose of this study was to identify these pathways, and during photoreceptor genesis, Notch signaling was investigated as the putative mediator of NeuroD function. Methods In embryos, genetic mosaic analysis was used to determine if NeuroD functions is cell- or non–cell-autonomous. Morpholino-induced NeuroD knockdown, CRISPR/Cas9 mutation, and pharmacologic and transgenic approaches were used, followed by in situ hybridization, immunocytochemistry, and quantitative RT-PCR (qRT-PCR), to identify mechanisms through which NeuroD functions. In adults, following photoreceptor ablation and NeuroD knockdown, similar methods as above were used to identify NeuroD function during photoreceptor regeneration. Results In embryos, NeuroD function is non–cell-autonomous, NeuroD knockdown increases Notch pathway gene expression, Notch inhibition rescues the NeuroD knockdown-induced deficiency in cell cycle exit but not photoreceptor maturation, and Notch activation and CRISPR/Cas9 mutation of neurod recapitulate NeuroD knockdown. In adults, NeuroD knockdown prevents cell cycle exit and photoreceptor regeneration and increases Notch pathway gene expression, and Notch inhibition rescues this phenotype. Conclusions These data demonstrate that during embryonic development, NeuroD governs photoreceptor genesis via non–cell-autonomous mechanisms and that, during photoreceptor development and regeneration, Notch signaling is a mechanistic link between NeuroD and cell cycle exit. In contrast, during embryonic development, NeuroD governs photoreceptor maturation via mechanisms

  12. Competition between Jagged-Notch and Endothelin1 Signaling Selectively Restricts Cartilage Formation in the Zebrafish Upper Face.

    PubMed

    Barske, Lindsey; Askary, Amjad; Zuniga, Elizabeth; Balczerski, Bartosz; Bump, Paul; Nichols, James T; Crump, J Gage

    2016-04-01

    The intricate shaping of the facial skeleton is essential for function of the vertebrate jaw and middle ear. While much has been learned about the signaling pathways and transcription factors that control facial patterning, the downstream cellular mechanisms dictating skeletal shapes have remained unclear. Here we present genetic evidence in zebrafish that three major signaling pathways - Jagged-Notch, Endothelin1 (Edn1), and Bmp - regulate the pattern of facial cartilage and bone formation by controlling the timing of cartilage differentiation along the dorsoventral axis of the pharyngeal arches. A genomic analysis of purified facial skeletal precursors in mutant and overexpression embryos revealed a core set of differentiation genes that were commonly repressed by Jagged-Notch and induced by Edn1. Further analysis of the pre-cartilage condensation gene barx1, as well as in vivo imaging of cartilage differentiation, revealed that cartilage forms first in regions of high Edn1 and low Jagged-Notch activity. Consistent with a role of Jagged-Notch signaling in restricting cartilage differentiation, loss of Notch pathway components resulted in expanded barx1 expression in the dorsal arches, with mutation of barx1 rescuing some aspects of dorsal skeletal patterning in jag1b mutants. We also identified prrx1a and prrx1b as negative Edn1 and positive Bmp targets that function in parallel to Jagged-Notch signaling to restrict the formation of dorsal barx1+ pre-cartilage condensations. Simultaneous loss of jag1b and prrx1a/b better rescued lower facial defects of edn1 mutants than loss of either pathway alone, showing that combined overactivation of Jagged-Notch and Bmp/Prrx1 pathways contribute to the absence of cartilage differentiation in the edn1 mutant lower face. These findings support a model in which Notch-mediated restriction of cartilage differentiation, particularly in the second pharyngeal arch, helps to establish a distinct skeletal pattern in the upper face.

  13. Competition between Jagged-Notch and Endothelin1 Signaling Selectively Restricts Cartilage Formation in the Zebrafish Upper Face

    PubMed Central

    Barske, Lindsey; Askary, Amjad; Zuniga, Elizabeth; Balczerski, Bartosz; Bump, Paul; Nichols, James T.; Crump, J. Gage

    2016-01-01

    The intricate shaping of the facial skeleton is essential for function of the vertebrate jaw and middle ear. While much has been learned about the signaling pathways and transcription factors that control facial patterning, the downstream cellular mechanisms dictating skeletal shapes have remained unclear. Here we present genetic evidence in zebrafish that three major signaling pathways − Jagged-Notch, Endothelin1 (Edn1), and Bmp − regulate the pattern of facial cartilage and bone formation by controlling the timing of cartilage differentiation along the dorsoventral axis of the pharyngeal arches. A genomic analysis of purified facial skeletal precursors in mutant and overexpression embryos revealed a core set of differentiation genes that were commonly repressed by Jagged-Notch and induced by Edn1. Further analysis of the pre-cartilage condensation gene barx1, as well as in vivo imaging of cartilage differentiation, revealed that cartilage forms first in regions of high Edn1 and low Jagged-Notch activity. Consistent with a role of Jagged-Notch signaling in restricting cartilage differentiation, loss of Notch pathway components resulted in expanded barx1 expression in the dorsal arches, with mutation of barx1 rescuing some aspects of dorsal skeletal patterning in jag1b mutants. We also identified prrx1a and prrx1b as negative Edn1 and positive Bmp targets that function in parallel to Jagged-Notch signaling to restrict the formation of dorsal barx1+ pre-cartilage condensations. Simultaneous loss of jag1b and prrx1a/b better rescued lower facial defects of edn1 mutants than loss of either pathway alone, showing that combined overactivation of Jagged-Notch and Bmp/Prrx1 pathways contribute to the absence of cartilage differentiation in the edn1 mutant lower face. These findings support a model in which Notch-mediated restriction of cartilage differentiation, particularly in the second pharyngeal arch, helps to establish a distinct skeletal pattern in the upper

  14. Integration of Orthogonal Signaling by the Notch and Dpp Pathways in Drosophila

    PubMed Central

    Stroebele, Elizabeth; Erives, Albert

    2016-01-01

    The transcription factor Suppressor of Hairless and its coactivator, the Notch intracellular domain, are polyglutamine (pQ)-rich factors that target enhancer elements and interact with other locally bound pQ-rich factors. To understand the functional repertoire of such enhancers, we identify conserved regulatory belts with binding sites for the pQ-rich effectors of both Notch and BMP/Dpp signaling, and the pQ-deficient tissue selectors Apterous (Ap), Scalloped (Sd), and Vestigial (Vg). We find that the densest such binding site cluster in the genome is located in the BMP-inducible nab locus, a homolog of the vertebrate transcriptional cofactors NAB1/NAB2. We report three major findings. First, we find that this nab regulatory belt is a novel enhancer driving dorsal wing margin expression in regions of peak phosphorylated Mad in wing imaginal discs. Second, we show that Ap is developmentally required to license the nab dorsal wing margin enhancer (DWME) to read out Notch and Dpp signaling in the dorsal compartment. Third, we find that the nab DWME is embedded in a complex of intronic enhancers, including a wing quadrant enhancer, a proximal wing disc enhancer, and a larval brain enhancer. This enhancer complex coordinates global nab expression via both tissue-specific activation and interenhancer silencing. We suggest that DWME integration of BMP signaling maintains nab expression in proliferating margin descendants that have divided away from Notch–Delta boundary signaling. As such, uniform expression of genes like nab and vestigial in proliferating compartments would typically require both boundary and nonboundary lineage-specific enhancers. PMID:26975664

  15. Dissecting and circumventing the requirement for RAM in CSL-dependent Notch signaling.

    PubMed

    Johnson, Scott E; Barrick, Douglas

    2012-01-01

    The Notch signaling pathway is an intercellular communication network vital to metazoan development. Notch activation leads to the nuclear localization of the intracellular portion (NICD) of the Notch receptor. Once in the nucleus, NICD binds the transcription factor CSL through a bivalent interaction involving the high-affinity RAM region and the lower affinity ANK domain, converting CSL from a transcriptionally-repressed to an active state. This interaction is believed to directly displace co-repressor proteins from CSL and recruit co-activator proteins. Here we investigate the consequences of this bivalent organization in converting CSL from the repressed to active form. One proposed function of RAM is to promote the weak ANK:CSL interaction; thus, fusion of CSL-ANK should bypass this function of RAM. We find that a CSL-ANK fusion protein is transcriptionally active in reporter assays, but that the addition of RAM in trans further increases transcriptional activity, suggesting another role of RAM in activation. A single F235L point substitution, which disrupts co-repressor binding to CSL, renders the CSL-ANK fusion fully active and refractory to further stimulation by RAM in trans. These results suggest that in the context of a mammalian CSL-ANK fusion protein, the main role of RAM is to displace co-repressor proteins from CSL.

  16. TQ inhibits hepatocellular carcinoma growth in vitro and in vivo via repression of Notch signaling

    PubMed Central

    Ke, Xiquan; Zhao, Yan; Lu, Xinlan; Wang, Zhe; Liu, Yuanyuan; Ren, Mudan; Lu, Guifang; Zhang, Dan; Sun, Zhenguo; Xu, Zhipeng; Song, Jee Hoon; Cheng, Yulan; Meltzer, Stephen J.; He, Shuixiang

    2015-01-01

    Thymoquinone (TQ) has been reported to possess anti-tumor activity in various types of cancer. However, its effects and molecular mechanism of action in hepatocellular carcinoma (HCC) are still not completely understood. We observed that TQ inhibited tumor cell growth in vitro, where treatment with TQ arrested the cell cycle in G1 by upregulating p21 and downregulating cyclinD1 and CDK2 expression; moreover, TQ induced apoptosis by decreasing expression of Bcl-2 and increasing expression of Bax. Simultaneously, TQ demonstrated a suppressive impact on the Notch pathway, where overexpression of NICD1 reversed the inhibitory effect of TQ on cell proliferation, thereby attenuating the repressive effects of TQ on the Notch pathway, cyclinD1, CDK2 and Bcl-2, and also diminishing upregulation of p21 and Bax. In a xenograft model, TQ inhibited HCC growth in nude mice; this inhibitory effect in vivo, as well as of HCC cell growth in vitro, was associated with a discernible decline in NICD1 and Bcl-2 levels and a dramatic rise in p21 expression. In conclusion, TQ inhibits HCC cell growth by inducing cell cycle arrest and apoptosis, achieving these effects by repression of the Notch signaling pathway, suggesting that TQ represents a potential preventive or therapeutic agent in HCC patients. PMID:26416455

  17. FGF SIGNALING EXPANDS EMBRYONIC CORTICAL SURFACE AREA BY REGULATING NOTCH-DEPENDENT NEUROGENESIS

    PubMed Central

    Rash, Brian G.; Lim, H. David; Breunig, Joshua J.; Vaccarino, Flora M.

    2011-01-01

    The processes regulating cortical surface area expansion during development and evolution are unknown. We show that loss of function of all Fibroblast Growth Factor Receptors (FgfR) expressed at the earliest stages of cortical development causes severe deficits in surface area growth by embryonic day (E) 12.5 in the mouse. In FgfR mutants, accelerated production of neurons led to severe loss of radial progenitors and premature termination of neurogenesis. Nevertheless, these mutants showed remarkably little change in cortical layer structure. Birthdating experiments indicated that a greater proportion of layer fates was generated during early neurogenic stages, revealing that FgfR activity normally slows the temporal progression of cortical layer fates. Electroporation of a dominant negative FgfR at E11.5 increased cortical neurogenesis in normal mice—an effect that was blocked by simultaneous activation of the Notch pathway. Together with changes in the expression of Notch pathway genes in FgfR mutant embryos, these findings indicate that Notch lies downstream of FgfR signaling in the same pathway regulating cortical neurogenesis and begin to establish a mechanism for regulating cortical surface expansion. PMID:22031906

  18. PlexinD1 Is a Novel Transcriptional Target and Effector of Notch Signaling in Cancer Cells

    PubMed Central

    Rehman, Michael; Capparuccia, Lorena

    2016-01-01

    The secreted semaphorin Sema3E controls cell migration and invasiveness in cancer cells. Sema3E-receptor, PlexinD1, is frequently upregulated in melanoma, breast, colon, ovarian and prostate cancers; however, the mechanisms underlying PlexinD1 upregulation and the downstream events elicited in tumor cells are still unclear. Here we show that the canonical RBPjk-dependent Notch signaling cascade controls PlexinD1 expression in primary endothelial and cancer cells. Transcriptional activation was studied by quantitative PCR and promoter activity reporter assays. We found that Notch ligands and constitutively activated intracellular forms of Notch receptors upregulated PlexinD1 expression; conversely RNAi-based knock-down, or pharmacological inhibition of Notch signaling by gamma-secretase inhibitors, downregulated PlexinD1 levels. Notably, both Notch1 and Notch3 expression positively correlates with PlexinD1 levels in prostate cancer, as well as in other tumor types. In prostate cancer cells, Sema3E-PlexinD1 axis was previously reported to regulate migration; however, implicated mechanisms were not elucidated. Here we show that in these cells PlexinD1 activity induces the expression of the transcription factor Slug, downregulates E-cadherin levels and enhances cell migration. Moreover, our mechanistic data identify PlexinD1 as a pivotal mediator of this signaling axis downstream of Notch in prostate cancer cells. In fact, on one hand, PlexinD1 is required to mediate cell migration and E-cadherin regulation elicited by Notch. On the other hand, PlexinD1 upregulation is sufficient to induce prostate cancer cell migration and metastatic potential in mice, leading to functional rescue in the absence of Notch. In sum, our work identifies PlexinD1 as a novel transcriptional target induced by Notch signaling, and reveals its role promoting prostate cancer cell migration and downregulating E-cadherin levels in Slug-dependent manner. Collectively, these findings suggest that

  19. Inhibition of notch signaling in glioblastoma targets cancer stem cells via an endothelial cell intermediate.

    PubMed

    Hovinga, Koos E; Shimizu, Fumiko; Wang, Rong; Panagiotakos, Georgia; Van Der Heijden, Maartje; Moayedpardazi, Hamideh; Correia, Ana Sofia; Soulet, Denis; Major, Tamara; Menon, Jayanthi; Tabar, Viviane

    2010-06-01

    Glioblastoma multiforme (GBM) is a highly heterogeneous malignant tumor. Recent data suggests the presence of a hierarchical organization within the GBM cell population that involves cancer cells with stem-like behavior, capable of repopulating the tumor and contributing to its resistance to therapy. Tumor stem cells are thought to reside within a vascular niche that provides structural and functional support. However, most GBM studies involve isolated tumor cells grown under various culture conditions. Here, we use a novel three-dimensional organotypic "explant" system of surgical GBM specimens that preserves cytoarchitecture and tumor stroma along with tumor cells. Notch inhibition in explants results in decreased proliferation and self-renewal of tumor cells but is also associated with a decrease in endothelial cells. When endothelial cells are selectively eliminated from the explants via a toxin conjugate, we also observed a decrease in self-renewal of tumor stem cells. These findings support a critical role for tumor endothelial cells in GBM stem cell maintenance, mediated at least in part by Notch signaling. The explant system further highlighted differences in the response to radiation between explants and isolated tumor neurospheres. Combination treatment with Notch blockade and radiation resulted in a substantial decrease in proliferation and in self-renewal in tumor explants while radiation alone was less effective. This data suggests that the Notch pathway plays a critical role in linking angiogenesis and cancer stem cell self-renewal and is thus a potential therapeutic target. Three-dimensional explant systems provide a novel approach for the study of tumor and microenvironment interactions.

  20. Ataxin 2-binding protein 1 is a context-specific positive regulator of Notch signaling during neurogenesis in Drosophila melanogaster.

    PubMed

    Shukla, Jay Prakash; Deshpande, Girish; Shashidhara, L S

    2017-03-01

    The role of the Notch pathway during the lateral inhibition that underlies binary cell fate choice is extensively studied, but the context specificity that generates diverse outcomes is less well understood. In the peripheral nervous system of Drosophila melanogaster, differential Notch signaling between cells of the proneural cluster orchestrates sensory organ specification. Here we report functional analysis of Drosophila Ataxin 2-binding protein 1 (A2BP1) during this process. Its human ortholog is linked to type 2 spinocerebellar ataxia and other complex neuronal disorders. Downregulation of Drosophila A2BP1 in the proneural cluster increases adult sensory bristle number, whereas its overexpression results in loss of bristles. We show that A2BP1 regulates sensory organ specification by potentiating Notch signaling. Supporting its direct involvement, biochemical analysis shows that A2BP1 is part of the Suppressor of Hairless [Su(H)] complex in the presence and absence of Notch. However, in the absence of Notch signaling, the A2BP1 interacting fraction of Su(H) does not associate with the repressor proteins Groucho and CtBP. We propose a model explaining the requirement of A2BP1 as a positive regulator of context-specific Notch activity.

  1. Common profiles of Notch signaling differentiate disease-free survival in luminal type A and triple negative breast cancer

    PubMed Central

    Orzechowska, Magdalena; Jędroszka, Dorota; Bednarek, Andrzej K

    2017-01-01

    Breast cancer (BC) is characterized by high heterogeneity regarding its biology and clinical characteristics. The Notch pathway regulates such processes as organ modeling and epithelial-to-mesenchymal transition (EMT). The aim of the study was to determine the effect of differential expression of Notch members on disease-free survival (DFS) in luminal type A (lumA) and triple negative (TN) BC. The differential expression of 19 Notch members was examined in a TCGA BC cohort. DFS analysis was performed using the log-rank test (p<0.05). Biological differences between DFS groups were determined with Gene Set Enrichment Analysis (GSEA) (tTest, FDR<0.25). Common expression profiles according to Notch signaling were examined using ExpressCluster (K-means, mean centered, Euclidean distance metric). The overexpression of HES1, LFNG and PSEN1 was found to be favorable for DFS in lumA, and lowered expression favorable for DFS in TN. GSEA analysis showed that differential Notch signaling is associated with cell cycle, tissue architecture and remodeling. Particularly, targets of E2F, early stage S phase transcription factor, were upregulated in the lumA unfavorable group and the TN favorable group differentiated on a basis of HES1 and PSEN1 expression. Summarizing, our analysis show significance of Notch signaling in BRCA progression through triggering EMT. Moreover, identification of numerous genes which overexpression is associated with disease recurrence may serve as a source of potential targets for a new anticancer therapy. PMID:27888801

  2. Pre-Clinical studies of Notch Signaling Inhibitor RO4929097 in Inflammatory Breast Cancer Cells

    PubMed Central

    Debeb, Bisrat G.; Cohen, Evan N.; Boley, Kimberly; Freiter, Erik M.; Li, Li; Robertson, Fredika M.; Reuben, James M.; Cristofanilli, Massimo; Buchholz, Thomas A.; Woodward, Wendy A.

    2015-01-01

    Basal breast cancer, common among patients presenting with inflammatory breast cancer, has been shown to be resistant to radiation and enriched in cancer stem cells. The Notch pathway plays an important role in self-renewal of breast cancer stem cells and contributes to inflammatory signaling that promotes the breast cancer stem cell phenotype. Herein we inhibited Notch signaling using a gamma secretase inhibitor, RO4929097, in an in vitro model that enriches for cancer initiating cells (3D clonogenic assay) and conventional 2D clonogenic assay to compare the effect on radiosensitization of the SUM149 and SUM190 inflammatory breast cancer (IBC) cell lines. RO4929097 downregulated the Notch target genes Hes1, Hey1 and HeyL and showed a significant reduction in anchorage independent growth in SUM190 and SUM149. However, the putative self-renewal assay mammosphere formation efficiency was increased with the drug. To assess radiosensitization of putative cancer stem cells, cells were exposed to increasing doses of radiation with or without 1uM RO4929097 in their standard (2D) and self-renewal enriching (3D) culture conditions. In the conventional 2D clonogenic assay, RO4929097 significantly sensitized SUM190 cells to ionizing radiation and has a modest radiosensitization effect in SUM149 cells. In the 3D clonogenic assays, however, a radioprotective effect was seen in both SUM149 and SUM190 cells at higher doses. Both cell lines express IL-6 and IL-8, cytokines known to mediate the efficacy of notch inhibition and to promote self-renewal of stem cells. We further showed that RO429097 inhibits normal T-cell synthesis of some inflammatory cytokines, including TNF-α, a potential mediator of IL-6 and IL-8 production in the microenvironment. These data suggest additional targeting agents may be required to selectively target IBC stem cells through notch inhibition, and that evaluation of microenvironmental influences may shed further light on the potential effects of this

  3. Stomaching Notch

    PubMed Central

    Yin, Xiaolei; Karp, Jeffrey M

    2015-01-01

    The self-renewal and differentiation of tissue stem cells must be tightly controlled. Unrestrained self-renewal leads to over-proliferation of stem cells, which may cause tumor formation, while uncontrolled differentiation leads to depletion of the stem cell pool. In this issue of The EMBO Journal, Demitrack et al (2015) show that the Notch pathway is a key regulator of Lgr5 antral stem cell self-renewal and differentiation. Notch signaling controls the proliferation and differentiation of stem cells as well as gastric tissue growth, while uncontrolled Notch activity in stem cells leads to polyp formation. PMID:26358838

  4. The NHR domains of Neuralized and related proteins: Beyond Notch signalling.

    PubMed

    Liu, Sili; Boulianne, Gabrielle L

    2017-01-01

    Neuralized Homology Repeats (NHRs) were first identified in Neuralized, an E3-ubiquitin ligase that plays a key role in the Notch signalling pathway. Since their original discovery, NHR domains have been shown to regulate protein-protein interactions in a broad range of developmental processes and in a wide variety of species from flies to humans. The NHR family of proteins can be categorized into three groups: (1) those that contain a RING finger, (2) those that contain a SOCS box and, (3) those that only have NHR domains. Here we review the structure and function of NHR domains in various cellular and developmental processes.

  5. Antagonism of EGFR and notch signalling in the reiterative recruitment of Drosophila adult chordotonal sense organ precursors.

    PubMed

    zur Lage, P; Jarman, A P

    1999-06-01

    The selection of Drosophila melanogaster sense organ precursors (SOPs) for sensory bristles is a progressive process: each neural equivalence group is transiently defined by the expression of proneural genes (proneural cluster), and neural fate is refined to single cells by Notch-Delta lateral inhibitory signalling between the cells. Unlike sensory bristles, SOPs of chordotonal (stretch receptor) sense organs are tightly clustered. Here we show that for one large adult chordotonal SOP array, clustering results from the progressive accumulation of a large number of SOPs from a persistent proneural cluster. This is achieved by a novel interplay of inductive epidermal growth factor-receptor (EGFR) and competitive Notch signals. EGFR acts in opposition to Notch signalling in two ways: it promotes continuous SOP recruitment despite lateral inhibition, and it attenuates the effect of lateral inhibition on the proneural cluster equivalence group, thus maintaining the persistent proneural cluster. SOP recruitment is reiterative because the inductive signal comes from previously recruited SOPs.

  6. The atypical cadherin Flamingo links Frizzled and Notch signaling in planar polarity establishment in the Drosophila eye.

    PubMed

    Das, Gishnu; Reynolds-Kenneally, Jessica; Mlodzik, Marek

    2002-05-01

    Planar cell polarity is established in the Drosophila eye through distinct fate specification of photoreceptors R3 and R4 by a two-tiered mechanism employing Fz and Notch signaling: Fz signaling specifies R3 and induces Dl to activate Notch in R4. We show that the atypical cadherin Flamingo (Fmi) plays critical, but distinct, roles in both R3 and R4. Fmi is first enriched at equatorial cell borders of R3/R4, positively interacting with Fz/Dsh. Subsequently, Fmi is upregulated in R4 by Notch and functions to downregulate Dl expression by antagonizing Fz signaling. This in turn amplifies and enforces the initial Fz-signaling bias in the R3/R4 pair. Our results reveal differences in the planar cell polarity genetic circuitry between the eye and the wing.

  7. Regulation of differentiation flux by Notch signalling influences the number of dopaminergic neurons in the adult brain.

    PubMed

    Trujillo-Paredes, Niurka; Valencia, Concepción; Guerrero-Flores, Gilda; Arzate, Dulce-María; Baizabal, José-Manuel; Guerra-Crespo, Magdalena; Fuentes-Hernández, Ayari; Zea-Armenta, Iván; Covarrubias, Luis

    2016-02-24

    Notch signalling is a well-established pathway that regulates neurogenesis. However, little is known about the role of Notch signalling in specific neuronal differentiation. Using Dll1 null mice, we found that Notch signalling has no function in the specification of mesencephalic dopaminergic neural precursor cells (NPCs), but plays an important role in regulating their expansion and differentiation into neurons. Premature neuronal differentiation was observed in mesencephalons of Dll1-deficient mice or after treatment with a Notch signalling inhibitor. Coupling between neurogenesis and dopaminergic differentiation was indicated from the coincident emergence of neuronal and dopaminergic markers. Early in differentiation, decreasing Notch signalling caused a reduction in NPCs and an increase in dopaminergic neurons in association with dynamic changes in the proportion of sequentially-linked dopaminergic NPCs (Msx1/2+, Ngn2+, Nurr1+). These effects in differentiation caused a significant reduction in the number of dopaminergic neurons produced. Accordingly, Dll1 haploinsufficient adult mice, in comparison with their wild-type littermates, have a consistent reduction in neuronal density that was particularly evident in the substantia nigra pars compacta. Our results are in agreement with a mathematical model based on a Dll1-mediated regulatory feedback loop between early progenitors and their dividing precursors that controls the emergence and number of dopaminergic neurons.

  8. Regulation of differentiation flux by Notch signalling influences the number of dopaminergic neurons in the adult brain

    PubMed Central

    Trujillo-Paredes, Niurka; Valencia, Concepción; Guerrero-Flores, Gilda; Arzate, Dulce-María; Baizabal, José-Manuel; Guerra-Crespo, Magdalena; Fuentes-Hernández, Ayari; Zea-Armenta, Iván; Covarrubias, Luis

    2016-01-01

    ABSTRACT Notch signalling is a well-established pathway that regulates neurogenesis. However, little is known about the role of Notch signalling in specific neuronal differentiation. Using Dll1 null mice, we found that Notch signalling has no function in the specification of mesencephalic dopaminergic neural precursor cells (NPCs), but plays an important role in regulating their expansion and differentiation into neurons. Premature neuronal differentiation was observed in mesencephalons of Dll1-deficient mice or after treatment with a Notch signalling inhibitor. Coupling between neurogenesis and dopaminergic differentiation was indicated from the coincident emergence of neuronal and dopaminergic markers. Early in differentiation, decreasing Notch signalling caused a reduction in NPCs and an increase in dopaminergic neurons in association with dynamic changes in the proportion of sequentially-linked dopaminergic NPCs (Msx1/2+, Ngn2+, Nurr1+). These effects in differentiation caused a significant reduction in the number of dopaminergic neurons produced. Accordingly, Dll1 haploinsufficient adult mice, in comparison with their wild-type littermates, have a consistent reduction in neuronal density that was particularly evident in the substantia nigra pars compacta. Our results are in agreement with a mathematical model based on a Dll1-mediated regulatory feedback loop between early progenitors and their dividing precursors that controls the emergence and number of dopaminergic neurons. PMID:26912775

  9. Myeloid-Specific Blockade of Notch Signaling Attenuates Choroidal Neovascularization through Compromised Macrophage Infiltration and Polarization in Mice

    PubMed Central

    Dou, Guo-Rui; Li, Na; Chang, Tian-Fang; Zhang, Ping; Gao, Xiang; Yan, Xian-Chun; Liang, Liang; Han, Hua; Wang, Yu-Sheng

    2016-01-01

    Macrophages have been recognized as an important inflammatory component in choroidal neovascularization (CNV). However, it is unclear how these cells are activated and polarized, how they affect angiogenesis and what the underlining mechanisms are during CNV. Notch signaling has been implicated in macrophage activation. Previously we have shown that inducible disruption of RBP-J, the critical transcription factor of Notch signaling, in adult mice results in enhanced CNV, but it is unclear what is the role of macrophage-specific Notch signaling in the development of CNV. In the current study, by using the myeloid specific RBP-J knockout mouse model combined with the laser-induced CNV model, we show that disruption of Notch signaling in macrophages displayed attenuated CNV growth, reduced macrophage infiltration and activation, and alleviated angiogenic response after laser induction. The inhibition of CNV occurred with reduced expression of VEGF and TNF-α in infiltrating inflammatory macrophages in myeloid specific RBP-J knockout mice. These changes might result in direct inhibition of EC lumen formation, as shown in an in vitro study. Therefore, clinical intervention of Notch signaling in CNV needs to pinpoint myeloid lineage to avoid the counteractive effects of global inhibition. PMID:27339903

  10. Notch1–STAT3–ETBR signaling axis controls reactive astrocyte proliferation after brain injury

    PubMed Central

    LeComte, Matthew D.; Shimada, Issei S.; Sherwin, Casey; Spees, Jeffrey L.

    2015-01-01

    Defining the signaling network that controls reactive astrogliosis may provide novel treatment targets for patients with diverse CNS injuries and pathologies. We report that the radial glial cell antigen RC2 identifies the majority of proliferating glial fibrillary acidic protein-positive (GFAP+) reactive astrocytes after stroke. These cells highly expressed endothelin receptor type B (ETBR) and Jagged1, a Notch1 receptor ligand. To study signaling in adult reactive astrocytes, we developed a model based on reactive astrocyte-derived neural stem cells isolated from GFAP-CreER-Notch1 conditional knockout (cKO) mice. By loss- and gain-of-function studies and promoter activity assays, we found that Jagged1/Notch1 signaling increased ETBR expression indirectly by raising the level of phosphorylated signal transducer and activator of transcription 3 (STAT3), a previously unidentified EDNRB transcriptional activator. Similar to inducible transgenic GFAP-CreER-Notch1-cKO mice, GFAP-CreER-ETBR-cKO mice exhibited a defect in reactive astrocyte proliferation after cerebral ischemia. Our results indicate that the Notch1–STAT3–ETBR axis connects a signaling network that promotes reactive astrocyte proliferation after brain injury. PMID:26124113

  11. Tsukushi expression is dependent on Notch signaling and oscillated in the presomitic mesoderm during chick somitogenesis.

    PubMed

    Acharjee, Uzzal Kumar; Gejima, Ryu; Felemban Athary Abdulhaleem, M; Riyadh, M Asrafuzzaman; Tanaka, Hideaki; Ohta, Kunimasa

    2015-09-25

    During somitogenesis, segmentation of the body axis occurs by epithelial somites budding off from the rostral end of the unsegmented presomitic mesoderm (PSM), and its molecular regulation is achieved by a molecular oscillator and signaling molecules. Tsukushi (TSK) is a unique secreted protein and involved in diverse biological cascades in vertebrate embryos by modulating several signaling pathways at the extracellular region. However, the involvement of TSK in somitogenesis remains unknown. In this study, we investigated the detailed expression patterns of TSK at different developmental stages of a chick embryo. Chick-TSK (C-TSK) is expressed in the PSM and shows an oscillation pattern with three phases. The oscillation pattern of C-TSK in the PSM is similar to that of c-Notch1 and c-hairy1, but not to c-Delta1. Our in vitro data showed that Notch signaling is necessary for the normal expression of C-TSK and that expression of C-TSK is an intrinsic property of the anterior PSM. These data suggest that TSK plays a role in chick somitogenesis.

  12. Preclinical profile of a potent gamma-secretase inhibitor targeting notch signaling with in vivo efficacy and pharmacodynamic properties.

    PubMed

    Luistro, Leopoldo; He, Wei; Smith, Melissa; Packman, Kathryn; Vilenchik, Maria; Carvajal, Daisy; Roberts, John; Cai, James; Berkofsky-Fessler, Windy; Hilton, Holly; Linn, Michael; Flohr, Alexander; Jakob-Røtne, Roland; Jacobsen, Helmut; Glenn, Kelli; Heimbrook, David; Boylan, John F

    2009-10-01

    Notch signaling is an area of great interest in oncology. RO4929097 is a potent and selective inhibitor of gamma-secretase, producing inhibitory activity of Notch signaling in tumor cells. The RO4929097 IC50 in cell-free and cellular assays is in the low nanomolar range with >100-fold selectivity with respect to 75 other proteins of various types (receptors, ion channels, and enzymes). RO4929097 inhibits Notch processing in tumor cells as measured by the reduction of intracellular Notch expression by Western blot. This leads to reduced expression of the Notch transcriptional target gene Hes1. RO4929097 does not block tumor cell proliferation or induce apoptosis but instead produces a less transformed, flattened, slower-growing phenotype. RO4929097 is active following oral dosing. Antitumor activity was shown in 7 of 8 xenografts tested on an intermittent or daily schedule in the absence of body weight loss or Notch-related toxicities. Importantly, efficacy is maintained after dosing is terminated. Angiogenesis reverse transcription-PCR array data show reduced expression of several key angiogenic genes. In addition, comparative microarray analysis suggests tumor cell differentiation as an additional mode of action. These preclinical results support evaluation of RO4929097 in clinical studies using an intermittent dosing schedule. A multicenter phase I dose escalation study in oncology is under way.

  13. Requirement of Split ends for Epigenetic Regulation of Notch Signal-Dependent Genes during Infection-Induced Hemocyte Differentiation▿ †

    PubMed Central

    Jin, Li Hua; Choi, Jung Kyoon; Kim, Byungil; Cho, Hwan Sung; Kim, Jihyun; Kim-Ha, Jeongsil; Kim, Young-Joon

    2009-01-01

    Drosophila producing a mutant form of the putative transcription coregulator, Split ends (Spen), originally identified in the analysis of neuronal development, display diverse immune defects. In order to understand the role of Spen in the innate immune response, we analyzed the transcriptional defects associated with spen mutant hemocytes and their relationship to the Notch signaling pathways. Spen is regulated by the Notch pathway in the lymph glands and is required for Notch-dependent activation of a large number of genes involved in energy metabolism and differentiation. Analysis of the epigenetic marks associated with Spen-dependent genes indicates that Spen performs its function as a coactivator by regulating chromatin modification. Intriguingly, expression of the Spen-dependent genes was transiently downregulated in a Notch-dependent manner by the Dif activated upon recognition of pathogen-associated molecules, demonstrating the existence of cross talk between hematopoietic regulation and the innate immune response. Our observations reveal a novel connection between the Notch and Toll/IMD signaling pathways and demonstrate a coactivating role for Spen in activating Notch-dependent genes in differentiating cells. PMID:19139277

  14. Requirement of Split ends for epigenetic regulation of Notch signal-dependent genes during infection-induced hemocyte differentiation.

    PubMed

    Jin, Li Hua; Choi, Jung Kyoon; Kim, Byungil; Cho, Hwan Sung; Kim, Jihyun; Kim-Ha, Jeongsil; Kim, Young-Joon

    2009-03-01

    Drosophila producing a mutant form of the putative transcription coregulator, Split ends (Spen), originally identified in the analysis of neuronal development, display diverse immune defects. In order to understand the role of Spen in the innate immune response, we analyzed the transcriptional defects associated with spen mutant hemocytes and their relationship to the Notch signaling pathways. Spen is regulated by the Notch pathway in the lymph glands and is required for Notch-dependent activation of a large number of genes involved in energy metabolism and differentiation. Analysis of the epigenetic marks associated with Spen-dependent genes indicates that Spen performs its function as a coactivator by regulating chromatin modification. Intriguingly, expression of the Spen-dependent genes was transiently downregulated in a Notch-dependent manner by the Dif activated upon recognition of pathogen-associated molecules, demonstrating the existence of cross talk between hematopoietic regulation and the innate immune response. Our observations reveal a novel connection between the Notch and Toll/IMD signaling pathways and demonstrate a coactivating role for Spen in activating Notch-dependent genes in differentiating cells.

  15. Notch3 signaling-mediated melanoma-endothelial crosstalk regulates melanoma stem-like cell homeostasis and niche morphogenesis.

    PubMed

    Hsu, Mei-Yu; Yang, Moon Hee; Schnegg, Caroline I; Hwang, Soonyean; Ryu, Byungwoo; Alani, Rhoda M

    2017-02-06

    Melanoma is among the most virulent cancers, owing to its propensity to metastasize and its resistance to current therapies. The treatment failure is largely attributed to tumor heterogeneity, particularly subpopulations possessing stem cell-like properties, ie, melanoma stem-like cells (MSLCs). Evidence indicates that the MSLC phenotype is malleable and may be acquired by non-MSLCs through phenotypic switching upon appropriate stimuli, the so-called 'dynamic stemness'. Since the phenotypic characteristics and functional integrity of MSLCs depend on their vascular niche, using a two-dimensional (2D) melanoma-endothelium co-culture model, where the MSLC niche is recapitulated in vitro, we identified Notch3 signaling pathway as a micro-environmental cue governing MSLC phenotypic plasticity via pathway-specific gene expression arrays. Accordingly, lentiviral shRNA-mediated Notch3 knockdown (KD) in melanoma cell lines exhibiting high levels of endogenous Notch3 led to retarded/abolished tumorigenicity in vivo through both depleting MSLC fractions, evinced by MSLC marker downregulation (eg, CD133 and CD271); and impeding the MSLC niche, corroborated by the attenuated tumor angiogenesis as well as vasculogenic mimicry. In contrast, Notch3 KD affected neither tumor growth nor MSLC subsets in a melanoma cell line with relatively low endogenous Notch3 expression. Thus, Notch3 signaling may facilitate MSLC plasticity and niche morphogenesis in a cell context-dependent manner. Our findings illustrate Notch3 as a molecular switch driving melanoma heterogeneity, and provide the biological rationale for Notch inhibition as a promising therapeutic option.Laboratory Investigation advance online publication, 6 February 2017; doi:10.1038/labinvest.2017.1.

  16. Thyroid Hormone-Induced Activation of Notch Signaling is Required for Adult Intestinal Stem Cell Development During Xenopus Laevis Metamorphosis.

    PubMed

    Hasebe, Takashi; Fujimoto, Kenta; Kajita, Mitsuko; Fu, Liezhen; Shi, Yun-Bo; Ishizuya-Oka, Atsuko

    2016-11-21

    In Xenopus laevis intestine during metamorphosis, the larval epithelial cells are removed by apoptosis, and the adult epithelial stem (AE) cells appear concomitantly. They proliferate and differentiate to form the adult epithelium (Ep). Thyroid hormone (TH) is well established to trigger this remodeling by regulating the expression of various genes including Notch receptor. To study the role of Notch signaling, we have analyzed the expression of its components, including the ligands (DLL and Jag), receptor (Notch), and targets (Hairy), in the metamorphosing intestine by real-time reverse transcription-polymerase chain reaction and in situ hybridization or immunohistochemistry. We show that they are up-regulated during both natural and TH-induced metamorphosis in a tissue-specific manner. Particularly, Hairy1 is specifically expressed in the AE cells. Moreover, up-regulation of Hairy1 and Hairy2b by TH was prevented by treating tadpoles with a γ-secretase inhibitor (GSI), which inhibits Notch signaling. More importantly, TH-induced up-regulation of LGR5, an adult intestinal stem cell marker, was suppressed by GSI treatment. Our results suggest that Notch signaling plays a role in stem cell development by regulating the expression of Hairy genes during intestinal remodeling. Furthermore, we show with organ culture experiments that prolonged exposure of tadpole intestine to TH plus GSI leads to hyperplasia of secretory cells and reduction of absorptive cells. Our findings here thus provide evidence for evolutionarily conserved role of Notch signaling in intestinal cell fate determination but more importantly reveal, for the first time, an important role of Notch pathway in the formation of adult intestinal stem cells during vertebrate development. Stem Cells 2016.

  17. The PDZ Protein Canoe/AF-6 Links Ras-MAPK, Notch and Wingless/Wnt Signaling Pathways by Directly Interacting with Ras, Notch and Dishevelled

    PubMed Central

    Carmena, Ana; Speicher, Stephan; Baylies, Mary

    2006-01-01

    Over the past few years, it has become increasingly apparent that signal transduction pathways are not merely linear cascades; they are organized into complex signaling networks that require high levels of regulation to generate precise and unique cell responses. However, the underlying regulatory mechanisms by which signaling pathways cross-communicate remain poorly understood. Here we show that the Ras-binding protein Canoe (Cno)/AF-6, a PDZ protein normally associated with cellular junctions, is a key modulator of Wingless (Wg)/Wnt, Ras-Mitogen Activated Protein Kinase (MAPK) and Notch (N) signaling pathways cross-communication. Our data show a repressive effect of Cno/AF-6 on these three signaling pathways through physical interactions with Ras, N and the cytoplasmic protein Dishevelled (Dsh), a key Wg effector. We propose a model in which Cno, through those interactions, actively coordinates, at the membrane level, Ras-MAPK, N and Wg signaling pathways during progenitor specification. PMID:17183697

  18. Hypoxia-NOTCH1-SOX2 signaling is important for maintaining cancer stem cells in ovarian cancer

    PubMed Central

    Jang, Il Ho; Choi, Eun Jung; Shin, Sang Hun; Lee, Su In; Kwon, Sang-Mo; Kim, Ki-Hyung; Suh, Dong-Soo; Kim, Jae Ho

    2016-01-01

    Hypoxia and NOTCH signaling have been reported to be associated with the self-renewal and drug resistance of cancer stem cells (CSCs). However, the molecular mechanisms by which hypoxia and NOTCH signaling stimulate the self-renewal and drug resistance of ovarian CSCs are poorly understood. In the present study, we identified SOX2 as a key transcription factor for CSC-like characteristics in the downstream of hypoxia-induced NOTCH signaling in epithelial ovarian cancer cells. Hypoxic treatment or overexpression of intracellular domain of NOTCH1 (NICD1) in ovarian cancer cells increased sphere formation, drug resistance, and expression of CSC-associated genes such as SOX2, ALDH, and ABC transporters. Hypoxic treatment increased the expression of NICD1, and hypoxic treatment or NICD1 overexpression increased SOX2 promoter activity, which was inhibited by deletion of HIF-1 or CSL binding sites. Furthermore, DAPT treatment decreased the effect of hypoxic treatment, and SOX2 knockdown decreased the effect of hypoxic treatment and NICD overexpression on sphere formation and drug resistance in established ovarian cancer cell lines and primary ovarian cancer cells. These results suggest that hypoxia-NOTCH1-SOX2 signaling axis is important for activation of ovarian CSCs, which may provide a novel opportunity for developing therapeutics to eradicate CSCs in ovarian cancer patients. PMID:27489349

  19. Postnatal dysregulation of Notch signal disrupts dendrite development of adult-born neurons in the hippocampus and contributes to memory impairment

    PubMed Central

    Ding, Xue-Feng; Gao, Xiang; Ding, Xin-Chun; Fan, Ming; Chen, Jinhui

    2016-01-01

    Deficits in the Notch pathway are involved in a number of neurologic diseases associated with mental retardation or/and dementia. The mechanisms by which Notch dysregulation are associated with mental retardation and dementia are poorly understood. We found that Notch1 is highly expressed in the adult-born immature neurons in the hippocampus of mice. Retrovirus mediated knockout of notch1 in single adult-born immature neurons decreases mTOR signaling and compromises their dendrite morphogenesis. In contrast, overexpression of Notch1 intracellular domain (NICD), to constitutively activate Notch signaling in single adult-born immature neurons, promotes mTOR signaling and increases their dendrite arborization. Using a unique genetic approach to conditionally and selectively knockout notch 1 in the postnatally born immature neurons in the hippocampus decreases mTOR signaling, compromises their dendrite morphogenesis, and impairs spatial learning and memory. Conditional overexpression of NICD in the postnatally born immature neurons in the hippocampus increases mTOR signaling and promotes dendrite arborization. These data indicate that Notch signaling plays a critical role in dendrite development of immature neurons in the postnatal brain, and dysregulation of Notch signaling in the postnatally born neurons disrupts their development and thus contributes to the cognitive deficits associated with neurological diseases. PMID:27173138

  20. A notch sweeter.

    PubMed

    Irvine, Kenneth D

    2008-01-25

    Notch is a key signaling protein mediating cell-fate decisions during development. In this issue, Acar et al. (2008) describe a new gene called rumi that is required for Notch signaling in Drosophila. This gene encodes an O-glucosyltransferase that attaches glucose sugars to serine residues in the multiple EGF domains of the extracellular region of Notch. This modification by Rumi likely influences Notch folding and trafficking.

  1. Oxidative stress and aberrant signaling in aging and cognitive decline

    PubMed Central

    Dröge, Wulf; Schipper, Hyman M

    2007-01-01

    Brain aging is associated with a progressive imbalance between antioxidant defenses and intracellular concentrations of reactive oxygen species (ROS) as exemplified by increases in products of lipid peroxidation, protein oxidation, and DNA oxidation. Oxidative conditions cause not only structural damage but also changes in the set points of redox-sensitive signaling processes including the insulin receptor signaling pathway. In the absence of insulin, the otherwise low insulin receptor signaling is strongly enhanced by oxidative conditions. Autophagic proteolysis and sirtuin activity, in turn, are downregulated by the insulin signaling pathway, and impaired autophagic activity has been associated with neurodegeneration. In genetic studies, impairment of insulin receptor signaling causes spectacular lifespan extension in nematodes, fruit flies, and mice. The predicted effects of age-related oxidative stress on sirtuins and autophagic activity and the corresponding effects of antioxidants remain to be tested experimentally. However, several correlates of aging have been shown to be ameliorated by antioxidants. Oxidative damage to mitochondrial DNA and the electron transport chain, perturbations in brain iron and calcium homeostasis, and changes in plasma cysteine homeostasis may altogether represent causes and consequences of increased oxidative stress. Aging and cognitive decline thus appear to involve changes at multiple nodes within a complex regulatory network. PMID:17517043

  2. Perineurial glia require Notch signaling during motor nerve development but not regeneration

    PubMed Central

    Binari, Laura A.; Lewis, Gwendolyn M.; Kucenas, Sarah

    2013-01-01

    Motor nerves play the critical role of shunting information out of the CNS to targets in the periphery. Their formation requires the coordinated development of distinct cellular components, including motor axons and the Schwann cells and perineurial glia that ensheath them. During nervous system assembly, these glial cells must migrate long distances and terminally differentiate, ensuring the efficient propagation of action potentials. Although we know quite a bit about the mechanisms that control Schwann cell development during this process, nothing is known about the mechanisms that mediate the migration and differentiation of perineurial glia. Using in vivo imaging in zebrafish, we demonstrate that Notch signaling is required for both perineurial migration and differentiation during nerve formation, but not regeneration. Interestingly, loss of Notch signaling in perineurial cells also causes a failure of Schwann cell differentiation, demonstrating that Schwann cells require perineurial glia for aspects of their own development. These studies describe a novel mechanism that mediates multiple aspects of perineurial development and reveal the critical importance of perineurial glia for Schwann cell maturation and nerve formation. PMID:23467342

  3. Dystroglycan is involved in skin morphogenesis downstream of the Notch signaling pathway

    PubMed Central

    Sirour, Cathy; Hidalgo, Magdalena; Bello, Valérie; Buisson, Nicolas; Darribère, Thierry; Moreau, Nicole

    2011-01-01

    Dystroglycan (Dg) is a transmembrane protein involved both in the assembly and maintenance of basement membrane structures essential for tissue morphogenesis, and the transmission of signals across the plasma membrane. We used a morpholino knockdown approach to investigate the function of Dg during Xenopus laevis skin morphogenesis. The loss of Dg disrupts epidermal differentiation by affecting the intercalation of multiciliated cells, deposition of laminin, and organization of fibronectin in the extracellular matrix (ECM). Depletion of Dg also affects cell–cell adhesion, as shown by the reduction of E-cadherin expression at the intercellular contacts, without affecting the distribution of β1 integrins. This was associated with a decrease of cell proliferation, a disruption of multiciliated-cell intercalation, and the down-regulation of the transcription factor P63, a marker of differentiated epidermis. In addition, we demonstrated that inhibition or activation of the Notch pathway prevents and promotes transcription of X-dg. Our study showed for the first time in vivo that Dg, in addition to organizing laminin in the ECM, also acts as a key signaling component in the Notch pathway. PMID:21680717

  4. Xanthohumol-Mediated Suppression of Notch1 Signaling Is Associated with Antitumor Activity in Human Pancreatic Cancer Cells.

    PubMed

    Kunnimalaiyaan, Selvi; Trevino, Jose; Tsai, Susan; Gamblin, T Clark; Kunnimalaiyaan, Muthusamy

    2015-06-01

    Pancreatic cancer remains a lethal disease with limited treatment options. At the time of diagnosis, approximately 80% of these patients present with unresectable tumors caused by either locally advanced lesions or progressive metastatic growth. Therefore, development of novel treatment strategies and new therapeutics is needed. Xanthohumol (XN) has emerged as a potential compound that inhibits various types of cancer, but the molecular mechanism underlying the effects of XN remains unclear. In the present study, we have assessed the efficacy of XN on pancreatic cancer cell lines (AsPC-1, PANC-1, L3.6pl, MiaPaCa-2, 512, and 651) against cell growth in real time and using colony-forming assays. Treatment with XN resulted in reduction in cellular proliferation in a dose- and time-dependent manner. The growth suppression effect of XN in pancreatic cancer cell lines is due to increased apoptosis via the inhibition of the Notch1 signaling pathway, as evidenced by reduction in Notch1, HES-1, and survivin both at mRNA as well as protein levels. Notch1 promoter reporter analysis after XN treatment indicated that XN downregulates Notch promoter activity. Importantly, overexpression of active Notch1 in XN-treated pancreatic cancer cells resulted in negation of growth suppression. Taken together, these findings demonstrate, for the first time, that the growth suppressive effect of XN in pancreatic cancer cells is mainly mediated by Notch1 reduction.

  5. Xanthohumol-mediated suppression of Notch1 signaling is associated with antitumor activity in human pancreatic cancer cells

    PubMed Central

    Kunnimalaiyaan, Selvi; Trevino, Jose; Tsai, Susan; Gamblin, T. Clark; Kunnimalaiyaan, Muthusamy

    2015-01-01

    Pancreatic cancer remains a lethal disease with limited treatment options. At the time of diagnosis, approximately 80% of these patients present with unresectable tumors caused by either locally advanced lesions or progressive metastatic growth. Therefore, development of novel treatment strategies and new therapeutics are needed. Xanthohumol (XN) has emerged as a potential compound that inhibits various types of cancer, but the molecular mechanism underlying the effects of XN remain unclear. In the present study, we have assessed the efficacy of XN on pancreatic cancer cell lines (AsPC-1, PANC-1, L3.6pl, MiaPaCa-2, 512, and 651) against cell growth in real time and using colony forming assays. Treatment with XN resulted in reduction in cellular proliferation in a dose and time dependent manner. The growth suppression effect of XN in pancreatic cancer cell lines is due to increased apoptosis via the inhibition of the Notch1 signaling pathway, as evidenced by reduction in Notch1, HES-1, and survivin both at mRNA as well as protein levels. Notch1 promoter reporter analysis after XN treatment indicated that XN down regulates Notch promoter activity. Importantly, overexpression of active Notch1 in XN-treated pancreatic cancer cells resulted in negation of growth suppression. Taken together, these findings demonstrate, for the first time, that the growth suppressive effect of XN in pancreatic cancer cells is mainly mediated by Notch1 reduction. PMID:25887885

  6. Emodin suppresses TGF-β1-induced epithelial-mesenchymal transition in alveolar epithelial cells through Notch signaling pathway.

    PubMed

    Gao, Rundi; Chen, Ruilin; Cao, Yu; Wang, Yuan; Song, Kang; Zhang, Ya; Yang, Junchao

    2017-03-01

    Pulmonary fibrosis is characterized by the destruction of lung tissue architecture and the formation of fibrous foci, currently has no satisfactory treatment. Emodin is a component of Chinese herb that has been reported to be medicament on pancreatic fibrosis and liver fibrosis. However, its role in pulmonary fibrosis has not been established yet. In the present study, we investigated the hypothesis that Emodin plays an inhibitory role in TGF-β1 induced epithelial-mesenchymal transition (EMT) of alveolar epithelial cell, and Emodin exerts its effect through the Notch signaling pathway. Emodin inhibits the proliferation of Rat alveolar type II epithelial cells RLE-6TN in a concentration-dependent manner; reduces the expression of Collagen I, α-SMA and Vimentin, promotes the expression of E-cadherin. Moreover, Emodin could regulate the expression patterns of the Notch signaling pathway-related factors and reduce the Notch-1 nucleus translocation. Knockdown of Notch-1 enhances the inhibitory effect of Emodin on TGF-β1-induced EMT in RLE-6TN cells. In conclusion, the data of the present study suggests that Emodin suppresses TGF-β1-induced EMT in alveolar epithelial cells through Notch signaling pathway and shows the potential to be effective in the treatment of pulmonary fibrosis.

  7. Acute Lymphoblastic Leukemia Cells Inhibit the Differentiation of Bone Mesenchymal Stem Cells into Osteoblasts In Vitro by Activating Notch Signaling

    PubMed Central

    Yang, Gui-Cun; Xu, You-Hua; Chen, Hong-Xia; Wang, Xiao-Jing

    2015-01-01

    The disruption of normal hematopoiesis has been observed in leukemia, but the mechanism is unclear. Osteoblasts originate from bone mesenchymal stem cells (BMSCs) and can maintain normal hematopoiesis. To investigate how leukemic cells inhibit the osteogenic differentiation of BMSCs and the role of Notch signaling in this process, we cocultured BMSCs with acute lymphoblastic leukemia (ALL) cells in osteogenic induction medium. The expression levels of Notch1, Hes1, and the osteogenic markers Runx2, Osteopontin (OPN), and Osteocalcin (OCN) were assessed by real-time RT-PCR and western blotting on day 3. Alkaline phosphatase (ALP) activity was analyzed using an ALP kit, and mineralization deposits were detected by Alizarin red S staining on day 14. And then we treated BMSCs with Jagged1 and anti-Jagged1 neutralizing Ab. The expression of Notch1, Hes1, and the abovementioned osteogenic differentiation markers was measured. Inhibition of the expression of Runx2, OPN, and OCN and reduction of ALP activity and mineralization deposits were observed in BMSCs cocultured with ALL cells, while Notch signal inhibiting rescued these effects. All these results indicated that ALL cells could inhibit the osteogenic differentiation of BMSCs by activating Notch signaling, resulting in a decreased number of osteoblastic cells, which may impair normal hematopoiesis. PMID:26339248

  8. Efficient differentiation of embryonic stem cells into mesodermal precursors by BMP, retinoic acid and Notch signalling.

    PubMed

    Torres, Josema; Prieto, Javier; Durupt, Fabrice C; Broad, Simon; Watt, Fiona M

    2012-01-01

    The ability to direct differentiation of mouse embryonic stem (ES) cells into specific lineages not only provides new insights into the pathways that regulate lineage selection but also has translational applications, for example in drug discovery. We set out to develop a method of differentiating ES cells into mesodermal cells at high efficiency without first having to induce embryoid body formation. ES cells were plated on a feeder layer of PA6 cells, which have membrane-associated stromal-derived inducing activity (SDIA), the molecular basis of which is currently unknown. Stimulation of ES/PA6 co-cultures with Bone Morphogenetic Protein 4 (BMP4) both favoured self-renewal of ES cells and induced differentiation into a Desmin and Nestin double positive cell population. Combined stimulation with BMP4 and all-trans Retinoic Acid (RA) inhibited self-renewal and resulted in 90% of cells expressing Desmin and Nestin. Quantitative reverse transcription-polymerase chain reaction (qPCR) analysis confirmed that the cells were of mesodermal origin and expressed markers of mesenchymal and smooth muscle cells. BMP4 activation of a MAD-homolog (Smad)-dependent reporter in undifferentiated ES cells was attenuated by co-stimulation with RA and co-culture with PA6 cells. The Notch ligand Jag1 was expressed in PA6 cells and inhibition of Notch signalling blocked the differentiation inducing activity of PA6 cells. Our data suggest that mesodermal differentiation is regulated by the level of Smad activity as a result of inputs from BMP4, RA and the Notch pathway.

  9. Efficient Differentiation of Embryonic Stem Cells into Mesodermal Precursors by BMP, Retinoic Acid and Notch Signalling

    PubMed Central

    Torres, Josema; Broad, Simon; Watt, Fiona M.

    2012-01-01

    The ability to direct differentiation of mouse embryonic stem (ES) cells into specific lineages not only provides new insights into the pathways that regulate lineage selection but also has translational applications, for example in drug discovery. We set out to develop a method of differentiating ES cells into mesodermal cells at high efficiency without first having to induce embryoid body formation. ES cells were plated on a feeder layer of PA6 cells, which have membrane-associated stromal-derived inducing activity (SDIA), the molecular basis of which is currently unknown. Stimulation of ES/PA6 co-cultures with Bone Morphogenetic Protein 4 (BMP4) both favoured self-renewal of ES cells and induced differentiation into a Desmin and Nestin double positive cell population. Combined stimulation with BMP4 and all-trans Retinoic Acid (RA) inhibited self-renewal and resulted in 90% of cells expressing Desmin and Nestin. Quantitative reverse transcription-polymerase chain reaction (qPCR) analysis confirmed that the cells were of mesodermal origin and expressed markers of mesenchymal and smooth muscle cells. BMP4 activation of a MAD-homolog (Smad)-dependent reporter in undifferentiated ES cells was attenuated by co-stimulation with RA and co-culture with PA6 cells. The Notch ligand Jag1 was expressed in PA6 cells and inhibition of Notch signalling blocked the differentiation inducing activity of PA6 cells. Our data suggest that mesodermal differentiation is regulated by the level of Smad activity as a result of inputs from BMP4, RA and the Notch pathway. PMID:22558462

  10. Convergence of a head-field selector Otx2 and Notch signaling: a mechanism for lens specification.

    PubMed

    Ogino, Hajime; Fisher, Marilyn; Grainger, Robert M

    2008-01-01

    Xenopus is ideal for systematic decoding of cis-regulatory networks because its evolutionary position among vertebrates allows one to combine comparative genomics with efficient transgenic technology in one system. Here, we have identified and analyzed the major enhancer of FoxE3 (Lens1), a gene essential for lens formation that is activated in the presumptive lens ectoderm (PLE) when commitment to the lens fate occurs. Deletion and mutation analyses of the enhancer based on comparison of Xenopus and mammalian sequences and in vitro and in vivo binding assays identified two essential transcriptional regulators: Otx2, a homeodomain protein expressed broadly in head ectoderm including the PLE, and Su(H), a nuclear signal transducer of Notch signaling. A Notch ligand, Delta2, is expressed in the optic vesicle adjacent to the PLE, and inhibition of its activity led to loss, or severe reduction, of FoxE3 expression followed by failure of placode formation. Ectopic activation of Notch signaling induced FoxE3 expression within head ectoderm expressing Otx2, and additional misexpression of Otx2 in trunk ectoderm extended the Notch-induced FoxE3 expression posteriorly. These data provide the first direct evidence of the involvement of Notch signaling in lens induction. The obligate integration of inputs of a field-selector (Otx2) and localized signaling (Notch) within target cis-regulatory elements might be a general mechanism of organ-field specification in vertebrates (as it is in Drosophila). This concept is also consistent with classical embryological studies of many organ systems involving a ;multiple-step induction'.

  11. Convergence of a head-field selector Otx2 and Notch signaling: a mechanism for lens specification

    PubMed Central

    Ogino, Hajime; Fisher, Marilyn; Grainger, Robert M.

    2014-01-01

    Summary Xenopus is ideal for systematic decoding of cis-regulatory networks because its evolutionary position among vertebrates allows one to combine comparative genomics with efficient transgenic technology in one system. Here we have identified and analyzed the major enhancer of FoxE3/Lens1, a gene essential for lens formation that is activated in the presumptive lens ectoderm (PLE) when commitment to the lens fate occurs. Deletion and mutation analyses of the enhancer based on comparison of Xenopus-mammalian sequences and in vitro and in vivo binding assays identified two essential transcriptional regulators; Otx2, a homeodomain protein expressed broadly in head ectoderm including the PLE, and Su(H), a nuclear signal transducer of Notch signaling. A Notch ligand, Delta2, is expressed in the optic vesicle adjacent to the PLE, and inhibition of its activity led to loss or severe reduction of FoxE3 expression followed by failure of placode formation. Ectopic activation of Notch signaling induced FoxE3 expression within head ectoderm expressing Otx2, and additional misexpression of Otx2 in trunk ectoderm extended the Notch-induced FoxE3 expression posteriorly. These data provide the first direct evidence of involvementof Notch signaling in lens induction. The obligate integration of inputs of a field-selector (Otx2) and localized signaling (Notch) within target cis-regulatory elements may be a general mechanism of organ-field specification in vertebrates (as in Drosophila). This concept is also consistent with classical embryological studies of many organ systems involving a “multiple-step induction”. PMID:18057103

  12. Notch signaling-mediated cell-to-cell interaction is dependent on E-cadherin adhesion in adult rat anterior pituitary.

    PubMed

    Batchuluun, Khongorzul; Azuma, Morio; Yashiro, Takashi; Kikuchi, Motoshi

    2017-04-01

    The rat anterior pituitary is composed of hormone-producing cells, non-hormone-producing cells (referred to as folliculostellate cells) and marginal layer cells. In the adult rat, progenitor cells of hormone-producing cells have recently been reported to be maintained within this non-hormone-producing cell population. In tissue, non-hormone-producing cells construct homophilic cell aggregates by the differential expression of the cell adhesion molecule E-cadherin. We have previously shown that Notch signaling, a known regulator of progenitor cells in a number of organs, is activated in the cell aggregates. We now investigate the relationship between Notch signaling and E-cadherin-mediated cell adhesion in the pituitary gland. Immunohistochemically, Notch signaling receptor Notch2 and the ligand Jagged1 were localized within E-cadherin-positive cells in the marginal cell layer and in the main part of the anterior lobe, whereas Notch1 was localized in E-cadherin-positive and -negative cells. Activation of Notch signaling within E-cadherin-positive cells was confirmed by immunostaining of the Notch target HES1. Notch2 and Jagged1 were always co-localized within the same cells suggesting that homologous cells have reciprocal effects in activating Notch signaling. When the E-cadherin function was inhibited by exposure to a monoclonal antibody (DECMA-1) in primary monolayer cell culture, the percentage of HES1-positive cells among Notch2-positive cells was less than half that of the control. The present results suggest that E-cadherin-mediated cell attachment is necessary for the activation of Notch signaling in the anterior pituitary gland but not for the expression of the Notch2 molecule.

  13. 17β-Estradiol Enhances Signalling Mediated by VEGF-A-Delta-Like Ligand 4-Notch1 Axis in Human Endothelial Cells

    PubMed Central

    Caliceti, Cristiana; Aquila, Giorgio; Pannella, Micaela; Morelli, Marco Bruno; Fortini, Cinzia; Pinton, Paolo; Bonora, Massimo; Hrelia, Silvana; Pannuti, Antonio; Miele, Lucio; Rizzo, Paola; Ferrari, Roberto

    2013-01-01

    Estrogens play a protective role in coronary artery disease. The mechanisms of action are still poorly understood, although a role for estrogens in stimulation of angiogenesis has been suggested. In several cell types, estrogens modulate the Notch pathway, which is involved in controlling angiogenesis downstream of vascular endothelial growth factor A (VEGF-A). The goal of our study was to establish whether estrogens modulate Notch activity in endothelial cells and the possible consequences on angiogenesis. Human umbilical vein endothelial cells (HUVECs) were treated with 17β-estradiol (E2) and the effects on Notch signalling were evaluated. E2 increased Notch1 processing as indicated by i) decreased levels of Notch1 transmembrane subunit ii) increased amount of Notch1 in nuclei iii) unaffected level of mRNA. Similarly, E2 increased the levels of the active form of Notch4 without altering Notch4 mRNA. Conversely, protein and mRNA levels of Notch2 were both reduced suggesting transcriptional repression of Notch2 by E2. Under conditions where Notch was activated by upregulation of Delta-like ligand 4 (Dll4) following VEGF-A treatment, E2 caused a further increase of the active form of Notch1, of the number of cells with nuclear Notch1 and of Hey2 mRNA. Estrogen receptor antagonist ICI 182.780 antagonized these effects suggesting that E2 modulation of Notch1 is mediated by estrogen receptors. E2 treatment abolished the increase in endothelial cells sprouting caused by Notch inhibition in a tube formation assay on 3D Matrigel and in mouse aortic ring explants. In conclusion, E2 affects several Notch pathway components in HUVECs, leading to an activation of the VEGF-A-Dll4-Notch1 axis and to a modulation of vascular branching when Notch signalling is inhibited. These results contribute to our understanding of the molecular mechanisms of cardiovascular protection exerted by estrogens by uncovering a novel role of E2 in the Notch signalling-mediated modulation of

  14. Emodin inhibits the proliferation of PC3 prostate cancer cells in vitro via the Notch signaling pathway.

    PubMed

    Deng, Gang; Ju, Xiang; Meng, Qi; Yu, Zhi-Jian; Ma, Li-Bin

    2015-09-01

    The aim of the present study was to examine the anticarcinogenic effects of emodin on the Notch signaling pathway and vascular endothelial growth factor (VEGF) in the PC3 androgen-independent prostate cancer cell line in vitro. The cell viability was assessed using an 3‑(4,5)‑dimethylthiahiazo(‑z‑y1)‑3,5‑di‑phenytetrazolium bromide assay. Cell apoptosis and cell cycle were detected using flow cytometry. Morphological alterations were observed using transmission electron microscopy. The mRNA and protein expression levels of Notch1, Jagged1, VEGF and bFGF were detected using reverse transcription‑quantitative polymerase chain reaction and western blotting, respectively. Laser scanning confocal microscope (LSCM) immunofluorescence analysis was performed to detect the levels of expression and the sub‑cellular localization of the Notch1 receptor protein. Growth of the PC3 cells was inhibited by emodin. Flow cytometry demonstrated that emodin induced apoptosis in the PC3 cells and arrested the cell cycle of the PC3 cells at the G2/M phase. The mRNA and protein expression of Notch1 in the PC3 cells was markedly increased, whereas the mRNA and protein expression levels of Jagged1, VEGF and bFGF were significantly decreased following exposure to emodin for 24 h. The LSCM assay revealed that the Notch1 was not only localized in the membrane and cytoplasm, but was also present in the nucleolus of the PC3 cells, and the expression of Notch1 in the nuclei gradually increased following treatment with emodin. These results demonstrated that emodin suppressed the growth of androgen‑independent prostate cancer cell lines and induced apoptosis and cell cycle arrest. The Notch signaling pathway was activated in the PC3 cells following exposure to emodin, which suggested that the Notch signaling pathway is involved in the prostate tumor‑suppressing mechanisms of emodin, the activation of which may depend on CBF1 protein in the nucleus by classic pathways. The

  15. Paradoxical effects of VEGF on synaptic activity partially involved in notch1 signaling in the mouse hippocampus.

    PubMed

    Yang, Jiajia; Yang, Chunxiao; Liu, Chunhua; Zhang, Tao; Yang, Zhuo

    2016-05-01

    It is well known that the neuronal effects of vascular endothelial growth factor (VEGF) include modulating learning and memory, plasticity of mature neurons, and synaptic transmission in addition to neurogenesis. However, there is conflicting evidence particularly of its role in the regulation of excitatory synaptic activity. In this study, application of the patch-clamp technique revealed that lower doses (10 and 50 ng/mL) of VEGF enhanced excitatory neurotransmission in hippocampal slices of mice through both presynaptic and postsynaptic mechanisms. However, the effects were reversed by higher doses of VEGF (>100 ng/mL), which inhibited excitatory neurotransmission via a presynaptic mechanism. These competing, concentration-dependent effects of VEGF suggested that different pathways were involved. The involvement of the Notch1 receptor was tested in the modulation of VEGF on synaptic activity by using heterozygous Notch1(+/-) mice. Notch1 knockdown did not influence the inhibitory effect of high VEGF doses (200 ng/mL) but reduced the enhancement effects of low concentration of VEGF (50 ng/mL) at the postsynaptic level, which might be due to the decreased level of VEGF receptor. The results indicate that the Notch1 receptor plays a role in VEGF-induced modulation of synaptic activity, which provides new insights into a complex VEGF/Notch signaling cross-talk. These findings set the groundwork for understanding new mechanisms of Notch signaling and the neurotrophic effects of VEGF, which is beneficial to develop new therapeutic targets to the VEGF/Notch axis and improve current treatments for neural diseases.

  16. Macrophage derived Wnt signalling opposes Notch signalling in a Numb mediated manner to specify HPC fate in chronic liver disease in human and mouse

    PubMed Central

    Boulter, Luke; Govaere, Olivier; Bird, Tom G; Radulescu, Sorina; Ramachandran, Prakash; Pellicoro, Antonella; Ridgway, Rachel A; Seo, Sang Soo; Spee, Bart; Van Rooijen, Nico; Sansom, Owen J.; Iredale, John P; Lowell, Sally; Roskams, Tania; Forbes, Stuart J

    2012-01-01

    During chronic injury, regeneration of the adult liver becomes impaired. In this context bipotent Hepatic Progenitor Cells (HPCs) become activated and can regenerate both cholangiocytes and hepatocytes. Notch and Wnt signalling during hepatic ontogeny are described, but their roles in HPC mediated liver regeneration are unclear. Here we show in human diseased liver and murine models of the ductular reaction with biliary and hepatocyte regeneration that Notch and Wnt signalling direct HPC specification within the activated myofibroblasts and macrophages HPC niche. During biliary regeneration, Numb is downregulated in HPCs, Jagged1 promotes biliary specification within HPCs. During hepatocyte regeneration, macrophage derived canonical Wnt signalling maintains Numb within HPCs, and Notch signalling is reduced promoting hepatocyte specification. This dominant Wnt state is stimulated through engulfment of hepatocyte debris by niche macrophages and can directly influence the HPCs. Macrophage Wnt3a expression in turn facilitates hepatocyte regeneration – thus exemplifying a novel positive feedback mechanism in adult parenchymal regeneration. PMID:22388089

  17. Serial specification of diverse neuroblast identities from a neurogenic placode by Notch and Egfr signaling

    PubMed Central

    Hwang, Helen J.; Rulifson, Eric

    2011-01-01

    We used the brain insulin-producing cell (IPC) lineage and its identified neuroblast (IPC NB) as a model to understand a novel example of serial specification of NB identities in the Drosophila dorsomedial protocerebral neuroectoderm. The IPC NB was specified from a small, molecularly identified group of cells comprising an invaginated epithelial placode. By progressive delamination of cells, the placode generated a series of NB identities, including the single IPC NB, a number of other canonical Type I NBs, and a single Type II NB that generates large lineages by transient amplification of neural progenitor cells. Loss of Notch function caused all cells of the placode to form as supernumerary IPC NBs, indicating that the placode is initially a fate equivalence group for the IPC NB fate. Loss of Egfr function caused all placodal cells to apoptose, except for the IPC NB, indicating a requirement of Egfr signaling for specification of alternative NB identities. Indeed, both derepressed Egfr activity in yan mutants and ectopic EGF activity produced supernumerary Type II NBs from the placode. Loss of both Notch and Egfr function caused all placode cells to become IPC NBs and survive, indicating that commitment to NB fate nullified the requirement of Egfr activity for placode cell survival. We discuss the surprising parallels between the serial specification of neural fates from this neurogenic placode and the fly retina. PMID:21653613

  18. Fine-tuning of Notch signaling sets the boundary of the organ of Corti and establishes sensory cell fates

    PubMed Central

    Basch, Martin L; Brown, Rogers M; Jen, Hsin-I; Semerci, Fatih; Depreux, Frederic; Edlund, Renée K; Zhang, Hongyuan; Norton, Christine R; Gridley, Thomas; Cole, Susan E; Doetzlhofer, Angelika; Maletic-Savatic, Mirjana; Segil, Neil; Groves, Andrew K

    2016-01-01

    The signals that induce the organ of Corti and define its boundaries in the cochlea are poorly understood. We show that two Notch modifiers, Lfng and Mfng, are transiently expressed precisely at the neural boundary of the organ of Corti. Cre-Lox fate mapping shows this region gives rise to inner hair cells and their associated inner phalangeal cells. Mutation of Lfng and Mfng disrupts this boundary, producing unexpected duplications of inner hair cells and inner phalangeal cells. This phenotype is mimicked by other mouse mutants or pharmacological treatments that lower but not abolish Notch signaling. However, strong disruption of Notch signaling causes a very different result, generating many ectopic hair cells at the expense of inner phalangeal cells. Our results show that Notch signaling is finely calibrated in the cochlea to produce precisely tuned levels of signaling that first set the boundary of the organ of Corti and later regulate hair cell development. DOI: http://dx.doi.org/10.7554/eLife.19921.001 PMID:27966429

  19. Notch-Jagged signalling can give rise to clusters of cells exhibiting a hybrid epithelial/mesenchymal phenotype

    PubMed Central

    Jolly, Mohit Kumar; Goldman, Aaron; Pietilä, Mika; Mani, Sendurai A.; Sengupta, Shiladitya; Ben-Jacob, Eshel; Levine, Herbert; Onuchic, Jose’ N.

    2016-01-01

    Metastasis can involve repeated cycles of epithelial-to-mesenchymal transition (EMT) and its reverse mesenchymal-to-epithelial transition. Cells can also undergo partial transitions to attain a hybrid epithelial/mesenchymal (E/M) phenotype that allows the migration of adhering cells to form a cluster of circulating tumour cells. These clusters can be apoptosis-resistant and possess an increased metastatic propensity as compared to the cells that undergo a complete EMT (mesenchymal cells). Hence, identifying the key players that can regulate the formation and maintenance of such clusters may inform anti-metastasis strategies. Here, we devise a mechanism-based theoretical model that links cell–cell communication via Notch-Delta-Jagged signalling with the regulation of EMT. We demonstrate that while both Notch-Delta and Notch-Jagged signalling can induce EMT in a population of cells, only Jagged-dominated Notch signalling, but not Delta-dominated signalling, can lead to the formation of clusters containing hybrid E/M cells. Our results offer possible mechanistic insights into the role of Jagged in tumour progression, and offer a framework to investigate the effects of other microenvironmental signals during metastasis. PMID:27170649

  20. Stem cell and lung cancer development: blaming the Wnt, Hh and Notch signalling pathway.

    PubMed

    García Campelo, María Rosario; Alonso Curbera, Guillermo; Aparicio Gallego, Guadalupe; Grande Pulido, Enrique; Antón Aparicio, Luis Miguel

    2011-02-01

    Primary lung cancer may arise from the central (bronchial) or peripheral (bronchiolo-alveolar) compartments. However the origins of the different histological types of primary lung cancer are not well understood. Stem cells are believed to be crucial players in tumour development and there is much interest in identifying those compartments that harbour stem cells involved in lung cancer. Although the role of stem cells in carcinogenesis is not well characterised, emerging evidence is providing new insights into this process. Numerous studies have indicated that lung cancer is not a result of a sudden transforming event but a multistep process in which a sequence of molecular changes result in genetic and morphological aberrations. The exact sequence of molecular events involved in lung carcinogenesis is not yet well understood, therefore deeper knowledge of the aberrant stem cell fate signalling pathway could be crucial in the development of new drugs against the advanced setting.

  1. Notch 1 and 3 receptor signaling modulates vascular smooth muscle cell growth, apoptosis, and migration via a CBF-1/RBP-Jk dependent pathway.

    PubMed

    Sweeney, Catherine; Morrow, David; Birney, Yvonne A; Coyle, Seamus; Hennessy, Colm; Scheller, Agnieszka; Cummins, Philip M; Walls, Dermot; Redmond, Eileen M; Cahill, Paul A

    2004-09-01

    Vascular smooth muscle cell (SMC) fate decisions (cell growth, migration, and apoptosis) are fundamental features in the pathogenesis of vascular disease. We investigated the role of Notch 1 and 3 receptor signaling in controlling adult SMC fate in vitro by establishing that hairy enhancer of split (hes-1 and -5) and related hrt's (hrt-1, -2, and -3) are direct downstream target genes of Notch 1 and 3 receptors in SMC and identified an essential role for nuclear protein CBF-1/RBP-Jk in their regulation. Constitutive expression of active Notch 1 and 3 receptors (Notch IC) resulted in a significant up-regulation of CBF-1/RBP-Jk-dependent promoter activity and Notch target gene expression concomitant with significant increases in SMC growth while concurrently inhibiting SMC apoptosis and migration. Moreover, inhibition of endogenous Notch mediated CBF-1/RBP-Jk regulated gene expression with a non-DNA binding mutant of CBF-1, a Notch IC deleted of its delta RAM domain and the Epstein-Barr virus encoded RPMS-1, in conjunction with pharmacological inhibitors of Notch IC receptor trafficking (brefeldin A and monensin), resulted in a significant decrease in cell growth while concomitantly increasing SMC apoptosis and migration. These findings suggest that endogenous Notch receptors and downstream target genes control vascular cell fate in vitro. Notch signaling, therefore, represents a novel therapeutic target for disease states in which changes in vascular cell fate occur in vivo.

  2. Involvement of aberrant calcium signalling in herpetic neuralgia.

    PubMed

    Warwick, Rebekah A; Hanani, Menachem

    2016-03-01

    Alpha-herpesviruses, herpes simplex viruses (HSV) and varicella zoster virus (VZV), are pathogens of the peripheral nervous system. After primary infection, these viruses establish latency within sensory ganglia, while retaining the ability to reactivate. Reactivation of VZV results in herpes zoster, a condition characterized by skin lesions that leads to post-herpetic neuralgia. Recurrent reactivations of HSV, which cause mucocutaneous lesions, may also result in neuralgia. During reactivation of alpha-herpesviruses, satellite glial cells (SGCs), which surround neurons in sensory ganglia, become infected with the replicating virus. SGCs are known to contribute to neuropathic pain in a variety of animal pain models. Here we investigated how infection of short-term cultures of mouse trigeminal ganglia with HSV-1 affects communication between SGCs and neurons, and how this altered communication may increase neuronal excitability, thus contributing to herpetic neuralgia. Mechanical stimulation of single neurons or SGCs resulted in intercellular calcium waves, which were larger in cultures infected with HSV-1. Two differences were observed between control and HSV-1 infected cultures that could account for this augmentation. Firstly, HSV-1 infection induced cell fusion among SGCs and neurons, which would facilitate the spread of calcium signals over farther distances. Secondly, using calcium imaging and intracellular electrical recordings, we found that neurons in the HSV-1 infected cultures exhibited augmented influx of calcium upon depolarization. These virally induced changes may not only cause more neurons in the sensory ganglia to fire action potentials, but may also increase neurotransmitter release at the presynaptic terminals in the spinal cord. They are therefore likely to be contributing factors to herpetic neuralgia.

  3. Activated Notch signaling is required for hepatitis B virus X protein to promote proliferation and survival of human hepatic cells.

    PubMed

    Wang, Fan; Zhou, Haiyan; Xia, Xiumei; Sun, Qian; Wang, Ying; Cheng, Bin

    2010-12-01

    Hepatitis B virus X protein (HBx) is a multifunctional oncoprotein which plays a crucial role in the pathogenesis of hepatocellular carcinoma (HCC). However, the exact mechanisms remain controversial. Here we show that HBx strongly stimulated cell growth, promoted cell cycle progression and inhibited apoptosis of human non-tumor hepatic cell line L02 cells. It also accelerated tumor formation of L02 cells in BALB/c nude mice. Furthermore, Notch signaling components were upregulated in HBx-expressing L02 cells compared to normal L02 cells. However, blocking Notch signaling with a γ-secretase inhibitor N-[N-(3,5-difluorophenacetyl)-l-alanyl]-S-phenylglycine t-butyl ester (DAPT) attenuated cell growth, shortened the S phase of cell cycle and promoted apoptosis of HBx-expressing L02 cell in a dose- and time-dependent manner, but normal L02 cells were not significantly affected by Notch signaling blocking. Therefore, our findings demonstrate that HBx could promote the growth of human non-tumor hepatic cell line L02 cells both in vitro and in vivo, which may require the activation of Notch signaling pathway.

  4. γ-Secretase inhibitor DAPT attenuates intimal hyperplasia of vein grafts by inhibition of Notch1 signaling.

    PubMed

    Xiao, Yong Guang; Wang, Wei; Gong, Dan; Mao, Zhi Fu

    2014-06-01

    The proliferation and high plasticity of vascular smooth muscle cells (vSMCs) are the major reasons for restenosis of vein grafts. N-[N-(3, 5-difluorophenacetyl)-l-alanyl]-S-phenylglycine t-butyl ester (DAPT), specific inhibitor of γ-secretase, has been shown to regulate vSMC proliferation and differentiation through the Notch signaling pathway, but the pathophysiological importance of these findings in venous grafts has not yet been determined. A rat vein graft model was employed wherein the left jugular vein was surgically interposed into the left common carotid artery. Daily subcutaneous injections of DAPT or placebo (DMSO) were administered postoperatively (control animals received no treatment). We showed that DAPT can inhibit restenosis of vein grafts by inhibiting vSMC proliferation and increasing apoptosis in vivo. Notch1 signaling was highly active during the development of intima thickening. By blocking the Notch signaling pathway, the γ-secretase inhibitor DAPT can significantly attenuated intima thickening. These changes in vein grafts coincided with enhanced binding of myocardin to the smooth muscle-specific protein SM22 and smooth muscle myosin heavy chain at the promoters of vSMC differentiation-specific genes. These studies showed that DAPT can restore the vSMC phenotype and inhibit vSMC proliferation through suppression of the Notch1 signaling pathway, and thus opens a new avenue for the treatment of restenosis in vein grafts.

  5. Top Notch cancer stem cells by paracrine NF-κB signaling in breast cancer.

    PubMed

    Zhang, Weizhou; Grivennikov, Sergei I

    2013-01-01

    Cancer stem cells are likely to play critical roles in metastasis, therapy resistance, and recurrence of hematological and solid malignancies. It is well known that the stem cell niche plays a key role for asymmetric division and homeostasis of normal stem cells, whereas cancer stem cells seem to use these niches. Among many pathways involved in self-renewal of cancer stem cells, nuclear factor-kappa B (NF-κB) signaling has been documented to promote their expansion in a cell-autonomous fashion. A recent study, however, suggests that paracrine NF-κB activation promotes the expansion of cancer stem cells through the activation of Notch in basal-type breast cancer cells.

  6. GDE2 regulates subtype-specific motor neuron generation through inhibition of Notch signaling.

    PubMed

    Sabharwal, Priyanka; Lee, Changhee; Park, Sungjin; Rao, Meenakshi; Sockanathan, Shanthini

    2011-09-22

    The specification of spinal interneuron and motor neuron identities initiates within progenitor cells, while motor neuron subtype diversification is regulated by hierarchical transcriptional programs implemented postmitotically. Here we find that mice lacking GDE2, a six-transmembrane protein that triggers motor neuron generation, exhibit selective losses of distinct motor neuron subtypes, specifically in defined subsets of limb-innervating motor pools that correlate with the loss of force-generating alpha motor neurons. Mechanistically, GDE2 is expressed by postmitotic motor neurons but utilizes extracellular glycerophosphodiester phosphodiesterase activity to induce motor neuron generation by inhibiting Notch signaling in neighboring motor neuron progenitors. Thus, neuronal GDE2 controls motor neuron subtype diversity through a non-cell-autonomous feedback mechanism that directly regulates progenitor cell differentiation, implying that subtype specification initiates within motor neuron progenitor populations prior to their differentiation into postmitotic motor neurons.

  7. Murine Jagged1/Notch signaling in the second heart field orchestrates Fgf8 expression and tissue-tissue interactions during outflow tract development

    PubMed Central

    High, Frances A.; Jain, Rajan; Stoller, Jason Z.; Antonucci, Nicole B.; Lu, Min Min; Loomes, Kathleen M.; Kaestner, Klaus H.; Pear, Warren S.; Epstein, Jonathan A.

    2009-01-01

    Notch signaling is vital for proper cardiovascular development and function in both humans and animal models. Indeed, mutations in either JAGGED or NOTCH cause congenital heart disease in humans and NOTCH mutations are associated with adult valvular disease. Notch typically functions to mediate developmental interactions between adjacent tissues. Here we show that either absence of the Notch ligand Jagged1 or inhibition of Notch signaling in second heart field tissues results in murine aortic arch artery and cardiac anomalies. In mid-gestation, these mutants displayed decreased Fgf8 and Bmp4 expression. Notch inhibition within the second heart field affected the development of neighboring tissues. For example, faulty migration of cardiac neural crest cells and defective endothelial-mesenchymal transition within the outflow tract endocardial cushions were observed. Furthermore, exogenous Fgf8 was sufficient to rescue the defect in endothelial-mesenchymal transition in explant assays of endocardial cushions following Notch inhibition within second heart field derivatives. These data support a model that relates second heart field, neural crest, and endocardial cushion development and suggests that perturbed Notch-Jagged signaling within second heart field progenitors accounts for some forms of congenital and adult cardiac disease. PMID:19509466

  8. Notch4-induced inhibition of endothelial sprouting requires the ankyrin repeats and involves signaling through RBP-Jkappa.

    PubMed

    MacKenzie, Farrell; Duriez, Patrick; Larrivée, Bruno; Chang, Linda; Pollet, Ingrid; Wong, Fred; Yip, Calvin; Karsan, Aly

    2004-09-15

    Notch proteins comprise a family of transmembrane receptors. Ligand activation of Notch releases the intracellular domain of the receptor that translocates to the nucleus and regulates transcription through the DNA-binding protein RBP-Jkappa. Previously, it has been shown that the Notch4 intracellular region (N4IC) can inhibit endothelial sprouting and angiogenesis. Here, N4IC deletion mutants were assessed for their ability to inhibit human microvascular endothelial cell (HMEC) sprouting with the use of a quantitative endothelial sprouting assay. Deletion of the ankyrin repeats, but not the RAM (RBP-Jkappa associated module) domain or C-terminal region (CT), abrogated the inhibition of fibroblast growth factor 2 (FGF-2)- and vascular endothelial growth factor (VEGF)-induced sprouting by Notch4, whereas the ankyrin repeats alone partially blocked sprouting. The ankyrin repeats were also the only domain required for up-regulation of RBP-Jkappa-dependent gene expression. Interestingly, enforced expression of the ankyrin domain alone was sufficient to up-regulate some, but not all, RBP-Jkappa-dependent genes. Although N4IC reduced VEGF receptor-2 (VEGFR-2) and vascular endothelial (VE)-cadherin expression, neither of these events is necessary and sufficient to explain N4IC-mediated inhibition of sprouting. A constitutively active RBP-Jkappa mutant significantly inhibited HMEC sprouting but not as strongly as N4IC. Thus, Notch4-induced inhibition of sprouting requires the ankyrin repeats and appears to involve RBP-Jkappa-dependent and -independent signaling.

  9. Overexpression of miR-34c inhibits high glucose-induced apoptosis in podocytes by targeting Notch signaling pathways.

    PubMed

    Liu, Xiang-Dong; Zhang, Lian-Yun; Zhu, Tie-Chui; Zhang, Rui-Fang; Wang, Shu-Long; Bao, Yan

    2015-01-01

    Recent findings have shown that microRNAs play critical roles in the pathogenesis of diabetic nephropathy. miR-34c has been found to inhibit fibrosis and the epithelial-mesenchymal transition of kidney cells. However, the role of miR-34c in diabetic nephropathy has not been well studied. The current study was designed to investigate the role and potential underlying mechanism of miR-34c in regulating diabetic nephropathy. After treating podocytes with high glucose (HG) in vitro, we found that miR-34c was downregulated and that overexpression of miR-34c inhibited HG-induced podocyte apoptosis. The direct interaction between miR-34c and the 3'-untranslated region (UTR) of Notch1 and Jagged1 was validated by dual-luciferase reporter assay. Moreover, Notch1 and Jagged1 as putative targets of miR-34c were downregulated by miR-34c overexpression in HG-treated podocytes. Overexpression of miR-34c inhibited HG-induced Notch signaling pathway activation, as indicated by decreased expression of the Notch intracellular domain (NICD) and downstream genes including Hes1 and Hey1. Furthermore, miR-34c overexpression increased the expression of the anti-apoptotic gene Bcl-2, and decreased the expression of the pro-apoptotic protein Bax and cleaved Caspase-3. Additionally, the phosphorylation of p53 was also downregulated by miR-34c overexpression. Taken together, our findings suggest that miR-34c overexpression inhibits the Notch signaling pathway by targeting Notch1 and Jaggged1 in HG-treated podocytes, representing a novel and potential therapeutic target for the treatment of diabetic nephropathy.

  10. Lineage-specific effects of Notch/Numb signaling in post-embryonic development of the Drosophila brain.

    PubMed

    Lin, Suewei; Lai, Sen-Lin; Yu, Huang-Hsiang; Chihara, Takahiro; Luo, Liqun; Lee, Tzumin

    2010-01-01

    Numb can antagonize Notch signaling to diversify the fates of sister cells. We report here that paired sister cells acquire different fates in all three Drosophila neuronal lineages that make diverse types of antennal lobe projection neurons (PNs). Only one in each pair of postmitotic neurons survives into the adult stage in both anterodorsal (ad) and ventral (v) PN lineages. Notably, Notch signaling specifies the PN fate in the vPN lineage but promotes programmed cell death in the missing siblings in the adPN lineage. In addition, Notch/Numb-mediated binary sibling fates underlie the production of PNs and local interneurons from common precursors in the lAL lineage. Furthermore, Numb is needed in the lateral but not adPN or vPN lineages to prevent the appearance of ectopic neuroblasts and to ensure proper self-renewal of neural progenitors. These lineage-specific outputs of Notch/Numb signaling show that a universal mechanism of binary fate decision can be utilized to govern diverse neural sibling differentiations.

  11. Inhibition of Notch signaling reduces the number of surviving Dclk1+ reserve crypt epithelial stem cells following radiation injury

    PubMed Central

    Qu, Dongfeng; May, Randal; Sureban, Sripathi M.; Weygant, Nathaniel; Chandrakesan, Parthasarathy; Ali, Naushad; Li, Linheng; Barrett, Terrence

    2013-01-01

    We have previously reported that doublecortin-like kinase 1 (Dclk1) is a putative intestinal stem cell (ISC) marker. In this report, we evaluated the use of Dclk1 as a marker of surviving ISCs in response to treatment with high-dose total body irradiation (TBI). Both apoptotic and mitotic Dclk1+ cells were observed 24 h post-TBI associated with a corresponding loss of intestinal crypts observed at 84 h post-TBI. Although the Notch signaling pathway plays an important role in regulating proliferation and lineage commitment within the intestine, its role in ISC function in response to severe genotoxic injury is not yet fully understood. We employed the microcolony assay to functionally assess the effects of Notch inhibition with difluorophenacetyl-l-alanyl-S-phenylglycine t-butyl ester (DAPT) on intestinal crypt stem cell survival following severe (>8 Gy) radiation injury. Following treatment with DAPT, we observed a nearly 50% reduction in the number of surviving Dclk1+ crypt epithelial cells at 24 h after TBI and similar reduction in the number of surviving small intestinal crypts at 84 h. These data indicate that inhibition of Notch signaling decreases ISC survival following radiation injury, suggesting that the Notch signaling pathway plays an important role in ISC-mediated crypt regeneration. These results also suggest that crypt epithelial cell Dclk1 expression can be used as one potential marker to evaluate the early survival of ISCs following severe radiation injury. PMID:24368703

  12. The Parkinson’s Disease-Associated Protein Kinase LRRK2 Modulates Notch Signaling through the Endosomal Pathway

    PubMed Central

    Imai, Yuzuru; Kobayashi, Yoshito; Inoshita, Tsuyoshi; Meng, Hongrui; Arano, Taku; Uemura, Kengo; Asano, Takeshi; Yoshimi, Kenji; Zhang, Chang-Liang; Matsumoto, Gen; Ohtsuka, Toshiyuki; Kageyama, Ryoichiro; Kiyonari, Hiroshi; Shioi, Go; Nukina, Nobuyuki; Hattori, Nobutaka; Takahashi, Ryosuke

    2015-01-01

    Leucine-rich repeat kinase 2 (LRRK2) is a key molecule in the pathogenesis of familial and idiopathic Parkinson’s disease (PD). We have identified two novel LRRK2-associated proteins, a HECT-type ubiquitin ligase, HERC2, and an adaptor-like protein with six repeated Neuralized domains, NEURL4. LRRK2 binds to NEURL4 and HERC2 via the LRRK2 Ras of complex proteins (ROC) domain and NEURL4, respectively. HERC2 and NEURL4 link LRRK2 to the cellular vesicle transport pathway and Notch signaling, through which the LRRK2 complex promotes the recycling of the Notch ligand Delta-like 1 (Dll1)/Delta (Dl) through the modulation of endosomal trafficking. This process negatively regulates Notch signaling through cis-inhibition by stabilizing Dll1/Dl, which accelerates neural stem cell differentiation and modulates the function and survival of differentiated dopaminergic neurons. These effects are strengthened by the R1441G ROC domain-mutant of LRRK2. These findings suggest that the alteration of Notch signaling in mature neurons is a component of PD etiology linked to LRRK2. PMID:26355680

  13. Latency-associated nuclear antigen of Kaposi sarcoma-associated herpesvirus promotes angiogenesis through targeting notch signaling effector Hey1.

    PubMed

    Wang, Xing; He, Zhiheng; Xia, Tian; Li, Xiaofan; Liang, Deguang; Lin, Xianzhi; Wen, Hao; Lan, Ke

    2014-04-01

    Notch signaling has been implicated in the pathogenesis of Kaposi sarcoma. Kaposi sarcoma is an angioproliferative neoplasm that originates from Kaposi sarcoma-associated herpesvirus (KSHV) infection. Previously, we showed that the KSHV LANA protein can stabilize intracellular Notch in KSHV-infected tumor cells and promote cell proliferation. However, whether Notch signaling functions in pathologic angiogenesis of Kaposi sarcoma remains largely unknown. Hey1, an essential downstream effector of the Notch signaling pathway, has been demonstrated to play a fundamental role in vascular development. In the present study, we performed whole transcriptome, paired-end sequencing on three patient-matched clinical Kaposi sarcoma specimens and their corresponding adjacent stroma samples, with an average depth of 42 million reads per sample. Dll4, Hey1, and HeyL displayed significant upregulation in Kaposi sarcoma. Further verification based on immunohistochemistry analysis demonstrated that Hey1 was indeed highly expressed in Kaposi sarcoma lesions. Using the Matrigel plug assay, we showed that downregulation of Hey1 and γ-secretase inhibitor treatment caused dramatic reduction in the formation of new blood vessels in mice. Interestingly, LANA was responsible for the elevated level of Hey1 through inhibition of its degradation. Importantly, Hey1 stabilized by LANA promoted the neoplastic vasculature. Taken together, our data suggest that hijacking of the proangiogenic property of Hey1 by LANA is an important strategy utilized by KSHV to achieve pathologic angiogenesis and that Hey1 is a potential therapeutic target in Kaposi sarcoma.

  14. Modulating of ocular inflammation with macrophage migration inhibitory factor is associated with notch signalling in experimental autoimmune uveitis.

    PubMed

    Yang, H; Zheng, S; Mao, Y; Chen, Z; Zheng, C; Li, H; Sumners, C; Li, Q; Yang, P; Lei, B

    2016-02-01

    The aim of this study was to examine whether macrophage migration inhibitory factor (MIF) could exaggerate inflammatory response in a mouse model of experimental autoimmune uveitis (EAU) and to explore the underlying mechanism. Mutant serotype 8 adeno-associated virus (AAV8) (Y733F)-chicken β-actin (CBA)-MIF or AAV8 (Y733F)-CBA-enhanced green fluorescent protein (eGFP) vector was delivered subretinally into B10.RIII mice, respectively. Three weeks after vector delivery, EAU was induced with a subcutaneous injection of a mixture of interphotoreceptor retinoid binding protein (IRBP) peptide with CFA. The levels of proinflammatory cytokines were detected by real-time polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA). Retinal function was evaluated with electroretinography (ERG). We found that the expression of MIF and its two receptors CD74 and CD44 was increased in the EAU mouse retina. Compared to AAV8.CBA.eGFP-injected and untreated EAU mice, the level of proinflammatory cytokines, the expression of Notch1, Notch4, delta-like ligand 4 (Dll4), Notch receptor intracellular domain (NICD) and hairy enhancer of split-1 (Hes-1) increased, but the ERG a- and b-wave amplitudes decreased in AAV8.CBA.MIF-injected EAU mice. The Notch inhibitor N-[N-(3,5-difluorophenacetyl)-l-alanyl]-S-phenylglycine t-butyl ester (DAPT) reduced the expression of NICD, Hes-1 and proinflammatory cytokines. Further, a MIF antagonist ISO-1 attenuated intraocular inflammation, and inhibited the differentiation of T helper type 1 (Th1) and Th17 in EAU mice. We demonstrated that over-expression of MIF exaggerated ocular inflammation, which was associated with the activation of the Notch signalling. The expression of both MIF and its receptors are elevated in EAU mice. Over-expression of MIF exaggerates ocular inflammation, and this exaggerated inflammation is associated with the activation of the Notch signalling and Notch pathway. Our data suggest that the MIF-Notch axis

  15. AP-1 clathrin adaptor and CG8538/Aftiphilin are involved in Notch signaling during eye development in Drosophila melanogaster.

    PubMed

    Kametaka, Satoshi; Kametaka, Ai; Yonekura, Shinichi; Haruta, Mineyuki; Takenoshita, Seiichi; Goto, Satoshi; Waguri, Satoshi

    2012-02-01

    Clathrin adaptor protein complex-1 (AP-1) and its accessory proteins play a role in the sorting of integral membrane proteins at the trans-Golgi network and endosomes. Their physiological functions in complex organisms, however, are not fully understood. In this study, we found that CG8538p, an uncharacterized Drosophila protein, shares significant structural and functional characteristics with Aftiphilin, a mammalian AP-1 accessory protein. The Drosophila Aftiphilin was shown to interact directly with the ear domain of γ-adaptin of Drosophila AP-1, but not with the GAE domain of Drosophila GGA. In S2 cells, Drosophila Aftiphilin and AP-1 formed a complex and colocalized at the Golgi compartment. Moreover, tissue-specific depletion of AP-1 or Aftiphilin in the developing eyes resulted in a disordered alignment of photoreceptor neurons in larval stage and roughened eyes with aberrant ommatidia in adult flies. Furthermore, AP-1-depleted photoreceptor neurons showed an intracellular accumulation of a Notch regulator, Scabrous, and downregulation of Notch by promoting its degradation in the lysosomes. These results suggest that AP-1 and Aftiphilin are cooperatively involved in the intracellular trafficking of Notch during eye development in Drosophila.

  16. Notch signaling regulates tumor-induced angiogenesis in SPARC-overexpressed neuroblastoma

    PubMed Central

    Gorantla, Bharathi; Bhoopathi, Praveen; Chetty, Chandramu; Gogineni, Venkateswara Rao; Sailaja, GS; Gondi, Christopher S.; Rao, Jasti S.

    2012-01-01

    Despite existing aggressive treatment modalities, the prognosis for advanced stage neuroblastoma remains poor with significant long-term illness in disease survivors. Advance stage disease features are associated with tumor vascularity, and as such, angiogenesis inhibitors may prove useful along with current therapies. The matricellular protein, secreted protein acidic and rich in cysteine (SPARC), is known to inhibit proliferation and migration of endothelial cells stimulated by growth factors. Here, we sought to determine the effect of SPARC on neuroblastoma tumor cell-induced angiogenesis and to decipher the molecular mechanisms involved in angiogenesis inhibition. Conditioned medium from SPARC-overexpressed neuroblastoma cells (pSPARC-CM) inhibited endothelial tube formation, cell proliferation, induced programmed cell death and suppressed expression of pro-angiogenic molecules such as VEGF, FGF, PDGF, and MMP-9 in endothelial cells. Further analyses revealed that pSPARC-CM-suppressed expression of growth factors was mediated by inhibition of the Notch signaling pathway, and cells cultured on conditioned medium from tumor cells that overexpress both Notch intracellular domain (NICD-CM) and SPARC resumed the pSPARC-CM-suppressed capillary tube formation and growth factor expression in vitro. Further, SPARC overexpression in neuroblastoma cells inhibited neo-vascularization in vivo in a mouse dorsal air sac model. Furthermore, SPARC overexpression-induced endothelial cell death was observed by co-localization studies with TUNEL assay and an endothelial marker, CD31, in xenograft tumor sections from SPARC-overexpressed mice. Our data collectively suggest that SPARC overexpression induces endothelial cell apoptosis and inhibits angiogenesis both in vitro and in vivo. PMID:22956186

  17. Embryonic Ethanol Exposure Dysregulates BMP and Notch Signaling, Leading to Persistent Atrio-Ventricular Valve Defects in Zebrafish.

    PubMed

    Sarmah, Swapnalee; Muralidharan, Pooja; Marrs, James A

    2016-01-01

    Fetal alcohol spectrum disorder (FASD), birth defects associated with ethanol exposure in utero, includes a wide spectrum of congenital heart defects (CHDs), the most prevalent of which are septal and conotruncal defects. Zebrafish FASD model was used to dissect the mechanisms underlying FASD-associated CHDs. Embryonic ethanol exposure (3-24 hours post fertilization) led to defects in atrio-ventricular (AV) valvulogenesis beginning around 37 hpf, a morphogenetic event that arises long after ethanol withdrawal. Valve leaflets of the control embryos comprised two layers of cells confined at the compact atrio-ventricular canal (AVC). Ethanol treated embryos had extended AVC and valve forming cells were found either as rows of cells spanning the AVC or as unorganized clusters near the AV boundary. Ethanol exposure reduced valve precursors at the AVC, but some ventricular cells in ethanol treated embryos exhibited few characteristics of valve precursors. Late staged larvae and juvenile fish exposed to ethanol during embryonic development had faulty AV valves. Examination of AVC morphogenesis regulatory networks revealed that early ethanol exposure disrupted the Bmp signaling gradient in the heart during valve formation. Bmp signaling was prominent at the AVC in controls, but ethanol-exposed embryos displayed active Bmp signaling throughout the ventricle. Ethanol exposure also led to mislocalization of Notch signaling cells in endocardium during AV valve formation. Normally, highly active Notch signaling cells were organized at the AVC. In ethanol-exposed embryos, highly active Notch signaling cells were dispersed throughout the ventricle. At later stages, ethanol-exposed embryos exhibited reduced Wnt/β-catenin activity at the AVC. We conclude that early embryonic ethanol exposure alters Bmp, Notch and other signaling activities during AVC differentiation leading to faulty valve morphogenesis and valve defects persist in juvenile fish.

  18. Embryonic Ethanol Exposure Dysregulates BMP and Notch Signaling, Leading to Persistent Atrio-Ventricular Valve Defects in Zebrafish

    PubMed Central

    Sarmah, Swapnalee; Muralidharan, Pooja

    2016-01-01

    Fetal alcohol spectrum disorder (FASD), birth defects associated with ethanol exposure in utero, includes a wide spectrum of congenital heart defects (CHDs), the most prevalent of which are septal and conotruncal defects. Zebrafish FASD model was used to dissect the mechanisms underlying FASD-associated CHDs. Embryonic ethanol exposure (3–24 hours post fertilization) led to defects in atrio-ventricular (AV) valvulogenesis beginning around 37 hpf, a morphogenetic event that arises long after ethanol withdrawal. Valve leaflets of the control embryos comprised two layers of cells confined at the compact atrio-ventricular canal (AVC). Ethanol treated embryos had extended AVC and valve forming cells were found either as rows of cells spanning the AVC or as unorganized clusters near the AV boundary. Ethanol exposure reduced valve precursors at the AVC, but some ventricular cells in ethanol treated embryos exhibited few characteristics of valve precursors. Late staged larvae and juvenile fish exposed to ethanol during embryonic development had faulty AV valves. Examination of AVC morphogenesis regulatory networks revealed that early ethanol exposure disrupted the Bmp signaling gradient in the heart during valve formation. Bmp signaling was prominent at the AVC in controls, but ethanol-exposed embryos displayed active Bmp signaling throughout the ventricle. Ethanol exposure also led to mislocalization of Notch signaling cells in endocardium during AV valve formation. Normally, highly active Notch signaling cells were organized at the AVC. In ethanol-exposed embryos, highly active Notch signaling cells were dispersed throughout the ventricle. At later stages, ethanol-exposed embryos exhibited reduced Wnt/β-catenin activity at the AVC. We conclude that early embryonic ethanol exposure alters Bmp, Notch and other signaling activities during AVC differentiation leading to faulty valve morphogenesis and valve defects persist in juvenile fish. PMID:27556898

  19. Cyclic AMP Response Element Binding Protein Mediates Pathological Retinal Neovascularization via Modulating DLL4-NOTCH1 Signaling

    PubMed Central

    Singh, Nikhlesh K.; Kotla, Sivareddy; Kumar, Raj; Rao, Gadiparthi N.

    2015-01-01

    Retinal neovascularization is the most common cause of moderate to severe vision loss in all age groups. Despite the use of anti-VEGFA therapies, this complication continues to cause blindness, suggesting a role for additional molecules in retinal neovascularization. Besides VEGFA and VEGFB, hypoxia induced VEGFC expression robustly. Based on this finding, we tested the role of VEGFC in pathological retinal angiogenesis. VEGFC induced proliferation, migration, sprouting and tube formation of human retinal microvascular endothelial cells (HRMVECs) and these responses require CREB-mediated DLL4 expression and NOTCH1 activation. Furthermore, down regulation of VEGFC levels substantially reduced tip cell formation and retinal neovascularization in vivo. In addition, we observed that CREB via modulating the DLL4-NOTCH1 signaling mediates VEGFC-induced tip cell formation and retinal neovascularization. In regard to upstream mechanism, we found that down regulation of p38β levels inhibited hypoxia-induced CREB-DLL4-NOTCH1 activation, tip cell formation, sprouting and retinal neovascularization. Based on these findings, it may be suggested that VEGFC besides its role in the regulation of lymphangiogenesis also plays a role in pathological retinal angiogenesis and this effect depends on p38β and CREB-mediated activation of DLL4-NOTCH1 signaling. PMID:26870802

  20. Notch1 promotes vasculogenic mimicry in hepatocellular carcinoma by inducing EMT signaling.

    PubMed

    Jue, Chen; Lin, Cui; Zhisheng, Zhang; Yayun, Qian; Feng, Jin; Min, Zhao; Haibo, Wang; Youyang, Shi; Hisamitsu, Tadashi; Shintaro, Ishikawa; Shiyu, Guo; Yanqing, Liu

    2017-01-10

    Hypervascularity is one of the main characteristics of hepatocellular carcinoma (HCC). However, the mechanisms of angiogenesis in HCC remain controversial. In this study, we investigate the role of Notch1 in angiogenesis of HCC. We found that Notch1 expression was correlated with formation of vasculogenic mimicry (VM) and expression of biomarkers of epithelial-to-mesenchymal transition (EMT) in the tumor specimens. Two HCC cell lines, HepG2 and MHCC97-H, with low and high Notch1 expression, respectively, were used to study the mechanism of VM formation both in vitro and in vivo. It was found that MHCC97-H cells, but not HepG2 cells form VM when they grow on matrigel in vitro. HepG2 cells gained the power of forming VM when they were overexpressed with Notch1, while knockdown Notch1 expression in MHCC97-H cells led to the loss of VM forming ability of the cells. Similar results were found in in vivo study. High expression of Notch1 in HepG2 promoted xenograft growth in nude mice, with abundant VM formation in the tumor samples. Moreover, we observed Notch1 was associated with the EMT and malignant behavior of hepatocellular carcinoma by analyzing clinical specimens, models for in vitro and in vivo experiments. HepG2 presented EMT phenomenon when induced by TGF-β1, accompanied by Notch1 activation while MHCC97-H with knockdown of Notch1 lost the responsiveness to TGF-β1 induction. Our results suggest that Notch1 promotes HCC progression through activating EMT pathway and forming VM. Our results will guide targeting Notch1 in new drug development.

  1. Small cell lung cancer, an epithelial to mesenchymal transition (EMT)-like cancer: significance of inactive Notch signaling and expression of achaete-scute complex homologue 1.

    PubMed

    Ito, Takaaki; Kudoh, Shinji; Ichimura, Takaya; Fujino, Kosuke; Hassan, Wael Ahmed Maher Abdo; Udaka, Naoko

    2017-01-01

    Small cell lung cancer (SCLC) is one of the most malignant neoplasms in common human cancers. The tumor is composed of small immature-looking cells with a round or fusiform shape, which possesses weak adhesion features among them, suggesting that SCLC shows the morphological characteristics of epithelial to mesenchymal transition (EMT). SCLC is characterized by high metastatic and recurrent rates, sensitivity to the initial chemotherapy, and easy acquirement of chemoresistance afterwards. These characters may be related to the EMT phenotype of SCLC. Notch signaling is an important signaling pathway, and could have roles in regulating neuroendocrine differentiation, proliferation, cell adhesion, EMT, and chemoresistance. Notch1 is usually absent in SCLC in vivo, but could appear after chemotherapy. Notch1 can enhance cell adhesion by induction of E-cadherin in SCLC, which indicates mesenchymal to epithelial transition. On the other hand, achaete-scute complex homologue 1 (ASCL1), negatively regulated by Notch signaling, is a lineage-specific gene of SCLC, and functions to promote neuroendocrine differentiation as well as EMT. ASCL1-transfected adenocarcinoma cell lines induced neuroendocrine phenotypes and lost epithelial cell features. SCLC is characterized by neuroendocrine differentiation and EMT-like features, which could be produced by inactive Notch signaling and ASCL1 expression. In addition, chemical and radiation treatments can activate Notch signaling, which suppress neuroendocrine differentiation and induces chemoradioresistance, accompanied by secession from EMT. Thus, the status of Notch signaling and ASCL1 expression may determine the cell behaviors of SCLC partly through modifying EMT phenotypes.

  2. miR-124 function during Ciona intestinalis neuronal development includes extensive interaction with the Notch signaling pathway.

    PubMed

    Chen, Jerry S; Pedro, Matthew San; Zeller, Robert W

    2011-11-01

    The nervous system-enriched microRNA miR-124 is necessary for proper nervous system development, although the mechanism remains poorly understood. Here, through a comprehensive analysis of miR-124 and its gene targets, we demonstrate that, in the chordate ascidian Ciona intestinalis, miR-124 plays an extensive role in promoting nervous system development. We discovered that feedback interaction between miR-124 and Notch signaling regulates the epidermal-peripheral nervous system (PNS) fate choice in tail midline cells. Notch signaling silences miR-124 in epidermal midline cells, whereas in PNS midline cells miR-124 silences Notch, Neuralized and all three Ciona Hairy/Enhancer-of-Split genes. Furthermore, ectopic expression of miR-124 is sufficient to convert epidermal midline cells into PNS neurons, consistent with a role in modulating Notch signaling. More broadly, genome-wide target extraction with validation using an in vivo tissue-specific sensor assay indicates that miR-124 shapes neuronal progenitor fields by downregulating non-neural genes, notably the muscle specifier Macho-1 and 50 Brachyury-regulated notochord genes, as well as several anti-neural factors including SCP1 and PTBP1. 3'UTR conservation analysis reveals that miR-124 targeting of SCP1 is likely to have arisen as a shared, derived trait in the vertebrate/tunicate ancestor and targeting of PTBP1 is conserved among bilaterians except for ecdysozoans, while extensive Notch pathway targeting appears to be Ciona specific. Altogether, our results provide a comprehensive insight into the specific mechanisms by which miR-124 promotes neuronal development.

  3. Eya1 controls cell polarity, spindle orientation, cell fate and Notch signaling in distal embryonic lung epithelium.

    PubMed

    El-Hashash, Ahmed H K; Turcatel, Gianluca; Al Alam, Denise; Buckley, Sue; Tokumitsu, Hiroshi; Bellusci, Saverio; Warburton, David

    2011-04-01

    Cell polarity, mitotic spindle orientation and asymmetric division play a crucial role in the self-renewal/differentiation of epithelial cells, yet little is known about these processes and the molecular programs that control them in embryonic lung distal epithelium. Herein, we provide the first evidence that embryonic lung distal epithelium is polarized with characteristic perpendicular cell divisions. Consistent with these findings, spindle orientation-regulatory proteins Insc, LGN (Gpsm2) and NuMA, and the cell fate determinant Numb are asymmetrically localized in embryonic lung distal epithelium. Interfering with the function of these proteins in vitro randomizes spindle orientation and changes cell fate. We further show that Eya1 protein regulates cell polarity, spindle orientation and the localization of Numb, which inhibits Notch signaling. Hence, Eya1 promotes both perpendicular division as well as Numb asymmetric segregation to one daughter in mitotic distal lung epithelium, probably by controlling aPKCζ phosphorylation. Thus, epithelial cell polarity and mitotic spindle orientation are defective after interfering with Eya1 function in vivo or in vitro. In addition, in Eya1(-/-) lungs, perpendicular division is not maintained and Numb is segregated to both daughter cells in mitotic epithelial cells, leading to inactivation of Notch signaling. As Notch signaling promotes progenitor cell identity at the expense of differentiated cell phenotypes, we test whether genetic activation of Notch could rescue the Eya1(-/-) lung phenotype, which is characterized by loss of epithelial progenitors, increased epithelial differentiation but reduced branching. Indeed, genetic activation of Notch partially rescues Eya1(-/-) lung epithelial defects. These findings uncover novel functions for Eya1 as a crucial regulator of the complex behavior of distal embryonic lung epithelium.

  4. EGFR and Notch signaling respectively regulate proliferative activity and multiple cell lineage differentiation of Drosophila gastric stem cells.

    PubMed

    Wang, Chenhui; Guo, Xingting; Xi, Rongwen

    2014-05-01

    Quiescent, multipotent gastric stem cells (GSSCs) in the copper cell region of adult Drosophila midgut can produce all epithelial cell lineages found in the region, including acid-secreting copper cells, interstitial cells and enteroendocrine cells, but mechanisms controlling their quiescence and the ternary lineage differentiation are unknown. By using cell ablation or damage-induced regeneration assays combined with cell lineage tracing and genetic analysis, here we demonstrate that Delta (Dl)-expressing cells in the copper cell region are the authentic GSSCs that can self-renew and continuously regenerate the gastric epithelium after a sustained damage. Lineage tracing analysis reveals that the committed GSSC daughter with activated Notch will invariably differentiate into either a copper cell or an interstitial cell, but not the enteroendocrine cell lineage, and loss-of-function and gain-of-function studies revealed that Notch signaling is both necessary and sufficient for copper cell/interstitial cell differentiation. We also demonstrate that elevated epidermal growth factor receptor (EGFR) signaling, which is achieved by the activation of ligand Vein from the surrounding muscle cells and ligand Spitz from progenitor cells, mediates the regenerative proliferation of GSSCs following damage. Taken together, we demonstrate that Dl is a specific marker for Drosophila GSSCs, whose cell cycle status is dependent on the levels of EGFR signaling activity, and the Notch signaling has a central role in controlling cell lineage differentiation from GSSCs by separating copper/interstitial cell lineage from enteroendocrine cell lineage.

  5. Chemotherapy enhances tumor vascularization via Notch signaling-mediated formation of tumor-derived endothelium in breast cancer.

    PubMed

    Zhang, Peng; He, Dongxu; Chen, Zhen; Pan, Qiongxi; Du, Fangfang; Zang, Xian; Wang, Yan; Tang, Chunlei; Li, Hong; Lu, He; Yao, Xiaoqiang; Jin, Jian; Ma, Xin

    2016-10-15

    It is believed that tumor cells can give rise to endothelial cells and tumor endothelium has a neoplastic origin. Yet, the stimuli and underlying mechanism remain unclear. Here, we demonstrate that adriamycin or paclitaxel, first-line chemotherapy agent, induced breast cancer cells to generate morphological, phenotypical and functional features of endothelial cells in vitro. In xenografts models, challenges from adriamycin or paclitaxel induced cancer cells to generate the majority of microvessels. Importantly, in breast cancer specimens from patients with neoadjuvant anthracycline-based or taxane-based chemotherapy, tumor-derived endothelial microvessels, lined by EGFR-amplified or/and TP53(+)-CD31(+) endothelial cells, was significantly higher in patients with progressive or stable disease (PD/SD) than in those with a partial or complete response (PR/CR). Further, exposure to the Notch signaling inhibitor and gene silencing studies showed that Notch signaling inhibition or silencing Nothc4/Dll3 decreased endothelial markers and function of tumor-derived endothelial cells under chemotherapy treatment, which may be through VEGFR3. Thus, our findings demonstrate that chemotherapy induces functional tumor-derived endothelial microvessels by mediating Notch signaling and VEGF signaling, and may provide new targets for anti-angiogenesis therapy in breast cancer.

  6. Targeting atypical protein kinase C iota reduces viability in glioblastoma stem-like cells via a notch signaling mechanism.

    PubMed

    Phillips, Emma; Lang, Verena; Bohlen, Jonathan; Bethke, Frederic; Puccio, Laura; Tichy, Diana; Herold-Mende, Christel; Hielscher, Thomas; Lichter, Peter; Goidts, Violaine

    2016-10-15

    In a previous study, Protein Kinase C iota (PRKCI) emerged as an important candidate gene for glioblastoma (GBM) stem-like cell (GSC) survival. Here, we show that PKCι is overexpressed and activated in patient derived GSCs compared with normal neural stem cells and normal brain lysate, and that silencing of PRKCI in GSCs causes apoptosis, along with loss of clonogenicity and reduced proliferation. Notably, PRKCI silencing reduces tumor growth in vivo in a xenograft mouse model. PKCι has been intensively studied as a therapeutic target in non-small cell lung cancer, resulting in the identification of an inhibitor, aurothiomalate (ATM), which disrupts the PKCι/ERK signaling axis. However, we show that, although sensitive to pharmacological inhibition via a pseudosubstrate peptide inhibitor, GSCs are much less sensitive to ATM, suggesting that PKCι acts along a different signaling axis in GSCs. Gene expression profiling of PRKCI-silenced GSCs revealed a novel role of the Notch signaling pathway in PKCι mediated GSC survival. A proximity ligation assay showed that Notch1 and PKCι are in close proximity in GSCs. Targeting PKCι in the context of Notch signaling could be an effective way of attacking the GSC population in GBM.

  7. 14-3-3{sigma} controls corneal epithelial cell proliferation and differentiation through the Notch signaling pathway

    SciTech Connect

    Xin, Ying; Lu, Qingxian; Li, Qiutang

    2010-02-19

    14-3-3{sigma} (also called stratifin) is specifically expressed in the stratified squamous epithelium and its function was recently shown to be linked to epidermal stratification and differentiation in the skin. In this study, we investigated its role in corneal epithelium cell proliferation and differentiation. We showed that the 14-3-3{sigma} mutation in repeated epilation (Er) mutant mice results in a dominant negative truncated protein. Primary corneal epithelial cells expressing the dominant negative protein failed to undergo high calcium-induced cell cycle arrest and differentiation. We further demonstrated that blocking endogenous 14-3-3{sigma} activity in corneal epithelial cells by overexpressing dominative negative 14-3-3{sigma} led to reduced Notch activity and Notch1/2 transcription. Significantly, expression of the active Notch intracellular domain overcame the block in epithelial cell differentiation in 14-3-3{sigma} mutant-expressing corneal epithelial cells. We conclude that 14-3-3{sigma} is critical for regulating corneal epithelial proliferation and differentiation by regulating Notch signaling activity.

  8. BRCA1 is a key regulator of breast differentiation through activation of Notch signalling with implications for anti-endocrine treatment of breast cancers

    PubMed Central

    Buckley, Niamh E.; Nic An tSaoir, Caoimhe B.; Blayney, Jaine K.; Oram, Lisa C.; Crawford, Nyree T.; D’Costa, Zenobia C.; Quinn, Jennifer E.; Kennedy, Richard D.; Harkin, D. Paul; Mullan, Paul B.

    2013-01-01

    Here, we show for the first time, that the familial breast/ovarian cancer susceptibility gene BRCA1 activates the Notch pathway in breast cells by transcriptional upregulation of Notch ligands and receptors in both normal and cancer cells. We demonstrate through chromatin immunoprecipitation assays that BRCA1 is localized to a conserved intronic enhancer region within the Notch ligand Jagged-1 (JAG1) gene, an event requiring ΔNp63. We propose that this BRCA1/ΔNp63-mediated induction of JAG1 may be important the regulation of breast stem/precursor cells, as knockdown of all three proteins resulted in increased tumoursphere growth and increased activity of stem cell markers such as Aldehyde Dehydrogenase 1 (ALDH1). Knockdown of Notch1 and JAG1 phenocopied BRCA1 knockdown resulting in the loss of Estrogen Receptor-α (ER-α) expression and other luminal markers. A Notch mimetic peptide could activate an ER-α promoter reporter in a BRCA1-dependent manner, whereas Notch inhibition using a γ-secretase inhibitor reversed this process. We demonstrate that inhibition of Notch signalling resulted in decreased sensitivity to the anti-estrogen drug Tamoxifen but increased expression of markers associated with basal-like breast cancer. Together, these findings suggest that BRCA1 transcriptional upregulation of Notch signalling is a key event in the normal differentiation process in breast tissue. PMID:23863842

  9. Angiopoietin-like proteins stimulate HSPC development through interaction with notch receptor signaling.

    PubMed

    Lin, Michelle I; Price, Emily N; Boatman, Sonja; Hagedorn, Elliott J; Trompouki, Eirini; Satishchandran, Sruthi; Carspecken, Charles W; Uong, Audrey; DiBiase, Anthony; Yang, Song; Canver, Matthew C; Dahlberg, Ann; Lu, Zhigang; Zhang, Cheng Cheng; Orkin, Stuart H; Bernstein, Irwin D; Aster, Jon C; White, Richard M; Zon, Leonard I

    2015-02-25

    Angiopoietin-like proteins (angptls) are capable of ex vivo expansion of mouse and human hematopoietic stem and progenitor cells (HSPCs). Despite this intriguing ability, their mechanism is unknown. In this study, we show that angptl2 overexpression is sufficient to expand definitive HSPCs in zebrafish embryos. Angptl1/2 are required for definitive hematopoiesis and vascular specification of the hemogenic endothelium. The loss-of-function phenotype is reminiscent of the notch mutant mindbomb (mib), and a strong genetic interaction occurs between angptls and notch. Overexpressing angptl2 rescues mib while overexpressing notch rescues angptl1/2 morphants. Gene expression studies in ANGPTL2-stimulated CD34(+) cells showed a strong MYC activation signature and myc overexpression in angptl1/2 morphants or mib restored HSPCs formation. ANGPTL2 can increase NOTCH activation in cultured cells and ANGPTL receptor interacted with NOTCH to regulate NOTCH cleavage. Together our data provide insight to the angptl-mediated notch activation through receptor interaction and subsequent activation of myc targets.

  10. Angiopoietin-like proteins stimulate HSPC development through interaction with notch receptor signaling

    PubMed Central

    Lin, Michelle I; Price, Emily N; Boatman, Sonja; Hagedorn, Elliott J; Trompouki, Eirini; Satishchandran, Sruthi; Carspecken, Charles W; Uong, Audrey; DiBiase, Anthony; Yang, Song; Canver, Matthew C; Dahlberg, Ann; Lu, Zhigang; Zhang, Cheng Cheng; Orkin, Stuart H; Bernstein, Irwin D; Aster, Jon C; White, Richard M; Zon, Leonard I

    2015-01-01

    Angiopoietin-like proteins (angptls) are capable of ex vivo expansion of mouse and human hematopoietic stem and progenitor cells (HSPCs). Despite this intriguing ability, their mechanism is unknown. In this study, we show that angptl2 overexpression is sufficient to expand definitive HSPCs in zebrafish embryos. Angptl1/2 are required for definitive hematopoiesis and vascular specification of the hemogenic endothelium. The loss-of-function phenotype is reminiscent of the notch mutant mindbomb (mib), and a strong genetic interaction occurs between angptls and notch. Overexpressing angptl2 rescues mib while overexpressing notch rescues angptl1/2 morphants. Gene expression studies in ANGPTL2-stimulated CD34+ cells showed a strong MYC activation signature and myc overexpression in angptl1/2 morphants or mib restored HSPCs formation. ANGPTL2 can increase NOTCH activation in cultured cells and ANGPTL receptor interacted with NOTCH to regulate NOTCH cleavage. Together our data provide insight to the angptl-mediated notch activation through receptor interaction and subsequent activation of myc targets. DOI: http://dx.doi.org/10.7554/eLife.05544.001 PMID:25714926

  11. High mobility group A1 protein acts as a new target of Notch1 signaling and regulates cell proliferation in T leukemia cells.

    PubMed

    Xi, Yang; Li, Yu-Sang; Tang, He-Bin

    2013-02-01

    Active mutations of Notch1 play pivotal roles during leukemogenesis, but the downstream targets and molecular mechanisms of activated Notch1 signaling have not yet been fully clarified. In this study, we detected the overexpression of the high mobility group A1 (HMGA1) and activation of Notch1 signaling in mouse thymic lymphomas. A direct regulation of Notch1 on HMGA1 transcription was demonstrated and two Notch1/RBPJ cobinding sites of T/CTCCCACA were found in HMGA1 promoter regions. It was the first time demonstrated that HMGA1 was the downstream target of Notch1 signaling. Moreover, knockdown of HMGA1 resulted in significantly impaired cell growth and decreased expressions of cyclin D and cyclin E in human T leukemia cells. The formation of complexes was also observed between HMGA1 and retinoblastoma (RB) protein indicating a mechanism of cell cycle regulation. These findings suggest that activated HMGA1 regulates cell proliferation through the Notch1 signaling pathway, which represents an important molecular pathway leading to leukemogenesis.

  12. Repressing Notch Signaling and Expressing TNFα Are Sufficient to Mimic Retinal Regeneration by Inducing Müller Glial Proliferation to Generate Committed Progenitor Cells

    PubMed Central

    Conner, Clay; Ackerman, Kristin M.; Lahne, Manuela; Hobgood, Joshua S.

    2014-01-01

    Retinal damage in teleosts, unlike mammals, induces robust Müller glia-mediated regeneration of lost neurons. We examined whether Notch signaling regulates Müller glia proliferation in the adult zebrafish retina and demonstrated that Notch signaling maintains Müller glia in a quiescent state in the undamaged retina. Repressing Notch signaling, through injection of the γ-secretase inhibitor RO4929097, stimulates a subset of Müller glia to reenter the cell cycle without retinal damage. This RO4929097-induced Müller glia proliferation is mediated by repressing Notch signaling because inducible expression of the Notch Intracellular Domain (NICD) can reverse the effect. This RO4929097-induced proliferation requires Ascl1a expression and Jak1-mediated Stat3 phosphorylation/activation, analogous to the light-damaged retina. Moreover, coinjecting RO4929097 and TNFα, a previously identified damage signal, induced the majority of Müller glia to reenter the cell cycle and produced proliferating neuronal progenitor cells that committed to a neuronal lineage in the undamaged retina. This demonstrates that repressing Notch signaling and activating TNFα signaling are sufficient to induce Müller glia proliferation that generates neuronal progenitor cells that differentiate into retinal neurons, mimicking the responses observed in the regenerating retina. PMID:25339752

  13. Pituitary Phenotypes of Mice Lacking the Notch Signalling Ligand Delta-Like 1 Homologue

    PubMed Central

    Cheung, L Y M; Rizzoti, K; Lovell-Badge, R; Tissier, P R

    2013-01-01

    The Notch signalling pathway ligand delta-like 1 homologue (Dlk1, also named Pref1) is expressed throughout the developing pituitary and becomes restricted to mostly growth hormone (GH) cells within the adult gland. We have investigated the role of Dlk1 in pituitary development and function from late embryogenesis to adulthood using a mouse model completely lacking the expression of Dlk1. We confirm that Dlk1-null mice are shorter and weigh less than wild-type littermates from late gestation, at parturition and in adulthood. A loss of Dlk1 leads to significant reduction in GH content throughout life, whereas other pituitary hormones are reduced to varying degrees depending on sex and age. Both the size of the pituitary and the proportion of hormone-producing cell populations are unchanged, suggesting that there is a reduction in hormone content per cell. In vivo challenge of mutant and wild-type littermates with growth hormone-releasing hormone and growth hormone-releasing hexapeptide shows that reduced GH secretion is unlikely to account for the reduced growth of Dlk1 knockout animals. These data suggest that loss of Dlk1 gives rise to minor pituitary defects manifesting as an age- and sex-dependent reduction in pituitary hormone contents. However, Dlk1 expression in other tissue is most likely responsible for the weight and length differences observed in mutant animals. PMID:23279263

  14. Subcellular localization of Suppressor of Hairless in Drosophila sense organ cells during Notch signalling.

    PubMed

    Gho, M; Lecourtois, M; Géraud, G; Posakony, J W; Schweisguth, F

    1996-06-01

    During imaginal development of Drosophila, Suppressor of Hairless [Su(H)], an evolutionarily conserved transcription factor that mediates intracellular signalling by the Notch (N) receptor, controls successive alternative cell fate decisions leading to the differentiation of multicellular sensory organs. We describe here the distribution of the Su(H) protein in the wing disc epithelium throughout development of adult sense organs. Su(H) was found to be evenly distributed in the nuclei of all imaginal disc cells during sensory organ precursor cells selection. Thus differential expression and/or subcellular localization of Su(H) is not essential for its function. Soon after division of the pIIa secondary precursor cell, Su(H) specifically accumulates in the nucleus of the future socket cell. At the onset of differentiation of the socket cell, Su(H) is also detected in the cytoplasm. In this differentiating cell, N and deltex participate in the cytoplasmic retention of Su(H). Still, Su(H) does not colocalize with N at the apical-lateral membranes. These observations suggest that N regulates in an indirect manner the cytoplasmic localization of Su(H) in the socket cell. Finally, the pIIb, shaft and socket cells are found to adopt invariant positions along the anteroposterior axis of the notum. This raises the possibility that tissue-polarity biases these N-mediated cell fate choices.

  15. The role of Notch signaling in muscle progenitor cell depletion and the rapid onset of histopathology in muscular dystrophy

    PubMed Central

    Mu, Xiaodong; Tang, Ying; Lu, Aiping; Takayama, Koji; Usas, Arvydas; Wang, Bing; Weiss, Kurt; Huard, Johnny

    2015-01-01

    Although it has been speculated that stem cell depletion plays a role in the rapid progression of the muscle histopathology associated with Duchenne Muscular Dystrophy (DMD), the molecular and cellular mechanisms responsible for stem cell depletion remain poorly understood. The rapid depletion of muscle stem cells has not been observed in the dystrophin-deficient model of DMD (mdx mouse), which may explain the relatively mild dystrophic phenotype observed in this animal model. In contrast, we have observed a rapid occurrence of stem cell depletion in the dystrophin/utrophin double knockout (dKO) mouse model, which exhibits histopathological features that more closely recapitulate the phenotype observed in DMD patients compared with the mdx mouse. Notch signaling has been found to be a key regulator of stem cell self-renewal and myogenesis in normal skeletal muscle; however, little is known about the role that Notch plays in the development of the dystrophic histopathology associated with DMD. Our results revealed an over-activation of Notch in the skeletal muscles of dKO mice, which correlated with sustained inflammation, impaired muscle regeneration and the rapid depletion and senescence of the muscle progenitor cells (MPCs, i.e. Pax7+ cells). Consequently, the repression of Notch in the skeletal muscle of dKO mice delayed/reduced the depletion and senescence of MPCs, and restored the myogenesis capacity while reducing inflammation and fibrosis. We suggest that the down-regulation of Notch could represent a viable approach to reduce the dystrophic histopathologies associated with DMD. PMID:25678553

  16. The role of Notch signaling in muscle progenitor cell depletion and the rapid onset of histopathology in muscular dystrophy.

    PubMed

    Mu, Xiaodong; Tang, Ying; Lu, Aiping; Takayama, Koji; Usas, Arvydas; Wang, Bing; Weiss, Kurt; Huard, Johnny

    2015-05-15

    Although it has been speculated that stem cell depletion plays a role in the rapid progression of the muscle histopathology associated with Duchenne Muscular Dystrophy (DMD), the molecular and cellular mechanisms responsible for stem cell depletion remain poorly understood. The rapid depletion of muscle stem cells has not been observed in the dystrophin-deficient model of DMD (mdx mouse), which may explain the relatively mild dystrophic phenotype observed in this animal model. In contrast, we have observed a rapid occurrence of stem cell depletion in the dystrophin/utrophin double knockout (dKO) mouse model, which exhibits histopathological features that more closely recapitulate the phenotype observed in DMD patients compared with the mdx mouse. Notch signaling has been found to be a key regulator of stem cell self-renewal and myogenesis in normal skeletal muscle; however, little is known about the role that Notch plays in the development of the dystrophic histopathology associated with DMD. Our results revealed an over-activation of Notch in the skeletal muscles of dKO mice, which correlated with sustained inflammation, impaired muscle regeneration and the rapid depletion and senescence of the muscle progenitor cells (MPCs, i.e. Pax7+ cells). Consequently, the repression of Notch in the skeletal muscle of dKO mice delayed/reduced the depletion and senescence of MPCs, and restored the myogenesis capacity while reducing inflammation and fibrosis. We suggest that the down-regulation of Notch could represent a viable approach to reduce the dystrophic histopathologies associated with DMD.

  17. Notch-1 signaling activates NF-κB in human breast carcinoma MDA-MB-231 cells via PP2A-dependent AKT pathway.

    PubMed

    Li, Li; Zhang, Jing; Xiong, Niya; Li, Shun; Chen, Yu; Yang, Hong; Wu, Chunhui; Zeng, Hongjuan; Liu, Yiyao

    2016-04-01

    Breast cancer has a high incidence in the world and is becoming a leading cause of death in female patients due to its high metastatic ability. High expression of Notch-1 and its ligand Jagged-1 correlates with poor prognosis in breast cancer. Our previous work has shown that Notch-1 signaling pathway upregulates NF-κB transcriptional activity and induces the adhesion, migration and invasion of human breast cancer cell line MDA-MB-231. However, the role of Notch-1 in NF-κB activation is still poorly understood. Here, we aim to understand the exact mechanism that Notch-1 regulates NF-κB activity. In MDA-MB-231 cells where Notch-1 is constitutively activated, the phosphorylation of p85 and AKT (Tyr308/Ser473) is upregulated, indicating PI3K/AKT pathway is activated. Notch-1 activation caused the increase of PP2A phosphorylation at Tyr307, indicating Notch-1 inhibits PP2A activity. NF-κB transcriptional activity was evaluated by dual-luciferase reporter assay, and the results showed that, while silencing of Notch-1, PP2A activity was upregulated and NF-κB activity was downregulated, whereas PP2A inhibitor okadaic acid (OA) restored NF-κB activity. Immunofluorescence and Western blots showed that OA treatment antagonized the decrease of p65 nuclear translocation caused by Notch-1 silencing. Moreover, OA treatment also upregulated MMP-2, MMP-9 and VEGF mRNA expression levels, indicating OA rescues Notch-1 silencing that caused low cell invasion. Taken together, our results suggest that Notch-1-activating PI3K/AKT/NF-κB pathway is PP2A dependent; PP2A may be a potential therapeutic target in breast cancer.

  18. Enhanced expression of ADCY1 underlies aberrant neuronal signalling and behaviour in a syndromic autism model

    PubMed Central

    Sethna, Ferzin; Feng, Wei; Ding, Qi; Robison, Alfred J.; Feng, Yue; Wang, Hongbing

    2017-01-01

    Fragile X syndrome (FXS), caused by the loss of functional FMRP, is a leading cause of autism. Neurons lacking FMRP show aberrant mRNA translation and intracellular signalling. Here, we identify that, in Fmr1 knockout neurons, type 1 adenylyl cyclase (Adcy1) mRNA translation is enhanced, leading to excessive production of ADCY1 protein and insensitivity to neuronal stimulation. Genetic reduction of Adcy1 normalizes the aberrant ERK1/2- and PI3K-mediated signalling, attenuates excessive protein synthesis and corrects dendritic spine abnormality in Fmr1 knockout mice. Genetic reduction of Adcy1 also ameliorates autism-related symptoms including repetitive behaviour, defective social interaction and audiogenic seizures. Moreover, peripheral administration of NB001, an experimental compound that preferentially suppresses ADCY1 activity over other ADCY subtypes, attenuates the behavioural abnormalities in Fmr1 knockout mice. These results demonstrate a connection between the elevated Adcy1 translation and abnormal ERK1/2 signalling and behavioural symptoms in FXS. PMID:28218269

  19. Modulation of aberrant CDK5 signaling rescues impaired neurogenesis in models of Alzheimer's disease.

    PubMed

    Crews, L; Patrick, C; Adame, A; Rockenstein, E; Masliah, E

    2011-02-10

    Recent studies show that in Alzheimer's disease (AD), alterations in neurogenesis contribute to the neurodegenerative process. Neurodegeneration in AD has been associated with aberrant signaling through the cyclin-dependent kinase-5 (CDK5) pathway via its activators p35/p25; however, the role of CDK5 in the mechanisms of defective adult neurogenesis in AD is unknown. First, to study AD-like abnormal activation of CDK5 signaling in an in vitro model of neurogenesis, neuronal progenitor cells (NPCs) were infected with a viral vector expressing p35, and exposed to amyloid-β protein (Aβ(1-42)). These conditions resulted in impaired maturation and neurite outgrowth in vitro, and these effects were reversed by pharmacological or genetic inhibition of CDK5. Similarly, neurogenesis was impaired in a transgenic mouse model of AD that expresses high levels of amyloid precursor protein (APP), and this effect was reversed in transgenic mice crossed with a CDK5 heterozygous-deficient mouse line. A similar rescue effect was observed in APP transgenic mice treated with Roscovitine, a pharmacological inhibitor of CDK5. Taken together, these data suggest that the CDK5 signaling pathway has a critical role in maintaining the integrity of NPCs and neuronal maturation in the adult hippocampus. Moreover, potential therapeutic approaches could focus on modulating the aberrant activity of CDK5 to target the neurogenic and neurodegenerative alterations in AD.

  20. Analysis of the Transcriptomes Downstream of Eyeless and the Hedgehog, Decapentaplegic and Notch Signaling Pathways in Drosophila melanogaster

    PubMed Central

    Nfonsam, Landry E.; Cano, Carlos; Mudge, Joann; Schilkey, Faye D.; Curtiss, Jennifer

    2012-01-01

    Tissue-specific transcription factors are thought to cooperate with signaling pathways to promote patterned tissue specification, in part by co-regulating transcription. The Drosophila melanogaster Pax6 homolog Eyeless forms a complex, incompletely understood regulatory network with the Hedgehog, Decapentaplegic and Notch signaling pathways to control eye-specific gene expression. We report a combinatorial approach, including mRNAseq and microarray analyses, to identify targets co-regulated by Eyeless and Hedgehog, Decapentaplegic or Notch. Multiple analyses suggest that the transcriptomes resulting from co-misexpression of Eyeless+signaling factors provide a more complete picture of eye development compared to previous efforts involving Eyeless alone: (1) Principal components analysis and two-way hierarchical clustering revealed that the Eyeless+signaling factor transcriptomes are closer to the eye control transcriptome than when Eyeless is misexpressed alone; (2) more genes are upregulated at least three-fold in response to Eyeless+signaling factors compared to Eyeless alone; (3) based on gene ontology analysis, the genes upregulated in response to Eyeless+signaling factors had a greater diversity of functions compared to Eyeless alone. Through a secondary screen that utilized RNA interference, we show that the predicted gene CG4721 has a role in eye development. CG4721 encodes a neprilysin family metalloprotease that is highly up-regulated in response to Eyeless+Notch, confirming the validity of our approach. Given the similarity between D. melanogaster and vertebrate eye development, the large number of novel genes identified as potential targets of Ey+signaling factors will provide novel insights to our understanding of eye development in D. melanogaster and humans. PMID:22952997

  1. NOTCH receptors in gastric and other gastrointestinal cancers: oncogenes or tumor suppressors?

    PubMed

    Huang, Tingting; Zhou, Yuhang; Cheng, Alfred S L; Yu, Jun; To, Ka Fai; Kang, Wei

    2016-12-09

    Gastric cancer (GC) ranks the most common cancer types and is one of the leading causes of cancer-related death. Due to delayed diagnosis and high metastatic frequency, 5-year survival rate of GC is rather low. It is a complex disease resulting from the interaction between environmental factors and host genetic alterations that deregulate multiple signaling pathways. The Notch signaling pathway, a highly conserved system in the regulation of the fate in several cell types, plays a pivotal role in cell differentiation, survival and proliferation. Notch is also one of the most commonly activated signaling pathways in tumors and its aberrant activation plays a key role in cancer advancement. Whether Notch cascade exerts oncogenic or tumor suppressive function in different cancer types depends on the cellular context. Mammals have four NOTCH receptors that modulate Notch pathway activity. In this review, we provide a comprehensive summary on the functional role of NOTCH receptors in gastric and other gastrointestinal cancers. Increasing knowledge of NOTCH receptors in gastrointestinal cancers will help us recognize the underlying mechanisms of Notch signaling and develop novel therapeutic strategies for GC.

  2. The vascular delta-like ligand-4 (DLL4)-Notch4 signaling correlates with angiogenesis in primary glioblastoma: an immunohistochemical study.

    PubMed

    Zhang, Jin-Feng; Chen, Yao; Qiu, Xian-Xin; Tang, Wen-Long; Zhang, Jian-Dong; Huang, Jian-Huang; Lin, Guo-Shi; Wang, Xing-Fu; Lin, Zhi-Xiong

    2016-03-01

    Delta-like ligand-4 (DLL4)-Notch signaling is known to play a pivotal role in the regulation of tumor angiogenesis. We had previously found that DLL4 was overexpressed, while Notch1 receptor, which binds to DLL4 during angiogenesis, was absent in the majority of human primary glioblastomas. Thus, DLL4-Notch signaling pathway in the regulation of tumor angiogenesis in primary glioblastoma remains unknown. Tumor tissues from 70 patients with primary glioblastoma were analyzed by immunohistochemistry for expression of components of DLL4-Notch signaling, vascular endothelial growth factor (VEGF), and microvessel density (MVD). Immunohistochemistry results showed that the positive staining of DLL4 and Notch4 was primarily distributed in tumor vascular endothelial cells but rarely detected in tumor cells. However, VEGF, hairy/enhancer of split-1 (HES1; a target gene of Notch signaling), and Notch1-3 expression was seen in both tumor vascular endothelial cells and tumor cells. Univariate analysis showed that the expression levels of VEGF and DLL4, HES1, and Notch4 in tumor endothelial cells were significantly associated with MVD in primary glioblastoma (P < 0.001). Binary logistic regression analysis showed that high expression levels of DLL4, HES1, and Notch4 in tumor endothelial cells were associated with a decrease of MVD in primary glioblastoma, while MVD increased with elevated VEGF expression in contrast. In addition, DLL4, Notch4, and HES1 expression were positively correlated in tumor vascular endothelial cells (P < 0.05). We conclude that the vascular DLL4-Notch4 signaling and VEGF signaling complementing each other plays an important role in the progression of tumor angiogenesis in primary glioblastoma. Graphical abstract A, positive staining of DLL4 in human kidney; B, positive staining of VEGF in human breast cancer; C, positive staining of CD34 in human lung cancer; D, positive staining of HES1 in human breast cancer; E-H, positive staining of Notch1