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Sample records for antagonizing wnt erk

  1. Wnt antagonism initiates cardiogenesis in Xenopus laevis

    PubMed Central

    Schneider, Valerie A.; Mercola, Mark

    2001-01-01

    Heart induction in Xenopus occurs in paired regions of the dorsoanterior mesoderm in response to signals from the Spemann organizer and underlying dorsoanterior endoderm. These tissues together are sufficient to induce heart formation in noncardiogenic ventral marginal zone mesoderm. Similarly, in avians the underlying definitive endoderm induces cardiogenesis in precardiac mesoderm. Heart-inducing factors in amphibians are not known, and although certain BMPs and FGFs can mimic aspects of cardiogenesis in avians, neither can induce the full range of activities elicited by the inducing tissues. Here we report that the Wnt antagonists Dkk-1 and Crescent can induce heart formation in explants of ventral marginal zone mesoderm. Other Wnt antagonists, including the frizzled domain-containing proteins Frzb and Szl, lacked this activity. Unlike Wnt antagonism, inhibition of BMP signaling did not promote cardiogenesis. Ectopic expression of GSK3β, which inhibits β-catenin-mediated Wnt signaling, also induced cardiogenesis in ventral mesoderm. Analysis of Wnt proteins expressed during gastrulation revealed that Wnt3A and Wnt8, but not Wnt5A or Wnt11, inhibited endogenous heart induction. These results indicate that diffusion of Dkk-1 and Crescent from the organizer initiate cardiogenesis in adjacent mesoderm by establishing a zone of low Wnt3A and Wnt8 activity. PMID:11159911

  2. Zebrafish Naked 1 and Naked 2 antagonize both canonical and non-canonical Wnt signaling

    PubMed Central

    Van Raay, Terence J.; Coffey, Robert J.; Solnica-Krezel, Lilianna

    2007-01-01

    Wnt signaling controls a wide range of developmental processes and its aberrant regulation can lead to disease. To better understand the regulation of this pathway, we identified zebrafish homologues of Naked Cuticle (Nkd), Nkd1 and Nkd2, which have previously been shown to inhibit canonical Wnt/β-catenin signaling. Zebrafish nkd1 expression increases substantially after the mid-blastula transition in a pattern mirroring that of activated canonical Wnt/β-catenin signaling, being expressed in both the ventrolateral blastoderm margin and also in the axial mesendoderm. In contrast, zebrafish nkd2 is maternally and ubiquitously expressed. Overexpression of Nkd1 or Nkd2a suppressed canonical Wnt/β-catenin signaling at multiple stages of early zebrafish development and also exacerbated the cyclopia and axial mesendoderm convergence and extension (C&E) defect in the non-canonical Wnt/PCP mutant silberblick (slb/wnt11). Thus, Nkds are sufficient to antagonize both canonical and non-canonical Wnt signaling. Reducing Nkd function using antisense morpholino oligonucleotides resulted in increased expression of canonical Wnt/β-catenin target genes. Finally, reducing Nkd1 function in slb mutants suppressed the axial mesendoderm C&E defect. These data indicate that zebrafish Nkd1 and Nkd2 function to limit both canonical and non-canonical Wnt signaling. PMID:17689523

  3. Canonical Wnt signaling is antagonized by noncanonical Wnt5a in hepatocellular carcinoma cells

    PubMed Central

    Yuzugullu, Haluk; Benhaj, Khemais; Ozturk, Nuri; Senturk, Serif; Celik, Emine; Toylu, Asli; Tasdemir, Nilgun; Yilmaz, Mustafa; Erdal, Esra; Akcali, Kamil Can; Atabey, Nese; Ozturk, Mehmet

    2009-01-01

    Background β-catenin mutations that constitutively activate the canonical Wnt signaling have been observed in a subset of hepatocellular carcinomas (HCCs). These mutations are associated with chromosomal stability, low histological grade, low tumor invasion and better patient survival. We hypothesized that canonical Wnt signaling is selectively activated in well-differentiated, but repressed in poorly differentiated HCCs. To this aim, we characterized differentiation status of HCC cell lines and compared their expression status of Wnt pathway genes, and explored their activity of canonical Wnt signaling. Results We classified human HCC cell lines into "well-differentiated" and "poorly differentiated" subtypes, based on the expression of hepatocyte lineage, epithelial and mesenchymal markers. Poorly differentiated cell lines lost epithelial and hepatocyte lineage markers, and overexpressed mesenchymal markers. Also, they were highly motile and invasive. We compared the expression of 45 Wnt pathway genes between two subtypes. TCF1 and TCF4 factors, and LRP5 and LRP6 co-receptors were ubiquitously expressed. Likewise, six Frizzled receptors, and canonical Wnt3 ligand were expressed in both subtypes. In contrast, canonical ligand Wnt8b and noncanonical ligands Wnt4, Wnt5a, Wnt5b and Wnt7b were expressed selectively in well- and poorly differentiated cell lines, respectively. Canonical Wnt signaling activity, as tested by a TCF reporter assay was detected in 80% of well-differentiated, contrary to 14% of poorly differentiated cell lines. TCF activity generated by ectopic mutant β-catenin was weak in poorly differentiated SNU449 cell line, suggesting a repressive mechanism. We tested Wnt5a as a candidate antagonist. It strongly inhibited canonical Wnt signaling that is activated by mutant β-catenin in HCC cell lines. Conclusion Differential expression of Wnt ligands in HCC cells is associated with selective activation of canonical Wnt signaling in well

  4. EGF-reduced Wnt5a transcription induces epithelial-mesenchymal transition via Arf6-ERK signaling in gastric cancer cells

    PubMed Central

    Zhang, Yujie; Du, Jun; Zheng, Jianchao; Liu, Jiaojing; Xu, Rui; Shen, Tian; Zhu, Yichao; Chang, Jun; Wang, Hong; Zhang, Zhihong; Meng, Fanqing; Wang, Yan; Chen, Yongchang; Xu, Yong; Gu, Luo

    2015-01-01

    Wnt5a, a ligand for activating the non-canonical Wnt signaling pathway, is commonly associated with Epithelial-to-mesenchymal transition (EMT) in cancer cell metastasis. Here, we show that downregulation of Wnt5a mRNA and protein by EGF is necessary for EGF-induced EMT in gastric cancer SGC-7901 cells. To further explore the mechanisms, we investigated the effect of EGF signaling on Wnt5a expression. EGF increased Arf6 and ERK activity, while blockade of Arf6 activation repressed ERK activity, up-regulated Wnt5a expression and repressed EMT in response to EGF. We also demonstrate that EGF inactivated Wnt5a transcription by direct recruitment of ERK to the Wnt5a promoter. On the other hand, inhibition of ERK phosphorylation resulted in decreased movement of ERK from the cytoplasm to the nucleus, following rescued Wnt5a mRNA and protein expression and favored an epithelial phenotype of SGC-7901 cells. In addition, we notice that kinase-dead, nuclear-localised ERK has inhibitory effect on Wnt5a transcription. Analysis of gastric cancer specimens revealed an inverse correlation between P-ERK and Wnt5a protein levels and an association between Wnt5a expression and better prognosis. These findings indicate that Wnt5a is a potential suppressor of EMT and identify a novel Arf6/ERK signaling pathway for EGF-regulated Wnt5a expression at transcriptional level of gastric cancer cells. PMID:25779663

  5. WNT5a is required for normal ovarian follicle development and antagonizes gonadotropin responsiveness in granulosa cells by suppressing canonical WNT signaling

    PubMed Central

    Abedini, Atefeh; Zamberlam, Gustavo; Lapointe, Evelyne; Tourigny, Catherine; Boyer, Alexandre; Paquet, Marilène; Hayashi, Kanako; Honda, Hiroaki; Kikuchi, Akira; Price, Christopher; Boerboom, Derek

    2015-01-01

    Whereas the roles of the canonical wingless-type MMTV (mouse mammary tumor virus) integration site family (WNT) signaling pathway in the regulation of ovarian follicle growth and steroidogenesis are now established, noncanonical WNT signaling in the ovary has been largely overlooked. Noncanonical WNTs, including WNT5a and WNT11, are expressed in granulosa cells (GCs) and are differentially regulated throughout follicle development, but their physiologic roles remain unknown. Using conditional gene targeting, we found that GC-specific inactivation of Wnt5a (but not Wnt11) results in the female subfertility associated with increased follicular atresia and decreased rates of ovulation. Microarray analyses have revealed that WNT5a acts to down-regulate the expression of FSH-responsive genes in vitro, and corresponding increases in the expression of these genes have been found in the GCs of conditional knockout mice. Unexpectedly, we found that WNT5a regulates its target genes not by signaling via the WNT/Ca2+ or planar cell polarity pathways, but rather by inhibiting the canonical pathway, causing both β-catenin (CTNNB1) and cAMP responsive element binding (CREB) protein levels to decrease via a glycogen synthase kinase-3β-dependent mechanism. We further found that WNT5a prevents follicle-stimulating hormone and luteinizing protein from up-regulating the CTNNB1 and CREB proteins and their target genes, indicating that WNT5a functions as a physiologic inhibitor of gonadotropin signaling. Together, these findings identify WNT5a as a key regulator of follicle development and gonadotropin responsiveness.—Abedini, A., Zamberlam, G., Lapointe, E., Tourigny, C., Boyer, A., Paquet, M., Hayashi, K., Honda, H., Kikuchi, A., Price, C., Boerboom, D. WNT5a is required for normal ovarian follicle development and antagonizes gonadotropin responsiveness in granulosa cells by suppressing canonical WNT signaling. PMID:26667040

  6. WNT5a is required for normal ovarian follicle development and antagonizes gonadotropin responsiveness in granulosa cells by suppressing canonical WNT signaling.

    PubMed

    Abedini, Atefeh; Zamberlam, Gustavo; Lapointe, Evelyne; Tourigny, Catherine; Boyer, Alexandre; Paquet, Marilène; Hayashi, Kanako; Honda, Hiroaki; Kikuchi, Akira; Price, Christopher; Boerboom, Derek

    2016-04-01

    Whereas the roles of the canonical wingless-type MMTV (mouse mammary tumor virus) integration site family (WNT) signaling pathway in the regulation of ovarian follicle growth and steroidogenesis are now established, noncanonical WNT signaling in the ovary has been largely overlooked. Noncanonical WNTs, including WNT5a and WNT11, are expressed in granulosa cells (GCs) and are differentially regulated throughout follicle development, but their physiologic roles remain unknown. Using conditional gene targeting, we found that GC-specific inactivation ofWnt5a(but notWnt11) results in the female subfertility associated with increased follicular atresia and decreased rates of ovulation. Microarray analyses have revealed that WNT5a acts to down-regulate the expression of FSH-responsive genesin vitro, and corresponding increases in the expression of these genes have been found in the GCs of conditional knockout mice. Unexpectedly, we found that WNT5a regulates its target genes not by signalingviathe WNT/Ca(2+)or planar cell polarity pathways, but rather by inhibiting the canonical pathway, causing both β-catenin (CTNNB1) and cAMP responsive element binding (CREB) protein levels to decreaseviaa glycogen synthase kinase-3β-dependent mechanism. We further found that WNT5a prevents follicle-stimulating hormone and luteinizing protein from up-regulating the CTNNB1 and CREB proteins and their target genes, indicating that WNT5a functions as a physiologic inhibitor of gonadotropin signaling. Together, these findings identify WNT5a as a key regulator of follicle development and gonadotropin responsiveness.-Abedini, A., Zamberlam, G., Lapointe, E., Tourigny, C., Boyer, A., Paquet, M., Hayashi, K., Honda, H., Kikuchi, A., Price, C., Boerboom, D. WNT5a is required for normal ovarian follicle development and antagonizes gonadotropin responsiveness in granulosa cells by suppressing canonical WNT signaling.

  7. Receptor for advanced glycation end products inhibits proliferation in osteoblast through suppression of Wnt, PI3K and ERK signaling

    SciTech Connect

    Li, Guofeng; Xu, Jingren; Li, Zengchun

    2012-07-13

    Highlights: Black-Right-Pointing-Pointer RAGE overexpression suppresses cell proliferation in MC3T3-E1 cells. Black-Right-Pointing-Pointer RAGE overexpression decreases Wnt/{beta}-catenin signaling. Black-Right-Pointing-Pointer RAGE overexpression decreases ERK and PI3K signaling. Black-Right-Pointing-Pointer Inhibition of Wnt signaling abolishes PI3K signaling restored by RAGE blockade. Black-Right-Pointing-Pointer Inhibition of Wnt signaling abolishes ERK signaling restored by RAGE blockade. -- Abstract: Expression of receptor for advanced glycation end products (RAGE) plays a crucial role in bone metabolism. However, the role of RAGE in the control of osteoblast proliferation is not yet evaluated. In the present study, we demonstrate that RAGE overexpression inhibits osteoblast proliferation in vitro. The negative regulation of RAGE on cell proliferation results from suppression of Wnt, PI3K and ERK signaling, and is restored by RAGE neutralizing antibody. Prevention of Wnt signaling using Sfrp1 or DKK1 rescues RAGE-decreased PI3K and ERK signaling and cell proliferation, indicating that the altered cell growth in RAGE overexpressing cells is in part secondary to alterations in Wnt signaling. Consistently, RAGE overexpression inhibits the expression of Wnt targets cyclin D1 and c-myc, which is partially reversed by RAGE blockade. Overall, these results suggest that RAGE inhibits osteoblast proliferation via suppression of Wnt, PI3K and ERK signaling, which provides novel mechanisms by which RAGE regulates osteoblast growth.

  8. Wnt signalling antagonizes stress granule assembly through a Dishevelled-dependent mechanism

    PubMed Central

    Sahoo, Pabitra K.; Murawala, Prayag; Sawale, Pravin T.; Sahoo, Manas R.; Tripathi, Mukesh M.; Gaikwad, Swati R.; Seshadri, Vasudevan; Joseph, Jomon

    2012-01-01

    Summary Cells often respond to diverse environmental stresses by inducing stress granules (SGs) as an adaptive mechanism. SGs are generally assembled as a result of aggregation of mRNAs stalled in a translational pre-initiation complex, mediated by a set of RNA-binding proteins such as G3BP and TIA-1. SGs may serve as triage centres for storage, translation re-initiation or degradation of specific mRNAs. However, the mechanism involved in the modulation of their assembly/disassembly is unclear. Here we report that Wnt signalling negatively regulates SG assembly through Dishevelled (Dvl), a cytoplasmic Wnt effector. Overexpression of Dvl2, an isoform of Dvl, leads to impairment of SG assembly through a DEP domain dependent mechanism. Intriguingly, the Dvl2 mutant K446M, which corresponds to an analogous mutation in Drosophila Dishevelled DEP domain (dsh1) that results in defective PCP pathway, fails to antagonize SG assembly. Furthermore, we show that Dvl2 exerts the antagonistic effect on SG assembly through a mechanism involving Rac1-mediated inhibition of RhoA. Dvl2 interacts with G3BP, a downstream component of Ras signalling involved in SG assembly, and functional analysis suggests a model wherein the Dvl-Rac1-RhoA axis regulates G3BP's SG-nucleating activity. Collectively, these results define an antagonistic effect of Wnt signalling on SG assembly, and reveal a novel role for Wnt/Dvl pathway in the modulation of mRNA functions. PMID:23213403

  9. In Hyperthermia Increased ERK and WNT Signaling Suppress Colorectal Cancer Cell Growth

    PubMed Central

    Bordonaro, Michael; Shirasawa, Senji; Lazarova, Darina L.

    2016-01-01

    Although neoplastic cells exhibit relatively higher sensitivity to hyperthermia than normal cells, hyperthermia has had variable success as an anti-cancer therapy. This variable outcome might be due to the fact that cancer cells themselves have differential degrees of sensitivity to high temperature. We hypothesized that the varying sensitivity of colorectal cancer (CRC) cells to hyperthermia depends upon the differential induction of survival pathways. Screening of such pathways revealed that Extracellular Signal-Regulated Kinase (ERK) signaling is augmented by hyperthermia, and the extent of this modulation correlates with the mutation status of V-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog (KRAS). Through clonal growth assays, apoptotic analyses and transcription reporter assays of CRC cells that differ only in KRAS mutation status we established that mutant KRAS cells are more sensitive to hyperthermia, as they exhibit sustained ERK signaling hyperactivation and increased Wingless/Integrated (WNT)/beta-catenin signaling. We propose that whereas increased levels of WNT and ERK signaling and a positive feedback between the two pathways is a major obstacle in anti-cancer therapy today, under hyperthermia the hyperinduction of the pathways and their positive crosstalk contribute to CRC cell death. Ascertaining the causative association between types of mutations and hyperthermia sensitivity may allow for a mutation profile-guided application of hyperthermia as an anti-cancer therapy. Since KRAS and WNT signaling mutations are prevalent in CRC, our results suggest that hyperthermia-based therapy might benefit a significant number, but not all, CRC patients. PMID:27187477

  10. Undariopsis peterseniana Promotes Hair Growth by the Activation of Wnt/β-Catenin and ERK Pathways

    PubMed Central

    Kang, Jung-Il; Kim, Min-Kyoung; Lee, Ji-Hyeok; Jeon, You-Jin; Hwang, Eun-Kyoung; Koh, Young-Sang; Hyun, Jin-Won; Kwon, Soon-Young; Yoo, Eun-Sook; Kang, Hee-Kyoung

    2017-01-01

    In this study, we investigated the effect and mechanism of Undariopsis peterseniana, an edible brown alga, on hair growth. The treatment of vibrissa follicles with U. peterseniana extract ex vivo for 21 days significantly increased the hair-fiber lengths. The U. peterseniana extract also significantly accelerated anagen initiation in vivo. Moreover, we found that U. peterseniana extract was able to open the KATP channel, which may contribute to increased hair growth. The U. peterseniana extract decreased 5α-reductase activity and markedly increased the proliferation of dermal papilla cells, a central regulator of the hair cycle. The U. peterseniana extract increased the levels of cell cycle proteins, such as Cyclin D1, phospho(ser780)-pRB, Cyclin E, phospho-CDK2, and CDK2. The U. peterseniana extract also increased the phosphorylation of ERK and the levels of Wnt/β-catenin signaling proteins such as glycogen synthase kinase-3β (GSK-3β) and β-catenin. These results suggested that the U. peterseniana extract had the potential to influence hair growth by dermal papilla cells proliferation through the activation of the Wnt/β-catenin and ERK pathways. We isolated a principal of the U. peterseniana extract, which was subsequently identified as apo-9′-fucoxanthinone, a trichogenic compound. The results suggested that U. peterseniana extract may have a pivotal role in the treatment of alopecia. PMID:28475144

  11. LGR5 regulates pro-survival MEK/ERK and proliferative Wnt/β-catenin signalling in neuroblastoma

    PubMed Central

    Melegh, Zsombor; Greenhough, Alexander; Malik, Sally; Szemes, Marianna; Park, Ji Hyun; Kaidi, Abderrahmane; Zhou, Li; Catchpoole, Daniel; Morgan, Rhys; Bates, David O.; Gabb, Peter J.; Malik, Karim

    2015-01-01

    LGR5 is a marker of normal and cancer stem cells in various tissues where it functions as a receptor for R-spondins and increases canonical Wnt signalling amplitude. Here we report that LGR5 is also highly expressed in a subset of high grade neuroblastomas. Neuroblastoma is a clinically heterogenous paediatric cancer comprising a high proportion of poor prognosis cases (~40%) which are frequently lethal. Unlike many cancers, Wnt pathway mutations are not apparent in neuroblastoma, although previous microarray analyses have implicated deregulated Wnt signalling in high-risk neuroblastoma. We demonstrate that LGR5 facilitates high Wnt signalling in neuroblastoma cell lines treated with Wnt3a and R-spondins, with SK-N-BE(2)-C, SK-N-NAS and SH-SY5Y cell-lines all displaying strong Wnt induction. These lines represent MYCN-amplified, NRAS and ALK mutant neuroblastoma subtypes respectively. Wnt3a/R-Spondin treatment also promoted nuclear translocation of β-catenin, increased proliferation and activation of Wnt target genes. Strikingly, short-interfering RNA mediated knockdown of LGR5 induces dramatic Wnt-independent apoptosis in all three cell-lines, accompanied by greatly diminished phosphorylation of mitogen/extracellular signal-regulated kinases (MEK1/2) and extracellular signal-regulated kinases (ERK1/2), and an increase of BimEL, an apoptosis facilitator downstream of ERK. Akt signalling is also decreased by a Rictor dependent, PDK1-independent mechanism. LGR5 expression is cell cycle regulated and LGR5 depletion triggers G1 cell-cycle arrest, increased p27 and decreased phosphorylated retinoblastoma protein. Our study therefore characterises new cancer-associated pathways regulated by LGR5, and suggest that targeting of LGR5 may be of therapeutic benefit for neuroblastomas with diverse etiologies, as well as other cancers expressing high LGR5. PMID:26517508

  12. LGR5 regulates pro-survival MEK/ERK and proliferative Wnt/β-catenin signalling in neuroblastoma.

    PubMed

    Vieira, Gabriella Cunha; Chockalingam, S; Melegh, Zsombor; Greenhough, Alexander; Malik, Sally; Szemes, Marianna; Park, Ji Hyun; Kaidi, Abderrahmane; Zhou, Li; Catchpoole, Daniel; Morgan, Rhys; Bates, David O; Gabb, Peter David; Malik, Karim

    2015-11-24

    LGR5 is a marker of normal and cancer stem cells in various tissues where it functions as a receptor for R-spondins and increases canonical Wnt signalling amplitude. Here we report that LGR5 is also highly expressed in a subset of high grade neuroblastomas. Neuroblastoma is a clinically heterogenous paediatric cancer comprising a high proportion of poor prognosis cases (~40%) which are frequently lethal. Unlike many cancers, Wnt pathway mutations are not apparent in neuroblastoma, although previous microarray analyses have implicated deregulated Wnt signalling in high-risk neuroblastoma. We demonstrate that LGR5 facilitates high Wnt signalling in neuroblastoma cell lines treated with Wnt3a and R-spondins, with SK-N-BE(2)-C, SK-N-NAS and SH-SY5Y cell-lines all displaying strong Wnt induction. These lines represent MYCN-amplified, NRAS and ALK mutant neuroblastoma subtypes respectively. Wnt3a/R-Spondin treatment also promoted nuclear translocation of β-catenin, increased proliferation and activation of Wnt target genes. Strikingly, short-interfering RNA mediated knockdown of LGR5 induces dramatic Wnt-independent apoptosis in all three cell-lines, accompanied by greatly diminished phosphorylation of mitogen/extracellular signal-regulated kinases (MEK1/2) and extracellular signal-regulated kinases (ERK1/2), and an increase of BimEL, an apoptosis facilitator downstream of ERK. Akt signalling is also decreased by a Rictor dependent, PDK1-independent mechanism. LGR5 expression is cell cycle regulated and LGR5 depletion triggers G1 cell-cycle arrest, increased p27 and decreased phosphorylated retinoblastoma protein. Our study therefore characterises new cancer-associated pathways regulated by LGR5, and suggest that targeting of LGR5 may be of therapeutic benefit for neuroblastomas with diverse etiologies, as well as other cancers expressing high LGR5.

  13. Sequential antagonism of early and late Wnt-signaling by zebrafish colgate promotes dorsal and anterior fates.

    PubMed

    Nambiar, Roopa M; Henion, Paul D

    2004-03-01

    The establishment of the vertebrate body plan involves patterning of the ectoderm, mesoderm, and endoderm along the dorsoventral and antero-posterior axes. Interactions among numerous signaling molecules from several multigene families, including Wnts, have been implicated in regulating these processes. Here we provide evidence that the zebrafish colgate(b382) (col) mutation results in increased Wnt signaling that leads to defects in dorsal and anterior development. col mutants display early defects in dorsoventral patterning manifested by a decrease in the expression of dorsal shield-specific markers and ectopic expression of ventrolaterally expressed genes during gastrulation. In addition to these early patterning defects, col mutants display a striking regional posteriorization within the neuroectoderm, resulting in a reduction in anterior fates and an expansion of posterior fates within the forebrain and midbrain-hindbrain regions. We are able to correlate these phenotypes to the overactivation of Wnt signaling in col mutants. The early dorsal and anterior patterning phenotypes of the col mutant embryos are selectively rescued by inactivation of Wnt8 function by morpholino translational interference. In contrast, the regionalized neuroectoderm posterioriorization phenotype is selectively rescued by morpholino-mediated inactivation of Wnt8b. These results suggest that col-mediated antagonism of early and late Wnt-signaling activity during gastrulation is normally required sequentially for both early dorsoventral patterning and the specification and patterning of regional fates within the anterior neuroectoderm.

  14. Tumor suppressor Fbxw7 antagonizes WNT signaling by targeting β-catenin for degradation in pancreatic cancer.

    PubMed

    Jiang, Jian-Xin; Sun, Cheng-Yi; Tian, She; Yu, Chao; Chen, Mei-Yuan; Zhang, Hao

    2016-10-01

    Pancreatic cancer is one of the deadliest solid malignancies associated with aberrant Wnt signaling activation. Fbxw7 mutations have been implicated in the development of pancreatic cancer, whereas the exact mechanism of this ubiquitin ligase as a tumor suppressor remains unclear in pancreatic carcinogenesis. Here, we describe that Fbxw7 is downregulated upon pancreatic cancer development. Depletion of Fbxw7 results in tumor suppression in pancreatic cancer cells, while Fbxw7 overexpression inhibits pancreatic cancer cell proliferation and invasion. Considering the negative correlation between Fbxw7 and β-catenin, we find that Fbxw7 antagonizes Wnt signaling through targeting β-catenin for its degradation. Moreover, the inhibitory effect of Fbxw7 on pancreatic cancer cell proliferation is mainly executed by the destruction of the Wnt/β-catenin signaling pathway. We also reveal that c-myc, a widely accepted target of Fbxw7, is also transcriptionally regulated by the Fbxw7/β-catenin axis in pancreatic cancer cells. Collectively, our results demonstrate that Fbxw7 is a novel regulator of Wnt/β-catenin signaling-dependent regulation of pancreatic cancer cell growth and invasion, and inactivation of Fbxw7 in pancreatic cancer tissues might be the reason for the aberrant activation of Wnt signaling.

  15. Antagonism of Muscarinic Acetylcholine Receptors Alters Synaptic ERK Phosphorylation in the Rat Forebrain.

    PubMed

    Mao, Li-Min; Wang, Henry H; Wang, John Q

    2016-12-28

    Acetylcholine (ACh) is a key transmitter in the mesocorticolimbic circuit. By interacting with muscarinic ACh receptors (mAChR) enriched in the circuit, ACh actively regulates various neuronal and synaptic activities. The extracellular signal-regulated kinase (ERK) is one of members of the mitogen-activated protein kinase family and is subject to the regulation by dopamine receptors, although the regulation of ERKs by limbic mAChRs is poorly understood. In this study, we investigated the role of mAChRs in the regulation of ERK phosphorylation (activation) in the mesocorticolimbic system of adult rat brains in vivo. We targeted a sub-pool of ERKs at synaptic sites. We found that a systemic injection of the mAChR antagonist scopolamine increased phosphorylation of synaptic ERKs in the striatum (caudate putamen and nucleus accumbens) and medial prefrontal cortex (mPFC). Increases in ERK phosphorylation in both forebrain regions were rapid and transient. Notably, pretreatment with a dopamine D1 receptor (D1R) antagonist SCH23390 blocked the scopolamine-stimulated ERK phosphorylation in these brain regions, while a dopamine D2 receptor antagonist eticlopride did not. Scopolamine and SCH23390 did not change the amount of total ERK proteins. These results demonstrate that mAChRs inhibit synaptic ERK phosphorylation in striatal and mPFC neurons under normal conditions. Blockade of this inhibitory mAChR tone leads to the upregulation of ERK phosphorylation likely through a mechanism involving the level of D1R activity.

  16. Palmitate Antagonizes Wnt/Beta-catenin Signaling in 3T3-L1 Pre-adipocytes

    USDA-ARS?s Scientific Manuscript database

    Long chain saturated free fatty acids such as palmitate (PA) produce insulin resistance, endoplasmic reticulum stress, and apoptosis in mature adipocytes and pre-adipocytes. In pre-adipocytes, saturated free fatty acids also promote adipogenic induction in the presence of adipogenic hormones. Wnt/be...

  17. Wnt5a-Dopamine D2 Receptor Interactions Regulate Dopamine Neuron Development via Extracellular Signal-regulated Kinase (ERK) Activation*

    PubMed Central

    Yoon, Sehyoun; Choi, Mi-hyun; Chang, Min Seok; Baik, Ja-Hyun

    2011-01-01

    The dopamine D2 receptor (D2R) plays an important role in mesencephalic dopaminergic neuronal development, particularly coupled with extracellular signal-regulated kinase (ERK) activation. Wnt5a protein is known to regulate the development of dopaminergic neurons. We analyzed the effect of Wnt5a on dopaminergic neuron development in mesencephalic primary cultures from wild-type (WT) and D2R knock-out (D2R−/−) mice. Treatment with Wnt5a increased the number and neuritic length of dopamine neurons in primary mesencephalic neuronal cultures from WT mice, but not from D2R−/− mice. The effect of Wnt5a was completely blocked by treatment with D2R antagonist or inhibitors of MAPK or EGFR. Wnt5a-mediated ERK activation in mesencephalic neuronal cultures was inhibited by treatment of D2R antagonist and EGFR inhibitors in WT mice. However, these regulations were not observed for D2R−/− mice. Co-immunoprecipitation and displacement of [3H]spiperone from D2R by Wnt5a demonstrated that Wnt5a could bind with D2R. This interaction was confirmed by GST pulldown assays demonstrating that the domain including transmembrane domain 4, second extracellular loop, and transmembrane domain 5 of D2R binds to Wnt5a. These results suggest that the interaction between D2R and Wnt5a has an important role in dopamine neuron development in association with EGFR and the ERK pathway. PMID:21454669

  18. Heterotrimeric G protein-dependent WNT-5A signaling to ERK1/2 mediates distinct aspects of microglia proinflammatory transformation

    PubMed Central

    2012-01-01

    Background WNT-5A signaling in the central nervous system is important for morphogenesis, neurogenesis and establishment of functional connectivity; the source of WNT-5A and its importance for cellular communication in the adult brain, however, are mainly unknown. We have previously investigated the inflammatory effects of WNT/β-catenin signaling in microglia in Alzheimer's disease. WNT-5A, however, generally recruits β-catenin-independent signaling. Thus, we aim here to characterize the role of WNT-5A and downstream signaling pathways for the inflammatory transformation of the brain's macrophages, the microglia. Methods Mouse brain sections were used for immunohistochemistry. Primary isolated microglia and astrocytes were employed to characterize the WNT-induced inflammatory transformation and underlying intracellular signaling pathways by immunoblotting, quantitative mRNA analysis, proliferation and invasion assays. Further, measurements of G protein activation by [γ-35 S]GTP binding, examination of calcium fluxes and cyclic AMP production were used to define intracellular signaling pathways. Results Astrocytes in the adult mouse brain express high levels of WNT-5A, which could serve as a novel astroglia-microglia communication pathway. The WNT-5A-induced proinflammatory microglia response is characterized by increased expression of inducible nitric oxide synthase, cyclooxygenase-2, cytokines, chemokines, enhanced invasive capacity and proliferation. Mapping of intracellular transduction pathways reveals that WNT-5A activates heterotrimeric Gi/o proteins to reduce cyclic AMP levels and to activate a Gi/o protein/phospholipase C/calcium-dependent protein kinase/extracellular signal-regulated kinase 1/2 (ERK1/2) axis. We show further that WNT-5A-induced ERK1/2 signaling is responsible for distinct aspects of the proinflammatory transformation, such as matrix metalloprotease 9/13 expression, invasion and proliferation. Conclusions Thus, WNT-5A-induced and G

  19. ZnRF3 Induces Apoptosis of Gastric Cancer Cells by Antagonizing Wnt and Hedgehog Signaling.

    PubMed

    Qin, Hongzhen; Cai, Aizhen; Xi, Hongqing; Yuan, Jing; Chen, Lin

    2015-11-01

    A large proportion of malignant cancers of the stomach are gastric adenocarcinoma type. In spite of many studies, the molecular basis for this cancer is still unclear. Deregulated cell proliferative signaling via Wnt/β-catenin and Hedgehog pathways is considered important in the pathogenesis of many cancers including the gastric cancer. Recent studies identified ZnRF3 protein, which is a E3-ubiquitin ligase and which is either deleted or mutated in cancers, to inhibit Wnt signaling. However, the significance of ZnRF3 in the control of gastric cancer and whether it also regulates Hedgehog signaling pathway, is not known. In the present study, we assessed the expression of ZnRF3 in gastric tumors and paracancerous tissues from 58 patients (44 male and 14 female) of different ages and related this to patient survival. We observed a clear relationship between ZnRF3 expression in paracancerous tissue and tumor size. Also, ZnRF3 expression was much higher in tumors from aged patients. Male patients showed higher mortality than the females. Mechanistic studies using normal gastric cells (GES1) and gastric cancer cells (MGC-803) infected with either AdZnRF3 or AdGFP viral vectors, revealed that ZnRF3 overexpression causes significantly more apoptosis and lowered proliferation of cancer cells. ZnRF3 overexpression led to greatly reduced levels of Lgr5, a component of Wnt signaling and also Gli1, a component of Hedgehog signaling. Thus, ZnRF3 negatively influences both the Wnt and Hedgehog proliferative pathways, and probably this way it negatively regulates cancer progression. These results suggest the importance of normal ZnRF3 function in checking the progression of cancer cell growth and indicate that a lack of this protein can lead to poorer clinical outcomes for gastric cancer patients.

  20. Activation of Wnt/β-Catenin in Ewing Sarcoma Cells Antagonizes EWS/ETS Function and Promotes Phenotypic Transition to More Metastatic Cell States.

    PubMed

    Pedersen, Elisabeth A; Menon, Rajasree; Bailey, Kelly M; Thomas, Dafydd G; Van Noord, Raelene A; Tran, Jenny; Wang, Hongwei; Qu, Ping Ping; Hoering, Antje; Fearon, Eric R; Chugh, Rashmi; Lawlor, Elizabeth R

    2016-09-01

    Ewing sarcomas are characterized by the presence of EWS/ETS fusion genes in the absence of other recurrent genetic alterations and mechanisms of tumor heterogeneity that contribute to disease progression remain unclear. Mutations in the Wnt/β-catenin pathway are rare in Ewing sarcoma but the Wnt pathway modulator LGR5 is often highly expressed, suggesting a potential role for the axis in tumor pathogenesis. We evaluated β-catenin and LGR5 expression in Ewing sarcoma cell lines and tumors and noted marked intra- and inter-tumor heterogeneity. Tumors with evidence of active Wnt/β-catenin signaling were associated with increased incidence of tumor relapse and worse overall survival. Paradoxically, RNA sequencing revealed a marked antagonism of EWS/ETS transcriptional activity in Wnt/β-catenin-activated tumor cells. Consistent with this, Wnt/β-catenin-activated cells displayed a phenotype that was reminiscent of Ewing sarcoma cells with partial EWS/ETS loss of function. Specifically, activation of Wnt/β-catenin induced alterations to the actin cytoskeleton, acquisition of a migratory phenotype, and upregulation of EWS/ETS-repressed genes. Notably, activation of Wnt/β-catenin signaling led to marked induction of tenascin C (TNC), an established promoter of cancer metastasis, and an EWS/ETS-repressed target gene. Loss of TNC function in Ewing sarcoma cells profoundly inhibited their migratory and metastatic potential. Our studies reveal that heterogeneous activation of Wnt/β-catenin signaling in subpopulations of tumor cells contributes to phenotypic heterogeneity and disease progression in Ewing sarcoma. Significantly, this is mediated, at least in part, by inhibition of EWS/ETS fusion protein function that results in derepression of metastasis-associated gene programs. Cancer Res; 76(17); 5040-53. ©2016 AACR. ©2016 American Association for Cancer Research.

  1. The Androgen Receptor Antagonizes Wnt/β-Catenin Signaling in Epidermal Stem Cells

    PubMed Central

    Kretzschmar, Kai; Cottle, Denny L; Schweiger, Pawel J; Watt, Fiona M

    2015-01-01

    Activation of Wnt/β-catenin signaling in adult mouse epidermis leads to expansion of the stem cell compartment and redirects keratinocytes in the interfollicular epidermis and sebaceous glands (SGs) to differentiate along the hair follicle (HF) lineages. Here we demonstrate that during epidermal development and homeostasis there is reciprocal activation of the androgen receptor (AR) and β-catenin in cells of the HF bulb. AR activation reduced β-catenin-dependent transcription, blocked β-catenin-induced induction of HF growth, and prevented β-catenin-mediated conversion of SGs into HFs. Conversely, AR inhibition enhanced the effects of β-catenin activation, promoting HF proliferation and differentiation, culminating in the formation of benign HF tumors and a complete loss of SG identity. We conclude that AR signaling has a key role in epidermal stem cell fate selection by modulating responses to β-catenin in adult mouse skin. PMID:26121213

  2. Antagonizing miR-218-5p attenuates Wnt signaling and reduces metastatic bone disease of triple negative breast cancer cells

    PubMed Central

    Taipaleenmäki, Hanna; Farina, Nicholas H.; van Wijnen, Andre J.; Stein, Janet L.

    2016-01-01

    Wnt signaling is implicated in bone formation and activated in breast cancer cells promoting primary and metastatic tumor growth. A compelling question is whether osteogenic miRNAs that increase Wnt activity for bone formation are aberrantly expressed in breast tumor cells to support metastatic bone disease. Here we report that miR-218-5p is highly expressed in bone metastases from breast cancer patients, but is not detected in normal mammary epithelial cells. Furthermore, inhibition of miR-218-5p impaired the growth of bone metastatic MDA-MB-231 cells in the bone microenvironment in vivo. These findings indicate a positive role for miR-218-5p in bone metastasis. Bioinformatic and biochemical analyses revealed a positive correlation between aberrant miR-218-5p expression and activation of Wnt signaling in breast cancer cells. Mechanistically, miR-218-5p targets the Wnt inhibitors Sclerostin (SOST) and sFRP-2, which highly enhances Wnt signaling. In contrast, delivery of antimiR-218-5p decreased Wnt activity and the expression of metastasis-related genes, including bone sialoprotein (BSP/IBSP), osteopontin (OPN/SPP1) and CXCR-4, implicating a Wnt/miR-218-5p regulatory network in bone metastatic breast cancer. Furthermore, miR-218-5p also mediates the Wnt-dependent up-regulation of PTHrP, a key cytokine promoting cancer-induced osteolysis. Antagonizing miR-218-5p reduced the expression of PTHrP and Rankl, inhibited osteoclast differentiation in vitro and in vivo, and prevented the development of osteolytic lesions in a preclinical metastasis model. We conclude that pathological elevation of miR-218-5p in breast cancer cells activates Wnt signaling to enhance metastatic properties of breast cancer cells and cancer-induced osteolytic disease, suggesting that miR-218-5p could be an attractive therapeutic target for preventing disease progression. PMID:27738322

  3. Receptor for Activated C Kinase 1 (RACK1) Promotes Dishevelled Protein Degradation via Autophagy and Antagonizes Wnt Signaling.

    PubMed

    Cheng, Minzhang; Xue, Hua; Cao, Weipeng; Li, Wenxia; Chen, Hua; Liu, Bofeng; Ma, Benyu; Yan, Xiaohua; Chen, Ye-Guang

    2016-06-10

    Wnt signaling plays a critical role in embryonic development, tissue homeostasis, and cancer development. Dishevelled (Dvl) is an essential and central component in Wnt signaling, and its stability and activity is tightly regulated. It has been shown that Dvl can be degraded via both the proteasome and autophagy-lysosome pathways. Here we report that receptor for activated C kinase 1 (RACK1) negatively regulates Dishevelled stability and Wnt signaling. RACK1 interacts with Dvl proteins and promotes their lysosomal degradation, and this effect is enhanced by autophagy induction. RACK1 also interacts with LC3 and enhances the association of LC3 with Dvl2, thereby leading to degradation of Dvl proteins through autophagy. These findings reveal a novel regulatory function of RACK1 in Wnt signaling by modulating Dvl stability. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  4. 4'-Demethylnobiletin, a bioactive metabolite of nobiletin enhancing PKA/ERK/CREB signaling, rescues learning impairment associated with NMDA receptor antagonism via stimulation of the ERK cascade.

    PubMed

    Al Rahim, Md; Nakajima, Akira; Saigusa, Daisuke; Tetsu, Naomi; Maruyama, Yuji; Shibuya, Masatoshi; Yamakoshi, Hiroyuki; Tomioka, Yoshihisa; Iwabuchi, Yoshiharu; Ohizumi, Yasushi; Yamakuni, Tohru

    2009-08-18

    The biochemical and pharmacological activities of nobiletin, including neurotrophic and memory-enhancing action, in both in vitro and in vivo systems are well established. However, whether its metabolites do have such beneficial effects like nobiletin remains to be examined. Here we, for the first time, report that 2-(4-hydroxy-3-methoxyphenyl)-5,6,7,8-tetramethoxychromen-4-one (4'-demethylnobiletin), a major metabolite of nobiletin identified in the urine of rats and mice, stimulates the phosphorylation of ERK and CREB and enhances CRE-mediated transcription by activating a PKA/MEK/ERK pathway, like nobiletin, in cultured hippocampal neurons. Since NMDA receptor-mediated ERK signaling is involved in memory processing, including associative memories, we also examined whether 4'-demethylnobiletin, by activating ERK signaling, could restore learning impairment. Chronic intraperitoneal (ip) treatment of the mice with 10 or 50 mg of 4'-demethylnobiletin/kg rescued the NMDA receptor antagonist MK-801-induced learning impairment, accompanied by improvement of the MK-801-induced decrease in the level of ERK phosphorylation in the hippocampus of the animals. Consistently, 4'-demethylnobiletin also restored MK-801-induced inhibition of NMDA-stimulated phosphorylation of not only ERK but also PKA substrates in cultured rat hippocampal neurons. Moreover, we actually detected 4'-demethylnobiletin in the brain of mice following acute ip administration, demonstrating that the metabolite can cross the blood-brain barrier to reach the brain and thereby exert its effects to reverse learning impairment. Therefore, these results suggest that 4'-demethylnobiletin, a bioactive metabolite of nobiletin, may serve as a potential therapeutic agent, at least, for memory disorders associated with a dysregulated NMDA receptor ERK signaling, like nobiletin.

  5. The human HECA interacts with cyclins and CDKs to antagonize Wnt-mediated proliferation and chemoresistance of head and neck cancer cells

    SciTech Connect

    Dowejko, Albert; Bauer, Richard; Bauer, Karin; Mueller-Richter, Urs D.A.; Reichert, Torsten E.

    2012-03-10

    antagonizes Wnt-mediated cell proliferation through interaction with major cell cycle factors. Black-Right-Pointing-Pointer Modulating HECA level confers benefits for engaging tumor cells with cisplatin.

  6. Novel Wnt Regulator NEL-Like Molecule-1 Antagonizes Adipogenesis and Augments Osteogenesis Induced by Bone Morphogenetic Protein 2

    PubMed Central

    Shen, Jia; James, Aaron W.; Zhang, Xinli; Pang, Shen; Zara, Janette N.; Asatrian, Greg; Chiang, Michael; Lee, Min; Khadarian, Kevork; Nguyen, Alan; Lee, Kevin S.; Siu, Ronald K.; Tetradis, Sotirios; Ting, Kang; Soo, Chia

    2017-01-01

    The differentiation factor NEL-like molecule-1 (NELL-1) has been reported as osteoinductive in multiple in vivo preclinical models. Bone morphogenetic protein (BMP)-2 is used clinically for skeletal repair, but in vivo administration can induce abnormal, adipose-filled, poor-quality bone. We demonstrate that NELL-1 combined with BMP2 significantly optimizes osteogenesis in a rodent femoral segmental defect model by minimizing the formation of BMP2-induced adipose-filled cystlike bone. In vitro studies using the mouse bone marrow stromal cell line M2-10B4 and human primary bone marrow stromal cells have confirmed that NELL-1 enhances BMP2-induced osteogenesis and inhibits BMP2-induced adipogenesis. Importantly, the ability of NELL-1 to direct BMP2-treated cells toward osteogenesis and away from adipogenesis requires intact canonical Wnt signaling. Overall, these studies establish the feasibility of combining NELL-1 with BMP2 to improve clinical bone regeneration and provide mechanistic insight into canonical Wnt pathway activity during NELL-1 and BMP2 osteogenesis. The novel abilities of NELL-1 to stimulate Wnt signaling and to repress adipogenesis may highlight new treatment approaches for bone loss in osteoporosis. PMID:26772960

  7. Akt Phosphorylates Wnt Coactivator and Chromatin Effector Pygo2 at Serine 48 to Antagonize Its Ubiquitin/Proteasome-mediated Degradation*

    PubMed Central

    Li, Qiuling; Li, Yuewei; Gu, Bingnan; Fang, Lei; Zhou, Pengbo; Bao, Shilai; Huang, Lan; Dai, Xing

    2015-01-01

    Pygopus 2 (Pygo2/PYGO2) is an evolutionarily conserved coactivator and chromatin effector in the Wnt/β-catenin signaling pathway that regulates cell growth and differentiation in various normal and malignant tissues. Although PYGO2 is highly overexpressed in a number of human cancers, the molecular mechanism underlying its deregulation is largely unknown. Here we report that Pygo2 protein is degraded through the ubiquitin/proteasome pathway and is posttranslationally stabilized through phosphorylation by activated phosphatidylinositol 3-kinase/Akt signaling. Specifically, Pygo2 is stabilized upon inhibition of the proteasome, and its intracellular level is regulated by Cullin 4 (Cul4) and DNA damage-binding protein 1 (DDB1), components of the Cul4-DDB1 E3 ubiquitin ligase complex. Furthermore, Pygo2 is phosphorylated at multiple residues, and Akt-mediated phosphorylation at serine 48 leads to its decreased ubiquitylation and increased stability. Finally, we provide evidence that Akt and its upstream growth factors act in parallel with Wnt to stabilize Pygo2. Taken together, our findings highlight chromatin regulator Pygo2 as a common node downstream of oncogenic Wnt and Akt signaling pathways and underscore posttranslational modification, particularly phosphorylation and ubiquitylation, as a significant mode of regulation of Pygo2 protein expression. PMID:26170450

  8. NDRG2 induced by oxidized LDL in macrophages antagonizes growth factor productions via selectively inhibiting ERK activation.

    PubMed

    Liu, Shumei; Yang, Pengyuan; Kang, Hui; Lu, Ling; Zhang, Yuefan; Pan, Jingwei; Rui, Yao-Cheng

    2010-02-01

    During atherogenesis, macrophage foam cells produce prodigious growth factors, cytokines, and chemokines, which play the central roles in inflammatory process in atherosclerotic plaque formation. In the present study, we identified a new protein marker, N-Myc downstream-regulated protein 2 (NDRG2), which is significantly up-regulated in oxidized low density lipoprotein (oxLDL) treated macrophages and in human atherosclerotic plaques. Over-expression and siRNA knockdown studies showed that NDRG2 is a negative regulator of platelet-derived growth factor (PDGF) and vascular endothelial growth factor (VEGF) productions in macrophages. Furthermore, we investigated the effects of NDRG2 on MAPK signal activation. Our results showed ERK1/2 activation, but not P38 or JNK1/2 activation, is responsible for regulation of NDRG2 on VEGF and PDGF productions. Consistent with the PDGF levels, the vascular smooth muscle cell (VSMC) proliferation was also regulated by the conditional medium of the oxLDL treated macrophages with NDRG2 knockdown or over-expression. Neutralizing anti-PDGF antibody can significantly inhibit the enhanced VSMC proliferation by macrophage medium with NDRG2 knockdown. Our present results demonstrate that NDRG2 participates in oxLDL-induced macrophage activation and modulates ERK1/2-dependent PDGF and VEGF production, which has potential application in atherogenesis. 2009 Elsevier B.V. All rights reserved.

  9. Enhancement of osteogenic differentiation of rat adipose tissue-derived mesenchymal stem cells by zinc sulphate under electromagnetic field via the PKA, ERK1/2 and Wnt/β-catenin signaling pathways

    PubMed Central

    Fathi, Ezzatollah; Farahzadi, Raheleh

    2017-01-01

    Zinc ion as an essential trace element and electromagnetic fields (EMFs) has been reported to be involved in the regulation of bone metabolism. The aim of this study was to elucidate the effects of zinc sulphate (ZnSO4) on the osteogenic differentiation of adipose tissue-derived mesenchymal stem cells (ADSCs) in the presence of EMF as a strategy in osteoporosis therapy. Alkaline phophatase (ALP) activity measurement, calcium assay and expression of several osteoblastic marker genes were examined to assess the effect of ZnSO4 on the osteogenic differentiation of ADSCs under EMF. The expression of cAMP and PKA was evaluated by ELISA. The expression of β-catenin, Wnt1, Wnt3a, low-density lipoprotein receptor-related protein 5 (LRP5) and reduced dickkopf1 (DKK1) genes were used to detect the Wnt/β-catenin pathway. It was found that ZnSO4, in the presence of EMF, resulted in an increase in the expression of osteogenic genes, ALP activity and calcium levels. EMF, in the presence of ZnSO4, increased the cAMP level and protein kinase A (PKA) activity. Treatment of ADSCs with (MAPK)/ERK kinase 1/2 inhibitor, or PKA inhibitor, significantly inhibited the promotion of osteogenic markers, indicating that the induction of osteogenesis was dependent on the ERK and PKA signaling pathways. Real-time PCR analysis showed that ZnSO4, in the presence of EMF, increased the mRNA expressions of β-catenin, Wnt1, Wnt3a, LRP5 and DKK1. In this study, it was shown that 0.432 μg/ml ZnSO4, in the presence of 50 Hz, 20 mT EMF, induced the osteogenic differentiation of ADSCs via PKA, ERK1/2 and Wnt/β-catenin signaling pathways. PMID:28339498

  10. Enhancement of osteogenic differentiation of rat adipose tissue-derived mesenchymal stem cells by zinc sulphate under electromagnetic field via the PKA, ERK1/2 and Wnt/β-catenin signaling pathways.

    PubMed

    Fathi, Ezzatollah; Farahzadi, Raheleh

    2017-01-01

    Zinc ion as an essential trace element and electromagnetic fields (EMFs) has been reported to be involved in the regulation of bone metabolism. The aim of this study was to elucidate the effects of zinc sulphate (ZnSO4) on the osteogenic differentiation of adipose tissue-derived mesenchymal stem cells (ADSCs) in the presence of EMF as a strategy in osteoporosis therapy. Alkaline phophatase (ALP) activity measurement, calcium assay and expression of several osteoblastic marker genes were examined to assess the effect of ZnSO4 on the osteogenic differentiation of ADSCs under EMF. The expression of cAMP and PKA was evaluated by ELISA. The expression of β-catenin, Wnt1, Wnt3a, low-density lipoprotein receptor-related protein 5 (LRP5) and reduced dickkopf1 (DKK1) genes were used to detect the Wnt/β-catenin pathway. It was found that ZnSO4, in the presence of EMF, resulted in an increase in the expression of osteogenic genes, ALP activity and calcium levels. EMF, in the presence of ZnSO4, increased the cAMP level and protein kinase A (PKA) activity. Treatment of ADSCs with (MAPK)/ERK kinase 1/2 inhibitor, or PKA inhibitor, significantly inhibited the promotion of osteogenic markers, indicating that the induction of osteogenesis was dependent on the ERK and PKA signaling pathways. Real-time PCR analysis showed that ZnSO4, in the presence of EMF, increased the mRNA expressions of β-catenin, Wnt1, Wnt3a, LRP5 and DKK1. In this study, it was shown that 0.432 μg/ml ZnSO4, in the presence of 50 Hz, 20 mT EMF, induced the osteogenic differentiation of ADSCs via PKA, ERK1/2 and Wnt/β-catenin signaling pathways.

  11. Wnt signaling in cancer

    PubMed Central

    Zhan, T; Rindtorff, N; Boutros, M

    2017-01-01

    Wnt signaling is one of the key cascades regulating development and stemness, and has also been tightly associated with cancer. The role of Wnt signaling in carcinogenesis has most prominently been described for colorectal cancer, but aberrant Wnt signaling is observed in many more cancer entities. Here, we review current insights into novel components of Wnt pathways and describe their impact on cancer development. Furthermore, we highlight expanding functions of Wnt signaling for both solid and liquid tumors. We also describe current findings how Wnt signaling affects maintenance of cancer stem cells, metastasis and immune control. Finally, we provide an overview of current strategies to antagonize Wnt signaling in cancer and challenges that are associated with such approaches. PMID:27617575

  12. Secreted and Transmembrane Wnt Inhibitors and Activators

    PubMed Central

    Cruciat, Cristina-Maria; Niehrs, Christof

    2013-01-01

    Signaling by the Wnt family of secreted glycoproteins plays important roles in embryonic development and adult homeostasis. Wnt signaling is modulated by a number of evolutionarily conserved inhibitors and activators. Wnt inhibitors belong to small protein families, including sFRP, Dkk, WIF, Wise/SOST, Cerberus, IGFBP, Shisa, Waif1, APCDD1, and Tiki1. Their common feature is to antagonize Wnt signaling by preventing ligand–receptor interactions or Wnt receptor maturation. Conversely, the Wnt activators, R-spondin and Norrin, promote Wnt signaling by binding to Wnt receptors or releasing a Wnt-inhibitory step. With few exceptions, these antagonists and agonists are not pure Wnt modulators, but also affect additional signaling pathways, such as TGF-β and FGF signaling. Here we discuss their interactions with Wnt ligands and Wnt receptors, their role in developmental processes, as well as their implication in disease. PMID:23085770

  13. Wnt/β-catenin and ERK pathway activation: A possible mechanism of photobiomodulation therapy with light-emitting diodes that regulate the proliferation of human outer root sheath cells.

    PubMed

    Kim, Jung E; Woo, Young J; Sohn, Ki M; Jeong, Kwan H; Kang, Hoon

    2017-09-25

    Outer root sheath cells (ORSCs) play important roles in maintaining hair follicle structure and provide support for the bulge area. The hair growth promoting effects of photobiomodulation therapy (PBMT) have been reported, but the mechanisms for this in human ORCs (hORSCs) have rarely been studied. The aim of this study was to investigate the effect of various wavelengths of light-emitting diode (LED) irradiation on human ORSCs (hORSCs). LED irradiation effects on hORSC proliferation and migration were examined with MTT assay, BrdU incorporation assay and migration assays. hORSCs were irradiated using four LED wavelengths (415, 525, 660, and 830 nm) with different low energy levels. LED irradiation effects on the expression of molecules associated with the Wnt/β-catenin signaling and ERK pathway, hair stem cell markers, and various growth factors and cytokines in hORSCs were examined with real-time PCR and Western blot assay. The effect of the LED-irradiated hORSCs on cell proliferation of human dermal papilla cells (hDPCs) was examined with co-culture and MTT assay. PBMT with LED light variably promoted hORSC proliferation and suppressed cell apoptosis depending on energy level. LED irradiation induced Wnt5a, Axin2, and Lef1 mRNA expression and β-catenin protein expression in hORSCs. Phosphorylation of ERK, c-Jun, and p38 in hORSCs was observed after LED light irradiation, and ERK inhibitor treatment before irradiation reduced ERK and c-Jun phosphorylation. Red light-treated hORSCs showed substantial increase in IL-6, IL-8, TNF-a, IGF-1, TGF-β1, and VEGF mRNA. Light irradiation at 660 and 830 nm projected onto hORSCs accelerated in vitro migration. LED-irradiated hORSCs increased hDPCs proliferation when they were co-cultured. The conditioned medium from LED-irradiated hORSCs was sufficient to stimulate hDPCs proliferation. These results demonstrate that LED light irradiation induced hORSC proliferation and migration and inhibited apoptosis in vitro. The

  14. E7449: A dual inhibitor of PARP1/2 and tankyrase1/2 inhibits growth of DNA repair deficient tumors and antagonizes Wnt signaling

    PubMed Central

    Wu, Jiayi; Chang, Paul; Kolber-Simonds, Donna; Ackermann, Karen; Twine, Natalie C.; Shie, Jue-Lon; Miu, Jingzang Tao; Huang, Kuan-Chun; Moniz, George A.; Nomoto, Kenichi

    2015-01-01

    Inhibition of Poly(ADP-ribose) Polymerase1 (PARP1) impairs DNA damage repair, and early generation PARP1/2 inhibitors (olaparib, niraparib, etc.) have demonstrated clinical proof of concept for cancer treatment. Here, we describe the development of the novel PARP inhibitor E7449, a potent PARP1/2 inhibitor that also inhibits PARP5a/5b, otherwise known as tankyrase1 and 2 (TNKS1 and 2), important regulators of canonical Wnt/β-catenin signaling. E7449 inhibits PARP enzymatic activity and additionally traps PARP1 onto damaged DNA; a mechanism previously shown to augment cytotoxicity. Cells deficient in DNA repair pathways beyond homologous recombination were sensitive to E7449 treatment. Chemotherapy was potentiated by E7449 and single agent had significant antitumor activity in BRCA-deficient xenografts. Additionally, E7449 inhibited Wnt/β-catenin signaling in colon cancer cell lines, likely through TNKS inhibition. Consistent with this possibility, E7449 stabilized axin and TNKS proteins resulting in β-catenin de-stabilization and significantly altered expression of Wnt target genes. Notably, hair growth mediated by Wnt signaling was inhibited by E7449. A pharmacodynamic effect of E7449 on Wnt target genes was observed in tumors, although E7449 lacked single agent antitumor activity in vivo, a finding typical for selective TNKS inhibitors. E7449 antitumor activity was increased through combination with MEK inhibition. Particularly noteworthy was the lack of toxicity, most significantly the lack of intestinal toxicity reported for other TNKS inhibitors. E7449 represents a novel dual PARP1/2 and TNKS1/2 inhibitor which has the advantage of targeting Wnt/β-catenin signaling addicted tumors. E7449 is currently in early clinical development. PMID:26513298

  15. Antagonizing canonical Wnt signaling pathway by recombinant human sFRP4 purified from E. coli and its implications in cancer therapy.

    PubMed

    Ghoshal, Archita; Ghosh, Siddhartha Sankar

    2016-07-01

    The Wnt signaling pathway plays a predominant role in aberrant proliferation in myriad of cancers. In non-cancerous cells, Wnts are blocked by the secreted frizzled-related proteins (sFRPs) that are generally downregulated in cancer cells. We have purified and characterized bacterially expressed glutathione S-transferase-tagged SFRP4 from a novel clone generated from human cell origin. Cervical cancer (HeLa) and lung cancer (A549) cells, in which Wnt and associated genes were found to be expressed, were treated with the purified recombinant sFRP4, which revealed a significant dose-dependent cell growth inhibition up to 40 %. The current investigation on functionality of this bacterially produced recombinant sFRP4 in arresting cancer cell proliferation is the first of its kind, where G2/M phase arrest and early apoptosis were evident. Increase in phosphorylated β-catenin in sFRP4 treatment indicated inhibition of Wnt pathway, which was further confirmed by downregulation of pro-proliferative genes, namely cyclin D1, c-myc, and survivin. Functional activity of recombinant sFRP4 was further exploited in co-therapy module with chemotherapeutic drugs to decipher molecular events. Collectively, our study on purified recombinant sFRP4 from bacterial host holds great promise in targeting Wnt signaling for exploring new strategies to combat cancer.

  16. Fenretinide (4-HPR) Targets Caspase-9, ERK 1/2 and the Wnt3a/β-Catenin Pathway in Medulloblastoma Cells and Medulloblastoma Cell Spheroids

    PubMed Central

    Bassani, Barbara; Bartolini, Desirèe; Pagani, Arianna; Principi, Elisa; Zollo, Massimo; Noonan, Douglas M.; Albini, Adriana; Bruno, Antonino

    2016-01-01

    Medulloblastoma (MB), a neuroectodermal tumor arising in the cerebellum, represents the most frequent childhood brain malignancy. Current treatments for MB combine radiation and chemotherapy and are often associated with relevant side effects; novel therapeutic strategies are urgently needed. N-(4-Hydroxyphenyl) retinamide (4-HPR, fenretinide), a synthetic analogue of all-trans retinoic acid, has emerged as a promising and well-tolerated cancer chemopreventive and chemotherapeutic agent for various neoplasms, from breast cancer to neuroblastoma. Here we investigated the effects of 4-HPR on MB cell lines and identified the mechanism of action for a potential use in therapy of MB. Flow cytometry analysis was performed to evaluate 4-HPR induction of apoptosis and oxygen reactive species (ROS) production, as well as cell cycle effects. Functional analysis to determine 4-HPR ability to interfere with MB cell migration and invasion were performed. Western Blot analysis were used to investigate the crucial molecules involved in selected signaling pathways associated with apoptosis (caspase-9 and PARP-1), cell survival (ERK 1/2) and tumor progression (Wnt3a and β-catenin). We show that 4-HPR induces caspase 9-dependent cell death in DAOY and ONS-76 cells, associated with increased ROS generation, suggesting that free radical intermediates might be directly involved. We observed 4-HPR induction of cell cycle arrest in G1/S phase, inactivated β-catenin, and inhibition of MB cell migration and invasion. We also evaluated the ability of 4-HPR to target MB cancer-stem/cancer-initiating cells, using an MB spheroids model, followed by flow cytometry and quantitative real-time PCR. 4-HPR treatment reduced DAOY and ONS-76 spheroid formation, in term of number and size. Decreased expression of the surface markers CD133+ and ABCG2+ as well as Oct-4 and Sox-2 gene expression were observed on BTICs treated with 4-HPR further reducing BITIC invasive activities. Finally, we analyzed 4

  17. Fenretinide (4-HPR) Targets Caspase-9, ERK 1/2 and the Wnt3a/β-Catenin Pathway in Medulloblastoma Cells and Medulloblastoma Cell Spheroids.

    PubMed

    Bassani, Barbara; Bartolini, Desirèe; Pagani, Arianna; Principi, Elisa; Zollo, Massimo; Noonan, Douglas M; Albini, Adriana; Bruno, Antonino

    2016-01-01

    Medulloblastoma (MB), a neuroectodermal tumor arising in the cerebellum, represents the most frequent childhood brain malignancy. Current treatments for MB combine radiation and chemotherapy and are often associated with relevant side effects; novel therapeutic strategies are urgently needed. N-(4-Hydroxyphenyl) retinamide (4-HPR, fenretinide), a synthetic analogue of all-trans retinoic acid, has emerged as a promising and well-tolerated cancer chemopreventive and chemotherapeutic agent for various neoplasms, from breast cancer to neuroblastoma. Here we investigated the effects of 4-HPR on MB cell lines and identified the mechanism of action for a potential use in therapy of MB. Flow cytometry analysis was performed to evaluate 4-HPR induction of apoptosis and oxygen reactive species (ROS) production, as well as cell cycle effects. Functional analysis to determine 4-HPR ability to interfere with MB cell migration and invasion were performed. Western Blot analysis were used to investigate the crucial molecules involved in selected signaling pathways associated with apoptosis (caspase-9 and PARP-1), cell survival (ERK 1/2) and tumor progression (Wnt3a and β-catenin). We show that 4-HPR induces caspase 9-dependent cell death in DAOY and ONS-76 cells, associated with increased ROS generation, suggesting that free radical intermediates might be directly involved. We observed 4-HPR induction of cell cycle arrest in G1/S phase, inactivated β-catenin, and inhibition of MB cell migration and invasion. We also evaluated the ability of 4-HPR to target MB cancer-stem/cancer-initiating cells, using an MB spheroids model, followed by flow cytometry and quantitative real-time PCR. 4-HPR treatment reduced DAOY and ONS-76 spheroid formation, in term of number and size. Decreased expression of the surface markers CD133+ and ABCG2+ as well as Oct-4 and Sox-2 gene expression were observed on BTICs treated with 4-HPR further reducing BITIC invasive activities. Finally, we analyzed 4

  18. Astaxanthin inhibits NF-κB and Wnt/β-catenin signaling pathways via inactivation of Erk/MAPK and PI3K/Akt to induce intrinsic apoptosis in a hamster model of oral cancer.

    PubMed

    Kavitha, K; Kowshik, J; Kishore, T Kranthi Kiran; Baba, Abdul Basit; Nagini, S

    2013-10-01

    The oncogenic transcription factors NF-κB and β-catenin, constitutively activated by upstream serine/threonine kinases control several cellular processes implicated in malignant transformation including apoptosis evasion. The aim of this study was to investigate the chemopreventive effects of astaxanthin, an antioxidant carotenoid, in the hamster buccal pouch (HBP) carcinogenesis model based on its ability to modulate NF-κB and Wnt signaling pathways and induce apoptosis. We determined the effect of dietary supplementation of astaxanthin on the oncogenic signaling pathways - NF-κB and Wnt/β-catenin, their upstream activator kinases - Erk/MAPK and PI-3K/Akt, and the downstream event - apoptosis evasion by real-time quantitative RT-PCR, western blot, and immunohistochemical analyses. We found that astaxanthin inhibits NF-κB and Wnt signaling by downregulating the key regulatory enzymes IKKβ and GSK-3β. Analysis of gene expression and docking interactions revealed that inhibition of these pathways may be mediated via inactivation of the upstream signaling kinases Erk/Akt by astaxanthin. Astaxanthin also induced caspase-mediated mitochondrial apoptosis by downregulating the expression of antiapoptotic Bcl-2, p-Bad, and survivin and upregulating proapoptotic Bax and Bad, accompanied by efflux of Smac/Diablo and cytochrome-c into the cytosol, and induced cleavage of poly (ADP-ribose) polymerase (PARP). The results provide compelling evidence that astaxanthin exerts chemopreventive effects by concurrently inhibiting phosphorylation of transcription factors and signaling kinases and inducing intrinsic apoptosis. Astaxanthin targets key molecules in oncogenic signaling pathways and induces apoptosis and is a promising candidate agent for cancer prevention and therapy. Copyright © 2013 Elsevier B.V. All rights reserved.

  19. Alternative Wnt Signaling Activates YAP/TAZ

    PubMed Central

    Park, Hyun Woo; Kim, Young Chul; Yu, Bo; Moroishi, Toshiro; Mo, Jung-Soon; Plouffe, Steven W.; Meng, Zhipeng; Lin, Kimberly C.; Yu, Fa-Xing; Alexander, Caroline M.; Wang, Cunyu; Guan, Kun-Liang

    2015-01-01

    SUMMARY The transcriptional co-activators YAP and TAZ are key regulators of organ size and tissue homeostasis, and their dysregulation contributes to human cancer. Here we discover YAP/TAZ as bona fide downstream effectors of the alternative Wnt signaling pathway. Wnt5a/b and Wnt3a induce YAP/TAZ activation independent of canonical Wnt/β-catenin signaling. Mechanistically, we delineate the ‘alternative Wnt-YAP/TAZ signaling axis’ that consists of Wnt - FZD/ROR - Gα12/13 - Rho GTPases -Lats1/2 to promote YAP/TAZ activation and TEAD-mediated transcription. YAP/TAZ mediate the biological functions of alternative Wnt signaling including gene expression, osteogenic differentiation, cell migration, and antagonism of Wnt/β-catenin signaling. Together, our work establishes YAP/TAZ as critical mediators of alternative Wnt signaling. PMID:26276632

  20. Oxidized ATM promotes abnormal proliferation of breast CAFs through maintaining intracellular redox homeostasis and activating the PI3K-AKT, MEK-ERK, and Wnt-β-catenin signaling pathways.

    PubMed

    Tang, Shifu; Hou, Yixuan; Zhang, Hailong; Tu, Gang; Yang, Li; Sun, Yifan; Lang, Lei; Tang, Xi; Du, Yan-E; Zhou, Mingli; Yu, Tenghua; Xu, Liyun; Wen, Siyang; Liu, Chunming; Liu, Manran

    2015-01-01

    Abnormal proliferation is one characteristic of cancer-associated fibroblasts (CAFs), which play a key role in tumorigenesis and tumor progression. Oxidative stress (OS) is the root cause of CAFs abnormal proliferation. ATM (ataxia-telangiectasia mutated protein kinase), an important redox sensor, is involved in DNA damage response and cellular homeostasis. Whether and how oxidized ATM regulating CAFs proliferation remains unclear. In this study, we show that there is a high level of oxidized ATM in breast CAFs in the absence of double-strand breaks (DSBs) and that oxidized ATM plays a critical role in CAFs proliferation. The effect of oxidized ATM on CAFs proliferation is mediated by its regulation of cellular redox balance and the activity of the ERK, PI3K-AKT, and Wnt signaling pathways. Treating cells with antioxidant N-acetyl-cysteine (NAC) partially rescues the proliferation defect of the breast CAFs caused by ATM deficiency. Administrating cells with individual or a combination of specific inhibitors of the ERK, PI3K-AKT, and Wnt signaling pathways mimics the effect of ATM deficiency on breast CAF proliferation. This is mainly ascribed to the β-catenin suppression and down-regulation of c-Myc, thus further leading to the decreased cyclinD1, cyclinE, and E2F1 expression and the enhanced p21(Cip1) level. Our results reveal an important role of oxidized ATM in the regulation of the abnormal proliferation of breast CAFs. Oxidized ATM could serve as a potential target for treating breast cancer.

  1. Wnt5a Suppresses Tumor Formation and Redirects Tumor Phenotype in MMTV-Wnt1 Tumors

    PubMed Central

    Easter, Stephanie L.; Mitchell, Elizabeth H.; Baxley, Sarah E.; Desmond, Renee; Frost, Andra R.; Serra, Rosa

    2014-01-01

    Wnt5a is a non-canonical signaling Wnt that has been implicated in tumor suppression. We previously showed that loss of Wnt5a in MMTV-PyVmT tumors resulted in a switch in tumor phenotype resulting in tumors with increased basal phenotype and high Wnt/β-catenin signaling. The object of this study was to test the hypothesis that Wnt5a can act to inhibit tumors formed by activation of Wnt/β-catenin signaling. To this end, we characterized tumor and non-tumor mammary tissue from MMTV-Wnt1 and double transgenic MMTV-Wnt1;MMTV-Wnt5a mice. Wnt5a containing mice demonstrated fewer tumors with increased latency when compared to MMTV-Wnt1 controls. Expression of markers for basal-like tumors was down-regulated in the tumors that formed in the presence of Wnt5a indicating a phenotypic switch. Reduced canonical Wnt signaling was detected in double transgenic tumors as a decrease in active β-catenin protein and a decrease in Axin2 mRNA transcript levels. In non-tumor tissues, over-expression of Wnt5a in MMTV-Wnt1 mammary glands resulted in attenuation of phenotypes normally observed in MMTV-Wnt1 glands including hyperbranching and increased progenitor and basal cell populations. Even though Wnt5a could antagonize Wnt/β-catenin signaling in primary mammary epithelial cells in culture, reduced Wnt/β-catenin signaling was not detected in non-tumor MMTV-Wnt1;Wnt5a tissue in vivo. The data demonstrate that Wnt5a suppresses tumor formation and promotes a phenotypic shift in MMTV-Wnt1 tumors. PMID:25401739

  2. Wnt5a suppresses tumor formation and redirects tumor phenotype in MMTV-Wnt1 tumors.

    PubMed

    Easter, Stephanie L; Mitchell, Elizabeth H; Baxley, Sarah E; Desmond, Renee; Frost, Andra R; Serra, Rosa

    2014-01-01

    Wnt5a is a non-canonical signaling Wnt that has been implicated in tumor suppression. We previously showed that loss of Wnt5a in MMTV-PyVmT tumors resulted in a switch in tumor phenotype resulting in tumors with increased basal phenotype and high Wnt/β-catenin signaling. The object of this study was to test the hypothesis that Wnt5a can act to inhibit tumors formed by activation of Wnt/β-catenin signaling. To this end, we characterized tumor and non-tumor mammary tissue from MMTV-Wnt1 and double transgenic MMTV-Wnt1;MMTV-Wnt5a mice. Wnt5a containing mice demonstrated fewer tumors with increased latency when compared to MMTV-Wnt1 controls. Expression of markers for basal-like tumors was down-regulated in the tumors that formed in the presence of Wnt5a indicating a phenotypic switch. Reduced canonical Wnt signaling was detected in double transgenic tumors as a decrease in active β-catenin protein and a decrease in Axin2 mRNA transcript levels. In non-tumor tissues, over-expression of Wnt5a in MMTV-Wnt1 mammary glands resulted in attenuation of phenotypes normally observed in MMTV-Wnt1 glands including hyperbranching and increased progenitor and basal cell populations. Even though Wnt5a could antagonize Wnt/β-catenin signaling in primary mammary epithelial cells in culture, reduced Wnt/β-catenin signaling was not detected in non-tumor MMTV-Wnt1;Wnt5a tissue in vivo. The data demonstrate that Wnt5a suppresses tumor formation and promotes a phenotypic shift in MMTV-Wnt1 tumors.

  3. TGF-β1 antagonizes TNF-α induced up-regulation of matrix metalloproteinase 3 in nucleus pulposus cells: role of the ERK1/2 pathway.

    PubMed

    Yang, Hao; Gao, Fei; Li, Xiang; Wang, Jianru; Liu, Hui; Zheng, Zhaomin

    2015-11-01

    Tumor necrosis factor-α (TNF-α) has been shown to have a catabolic effect on intervertebral disc degeneration (IVDD), including increasing MMP3 expression and subsequent extracellular matrix (ECM) degradation. In contrast, transforming growth factor-β1 (TGF-β1) has an anabolic effect on nucleus pulposus (NP) cells. However, the anti-catabolic effect of TGF-β1 under inflammatory condition is unknown. The aim of this study was to demonstrate whether TGF-β1 can reverse TNF-α-induced MMP3 increase in NP cells and to further investigate the underlying mechanisms. The transcriptional activity, gene expression, and protein levels of MMP3 were measured by luciferase reporter assay, qRT-PCR and western blot, respectively. TNF-α increased MMP3 expression in rat NP cells time and dose dependently. TGF-β1 could abolish TNF-α-mediated up-regulation of collagen I and MMP3 expression, and down-regulate aggrecan and collagen II expression. The ERK1/2 signaling pathway was activated after exposure to TGF-β1. Treatment with ERK1/2 inhibitors (PD98059 and U0126) abolished the antagonistic effect of TGF-β1 on TNF-α mediated catabolic responses. These findings provide novel evidence supporting the anti-catabolic role of TGF-β1 in IVDD, which is important for the potential clinical application of TGF-β1 in disc degenerative disorders.

  4. Ubiquitin-independent proteasomal degradation of Fra-1 is antagonized by Erk1/2 pathway-mediated phosphorylation of a unique C-terminal destabilizer.

    PubMed

    Basbous, Jihane; Chalbos, Dany; Hipskind, Robert; Jariel-Encontre, Isabelle; Piechaczyk, Marc

    2007-06-01

    Fra-1, a transcription factor that is phylogenetically and functionally related to the proto-oncoprotein c-Fos, controls many essential cell functions. It is expressed in many cell types, albeit with differing kinetics and abundances. In cells reentering the cell cycle, Fra-1 expression is transiently stimulated albeit later than that of c-Fos and for a longer time. Moreover, Fra-1 overexpression is found in cancer cells displaying high Erk1/2 activity and has been linked to tumorigenesis. One crucial point of regulation of Fra-1 levels is controlled protein degradation, the mechanism of which remains poorly characterized. Here, we have combined genetic, pharmacological, and signaling studies to investigate this process in nontransformed cells and to elucidate how it is altered in cancer cells. We report that the intrinsic instability of Fra-1 depends on a single destabilizer contained within the C-terminal 30 to 40 amino acids. Two serines therein, S252 and S265, are phosphorylated by kinases of the Erk1/2 pathway, which compromises protein destruction upon both normal physiological induction and tumorigenic constitutive activation of this cascade. Our data also indicate that Fra-1, like c-Fos, belongs to a small group of proteins that may, under certain circumstances, undergo ubiquitin-independent degradation by the proteasome. Our work reveals both similitudes and differences between Fra-1 and c-Fos degradation mechanisms. In particular, the presence of a single destabilizer within Fra-1, instead of two that are differentially regulated in c-Fos, explains the much faster turnover of the latter when cells traverse the G(0)/G(1)-to-S-phase transition. Finally, our study offers further insights into the signaling-regulated expression of the other Fos family proteins.

  5. Liposomal Packaging Generates Wnt Protein with In Vivo Biological Activity

    PubMed Central

    Zhao, Ludan; Kim, Jae-Beom; ten Berge, Derk; Ponnusamy, Karthik; Carre, A. Lyonel; Dudek, Henryk; Zachlederova, Marie; McElhaney, Michael; Brunton, Shirley; Gunzner, Janet; Callow, Marinella; Polakis, Paul; Costa, Mike; Zhang, Xiaoyan M.; Helms, Jill A.; Nusse, Roel

    2008-01-01

    Wnt signals exercise strong cell-biological and regenerative effects of considerable therapeutic value. There are, however, no specific Wnt agonists and no method for in vivo delivery of purified Wnt proteins. Wnts contain lipid adducts that are required for activity and we exploited this lipophilicity by packaging purified Wnt3a protein into lipid vesicles. Rather than being encapsulated, Wnts are tethered to the liposomal surface, where they enhance and sustain Wnt signaling in vitro. Molecules that effectively antagonize soluble Wnt3a protein but are ineffective against the Wnt3a signal presented by a cell in a paracrine or autocrine manner are also unable to block liposomal Wnt3a activity, suggesting that liposomal packaging mimics the biological state of active Wnts. When delivered subcutaneously, Wnt3a liposomes induce hair follicle neogenesis, demonstrating their robust biological activity in a regenerative context. PMID:18698373

  6. Kallistatin induces breast cancer cell apoptosis and autophagy by modulating Wnt signaling and microRNA synthesis

    PubMed Central

    Li, Pengfei; Guo, Youming; Bledsoe, Grant; Yang, Zhirong; Chao, Lee; Chao, Julie

    2016-01-01

    Kallistatin is an endogenous protein that regulates differential signaling pathways and biological functions. Our previous studies showed that kallistatin gene therapy inhibited angiogenesis, tumor growth and metastasis in mice, and kallistatin protein suppressed Wnt-mediated growth, migration and invasion by blocking Wnt/β-catenin signaling pathway in breast cancer cells. In this study, we show that kallistatin reduced cell viability, and increased apoptotic cell death and caspase-3 activity in MDA-MB-231 breast cancer cells. Kallistatin also induced cancer cell autophagy, as evidenced by increased LC3B levels and elevated Atg5 and Beclin-1 expression; however, co-administration of Wnt or PPARγ antagonist GW9662 abolished these effects. Moreover, kallistatin via its heparin-binding site antagonized Wnt3a-induced cancer cell proliferation and increased PPARγ expression. Kallistatin inhibited oncogenic miR-21 synthesis associated with reduced Akt phosphorylation and Bcl-2 synthesis, but increased BAX expression. Kallistatin via PKC-ERK activation reduced miR-203 levels, leading to increased expression of suppressor of cytokine signaling 3 (SOCS3), a tumor suppressor. Conversely, kallistatin stimulated expression of the tumorigenic suppressors miR-34a and p53. Kallistatin’s active site is essential for suppressing miR-21 and miR-203, and stimulating miR-34a and SOCS3 expression. This is the first study to demonstrate that kallistatin’s heparin-binding site is essential for inhibiting Wnt-mediated effects, and its active site plays a key role in regulating miR-21, miR-203, miR-34a and SOCS3 synthesis in breast cancer cells. These findings reveal novel mechanisms of kallistatin in inducing apoptosis and autophagy in breast cancer cells, thus inhibiting tumor progression by regulation of Wnt/PPARγ signaling, as well as miR-21, miR-203 and miR-34a synthesis. PMID:26790955

  7. ERK1 and ERK2 mitogen-activated protein kinases affect Ras-dependent cell signaling differentially

    PubMed Central

    Vantaggiato, Chiara; Formentini, Ivan; Bondanza, Attilio; Bonini, Chiara; Naldini, Luigi; Brambilla, Riccardo

    2006-01-01

    Background The mitogen-activated protein (MAP) kinases p44ERK1 and p42ERK2 are crucial components of the regulatory machinery underlying normal and malignant cell proliferation. A currently accepted model maintains that ERK1 and ERK2 are regulated similarly and contribute to intracellular signaling by phosphorylating a largely common subset of substrates, both in the cytosol and in the nucleus. Results Here, we show that ablation of ERK1 in mouse embryo fibroblasts and NIH 3T3 cells by gene targeting and RNA interference results in an enhancement of ERK2-dependent signaling and in a significant growth advantage. By contrast, knockdown of ERK2 almost completely abolishes normal and Ras-dependent cell proliferation. Ectopic expression of ERK1 but not of ERK2 in NIH 3T3 cells inhibits oncogenic Ras-mediated proliferation and colony formation. These phenotypes are independent of the kinase activity of ERK1, as expression of a catalytically inactive form of ERK1 is equally effective. Finally, ectopic expression of ERK1 but not ERK2 is sufficient to attenuate Ras-dependent tumor formation in nude mice. Conclusion These results reveal an unexpected interplay between ERK1 and ERK2 in transducing Ras-dependent cell signaling and proliferation. Whereas ERK2 seems to have a positive role in controlling normal and Ras-dependent cell proliferation, ERK1 probably affects the overall signaling output of the cell by antagonizing ERK2 activity. PMID:16805921

  8. Long-Term Memory Deficits are Associated with Elevated Synaptic ERK1/2 Activation and Reversed by mGluR5 Antagonism in an Animal Model of Autism

    PubMed Central

    Seese, Ronald R; Maske, Anna R; Lynch, Gary; Gall, Christine M

    2014-01-01

    A significant proportion of patients with autism exhibit some degree of intellectual disability. The BTBR T+ Itpr3tf/J mouse strain exhibits behaviors that align with the major diagnostic criteria of autism. To further evaluate the BTBR strain's cognitive impairments, we quantified hippocampus-dependent object location memory (OLM) and found that one-third of the BTBR mice exhibited robust memory, whereas the remainder did not. Fluorescence deconvolution tomography was used to test whether synaptic levels of activated extracellular signal-regulated kinase 1/2 (ERK1/2), a protein that contributes importantly to plasticity, correlate with OLM scores in individual mice. In hippocampal field CA1, the BTBRs had fewer post-synaptic densities associated with high levels of phosphorylated (p-) ERK1/2 as compared with C57BL/6 mice. Although counts of p-ERK1/2 immunoreactive synapses did not correlate with OLM performance, the intensity of synaptic p-ERK1/2 immunolabeling was negatively correlated with OLM scores across BTBRs. Metabotropic glutamate receptor (mGluR) 5 signaling activates ERK1/2. Therefore, we tested whether treatment with the mGluR5 antagonist MPEP normalizes synaptic and learning measures in BTBR mice: MPEP facilitated OLM and decreased synaptic p-ERK1/2 immunolabeling intensity without affecting numbers of p-ERK1/2+ synapses. In contrast, semi-chronic ampakine treatment, which facilitates memory in other models of cognitive impairment, had no effect on OLM in BTBRs. These results suggest that intellectual disabilities associated with different neurodevelopmental disorders on the autism spectrum require distinct therapeutic strategies based on underlying synaptic pathology. PMID:24448645

  9. Fgf9 inhibition of meiotic differentiation in spermatogonia is mediated by Erk-dependent activation of Nodal-Smad2/3 signaling and is antagonized by Kit Ligand.

    PubMed

    Tassinari, V; Campolo, F; Cesarini, V; Todaro, F; Dolci, S; Rossi, P

    2015-03-12

    Both fibroblast growth factor 9 (Fgf9) and Kit Ligand (Kl) signal through tyrosine kinase receptors, yet they exert opposite effects on meiotic differentiation in postnatal spermatogonia, Fgf9 acting as a meiosis-inhibiting substance and Kl acting as a promoter of the differentiation process. To understand the molecular mechanisms that might underlie this difference, we tried to dissect the intracellular signaling elicited by these two growth factors. We found that both Fgf9 and Kl stimulate Erk1/2 activation in Kit+ (differentiating) spermatogonia, even though with different time courses, whereas Kl, but not Fgf9, elicits activation of the Pi3k-Akt pathway. Sustained Erk1/2 activity promoted by Fgf9 is required for induction of the autocrine Cripto-Nodal-Smad2/3 signaling loop in these cells. Nodal signaling, in turn, is essential to mediate Fgf9 suppression of the meiotic program, including inhibition of Stra8 and Scp3 expression and induction of the meiotic gatekeeper Nanos2. On the contrary, sustained activation of the Pi3k-Akt pathway is required for the induction of Stra8 expression elicited by Kl and retinoic acid. Moreover, we found that Kl treatment impairs Nodal mRNA expression and Fgf9-mediated Nanos2 induction, reinforcing the antagonistic effect of these two growth factors on the meiotic fate of male germ cells.

  10. Noncanonical Wnt Signaling Maintains Hematopoietic Stem Cells in the Niche

    PubMed Central

    Sugimura, Ryohichi; He, Xi C.; Venkatraman, Aparna; Arai, Fumio; Box, Andrew; Semerad, Craig; Haug, Jeffrey S.; Peng, Lai; Zhong, Xiao-bo; Suda, Toshio; Li, Linheng

    2015-01-01

    SUMMARY Wnt signaling is involved in self-renewal and maintenance of hematopoietic stem cells (HSCs); however, the particular role of noncanonical Wnt signaling in regulating HSCs in vivo is largely unknown. Here, we show Flamingo (Fmi) and Frizzled (Fz) 8, members of noncanonical Wnt signaling, both express in and functionally maintain quiescent long-term HSCs. Fmi regulates Fz8 distribution at the interface between HSCs and N-cadherin+ osteoblasts (N-cad+OBs that enrich osteoprogenitors) in the niche. We further found that N-cad+OBs predominantly express noncanonical Wnt ligands and inhibitors of canonical Wnt signaling under homeostasis. Under stress, noncanonical Wnt signaling is attenuated and canonical Wnt signaling is enhanced in activation of HSCs. Mechanistically, noncanonical Wnt signaling mediated by Fz8 suppresses the Ca2+-NFAT- IFNγ pathway, directly or indirectly through the CDC42-CK1α complex and also antagonizes canonical Wnt signaling in HSCs. Taken together, our findings demonstrate that noncanonical Wnt signaling maintains quiescent long-term HSCs through Fmi and Fz8 interaction in the niche. PMID:22817897

  11. Distinct functions for ERKs?

    PubMed Central

    Lloyd, Alison C

    2006-01-01

    The Ras/Raf/MEK/ERK signaling pathway is one of the best understood signal routes in cells. Recent studies add complexity to this cascade by indicating that the two ERK kinases, ERK1 (p44ERK1) and ERK2 (p42ERK2), may have distinct functions. PMID:16879721

  12. Wnt5a Regulates Midbrain Dopaminergic Axon Growth and Guidance

    PubMed Central

    Blakely, Brette D.; Bye, Christopher R.; Fernando, Chathurini V.; Horne, Malcolm K.; Macheda, Maria L.; Stacker, Steven A.; Arenas, Ernest; Parish, Clare L.

    2011-01-01

    During development, precise temporal and spatial gradients are responsible for guiding axons to their appropriate targets. Within the developing ventral midbrain (VM) the cues that guide dopaminergic (DA) axons to their forebrain targets remain to be fully elucidated. Wnts are morphogens that have been identified as axon guidance molecules. Several Wnts are expressed in the VM where they regulate the birth of DA neurons. Here, we describe that a precise temporo-spatial expression of Wnt5a accompanies the development of nigrostriatal projections by VM DA neurons. In mice at E11.5, Wnt5a is expressed in the VM where it was found to promote DA neurite and axonal growth in VM primary cultures. By E14.5, when DA axons are approaching their striatal target, Wnt5a causes DA neurite retraction in primary cultures. Co-culture of VM explants with Wnt5a-overexpressing cell aggregates revealed that Wnt5a is capable of repelling DA neurites. Antagonism experiments revealed that the effects of Wnt5a are mediated by the Frizzled receptors and by the small GTPase, Rac1 (a component of the non-canonical Wnt planar cell polarity pathway). Moreover, the effects were specific as they could be blocked by Wnt5a antibody, sFRPs and RYK-Fc. The importance of Wnt5a in DA axon morphogenesis was further verified in Wnt5a−/− mice, where fasciculation of the medial forebrain bundle (MFB) as well as the density of DA neurites in the MFB and striatal terminals were disrupted. Thus, our results identify a novel role of Wnt5a in DA axon growth and guidance. PMID:21483795

  13. Discovery of Klotho peptide antagonists against Wnt3 and Wnt3a target proteins using combination of protein engineering, protein-protein docking, peptide docking and molecular dynamics simulations.

    PubMed

    Mirza, Shaher Bano; Ekhteiari Salmas, Ramin; Fatmi, M Qaiser; Durdagi, Serdar

    2017-12-01

    The Klotho is known as lifespan enhancing protein involved in antagonizing the effect of Wnt proteins. Wnt proteins are stem cell regulators, and uninterrupted exposure of Wnt proteins to the cell can cause stem and progenitor cell senescence, which may lead to aging. Keeping in mind the importance of Klotho in Wnt signaling, in silico approaches have been applied to study the important interactions between Klotho and Wnt3 and Wnt3a (wingless-type mouse mammary tumor virus (MMTV) integration site family members 3 and 3a). The main aim of the study is to identify important residues of the Klotho that help in designing peptides which can act as Wnt antagonists. For this aim, a protein engineering study is performed for Klotho, Wnt3 and Wnt3a. During the theoretical analysis of homology models, unexpected role of number of disulfide bonds and secondary structure elements has been witnessed in case of Wnt3 and Wnt3a proteins. Different in silico experiments were carried out to observe the effect of correct number of disulfide bonds on 3D protein models. For this aim, total of 10 molecular dynamics (MD) simulations were carried out for each system. Based on the protein-protein docking simulations of selected protein models of Klotho with Wnt3 and Wnt3a, different peptides derived from Klotho have been designed. Wnt3 and Wnt3a proteins have three important domains: Index finger, N-terminal domain and a patch of ∼10 residues on the solvent exposed surface of palm domain. Protein-peptide docking of designed peptides of Klotho against three important domains of palmitoylated Wnt3 and Wnt3a yields encouraging results and leads better understanding of the Wnt protein inhibition by proposed Klotho peptides. Further in vitro studies can be carried out to verify effects of novel designed peptides as Wnt antagonists.

  14. Amino acid limitation induces down-regulation of WNT5a at transcriptional level

    SciTech Connect

    Wang Zuguang; Chen Hong

    2009-01-23

    An aberrant WNT signaling contributes to the development and progression of multiple cancers. WNT5a is one of the WNT signaling molecules. This study was designed to test the hypothesis that amino acid deprivation induces changes in the WNT signaling pathway in colon cancer cells. Results showed that targets of the amino acid response pathway, ATF3 and p21, were induced in the human colon cancer cell line SW480 during amino acid limitation. There was a significant decrease in the WNT5a mRNA level following amino acid deprivation. The down-regulation of WNT5a mRNA by amino acid deprivation is not due to mRNA destabilization. There is a reduction of nuclear {beta}-catenin protein level by amino acid limitation. Under amino acid limitation, phosphorylation of ERK1/2 was increased and the blockage of ERK1/2 by the inhibitor U0126 partially restored WNT5a mRNA level. In conclusion, amino acid limitation in colon cancer cells induces phosphorylation of ERK1/2, which then down-regulates WNT5a expression.

  15. Wnt5a as an Effector of TGFβ in Mammary Development and Cancer

    PubMed Central

    Easter, Stephanie L.; Jiang, Wen; Baxley, Sarah E.

    2011-01-01

    Wnt5a is a member of the Wingless-related/MMTV-integration family of secreted growth factors, which are involved in a wide range of cellular processes. Wnt signaling can be broadly divided into two categories the canonical, β-catenin-dependent pathway and the non-canonical β-catenin-independent pathway. Wnt5a is a non-canonical signaling member of the Wnt family. Loss of Wnt5a is associated with early relapse of invasive breast cancer, increased metastasis, and poor survival in humans. It has been shown that TGF-β directly regulates expression of Wnt5a in mammary gland and that Wnt5a mediates the effects of TGF-β on branching during mammary gland development. Here we review the evidence suggesting Wnt5a acts as an effector of TGF-β actions in breast cancer. It is suggested that the tumor suppressive functions of TGF-β involve Wnt5a-mediated antagonism of Wnt/β-catenin signaling and limiting the stem cell population. Interactions between TGF-β and Wnt5a in metastasis appear to be more complex, and may depend on specific cues from the microenvironment as well as activation of specific intracellular signaling pathways. PMID:21416313

  16. Differential antigen expression and aberrant signaling via PI3/AKT, MAP/ERK, JAK/STAT, and Wnt/β catenin pathways in Lin-/CD38-/CD34+ cells in acute myeloid leukemia.

    PubMed

    Garg, Swati; Shanmukhaiah, Chandrakala; Marathe, Supreet; Mishra, Prashant; Babu Rao, Vunditi; Ghosh, Kanjaksha; Madkaikar, Manisha

    2016-03-01

    Acute myeloid leukemia is often called as stem cell disease that presents with treatment failure and poor disease outcome. Leukemic stem cells in acute myeloid leukemia (AML) are enriched in Lineage-/CD38-/CD34+ compartment of CD34-positive AML. Many markers important for stem cell biology have been reported for their association with leukemic stem cell population, but what remains clinically most important is a rapid identification of prognostic information. In this study, we evaluated four signal transduction pathways and thirteen markers on Lin-/CD38-/CD34+ population in AML. Expressions were compared in different AML subtypes, survival, and treatment outcome groups. We observed that markers important in homing, cell quiescence, and signal propagation such as CD44, CD96, CD90, WT-1, CD123 and CD25 were most significantly differentially expressed on Lin-/CD38-/CD34+ population in AML from their normal counterparts (P < 0.05, Mann-Whitney). Constitutive activation of phospho ERK, AKT, and STAT5 in these cells was associated with poor outcome. Also, an increased frequency of putative leukemic stem cell population shows negative impact on treatment outcome and overall survival, suggesting that initial evaluation of AML samples for pLSC frequency and constitutively activated signaling pathway can provide prognostic and therapeutic information at the time of diagnosis. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  17. Structure-based Discovery of Novel Small Molecule Wnt Signaling Inhibitors by Targeting the Cysteine-rich Domain of Frizzled*

    PubMed Central

    Lee, Ho-Jin; Bao, Ju; Miller, Ami; Zhang, Chi; Wu, Jibo; Baday, Yiressy C.; Guibao, Cristina; Li, Lin; Wu, Dianqing; Zheng, Jie J.

    2015-01-01

    Frizzled is the earliest discovered glycosylated Wnt protein receptor and is critical for the initiation of Wnt signaling. Antagonizing Frizzled is effective in inhibiting the growth of multiple tumor types. The extracellular N terminus of Frizzled contains a conserved cysteine-rich domain that directly interacts with Wnt ligands. Structure-based virtual screening and cell-based assays were used to identify five small molecules that can inhibit canonical Wnt signaling and have low IC50 values in the micromolar range. NMR experiments confirmed that these compounds specifically bind to the Wnt binding site on the Frizzled8 cysteine-rich domain with submicromolar dissociation constants. Our study confirms the feasibility of targeting the Frizzled cysteine-rich domain as an effective way of regulating canonical Wnt signaling. These small molecules can be further optimized into more potent therapeutic agents for regulating abnormal Wnt signaling by targeting Frizzled. PMID:26504084

  18. Restoration of WNT4 inhibits cell growth in leukemia-derived cell lines

    PubMed Central

    2013-01-01

    cells corroborated this observation. Interestingly, restoration of WNT4 expression in BJAB cells increased the accumulation of cells in G1 phase, and did not induce activation of canonical WNT/β-catenin target genes. Conclusions Our findings suggest that the WNT4 ligand plays a role in regulating the cell growth of leukemia-derived cells by arresting cells in the G1 cell cycle phase in an FZD6-independent manner, possibly through antagonizing the canonical WNT/β-catenin signaling pathway. PMID:24274766

  19. Molecular cloning, characterization and expression analysis of Wnt4, Wnt5, Wnt6, Wnt7, Wnt10 and Wnt16 from Litopenaeus vannamei.

    PubMed

    Zhang, Shuang; Li, Chao-Zheng; Yang, Qi-Hui; Dong, Xiao-Hui; Chi, Shu-Yan; Liu, Hong-Yu; Shi, Li-Li; Tan, Bei-Ping

    2016-07-01

    The Wnt (Wg-type MMTV integration site) signaling represents as the negative regulator of virus-induced innate immune responses. Wnt genes act as ligands to activate the Wnt signaling. To know more about the information of Wnt genes in invertebrates, Litopenaeus vannamei Wnt genes (LvWnts) were identified and characterized. In this study, Six Wnt genes (LvWnt4, LvWnt5, LvWnt6, LvWnt7, LvWnt10 and LvWnt16) were obtained in L. vannamei. The complete cDNAs open reading frames (ORF) of LvWnt4, LvWnt5, LvWnt6, LvWnt7, LvWnt10 and LvWnt16 were 1077 bp, 1107 bp, 1350 bp, 1047 bp, 1509 bp and 1158 bp (GenBank accession no. KU169896, KU169897, KU169898, KU169899, KU169900 and KU169901), encoding 358, 368, 449, 348, 502 and 385 amino acid (aa) residues respectively. All the six members of LvWnts contain a Wnt1 domain, which is considered as an important feature of Wnt gene family. ClustalW analysis with amino acid sequences revealed that the proportion of identity with other species was more than 48% for all the LvWnts except LvWnt10 (36-41%). The phylogenetic relationship analysis illustrated that different subtype of Wnts formed their own separate branches and were placed in branch of invertebrates respectively with strong bootstrap support. The constitutive expressions of LvWnts were confirmed by RT-PCR in all the examined five developmental stages and eleven tissues of L. vannamei with different express patterns. LvWnt4, LvWnt5 and LvWnt10 were expressed highest in nerve while LvWnt6, LvWnt7 and LvWnt16 were expressed highest in intestine, stomach and gill, respectively. In addition, all the LvWnts were regulated by white spot syndrome virus (WSSV) challenges at different levels in hepatopancreas, gill and hemocytes, suggesting that Wnt genes may play a role in the defense against pathogenic virus infection in innate immune of L. vannamei.

  20. Separate and distinctive roles for Wnt5a in tongue, lingual tissue and taste papilla development

    PubMed Central

    Liu, Hong-Xiang; Grosse, Ann S.; Iwatsuki, Ken; Mishina, Yuji; Gumucio, Deborah L.; Mistretta, Charlotte M.

    2012-01-01

    Although canonical Wnt signaling is known to regulate taste papilla induction and numbers, roles for noncanonical Wnt pathways in tongue and taste papilla development have not been explored. With mutant mice and whole tongue organ cultures we demonstrate that Wnt5a protein and message are within anterior tongue mesenchyme across embryo stages from the initiation of tongue formation, through papilla placode appearance and taste papilla development. The Wnt5a mutant tongue is severely shortened, with an ankyloglossia, and lingual mesenchyme is disorganized. However, fungiform papilla morphology, number and innervation are preserved, as is expression of the papilla marker, Shh. These data demonstrate that the genetic regulation for tongue size and shape can be separated from that directing lingual papilla development. Preserved number of papillae in a shortened tongue results in an increased density of fungiform papillae in the mutant tongues. In tongue organ cultures, exogenous Wnt5a profoundly suppresses papilla formation and simultaneously decreases canonical Wnt signaling as measured by the TOPGAL reporter. These findings suggest that Wnt5a antagonizes canonical Wnt signaling to dictate papilla number and spacing. In all, distinctive roles for Wnt5a in tongue size, fungiform papilla patterning and development are shown and a necessary balance between non-canonical and canonical Wnt paths in regulating tongue growth and fungiform papillae is proposed in a model, through the Ror2 receptor. PMID:22024319

  1. Noncanonical Wnt signaling promotes osteoclast differentiation and is facilitated by the human immunodeficiency virus protease inhibitor ritonavir

    SciTech Connect

    Santiago, Francisco; Oguma, Junya; Brown, Anthony M.C.; Laurence, Jeffrey

    2012-01-06

    Highlights: Black-Right-Pointing-Pointer First demonstration of direct role for noncanonical Wnt in osteoclast differentiation. Black-Right-Pointing-Pointer Demonstration of Ryk as a Wnt5a/b receptor in inhibition of canonical Wnt signaling. Black-Right-Pointing-Pointer Modulation of noncanonical Wnt signaling by a clinically important drug, ritonavir. Black-Right-Pointing-Pointer Establishes a mechanism for an important clinical problem: HIV-associated bone loss. -- Abstract: Wnt proteins that signal via the canonical Wnt/{beta}-catenin pathway directly regulate osteoblast differentiation. In contrast, most studies of Wnt-related effects on osteoclasts involve indirect changes. While investigating bone mineral density loss in the setting of human immunodeficiency virus (HIV) infection and its treatment with the protease inhibitor ritonavir (RTV), we observed that RTV decreased nuclear localization of {beta}-catenin, critical to canonical Wnt signaling, in primary human and murine osteoclast precursors. This occurred in parallel with upregulation of Wnt5a and Wnt5b transcripts. These Wnts typically stimulate noncanonical Wnt signaling, and this can antagonize the canonical Wnt pathway in many cell types, dependent upon Wnt receptor usage. We now document RTV-mediated upregulation of Wnt5a/b protein in osteoclast precursors. Recombinant Wnt5b and retrovirus-mediated expression of Wnt5a enhanced osteoclast differentiation from human and murine monocytic precursors, processes facilitated by RTV. In contrast, canonical Wnt signaling mediated by Wnt3a suppressed osteoclastogenesis. Both RTV and Wnt5b inhibited canonical, {beta}-catenin/T cell factor-based Wnt reporter activation in osteoclast precursors. RTV- and Wnt5-induced osteoclast differentiation were dependent upon the receptor-like tyrosine kinase Ryk, suggesting that Ryk may act as a Wnt5a/b receptor in this context. This is the first demonstration of a direct role for Wnt signaling pathways and Ryk in

  2. Updating the Wnt pathways

    PubMed Central

    Yu, Jia; Virshup, David M.

    2014-01-01

    In the three decades since the discovery of the Wnt1 proto-oncogene in virus-induced mouse mammary tumours, our understanding of the signalling pathways that are regulated by the Wnt proteins has progressively expanded. Wnts are involved in an complex signalling network that governs multiple biological processes and cross-talk with multiple additional signalling cascades, including the Notch, FGF (fibroblast growth factor), SHH (Sonic hedgehog), EGF (epidermal growth factor) and Hippo pathways. The Wnt signalling pathway also illustrates the link between abnormal regulation of the developmental processes and disease manifestation. Here we provide an overview of Wnt-regulated signalling cascades and highlight recent advances. We focus on new findings regarding the dedicated Wnt production and secretion pathway with potential therapeutic targets that might be beneficial for patients with Wnt-related diseases. PMID:25208913

  3. Secreted Frizzled-related protein-2 (sFRP2) augments canonical Wnt3a-induced signaling

    SciTech Connect

    Marschall, Zofia von; Fisher, Larry W.

    2010-09-24

    Research highlights: {yields} sFRP2 enhances the Wnt3a-induced {beta}-catenin stabilization and its nuclear translocation. {yields} sFRP2 enhances LRP6 phosphorylation and Wnt3a/{beta}-catenin transcriptional reporter activity. {yields} Dickkopf-1 (DKK1) fully antagonizes both Wnt3a/sFRP2-induced LRP6 phosphorylation and transcriptional activity. {yields} sFRP2 enhances expression of genes known to be regulated by Wnt3a signaling. -- Abstract: Secreted Frizzled-related proteins (sFRP) are involved in embryonic development as well as pathological conditions including bone and myocardial disorders and cancer. Because of their sequence homology with the Wnt-binding domain of Frizzled, they have generally been considered antagonists of canonical Wnt signaling. However, additional activities of various sFRPs including both synergism and mimicry of Wnt signaling as well as functions other than modulation of Wnt signaling have been reported. Using human embryonic kidney cells (HEK293A), we found that sFRP2 enhanced Wnt3a-dependent phosphorylation of LRP6 as well as both cytosolic {beta}-catenin levels and its nuclear translocation. While addition of recombinant sFRP2 had no activity by itself, Top/Fop luciferase reporter assays showed a dose-dependent increase of Wnt3a-mediated transcriptional activity. sFRP2 enhancement of Wnt3a signaling was abolished by treatment with the Wnt antagonist, Dickkopf-1 (DKK1). Wnt-signaling pathway qPCR arrays showed that sFRP2 enhanced the Wnt3a-mediated transcriptional up-regulation of several genes regulated by Wnt3a including its antagonists, DKK1, and Naked cuticle-1 homolog (NKD1). These results support sFRP2's role as an enhancer of Wnt/{beta}-catenin signaling, a result with biological impact for both normal development and diverse pathologies such as tumorigenesis.

  4. Wnt5a is associated with right ventricular dysfunction and adverse outcome in dilated cardiomyopathy.

    PubMed

    Abraityte, Aurelija; Lunde, Ida G; Askevold, Erik T; Michelsen, Annika E; Christensen, Geir; Aukrust, Pål; Yndestad, Arne; Fiane, Arnt; Andreassen, Arne; Aakhus, Svend; Dahl, Christen P; Gullestad, Lars; Broch, Kaspar; Ueland, Thor

    2017-06-14

    The Wingless (Wnt) pathway has been implicated in the pathogenesis of dilated cardiomyopathy (DCM). To explore the role of Wnt modulators Wnt5a and sFRP3 in DCM patients we analyzed the expression of Wnt5a and sFRP3 in plasma and myocardium of DCM patients and evaluated their effects on NFAT luciferase activity in neonatal mouse cardiomyocytes. Elevated circulating Wnt5a (n = 102) was associated with increased pulmonary artery pressures, decreased right ventricular function and adverse outcome, with a stronger association in more severely affected patients. A higher Wnt5a/sFRP3 ratio (n = 25) was found in the right ventricle vs. the left ventricle and was correlated with NFAT activation as well as pulmonary artery pressures. Wnt5a induced NFAT activation and sFRP3 release in cardiomyocytes in vitro, while sFRP3 antagonized Wnt5a. Wnt5a is associated with right ventricular dysfunction and adverse outcome in DCM patients and may promote the progression of DCM through NFAT signaling.

  5. Adenovirus-mediated Wnt5a expression inhibits the telogen-to-anagen transition of hair follicles in mice.

    PubMed

    Xing, Yi-Zhan; Wang, Rui-Min; Yang, Ke; Guo, Hai-Ying; Deng, Fang; Li, Yu-Hong; Ye, Ji-Xing; He, Long; Lian, Xiao-Hua; Yang, Tian

    2013-01-01

    The canonical Wnt/β-catenin pathway plays an important role in hair cycle induction. Wnt5a is a non-canonical Wnt family member that generally antagonizes canonical Wnt signaling in other systems. In hair follicles, Wnt5a and canonical Wnt are both expressed in cells in the telogen stage. Wnt5a has been shown to be critical for controlling hair cell fate. However, the role that Wnt5a plays in the transition from the telogen to anagen stage is unknown. In this study, using whole-mount in situ hybridization, we show that Wnt5a is produced by several other cell types, excluding dermal papilla cells, throughout the hair cycle. For example, Wnt5a is expressed in bulge and secondary hair germ cells in the telogen stage. Our studies focused on the depilated 8-week-old mouse as a synchronized model of hair growth. Interestingly, overexpression of adenovirus Wnt5a in the dorsal skin of mice led to the elongation of the telogen stage and inhibition of the initiation of the anagen stage. However, following an extended period of time, four pelage hair types grew from hairless skin that was induced by Wnt5a, and the structure of these new hair shafts was normal. Using microarray analysis and quantitative arrays, we showed that the expression of β-catenin and some target genes of canonical Wnt signaling decreased after Wnt5a treatment. These data demonstrate that Wnt5a may inhibit the telogen stage to maintain a quiescent state of the hair follicle.

  6. WNT3 Inhibits Cerebellar Granule Neuron Progenitor Proliferation and Medulloblastoma Formation via MAPK Activation

    PubMed Central

    Ayrault, Olivier; Kim, Jee Hae; Zhu, Xiaodong; Murphy, David A.; Van Aelst, Linda; Roussel, Martine F.; Hatten, Mary E.

    2013-01-01

    During normal cerebellar development, the remarkable expansion of granule cell progenitors (GCPs) generates a population of granule neurons that outnumbers the total neuronal population of the cerebral cortex, and provides a model for identifying signaling pathways that may be defective in medulloblastoma. While many studies focus on identifying pathways that promote growth of GCPs, a critical unanswered question concerns the identification of signaling pathways that block mitogenic stimulation and induce early steps in differentiation. Here we identify WNT3 as a novel suppressor of GCP proliferation during cerebellar development and an inhibitor of medulloblastoma growth in mice. WNT3, produced in early postnatal cerebellum, inhibits GCP proliferation by down-regulating pro-proliferative target genes of the mitogen Sonic Hedgehog (SHH) and the bHLH transcription factor Atoh1. WNT3 suppresses GCP growth through a non-canonical Wnt signaling pathway, activating prototypic mitogen-activated protein kinases (MAPKs), the Ras-dependent extracellular-signal-regulated kinases 1/2 (ERK1/2) and ERK5, instead of the classical β-catenin pathway. Inhibition of MAPK activity using a MAPK kinase (MEK) inhibitor reversed the inhibitory effect of WNT3 on GCP proliferation. Importantly, WNT3 inhibits proliferation of medulloblastoma tumor growth in mouse models by a similar mechanism. Thus, the present study suggests a novel role for WNT3 as a regulator of neurogenesis and repressor of neural tumors. PMID:24303070

  7. ARF6-Regulated Endocytosis of Growth Factor Receptors Links Cadherin-Based Adhesion to Canonical Wnt Signaling in Epithelia

    PubMed Central

    Pellon-Cardenas, Oscar; Clancy, James; Uwimpuhwe, Henriette

    2013-01-01

    Wnt signaling has an essential role in embryonic development as well as stem/progenitor cell renewal, and its aberrant activation is implicated in many diseases, including several cancers. β-Catenin is a critical component of Wnt-mediated transcriptional activation. Here we show that ARF6 activation during canonical Wnt signaling promotes the intracellular accumulation of β-catenin via a mechanism that involves the endocytosis of growth factor receptors and robust activation of extracellular signal-regulated kinase (ERK). ERK promotes casein kinase 2-mediated phosphorylation of α-catenin, leading to destabilization of the adherens junctions and a subsequent increase in cytoplasmic pools of active β-catenin and E-cadherin. ERK also phosphorylates LRP6 to amplify the Wnt transduction pathway. The aforementioned Wnt-ERK signaling pathway initiates lumen filling of epithelial cysts by promoting cell proliferation in three-dimensional cell cultures. This study elucidates a mechanism responsible for the switch in β-catenin functions in cell adhesion at the adherens junctions and Wnt-induced nuclear signaling. PMID:23716594

  8. ARF6-regulated endocytosis of growth factor receptors links cadherin-based adhesion to canonical Wnt signaling in epithelia.

    PubMed

    Pellon-Cardenas, Oscar; Clancy, James; Uwimpuhwe, Henriette; D'Souza-Schorey, Crislyn

    2013-08-01

    Wnt signaling has an essential role in embryonic development as well as stem/progenitor cell renewal, and its aberrant activation is implicated in many diseases, including several cancers. β-Catenin is a critical component of Wnt-mediated transcriptional activation. Here we show that ARF6 activation during canonical Wnt signaling promotes the intracellular accumulation of β-catenin via a mechanism that involves the endocytosis of growth factor receptors and robust activation of extracellular signal-regulated kinase (ERK). ERK promotes casein kinase 2-mediated phosphorylation of α-catenin, leading to destabilization of the adherens junctions and a subsequent increase in cytoplasmic pools of active β-catenin and E-cadherin. ERK also phosphorylates LRP6 to amplify the Wnt transduction pathway. The aforementioned Wnt-ERK signaling pathway initiates lumen filling of epithelial cysts by promoting cell proliferation in three-dimensional cell cultures. This study elucidates a mechanism responsible for the switch in β-catenin functions in cell adhesion at the adherens junctions and Wnt-induced nuclear signaling.

  9. Substrate-dependent Wnt signaling in MSC differentiation within biomaterial-derived 3D spheroids.

    PubMed

    Hsu, Shan-hui; Huang, Guo-Shiang

    2013-07-01

    A unique biomaterial-based system was developed to generate dynamic three-dimensional (3D) multicellular spheroids of mesenchymal stem cells (MSCs). MSCs were cultured on transparent membranes made of chitosan or those further grafted with hyaluronan (HA) in different densities. MSCs vigorously migrated and were self-assembled into highly mobile 3D spheroids with substrate-dependent upregulation of adhesion molecule N-cadherin. MSC spheroids showed increased expression of Wnt genes/proteins and substrate-dependent cell fate. The correlation of differentiation capacities with Wnt signaling and crosstalk with other pathways such as ERK1/2 or Smad2/3 were observed for MSC spheroids but not for the conventional 2D cultured cells. Wnt3a-mediated canonical Wnt signaling was more active for MSC spheroids derived on chitosan, which were prone to osteogenesis. Wnt5a-mediated non-canonical Wnt signaling was more active for MSC spheroids derived on HA-grafted chitosan, which were prone to chondrogenesis. In particular, the relative importance of Wnt5a-mediated non-canonical vs. Wnt3a-mediated canonical Wnt signals in determining the cell fate was controlled by the grafting density of HA on chitosan. Treatment with the inhibitor of canonical Wnt-associated signaling molecules suppressed the osteogenesis of MSC spheroids on chitosan. This study demonstrates that Wnt signaling of MSCs is distinct in 3D environment and is substrate-dependent. The convenient 3D platform may be used to examine the role of Wnt signaling in controlling MSC fate under different extracellular environments, and potentially applied to study stem cell behavior in regenerative medicine, normal development, and cancer. Copyright © 2013 Elsevier Ltd. All rights reserved.

  10. Wnt signaling in cnidarians.

    PubMed

    Holstein, Thomas W

    2008-01-01

    Cnidarians are an ancient group of animals at the base of metazoan evolution. They exhibit a simple body plan with only one well-defined body axis and a small number of cell types. Cnidarians are also well known for their enormous regeneration capacity. Recent work in the freshwater polyp Hydra and in the sea anemone Nematostella has identified an unexpectedly high level of genetic complexity of wnt genes. Canonical Wnt signaling acts in pattern formation and regeneration of Hydra and also in gastrulation and early embryogenesis of Nematostella. Vertebrate-specific Wnt-antagonists were also identified from cnidarians and exhibit similar conserved functions. The simple cnidarian body plan and the now available genomes from Hydra and Nematostella, together with new functional approaches, make these animals an attractive model for studying the basic functions of canonical and non-canonical Wnt signaling.

  11. Wnt signaling and osteoporosis

    PubMed Central

    Manolagas, Stavros C.

    2014-01-01

    Major advances in understanding basic bone biology and the cellular and molecular mechanisms responsible for the development of osteoporosis, over the last 20 years, have dramatically altered the management of this disease. The purpose of this mini-review is to highlight the seminal role of Wnt signaling in bone homeostasis and disease and the emergence of novel osteoporosis therapies by targeting Wnt signaling with drugs. PMID:24815296

  12. The Wnt pathway limits BMP signaling outside of the germline stem cell niche in Drosophila ovaries.

    PubMed

    Mottier-Pavie, Violaine I; Palacios, Victor; Eliazer, Susan; Scoggin, Shane; Buszczak, Michael

    2016-09-01

    The mechanisms that modulate and limit the signaling output of adult stem cell niches remain poorly understood. To gain further insights into how these microenvironments are regulated in vivo, we performed a candidate gene screen designed to identify factors that restrict BMP signal production to the cap cells that comprise the germline stem cell (GSC) niche of Drosophila ovaries. Through these efforts, we found that disruption of Wnt4 and components of the canonical Wnt pathway results in a complex germ cell phenotype marked by an expansion of GSC-like cells, pre-cystoblasts and cystoblasts in young females. This phenotype correlates with an increase of decapentaplegic (dpp) mRNA levels within escort cells and varying levels of BMP responsiveness in the germline. Further genetic experiments show that Wnt4, which exhibits graded expression in somatic cells of germaria, activates the Wnt pathway in posteriorly positioned escort cells. The activation of the Wnt pathway appears to be limited by the BMP pathway itself, as loss of Mad in escort cells results in the expansion of Wnt pathway activation. Wnt pathway activity changes within germaria during the course of aging, coincident with changes in dpp production. These data suggest that mutual antagonism between the BMP and Wnt pathways in somatic cells helps to regulate germ cell differentiation. Copyright © 2016 Elsevier Inc. All rights reserved.

  13. Nicotine may promote tongue squamous cell carcinoma progression by activating the Wnt/β-catenin and Wnt/PCP signaling pathways.

    PubMed

    Wang, Chengze; Xu, Xin; Jin, Hairu; Liu, Gangli

    2017-05-01

    To investigate the effects and the possible underlying mechanisms of nicotine stimulation on tongue squamous cell carcinoma (TSCC) progression, a TSCC cell line Cal27 and 34 samples of paraffin-embedded TSCC were examined. Immunofluorescence, western blot analysis, and TOP/FOP flash, CCK-8, wound healing and Transwell invasion assays were used to evaluate Cal27 in response to nicotine stimulation. We also investigated expression levels of related proteins of Wnt/β-catenin and Wnt/PCP pathways in paraffin-embedded TSCC samples with or without a history of smoking by immunohistochemistry. Nicotine stimulation can promote proliferation, migration, and invasion of TSCC cells in vitro, downregulate E-cadherin, and activate the Wnt/β-catenin and Wnt/PCP pathways, which could be antagonized by the α7 nicotine acetylcholine receptor (α7 nAChR) inhibitor α-BTX. Moreover, the expression levels of β-catenin, Wnt5a and Ror2 were higher in TSCC patients with a history of smoking than those without a history of smoking. Our results suggest nicotine may promote tongue squamous carcinoma cells progression by activating the Wnt/β-catenin and Wnt/PCP signaling pathways and may play a significant role in the progression and metastasis of smoking-related TSCC.

  14. TLR4 Activation Promotes Bone Marrow MSC Proliferation and Osteogenic Differentiation via Wnt3a and Wnt5a Signaling.

    PubMed

    He, Xiaoqing; Wang, Hai; Jin, Tao; Xu, Yongqing; Mei, Liangbin; Yang, Jun

    2016-01-01

    Mesenchymal stem cells (MSCs) from adult bone marrow maintain their self-renewal ability and the ability to differentiate into osteoblast. Thus, adult bone marrow MSCs play a key role in the regeneration of bone tissue. Previous studies indicated that TLR4 is expressed in MSCs and is critical in regulating the fate decision of MSCs. However, the exact functional role and underlying mechanisms of how TLR4 regulate bone marrow MSC proliferation and differentiation are unclear. Here, we found that activated TLR4 by its ligand LPS promoted the proliferation and osteogenic differentiation of MSCs in vitro. TLR4 activation by LPS also increased cytokine IL-6 and IL-1β production in MSCs. In addition, LPS treatment has no effect on inducing cell death of MSCs. Deletion of TLR4 expression in MSCs completely eliminated the effects of LPS on MSC proliferation, osteogenic differentiation and cytokine production. We also found that the mRNA and protein expression of Wnt3a and Wnt5a, two important factors in regulating MSC fate decision, was upregulated in a TLR4-dependent manner. Silencing Wnt3a with specific siRNA remarkably inhibited TLR4-induced MSC proliferation, while Wnt5a specific siRNA treatment significantly antagonized TLR4-induced MSC osteogenic differentiation. These results together suggested that TLR4 regulates bone marrow MSC proliferation and osteogenic differentiation through Wnt3a and Wnt5a signaling. These finding provide new data to understand the role and the molecular mechanisms of TLR4 in regulating bone marrow MSC functions. These data also provide new insight in developing new therapy in bone regeneration using MSCs by modulating TLR4 and Wnt signaling activity.

  15. Adipogenesis and WNT signalling

    PubMed Central

    Christodoulides, Constantinos; Lagathu, Claire; Sethi, Jaswinder K.; Vidal-Puig, Antonio

    2015-01-01

    An inability of adipose tissue to expand consequent to exhausted capacity to recruit new adipocytes might underlie the association between obesity and insulin resistance. Adipocytes arise from mesenchymal precursors whose commitment and differentiation along the adipocytic lineage is tightly regulated. These regulatory factors mediate cross-talk between adipose cells, ensuring that adipocyte growth and differentiation are coupled to energy storage demands. The WNT family of autocrine and paracrine growth factors regulates adult tissue maintenance and remodelling and, consequently, is well suited to mediate adipose cell communication. Indeed, several recent reports, summarized in this review, implicate WNT signalling in regulating adipogenesis. Manipulating the WNT pathway to alter adipose cellular makeup, therefore, constitutes an attractive drug-development target to combat obesity-associated metabolic complications. PMID:19008118

  16. ERK Signals: Scaffolding Scaffolds?

    PubMed Central

    Casar, Berta; Crespo, Piero

    2016-01-01

    ERK1/2 MAP Kinases become activated in response to multiple intra- and extra-cellular stimuli through a signaling module composed of sequential tiers of cytoplasmic kinases. Scaffold proteins regulate ERK signals by connecting the different components of the module into a multi-enzymatic complex by which signal amplitude and duration are fine-tuned, and also provide signal fidelity by isolating this complex from external interferences. In addition, scaffold proteins play a central role as spatial regulators of ERKs signals. In this respect, depending on the subcellular localization from which the activating signals emanate, defined scaffolds specify which substrates are amenable to be phosphorylated. Recent evidence has unveiled direct interactions among different scaffold protein species. These scaffold-scaffold macro-complexes could constitute an additional level of regulation for ERK signals and may serve as nodes for the integration of incoming signals and the subsequent diversification of the outgoing signals with respect to substrate engagement. PMID:27303664

  17. Wnt Isoform-Specific Interactions with Coreceptor Specify Inhibition or Potentiation of Signaling by LRP6 Antibodies

    PubMed Central

    Gong, Yan; Bourhis, Eric; Chiu, Cecilia; Stawicki, Scott; DeAlmeida, Venita I.; Liu, Bob Y.; Phamluong, Khanhky; Cao, Tim C.; Carano, Richard A. D.; Ernst, James A.; Solloway, Mark; Rubinfeld, Bonnee; Hannoush, Rami N.; Wu, Yan; Polakis, Paul; Costa, Mike

    2010-01-01

    β-catenin-dependent Wnt signaling is initiated as Wnt binds to both the receptor FZD and coreceptor LRP5/6, which then assembles a multimeric complex at the cytoplasmic membrane face to recruit and inactivate the kinase GSK3. The large number and sequence diversity of Wnt isoforms suggest the possibility of domain-specific ligand-coreceptor interactions, and distinct binding sites on LRP6 for Wnt3a and Wnt9b have recently been identified in vitro. Whether mechanistically different interactions between Wnts and coreceptors might mediate signaling remains to be determined. It is also not clear whether coreceptor homodimerization induced extracellularly can activate Wnt signaling, as is the case for receptor tyrosine kinases. We generated monoclonal antibodies against LRP6 with the unexpected ability to inhibit signaling by some Wnt isoforms and potentiate signaling by other isoforms. In cell culture, two antibodies characterized further show reciprocal activities on most Wnts, with one antibody antagonizing and the other potentiating. We demonstrate that these antibodies bind to different regions of LRP6 protein, and inhibition of signaling results from blocking Wnt binding. Antibody-mediated dimerization of LRP6 can potentiate signaling only when a Wnt isoform is also able to bind the complex, presumably recruiting FZD. Endogenous autocrine Wnt signaling in different tumor cell lines can be either antagonized or enhanced by the LRP6 antibodies, indicating expression of different Wnt isoforms. As anticipated from the roles of Wnt signaling in cancer and bone development, antibody activities can also be observed in mice for inhibition of tumor growth and in organ culture for enhancement of bone mineral density. Collectively, our results indicate that separate binding sites for different subsets of Wnt isoforms determine the inhibition or potentiation of signaling conferred by LRP6 antibodies. This complexity of coreceptor-ligand interactions may allow for

  18. Fritz: a secreted frizzled-related protein that inhibits Wnt activity.

    PubMed

    Mayr, T; Deutsch, U; Kühl, M; Drexler, H C; Lottspeich, F; Deutzmann, R; Wedlich, D; Risau, W

    1997-04-01

    Signaling molecules of the Wnt gene family are involved in the regulation of dorso-ventral, segmental and tissue polarity in Xenopus and Drosophila embryos. Members of the frizzled gene family, such as Drosophila frizzled-2 and rat frizzled-1, have been shown encode Wnt binding activity and to engage intracellular signal transduction molecules known to be part of the Wnt signaling pathway. Here we describe the cloning and characterization of Fritz, a mouse (mfiz) and human (hfiz) gene which codes for a secreted protein that is structurally related to the extracellular portion of the frizzled genes from Drosophila and vertebrates. The Fritz protein antagonizes Wnt function when both proteins are ectopically expressed in Xenopus embryos. In early gastrulation, mouse fiz mRNA is expressed in all three germ layers. Later in embryogenesis fiz mRNA is found in the central and peripheral nervous systems, nephrogenic mesenchyme and several other tissues, all of which are sites where Wnt proteins have been implicated in tissue patterning. We propose a model in which Fritz can interfere with the activity of Wnt proteins via their cognate frizzled receptors and thereby modulate the biological responses to Wnt activity in a multitude of tissue sites.

  19. Inversin, Wnt signaling and primary cilia.

    PubMed

    Lienkamp, Soeren; Ganner, Athina; Walz, Gerd

    2012-02-01

    Mutations of the ankyrin-repeat protein Inversin, a member of a diverse family of more than 12 proteins, cause nephronophthisis (NPH), an autosomal recessive cystic kidney disease associated with extra-renal manifestations such as retinitis pigmentosa, cerebellar aplasia and situs inversus. Most NPH gene products (NPHPs) localize to the cilium, and appear to control the transport of cargo protein to the cilium by forming functional networks. Inversin interacts with NPHP1 and NPHP3, and shares with NPHP4 the ability to antagonize Dishevelled-stimulated canonical Wnt signaling, potentially through recruitment of the Anaphase Promoting Complex (APC/C). However, Dishevelled antagonism may be confined towards the basal body, thereby polarizing motile cilia on the cells of the ventral node and respiratory tract. Inversin is essential for recruiting Dishevelled to the plasma membrane in response to activated Frizzled, a crucial step in planar cell polarity signaling. During vertebrate pronephros development, the Inversin-mediated translocation of Dishevelled appears to orchestrate the migration of cells and differentiation of segments that correspond to the mammalian loop of Henle. Thus, defective tubule migration and elongation may contribute to concentration defects and cause cyst formation in patients with NPH. Copyright © 2011 International Society of Differentiation. Published by Elsevier B.V. All rights reserved.

  20. Mutational Analysis of Sclerostin Shows Importance of the Flexible Loop and the Cystine-Knot for Wnt-Signaling Inhibition

    PubMed Central

    Boschert, Verena; van Dinther, Maarten; Weidauer, Stella; van Pee, Katharina; Muth, Eva-Maria; ten Dijke, Peter; Mueller, Thomas D.

    2013-01-01

    The cystine-knot containing protein Sclerostin is an important negative regulator of bone growth and therefore represents a promising therapeutic target. It exerts its biological task by inhibiting the Wnt (wingless and int1) signaling pathway, which participates in bone formation by promoting the differentiation of mesenchymal stem cells to osteoblasts. The core structure of Sclerostin consists of three loops with the first and third loop (Finger 1 and Finger 2) forming a structured β-sheet and the second loop being unstructured and highly flexible. Biochemical data showed that the flexible loop is important for binding of Sclerostin to Wnt co-receptors of the low-density lipoprotein related-protein family (LRP), by interacting with the Wnt co-receptors LRP5 or -6 it inhibits Wnt signaling. To further examine the structural requirements for Wnt inhibition, we performed an extensive mutational study within all three loops of the Sclerostin core domain involving single and multiple mutations as well as truncation of important regions. By this approach we could confirm the importance of the second loop and especially of amino acids Asn92 and Ile94 for binding to LRP6. Based on a Sclerostin variant found in a Turkish family suffering from Sclerosteosis we generated a Sclerostin mutant with cysteines 84 and 142 exchanged thereby removing the third disulfide bond of the cystine-knot. This mutant binds to LRP6 with reduced binding affinity and also exhibits a strongly reduced inhibitory activity against Wnt1 thereby showing that also elements outside the flexible loop are important for inhibition of Wnt by Sclerostin. Additionally, we examined the effect of the mutations on the inhibition of two different Wnt proteins, Wnt3a and Wnt1. We could detect clear differences in the inhibition of these proteins, suggesting that the mechanism by which Sclerostin antagonizes Wnt1 and Wnt3a is fundamentally different. PMID:24312339

  1. Distinct roles and differential expression levels of Wnt5a mRNA isoforms in colorectal cancer cells

    PubMed Central

    Huang, Tsui-Chin; Huang, Chi-Chen; Ko, Chiung-Yuan; Lee, Yi-Chao; Lin, Ding-Yen; Cheng, Ya-Wen

    2017-01-01

    The canonical Wnt/β-catenin pathway is constitutively activated in more than 90% of colorectal cancer (CRC) cases in which β-catenin contributes to CRC cell growth and survival. In contrast to the Wnt/β-catenin pathway, the non-canonical Wnt pathway can antagonize functions of the canonical Wnt/β-catenin pathway. Wnt5a is a key factor in the non-canonical Wnt pathway, and it plays diverse roles in different types of cancers. It was shown that reintroducing Wnt5a into CRC cells resulted in inhibited cell proliferation and impaired cell motility. However, contradictory results were reported describing increased Wnt5a expression being associated with a poor prognosis of CRC patients. Recently, it was shown that the diverse roles of Wnt5a are due to two distinct roles of Wnt5a isoforms. However, the exact roles and functions of the Wnt5a isoforms in CRC remain largely unclear. The present study for the first time showed the ambiguous role of Wnt5a in CRC was due to the encoding of distinct roles of the various Wnt5a mRNA isoforms. A relatively high expression level of the Wnt5a-short (S) isoform transcript and a low expression level of the Wnt5a-long (L) isoform transcript were detected in CRC cell lines and specimens. In addition, high expression levels of the Wnt5a-S mRNA isoform and low expression levels of the Wnt5a-L mRNA isoform were significantly positively correlated with tumor depth of CRC patients. Furthermore, knockdown of the endogenous expression of the Wnt5a-S mRNA isoform in HCT116 cells drastically inhibited their growth ability by inducing apoptosis through induction of FASLG expression and reduction of TNFRSF11B expression. Moreover, reactivation of methylation inactivation of the Wnt5a-L mRNA isoform by treatment with 5-azacytidine (5-Aza) enhanced the siWnt5a-S isoform's ability to induce apoptosis. Finally, we showed that the simultaneous reactivation of Wnt5a-L mRNA isoform and knockdown of Wnt5a-S mRNA isoform expression enhanced siWnt5a

  2. Recent identification of an ERK signal gradient governing planarian regeneration.

    PubMed

    Agata, Kiyokazu; Tasaki, Junichi; Nakajima, Elizabeth; Umesono, Yoshihiko

    2014-06-01

    Planarians have strong regenerative abilities derived from their adult pluripotent stem cell (neoblast) system. However, the molecular mechanisms involved in planarian regeneration have long remained a mystery. In particular, no anterior-specifying factor(s) could be found, although Wnt family proteins had been successfully identified as posterior-specifying factors during planarian regeneration (Gurley et al., 2008; Petersen and Reddien, 2008). A recent textbook of developmental biology therefore proposes a Wnt antagonist as a putative anterior factor (Gilbert, 2013). That is, planarian regeneration was supposed to be explained by a single decreasing gradient of the β-catenin signal from tail to head. However, recently we succeeded in demonstrating that in fact the extracellular-signal regulated kinases (ERK) form a decreasing gradient from head to tail to direct the reorganization of planarian body regionality after amputation (Umesono et al., 2013). Copyright © 2014 Elsevier GmbH. All rights reserved.

  3. Wnt gene loss in flatworms.

    PubMed

    Riddiford, Nick; Olson, Peter D

    2011-10-01

    Wnt genes encode secreted glycoproteins that act in cell-cell signalling to regulate a wide array of developmental processes, ranging from cellular differentiation to axial patterning. Discovery that canonical Wnt/β-catenin signalling is responsible for regulating head/tail specification in planarian regeneration has recently highlighted their importance in flatworm (phylum Platyhelminthes) development, but examination of their roles in the complex development of the diverse parasitic groups has yet to be conducted. Here, we characterise Wnt genes in the model tapeworm Hymenolepis microstoma and mine genomic resources of free-living and parasitic species for the presence of Wnts and downstream signalling components. We identify orthologs through a combination of BLAST and phylogenetic analyses, showing that flatworms have a highly reduced and dispersed complement that includes orthologs of only five subfamilies (Wnt1, Wnt2, Wnt4, Wnt5 and Wnt11) and fewer paralogs in parasitic flatworms (5-6) than in planarians (9). All major signalling components are identified, including antagonists and receptors, and key binding domains are intact, indicating that the canonical (Wnt/β-catenin) and non-canonical (planar cell polarity and Wnt/Ca(2+)) pathways are functional. RNA-Seq data show expression of all Hymenolepis Wnts and most downstream components in adults and larvae with the notable exceptions of wnt1, expressed only in adults, and wnt2 expressed only in larvae. The distribution of Wnt subfamilies in animals corroborates the idea that the last common ancestor of the Cnidaria and Bilateria possessed all contemporary Wnts and highlights the extent of gene loss in flatworms.

  4. Wnt5A regulates ABCB1 expression in multidrug-resistant cancer cells through activation of the non-canonical PKA/β-catenin pathway

    PubMed Central

    Hung, Tsai-Hsien; Hsu, Sheng-Chi; Cheng, Ching-Yi; Choo, Kong-Bung; Tseng, Ching-Ping; Chen, Tse-Ching; Lan, Ying-Wei; Huang, Tsung-Teng; Lai, Hsin-Chih; Chen, Chuan-Mu; Chong, Kowit-Yu

    2014-01-01

    Multidrug resistance in cancer cells arises from altered drug permeability of the cell. We previously reported activation of the Wnt pathway in ABCB1-overexpressed human uterus sarcoma drug-resistant MES-SA/Dx5 cells through active β-catenin and associated transactivation activities, and upregulation of Wnt-targeting genes. In this study, Wnt5A was found to be significantly upregulated in MES-SA/Dx5 and MCF7/ADR2 cells, suggesting an important role for the Wnt5A signaling pathway in cancer drug resistance. Higher cAMP response elements and Tcf/Lef transcription activities were shown in the drug-resistant cancer cells. However, expression of Wnt target genes and CRE activities was downregulated in Wnt5A shRNA stably-transfected MES-SA/Dx5 cells. Cell viability of the drug-resistant cancer cells was also reduced by doxorubicin treatment and Wnt5A shRNA transfection, or by Wnt5A depletion. The in vitro data were supported by immunohistochemical analysis of 24 paired breast cancer biopsies obtained pre- and post-chemotherapeutic treatment. Wnt5A, VEGF and/or ABCB1 were significantly overexpressed after treatment, consistent with clinical chemoresistance. Taken together, the Wnt5A signaling pathway was shown to contribute to regulating the drug-resistance protein ABCB1 and β-catenin-related genes in antagonizing the toxic effects of doxorubicin in the MDR cell lines and in clinical breast cancer samples. PMID:25401518

  5. Novel β-carbolines against colorectal cancer cell growth via inhibition of Wnt/β-catenin signaling

    PubMed Central

    Li, X; Bai, B; Liu, L; Ma, P; Kong, L; Yan, J; Zhang, J; Ye, Z; Zhou, H; Mao, B; Zhu, H; Li, Y

    2015-01-01

    Wnt signaling pathway is aberrantly activated in a variety of cancers, especially in colorectal cancer (CRC), because of mutations in the genes encoding adenomatous polyposis coli (APC), β-catenin and Axin. Small-molecule antagonists of Wnt/β-catenin signaling are attractive candidates for developing effective therapeutics for CRC. In this study, we have identified a novel Wnt signaling inhibitor, isopropyl 9-ethyl-1- (naphthalen-1-yl)-9H-pyrido[3,4-b]indole-3- carboxylate (Z86). Z86 inhibited Wnt reporter activities and the expression of endogenous Wnt signaling target genes in mammalian cells and antagonized the second axis formation of Xenopus embryos induced by Wnt8. We showed that Z86 treatment inhibits GSK3β (Ser9) phosphorylation, leading to its overactivation and promoting the phosphorylation and degradation of β-catenin. In vitro, Z86 selectively inhibited the growth of CRC cells with constitutive Wnt signaling and caused obvious G1-phase arrest of the cell cycle. Notably, in a nude mouse model, Z86 inhibited dramatically the xenografted tumor growth of CRC. Daily intraperitoneal injection of Z86 at 5 mg/kg resulted in >70% reduction in the tumor weight of HCT116 cell origin that was associated with decreased GSK3β (Ser9) phosphorylation and increased β-catenin phosphorylation. Taken together, our findings provide a novel promising chemotype for CRC therapeutics development targeting the canonical Wnt signaling. PMID:27551464

  6. Novel β-carbolines against colorectal cancer cell growth via inhibition of Wnt/β-catenin signaling.

    PubMed

    Li, X; Bai, B; Liu, L; Ma, P; Kong, L; Yan, J; Zhang, J; Ye, Z; Zhou, H; Mao, B; Zhu, H; Li, Y

    2015-01-01

    Wnt signaling pathway is aberrantly activated in a variety of cancers, especially in colorectal cancer (CRC), because of mutations in the genes encoding adenomatous polyposis coli (APC), β-catenin and Axin. Small-molecule antagonists of Wnt/β-catenin signaling are attractive candidates for developing effective therapeutics for CRC. In this study, we have identified a novel Wnt signaling inhibitor, isopropyl 9-ethyl-1- (naphthalen-1-yl)-9H-pyrido[3,4-b]indole-3- carboxylate (Z86). Z86 inhibited Wnt reporter activities and the expression of endogenous Wnt signaling target genes in mammalian cells and antagonized the second axis formation of Xenopus embryos induced by Wnt8. We showed that Z86 treatment inhibits GSK3β (Ser9) phosphorylation, leading to its overactivation and promoting the phosphorylation and degradation of β-catenin. In vitro, Z86 selectively inhibited the growth of CRC cells with constitutive Wnt signaling and caused obvious G1-phase arrest of the cell cycle. Notably, in a nude mouse model, Z86 inhibited dramatically the xenografted tumor growth of CRC. Daily intraperitoneal injection of Z86 at 5 mg/kg resulted in >70% reduction in the tumor weight of HCT116 cell origin that was associated with decreased GSK3β (Ser9) phosphorylation and increased β-catenin phosphorylation. Taken together, our findings provide a novel promising chemotype for CRC therapeutics development targeting the canonical Wnt signaling.

  7. Wnt Signaling and Injury Repair

    PubMed Central

    Whyte, Jemima L.; Smith, Andrew A.; Helms, Jill A.

    2012-01-01

    Wnt signaling is activated by wounding and participates in every subsequent stage of the healing process from the control of inflammation and programmed cell death, to the mobilization of stem cell reservoirs within the wound site. In this review we summarize recent data elucidating the roles that the Wnt pathway plays in the injury repair process. These data provide a foundation for potential Wnt-based therapeutic strategies aimed at stimulating tissue regeneration. PMID:22723493

  8. Lighting Up ERK Activity.

    PubMed

    Shilo, Ben-Zion; Barkai, Naama

    2017-01-23

    Activation of extracellular signal regulated kinase (ERK) is used by many signaling pathways to control tissue patterning in a broad range of multicellular organisms. In this issue of Developmental Cell, Johnson et al. (2017) provide an optogenetic approach to manipulate this pathway with high precision and explore its signaling code. Copyright © 2017 Elsevier Inc. All rights reserved.

  9. Evolutionary inevitability of sexual antagonism.

    PubMed

    Connallon, Tim; Clark, Andrew G

    2014-02-07

    Sexual antagonism, whereby mutations are favourable in one sex and disfavourable in the other, is common in natural populations, yet the root causes of sexual antagonism are rarely considered in evolutionary theories of adaptation. Here, we explore the evolutionary consequences of sex-differential selection and genotype-by-sex interactions for adaptation in species with separate sexes. We show that sexual antagonism emerges naturally from sex differences in the direction of selection on phenotypes expressed by both sexes or from sex-by-genotype interactions affecting the expression of such phenotypes. Moreover, modest sex differences in selection or genotype-by-sex effects profoundly influence the long-term evolutionary trajectories of populations with separate sexes, as these conditions trigger the evolution of strong sexual antagonism as a by-product of adaptively driven evolutionary change. The theory demonstrates that sexual antagonism is an inescapable by-product of adaptation in species with separate sexes, whether or not selection favours evolutionary divergence between males and females.

  10. Wnt5a Signals through DVL1 to Repress Ribosomal DNA Transcription by RNA Polymerase I.

    PubMed

    Dass, Randall A; Sarshad, Aishe A; Carson, Brittany B; Feenstra, Jennifer M; Kaur, Amanpreet; Obrdlik, Ales; Parks, Matthew M; Prakash, Varsha; Love, Damon K; Pietras, Kristian; Serra, Rosa; Blanchard, Scott C; Percipalle, Piergiorgio; Brown, Anthony M C; Vincent, C Theresa

    2016-08-01

    Ribosome biogenesis is essential for cell growth and proliferation and is commonly elevated in cancer. Accordingly, numerous oncogene and tumor suppressor signaling pathways target rRNA synthesis. In breast cancer, non-canonical Wnt signaling by Wnt5a has been reported to antagonize tumor growth. Here, we show that Wnt5a rapidly represses rDNA gene transcription in breast cancer cells and generates a chromatin state with reduced transcription of rDNA by RNA polymerase I (Pol I). These effects were specifically dependent on Dishevelled1 (DVL1), which accumulates in nucleolar organizer regions (NORs) and binds to rDNA regions of the chromosome. Upon DVL1 binding, the Pol I transcription activator and deacetylase Sirtuin 7 (SIRT7) releases from rDNA loci, concomitant with disassembly of Pol I transcription machinery at the rDNA promoter. These findings reveal that Wnt5a signals through DVL1 to suppress rRNA transcription. This provides a novel mechanism for how Wnt5a exerts tumor suppressive effects and why disruption of Wnt5a signaling enhances mammary tumor growth in vivo.

  11. Dishevelled attenuates the repelling activity of Wnt signaling during neurite outgrowth in Caenorhabditis elegans

    PubMed Central

    Zheng, Chaogu; Diaz-Cuadros, Margarete; Chalfie, Martin

    2015-01-01

    Wnt proteins regulate axonal outgrowth along the anterior–posterior axis, but the intracellular mechanisms that modulate the strength of Wnt signaling in axon guidance are largely unknown. Using the Caenorhabditis elegans mechanosensory PLM neurons, we found that posteriorly enriched LIN-44/Wnt acts as a repellent to promote anteriorly directed neurite outgrowth through the LIN-17/Frizzled receptor, instead of controlling neuronal polarity as previously thought. Dishevelled (Dsh) proteins DSH-1 and MIG-5 redundantly mediate the repulsive activity of the Wnt signals to induce anterior outgrowth, whereas DSH-1 also provides feedback inhibition to attenuate the signaling to allow posterior outgrowth against the Wnt gradient. This inhibitory function of DSH-1, which requires its dishevelled, Egl-10, and pleckstrin (DEP) domain, acts by promoting LIN-17 phosphorylation and is antagonized by planar cell polarity signaling components Van Gogh (VANG-1) and Prickle (PRKL-1). Our results suggest that Dsh proteins both respond to Wnt signals to shape neuronal projections and moderate its activity to fine-tune neuronal morphology. PMID:26460008

  12. Resveratrol augments the canonical Wnt signaling pathway in promoting osteoblastic differentiation of multipotent mesenchymal cells

    SciTech Connect

    Zhou, Haibin; Shang, Linshan; Li, Xi; Zhang, Xiyu; Gao, Guimin; Guo, Chenhong; Chen, Bingxi; Liu, Qiji; Gong, Yaoqin; Shao, Changshun

    2009-10-15

    Resveratrol has been shown to possess many health-benefiting effects, including the promotion of bone formation. In this report we investigated the mechanism by which resveratrol promotes osteoblastic differentiation from pluripotent mesenchymal cells. Since Wnt signaling is well documented to induce osteoblastogenesis and bone formation, we characterized the factors involved in Wnt signaling in response to resveratrol treatment. Resveratrol treatment of mesenchymal cells led to an increase in stabilization and nuclear accumulation of {beta}-catenin dose-dependently and time-dependently. As a consequence of the increased nuclear accumulation of {beta}-catenin, the ability to activate transcription of {beta}-catenin-TCF/LEF target genes that are required for osteoblastic differentiation was upregulated. However, resveratrol did not affect the initial step of the Wnt signaling pathway, as resveratrol was as effective in upregulating the activity of {beta}-catenin in cells in which Lrp5 was knocked down as in control cells. In addition, while conditioned medium enriched in Wnt signaling antagonist Dkk1 was able to inhibit Wnt3a-induced {beta}-catenin upregulation, this inhibitory effect can be abolished in resveratrol-treated cells. Furthermore, we showed that the level of glycogen synthase kinase 3{beta} (GSK-3{beta}), which phosphorylates and destabilizes {beta}-catenin, was reduced in response to resveratrol treatment. The phosphorylation of GSK-3{beta} requires extracellular signal-regulated kinase (ERK)1/2. Together, our data indicate that resveratrol promotes osteoblastogenesis and bone formation by augmenting Wnt signaling.

  13. Discovering small molecules that promote cardiomyocyte generation by modulating Wnt signaling

    PubMed Central

    Ni, Terri T.; Rellinger, Eric J.; Mukherjee, Amrita; Stephens, Lauren; Thorne, Cutris A; Kim, Kwangho; Hu, Jiangyong; Xie, Shuying; Lee, Ethan; Marnett, Larry; Hatzopoulos, Antonis K.; Zhong, Tao P.

    2011-01-01

    Summary We have developed a robust in vivo small molecule screen that modulates heart size and cardiomyocyte generation in zebrafish. Three structurally-related compounds (Cardionogen-1 to -3) identified from our screen enlarge the size of the developing heart via myocardial hyperplasia. Increased cardiomyocyte number in Cardionogen-treated embryos is due to expansion of cardiac progenitor cells. In zebrafish embryos and murine embryonic stem (ES) cells, Cardionogen treatment promotes cardiogenesis during and after gastrulation, whereas inhibits heart formation before gastrulation. Cardionogen-induced effects can be antagonized by increasing Wnt/β-catenin signaling activity. We demonstrate that Cardionogen inhibits Wnt/β-catenin-dependent transcription in murine ES cells and zebrafish embryos. Cardionogen can rescue Wnt8-induced cardiomyocyte deficiency and heart-specific phenotypes during development. These findings demonstrate that in vivo small molecule screens targeted on heart size can discover compounds with cardiomyogenic effects and identify underlying target pathways. PMID:22195568

  14. Transcriptional silencing of the Wnt-antagonist DKK1 by promoter methylation is associated with enhanced Wnt signaling in advanced multiple myeloma.

    PubMed

    Kocemba, Kinga A; Groen, Richard W J; van Andel, Harmen; Kersten, Marie José; Mahtouk, Karène; Spaargaren, Marcel; Pals, Steven T

    2012-01-01

    The Wnt/β-catenin pathway plays a crucial role in the pathogenesis of various human cancers. In multiple myeloma (MM), aberrant auto-and/or paracrine activation of canonical Wnt signaling promotes proliferation and dissemination, while overexpression of the Wnt inhibitor Dickkopf1 (DKK1) by MM cells contributes to osteolytic bone disease by inhibiting osteoblast differentiation. Since DKK1 itself is a target of TCF/β-catenin mediated transcription, these findings suggest that DKK1 is part of a negative feedback loop in MM and may act as a tumor suppressor. In line with this hypothesis, we show here that DKK1 expression is low or undetectable in a subset of patients with advanced MM as well as in MM cell lines. This absence of DKK1 is correlated with enhanced Wnt pathway activation, evidenced by nuclear accumulation of β-catenin, which in turn can be antagonized by restoring DKK1 expression. Analysis of the DKK1 promoter revealed CpG island methylation in several MM cell lines as well as in MM cells from patients with advanced MM. Moreover, demethylation of the DKK1 promoter restores DKK1 expression, which results in inhibition of β-catenin/TCF-mediated gene transcription in MM lines. Taken together, our data identify aberrant methylation of the DKK1 promoter as a cause of DKK1 silencing in advanced stage MM, which may play an important role in the progression of MM by unleashing Wnt signaling.

  15. Activation of the ERK pathway in osteoblastic cells, role of gremlin and BMP-2.

    PubMed

    Zanotti, Stefano; Smerdel-Ramoya, Anna; Stadmeyer, Lisa; Canalis, Ernesto

    2008-07-01

    Gremlin is a glycoprotein that binds and antagonizes the actions of bone morphogenetic proteins (BMPs) -2, -4, and -7. Gremlin appears to activate the extracellular regulated kinase (ERK) pathway in endothelial and tumor cells, and as a consequence to have direct cellular effects. To determine whether gremlin has direct effects in osteoblasts, independent of its BMP binding activity, we examined its effects in ST-2 murine stromal cell lines and in primary cultures of murine calvarial osteoblasts. Gremlin did not activate Signaling mothers against decapentaplegic (Smad), and suppressed the BMP-2 induced Smad 1/5/8 phosphorylation and the transactivation of the BMP/Smad reporter construct 12xSBE-Oc-pGL3, confirming its BMPs antagonizing activity. Neither gremlin nor BMP-2 induced ERK 1/2 activation in ST-2 cells or calvarial osteoblasts. Moreover, slight changes in culture conditions induced the phosphorylation of ERK independent from BMP or gremlin exposure. In conclusion, gremlin inhibits BMP-2 signaling and activity, and does not have independent actions on ERK signaling in osteoblasts. Consequently, gremlin activity in osteoblasts can be attributed only to its BMP antagonizing effects. 2008 Wiley-Liss, Inc.

  16. Bone Morphogenetic Protein Antagonist Noggin Promotes Skin Tumorigenesis via Stimulation of the Wnt and Shh Signaling Pathways

    PubMed Central

    Sharov, Andrey A.; Mardaryev, Andrei N.; Sharova, Tatyana Y.; Grachtchouk, Marina; Atoyan, Ruzanna; Byers, H. Randolph; Seykora, John T.; Overbeek, Paul; Dlugosz, Andrzej; Botchkarev, Vladimir A.

    2009-01-01

    Bone morphogenetic proteins (BMPs) play pivotal roles in the regulation of skin development. To study the role of BMPs in skin tumorigenesis, BMP antagonist noggin was used to generate keratin 14-targeted transgenic mice. In contrast to wild-type mice, transgenic mice developed spontaneous hair follicle-derived tumors, which resemble human trichofolliculoma. Global gene expression profiles revealed that in contrast to anagen hair follicles of wild-type mice, tumors of transgenic mice showed stage-dependent increases in the expression of genes encoding the selected components of Wnt and Shh pathways. Specifically, expression of the Wnt ligands increased at the initiation stage of tumor formation, whereas expression of the Wnt antagonist and tumor suppressor Wnt inhibitory factor-1 decreased, as compared with fully developed tumors. In contrast, expression of the components of Shh pathway increased in fully developed tumors, as compared with the tumor placodes. Consistent with the expression data, pharmacological treatment of transgenic mice with Wnt and Shh antagonists resulted in the stage-dependent inhibition of tumor initiation, and progression, respectively. Furthermore, BMP signaling stimulated Wnt inhibitory factor-1 expression and promoter activity in cultured tumor cells and HaCaT keratinocytes, as well as inhibited Shh expression, as compared with the corresponding controls. Thus, tumor suppressor activity of the BMPs in skin epithelium depends on the local concentrations of noggin and is mediated at least in part via stage-dependent antagonizing of Wnt and Shh signaling pathways. PMID:19700758

  17. Wnt/Ryk signaling contributes to neuropathic pain by regulating sensory neuron excitability and spinal synaptic plasticity in rats.

    PubMed

    Liu, Su; Liu, Yue-Peng; Huang, Zhi-Jiang; Zhang, Yan-Kai; Song, Angela A; Ma, Ping-Chuan; Song, Xue-Jun

    2015-12-01

    Treating neuropathic pain continues to be a major clinical challenge and underlying mechanisms of neuropathic pain remain elusive. We have recently demonstrated that Wnt signaling, which is important in developmental processes of the nervous systems, plays critical roles in the development of neuropathic pain through the β-catenin-dependent pathway in the spinal cord and the β-catenin-independent pathway in primary sensory neurons after nerve injury. Here, we report that Wnt signaling may contribute to neuropathic pain through the atypical Wnt/Ryk signaling pathway in rats. Sciatic nerve injury causes a rapid-onset and long-lasting expression of Wnt3a, Wnt5b, and Ryk receptors in primary sensory neurons, and dorsal horn neurons and astrocytes. Spinal blocking of the Wnt/Ryk receptor signaling inhibits the induction and persistence of neuropathic pain without affecting normal pain sensitivity and locomotor activity. Blocking activation of the Ryk receptor with anti-Ryk antibody, in vivo or in vitro, greatly suppresses nerve injury-induced increased intracellular Ca and hyperexcitability of the sensory neurons, and also the enhanced plasticity of synapses between afferent C-fibers and the dorsal horn neurons, and activation of the NR2B receptor and the subsequent Ca-dependent signals CaMKII, Src, ERK, PKCγ, and CREB in sensory neurons and the spinal cord. These findings indicate a critical mechanism underlying the pathogenesis of neuropathic pain and suggest that targeting the Wnt/Ryk signaling may be an effective approach for treating neuropathic pain.

  18. Wnt trafficking: new insights into Wnt maturation, secretion and spreading.

    PubMed

    Port, Fillip; Basler, Konrad

    2010-10-01

    Proteins of the Wnt family are secreted signaling molecules that regulate multiple processes in animal development and control tissue homeostasis in the adult. Wnts spread over considerable distances to regulate gene expression in cells located at distant sites. Paradoxically, Wnts are poorly mobile because of their posttranslational modification with lipids. Recent evidence suggests that several pathways exist that are capable of transforming hydrophobic, insoluble Wnts into long-range signaling molecules. Furthermore, the discovery of Wntless as a protein specifically required for the secretion of Wnt suggests that Wnt trafficking through the secretory pathway is already under special scrutiny. Here, we review recent data on the molecular machinery that controls Wnt secretion and discuss how Wnts can be mobilized for long-range signaling.

  19. Cordycepin promotes apoptosis by modulating the ERK-JNK signaling pathway via DUSP5 in renal cancer cells

    PubMed Central

    Hwang, Jung-Hoo; Joo, Jong Cheon; Kim, Dae Joon; Jo, Eunbi; Yoo, Hwa-Seung; Lee, Kyung-Bok; Park, Soo Jung; Jang, Ik-Soon

    2016-01-01

    Constitutive activation of extracellular signal regulated kinase (ERK)-Jun NH2-terminal kinase (JNK) signaling commonly occurs in tumors. The activation of ERK promotes cell proliferation, whereas that of JNK induces cell apoptosis. However, the apoptotic mechanism of ERK-JNK signaling in cancer is not well understood. Recently, we identified that apoptosis and activation of the JNK signaling pathway were induced after cordycepin treatment in human renal cancer, suggesting that JNK signaling might contribute to TK-10 cell apoptosis. We investigated the apoptotic effects of cordycepin by evaluating the activation of the ERK-JNK signaling pathway in renal cancer TK-10 cells. We found that cordycepin downregulated ERK and DUSP5, upregulated phosphorylated-JNK (p-JNK), and induced apoptosis. Moreover, we showed that siRNA-mediated inhibition of ERK downregulated DUSP5, whereas ERK overexpression upregulated DUSP5, and that DUSP5 knockdown by siRNA upregulated p-JNK. The JNK-specific inhibitor SP600125 upregulated nuclear translocation of β-catenin, and downregulated Dickkopf-1 (Dkk1), which has been shown to be a potent inhibitor of Wnt signaling. Dkk1 knockdown by siRNA upregulated nuclear β-catenin, suggesting the involvement of the Wnt/β-catenin signaling pathway. DUSP5 overexpression in TK-10 cells decreased p-JNK and increased nuclear β-catenin. The decreased Bax activation markedly protected against cordycepin-induced apoptosis. Bax subfamily proteins induced apoptosis through caspase-3. Taken together, we show that JNK signaling activation by cordycepin mediated ERK inhibition, which might have induced Bax translocation and caspase-3 activation via regulation of DUSP5 in TK-10 cells, thereby promoting the apoptosis of TK-10 cells. Targeting ERK-JNK signaling via the apoptotic effects of cordycepin could be a potential therapeutic strategy to treat renal cancer. PMID:27648363

  20. Hmga2 is required for canonical WNT signaling during lung development

    PubMed Central

    2014-01-01

    Background The high-mobility-group (HMG) proteins are the most abundant non-histone chromatin-associated proteins. HMG proteins are present at high levels in various undifferentiated tissues during embryonic development and their levels are strongly reduced in the corresponding adult tissues, where they have been implicated in maintaining and activating stem/progenitor cells. Here we deciphered the role of the high-mobility-group AT-hook protein 2 (HMGA2) during lung development by analyzing the lung of Hmga2-deficient mice (Hmga2 −/− ). Results We found that Hmga2 is expressed in the mouse embryonic lung at the distal airways. Analysis of Hmga2 −/− mice showed that Hmga2 is required for proper cell proliferation and distal epithelium differentiation during embryonic lung development. Hmga2 knockout led to enhanced canonical WNT signaling due to an increased expression of secreted WNT glycoproteins Wnt2b, Wnt7b and Wnt11 as well as a reduction of the WNT signaling antagonizing proteins GATA-binding protein 6 and frizzled homolog 2. Analysis of siRNA-mediated loss-of-function experiments in embryonic lung explant culture confirmed the role of Hmga2 as a key regulator of distal lung epithelium differentiation and supported the causal involvement of enhanced canonical WNT signaling in mediating the effect of Hmga2-loss-of-fuction. Finally, we found that HMGA2 directly regulates Gata6 and thereby modulates Fzd2 expression. Conclusions Our results support that Hmga2 regulates canonical WNT signaling at different points of the pathway. Increased expression of the secreted WNT glycoproteins might explain a paracrine effect by which Hmga2-knockout enhanced cell proliferation in the mesenchyme of the developing lung. In addition, HMGA2-mediated direct regulation of Gata6 is crucial for fine-tuning the activity of WNT signaling in the airway epithelium. Our results are the starting point for future studies investigating the relevance of Hmga2-mediated regulation of

  1. WNT16 antagonises excessive canonical WNT activation and protects cartilage in osteoarthritis.

    PubMed

    Nalesso, Giovanna; Thomas, Bethan Lynne; Sherwood, Joanna Claire; Yu, Jing; Addimanda, Olga; Eldridge, Suzanne Elizabeth; Thorup, Anne-Sophie; Dale, Leslie; Schett, Georg; Zwerina, Jochen; Eltawil, Noha; Pitzalis, Costantino; Dell'Accio, Francesco

    2017-01-01

    Both excessive and insufficient activation of WNT signalling results in cartilage breakdown and osteoarthritis. WNT16 is upregulated in the articular cartilage following injury and in osteoarthritis. Here, we investigate the function of WNT16 in osteoarthritis and the downstream molecular mechanisms. Osteoarthritis was induced by destabilisation of the medial meniscus in wild-type and WNT16-deficient mice. Molecular mechanisms and downstream effects were studied in vitro and in vivo in primary cartilage progenitor cells and primary chondrocytes. The pathway downstream of WNT16 was studied in primary chondrocytes and using the axis duplication assay in Xenopus. WNT16-deficient mice developed more severe osteoarthritis with reduced expression of lubricin and increased chondrocyte apoptosis. WNT16 supported the phenotype of cartilage superficial-zone progenitor cells and lubricin expression. Increased osteoarthritis in WNT16-deficient mice was associated with excessive activation of canonical WNT signalling. In vitro, high doses of WNT16 weakly activated canonical WNT signalling, but, in co-stimulation experiments, WNT16 reduced the capacity of WNT3a to activate the canonical WNT pathway. In vivo, WNT16 rescued the WNT8-induced primary axis duplication in Xenopus embryos. In osteoarthritis, WNT16 maintains a balanced canonical WNT signalling and prevents detrimental excessive activation, thereby supporting the homeostasis of progenitor cells. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/.

  2. Vitamin D Is a Multilevel Repressor of Wnt/β-Catenin Signaling in Cancer Cells

    PubMed Central

    Larriba, María Jesús; González-Sancho, José Manuel; Barbáchano, Antonio; Niell, Núria; Ferrer-Mayorga, Gemma; Muñoz, Alberto

    2013-01-01

    The Wnt/β-catenin signaling pathway is abnormally activated in most colorectal cancers and in a proportion of other neoplasias. This activation initiates or contributes to carcinogenesis by regulating the expression of a large number of genes in tumor cells. The active vitamin D metabolite 1α,25-dihydroxyvitamin D3 (1,25(OH)2D3) inhibits Wnt/β-catenin signaling by several mechanisms at different points along the pathway. Additionally, paracrine actions of 1,25(OH)2D3 on stromal cells may also repress this pathway in neighbouring tumor cells. Here we review the molecular basis for the various mechanisms by which 1,25(OH)2D3 antagonizes Wnt/β-catenin signaling, preferentially in human colon carcinoma cells, and the consequences of this inhibition for the phenotype and proliferation rate. The effect of the vitamin D system on Wnt/β-catenin signaling and tumor growth in animal models will also be commented in detail. Finally, we revise existing data on the relation between vitamin D receptor expression and vitamin D status and the expression of Wnt/β-catenin pathway genes and targets in cancer patients. PMID:24202444

  3. Gene regulatory networks mediating canonical Wnt signal-directed control of pluripotency and differentiation in embryo stem cells.

    PubMed

    Zhang, Xiaoxiao; Peterson, Kevin A; Liu, X Shirley; McMahon, Andrew P; Ohba, Shinsuke

    2013-12-01

    Canonical Wnt signaling supports the pluripotency of embryonic stem cells (ESCs) but also promotes differentiation of early mammalian cell lineages. To explain these paradoxical observations, we explored the gene regulatory networks at play. Canonical Wnt signaling is intertwined with the pluripotency network comprising Nanog, Oct4, and Sox2 in mouse ESCs. In defined media supporting the derivation and propagation of ESCs, Tcf3 and β-catenin interact with Oct4; Tcf3 binds to Sox motif within Oct-Sox composite motifs that are also bound by Oct4-Sox2 complexes. Furthermore, canonical Wnt signaling upregulates the activity of the Pou5f1 distal enhancer via the Sox motif in ESCs. When viewed in the context of published studies on Tcf3 and β-catenin mutants, our findings suggest Tcf3 counters pluripotency by competition with Sox2 at these sites, and Tcf3 inhibition is blocked by β-catenin entry into this complex. Wnt pathway stimulation also triggers β-catenin association at regulatory elements with classic Lef/Tcf motifs associated with differentiation programs. The failure to activate these targets in the presence of a mitogen-activated protein kinase kinase (MEK)/extracellular signal-regulated kinase (ERK) inhibitor essential for ESC culture suggests MEK/ERK signaling and canonical Wnt signaling combine to promote ESC differentiation.

  4. Wnt5a and Wnt11 are essential for second heart field progenitor development

    PubMed Central

    Cohen, Ethan David; Miller, Mayumi F.; Wang, Zichao; Moon, Randall T.; Morrisey, Edward E.

    2012-01-01

    Wnt/β-catenin has a biphasic effect on cardiogenesis, promoting the induction of cardiac progenitors but later inhibiting their differentiation. Second heart field progenitors and expression of the second heart field transcription factor Islet1 are inhibited by the loss of β-catenin, indicating that Wnt/β-catenin signaling is necessary for second heart field development. However, expressing a constitutively active β-catenin with Islet1-Cre also inhibits endogenous Islet1 expression, reflecting the inhibitory effect of prolonged Wnt/β-catenin signaling on second heart field development. We show that two non-canonical Wnt ligands, Wnt5a and Wnt11, are co-required to regulate second heart field development in mice. Loss of Wnt5a and Wnt11 leads to a dramatic loss of second heart field progenitors in the developing heart. Importantly, this loss of Wnt5a and Wnt11 is accompanied by an increase in Wnt/β-catenin signaling, and ectopic Wnt5a/Wnt11 inhibits β-catenin signaling and promotes cardiac progenitor development in differentiating embryonic stem cells. These data show that Wnt5a and Wnt11 are essential regulators of the response of second heart field progenitors to Wnt/β-catenin signaling and that they act by restraining Wnt/β-catenin signaling during cardiac development. PMID:22569553

  5. Targeting Wnt signaling at the neuroimmune interface for dopaminergic neuroprotection/repair in Parkinson’s disease

    PubMed Central

    L’Episcopo, Francesca; Tirolo, Cataldo; Caniglia, Salvo; Testa, Nuccio; Morale, Maria Concetta; Serapide, Maria Francesca; Pluchino, Stefano; Marchetti, Bianca

    2014-01-01

    During the past three decades, the Wingless-type MMTV integration site (Wnt) signaling cascade has emerged as an essential system regulating multiple processes in developing and adult brain. Accumulating evidence points to a dysregulation of Wnt signaling in major neurodegenerative pathologies including Parkinson’s disease (PD), a common neurodegenerative disorder characterized by the progressive loss of midbrain dopaminergic (mDA) neurons and deregulated activation of astrocytes and microglia. This review highlights the emerging link between Wnt signaling and key inflammatory pathways during mDA neuron damage/repair in PD progression. In particular, we summarize recent evidence documenting that aging and neurotoxicant exposure strongly antagonize Wnt/β-catenin signaling in mDA neurons and subventricular zone (SVZ) neuroprogenitors via astrocyte–microglial interactions. Dysregulation of the crosstalk between Wnt/β-catenin signaling and anti-oxidant/anti-inflammatory pathways delineate novel mechanisms driving the decline of SVZ plasticity with age and the limited nigrostriatal dopaminergic self-repair in PD. These findings hold a promise in developing therapies that target Wnt/β-catenin signaling to enhance endogenous restoration and neuronal outcome in age-dependent diseases, such as PD. PMID:24431301

  6. Wnt5a attenuates Wnt3a-induced alkaline phosphatase expression in dental follicle cells

    SciTech Connect

    Sakisaka, Yukihiko; Tsuchiya, Masahiro; Nakamura, Takashi; Tamura, Masato; Shimauchi, Hidetoshi; Nemoto, Eiji

    2015-08-01

    Wnt signaling regulates multiple cellular events such as cell proliferation, differentiation, and apoptosis through β-catenin-dependent canonical and β-catenin-independent noncanonical pathways. Canonical Wnt/β-catenin signaling can promote the differentiation of dental follicle cells, putative progenitor cells for cementoblasts, osteoblasts, and periodontal ligament cells, toward a cementoblast/osteoblast phenotype during root formation, but little is known about the biological significance of noncanonical Wnt signaling in this process. We identified the expression of Wnt5a, a representative noncanonical Wnt ligand, in tooth root lining cells (i.e. precementoblasts/cementoblasts) and dental follicle cells during mouse tooth root development, as assessed by immunohistochemistry. Silencing expression of the Wnt5a gene in a dental follicle cell line resulted in enhancement of the Wnt3a (a representative canonical Wnt ligand)-mediated increase in alkaline phosphatase (ALP) expression. Conversely, treatment with recombinant Wnt5a inhibited the increase in ALP expression, suggesting that Wnt5a signaling functions as a negative regulator of canonical Wnt-mediated ALP expression of dental follicle cells. Wnt5a did not affect the nuclear translocation of β-catenin as well as β-catenin-mediated transcriptional activation of T-cell factor (Tcf) triggered by Wnt3a, suggesting that Wnt5a inhibits the downstream part of the β-catenin-Tcf pathway. These findings suggest the existence of a feedback mechanism between canonical and noncanonical Wnt signaling during the differentiation of dental follicle cells. - Highlights: • Dental follicle cells express Wnt5a during tooth root development. • Silencing of Wnt5a enhances Wnt3a-mediated ALP expression of dental follicle cells. • Conversely, treatment with rWnt5a inhibited the increase in ALP expression. • Wnt5a functions as a negative regulator of Wnt3a-mediated ALP expression.

  7. Rapamycin antagonizes TNF induction of VCAM-1 on endothelial cells by inhibiting mTORC2

    PubMed Central

    Wang, Chen; Qin, Lingfeng; Manes, Thomas D.; Kirkiles-Smith, Nancy C.; Tellides, George

    2014-01-01

    Recruitment of circulating leukocytes into inflamed tissues depends on adhesion molecules expressed by endothelial cells (ECs). Here we report that rapamycin pretreatment reduced the ability of TNF-treated ECs to capture T cells under conditions of venular flow. This functional change was caused by inhibition of TNF-induced expression of vascular cell adhesion molecule-1 (VCAM-1) and could be mimicked by knockdown of mammalian target of rapamycin (mTOR) or rictor, but not raptor, implicating mTORC2 as the target of rapamycin for this effect. Mechanistically, mTORC2 acts through Akt to repress Raf1-MEK1/2-ERK1/2 signaling, and inhibition of mTORC2 consequently results in hyperactivation of ERK1/2. Increased ERK1/2 activity antagonizes VCAM-1 expression by repressing TNF induction of the transcription factor IRF-1. Preventing activation of ERK1/2 reduced the ability of rapamycin to inhibit TNF-induced VCAM-1 expression. In vivo, rapamycin inhibited mTORC2 activity and potentiated activation of ERK1/2. These changes correlated with reduced endothelial expression of TNF-induced VCAM-1, which was restored via pharmacological inhibition of ERK1/2. Functionally, rapamycin reduced infiltration of leukocytes into renal glomeruli, an effect which was partially reversed by inhibition of ERK1/2. These data demonstrate a novel mechanism by which rapamycin modulates the ability of vascular endothelium to mediate inflammation and identifies endothelial mTORC2 as a potential therapeutic target. PMID:24516119

  8. Wnt signaling requires retromer-dependent recycling of MIG-14/Wntless in Wnt-producing cells.

    PubMed

    Yang, Pei-Tzu; Lorenowicz, Magdalena J; Silhankova, Marie; Coudreuse, Damien Y M; Betist, Marco C; Korswagen, Hendrik C

    2008-01-01

    Wnt proteins are secreted signaling molecules that play a central role in development and adult tissue homeostasis. We have previously shown that Wnt signaling requires retromer function in Wnt-producing cells. The retromer is a multiprotein complex that mediates endosome-to-Golgi transport of specific sorting receptors. MIG-14/Wls is a conserved transmembrane protein that binds Wnt and is required in Wnt-producing cells for Wnt secretion. Here, we demonstrate that in the absence of retromer function, MIG-14/Wls is degraded in lysosomes and becomes limiting for Wnt signaling. We show that retromer-dependent recycling of MIG-14/Wls is part of a trafficking pathway that retrieves MIG-14/Wls from the plasma membrane. We propose that MIG-14/Wls cycles between the Golgi and the plasma membrane to mediate Wnt secretion. Regulation of this transport pathway may enable Wnt-producing cells to control the range of Wnt signaling in the tissue.

  9. Wnt your brain be inflamed? Yes, it Wnt!

    PubMed Central

    Marchetti, Bianca; Pluchino, Stefano

    2013-01-01

    The roles of Wnts in neural development, synaptogenesis, and cancer are generally well characterized. Nonetheless, evidence exists that interactions between the immune and nervous systems control major brain regenerative processes ranging from physiological or pathological (reparative) regeneration to neurogenesis and synaptic plasticity. Recent studies describe deregulated Wnt-Fzd signaling in degenerative and inflammatory central nervous system (CNS) disorders, and the expression of Wnt signaling components in the immune system, and in immune-like cells of the mammalian CNS. This would suggest a likely involvement of Wnts in inflammation-driven brain damage and inflammation-directed brain repair. Here, we review how Wnts modulate neuroimmune interactions and offer a perspective on the most challenging therapeutic opportunities for those CNS diseases where injury-reactive Wnt-flavored inflammation precedes secondary neurodegeneration. PMID:23312954

  10. WNT-1 Signaling in Mammary Carcinogenesis

    DTIC Science & Technology

    2002-04-01

    segment polarity gene whose mutant phenotype resembles that of the wingless (Drosophila Wnt-1) mutation (3). arrow encodes a transmembrane receptor...and function ofSpemann’s organizer. Annu. Rev. C Drv. of those caused by mutations in individual Wnt genes . Further- Biaol 13, 611-667 (1997). more, we... mutations of multiple Wnt genes [31]. In the 0.5 nM and thus is significantly higher than Wnt-Fz bind- Xenopus embryo, inhibition of LRP6 function

  11. WNT10A — EDRN Public Portal

    Cancer.gov

    WNT10A, a probable developmental protein, is a ligand for members of the frizzled family of seven transmembrane receptors. It may be a signaling molecule important in CNS development. WNT10A is a member of the WNT family. The WNT gene family consists of structurally related genes which encode secreted signaling proteins. These proteins have been implicated in oncogenesis and in several developmental processes, including regulation of cell fate and patterning during embryogenesis.

  12. ERK1 and ERK2 present functional redundancy in tetrapods despite higher evolution rate of ERK1.

    PubMed

    Buscà, Roser; Christen, Richard; Lovern, Matthew; Clifford, Alexander M; Yue, Jia-Xing; Goss, Greg G; Pouysségur, Jacques; Lenormand, Philippe

    2015-09-03

    The Ras/Raf/MEK/ERK signaling pathway is involved in essential cell processes and it is abnormally activated in ~30 % of cancers and cognitive disorders. Two ERK isoforms have been described, ERK1 and ERK2; ERK2 being regarded by many as essential due to the embryonic lethality of ERK2 knock-out mice, whereas mice lacking ERK1 are viable and fertile. The controversial question of why we have two ERKs and whether they have differential functions or display functional redundancy has not yet been resolved. To investigate this question we used a novel approach based on comparing the evolution of ERK isoforms' sequences and protein expression across vertebrates. We gathered and cloned erk1 and erk2 coding sequences and we examined protein expression of isoforms in brain extracts in all major clades of vertebrate evolution. For the first time, we measured each isoforms' relative protein level in phylogenetically distant animals using anti-phospho antibodies targeting active ERKs. We demonstrate that squamates (lizards, snakes and geckos), despite having both genes, do not express ERK2 protein whereas other tetrapods either do not express ERK1 protein or have lost the erk1 gene. To demonstrate the unexpected squamates' lack of ERK2 expression, we targeted each ERK isoform in lizard primary fibroblasts by specific siRNA-mediated knockdown. We also found that undetectable expression of ERK2 in lizard is compensated by a greater strength of lizard's erk1 promoter. Finally, phylogenetic analysis revealed that ERK1 amino acids sequences evolve faster than ERK2's likely due to genomic factors, including a large difference in gene size, rather than from functional differences since amino acids essential for function are kept invariant. ERK isoforms appeared by a single gene duplication at the onset of vertebrate evolution at least 400 Mya. Our results demonstrate that tetrapods can live by expressing either one or both ERK isoforms, supporting the notion that ERK1/2 act

  13. Opposing effects of Elk-1 multisite phosphorylation shape its response to ERK activation.

    PubMed

    Mylona, Anastasia; Theillet, Francois-Xavier; Foster, Charles; Cheng, Tammy M; Miralles, Francesc; Bates, Paul A; Selenko, Philipp; Treisman, Richard

    2016-10-14

    Multisite phosphorylation regulates many transcription factors, including the serum response factor partner Elk-1. Phosphorylation of the transcriptional activation domain (TAD) of Elk-1 by the protein kinase ERK at multiple sites potentiates recruitment of the Mediator transcriptional coactivator complex and transcriptional activation, but the roles of individual phosphorylation events had remained unclear. Using time-resolved nuclear magnetic resonance spectroscopy, we found that ERK2 phosphorylation proceeds at markedly different rates at eight TAD sites in vitro, which we classified as fast, intermediate, and slow. Mutagenesis experiments showed that phosphorylation of fast and intermediate sites promoted Mediator interaction and transcriptional activation, whereas modification of slow sites counteracted both functions, thereby limiting Elk-1 output. Progressive Elk-1 phosphorylation thus ensures a self-limiting response to ERK activation, which occurs independently of antagonizing phosphatase activity.

  14. Wnt signaling in the murine diastema

    PubMed Central

    Porntaveetus, Thantrira; Ohazama, Atsushi; Choi, Hong Y.; Herz, Joachim

    2012-01-01

    The correct number and shape of teeth are critical factors for an aesthetic and functional dentition. Understanding the molecular mechanisms regulating tooth number and shape are therefore important in orthodontics. Mice have only one incisor and three molars in each jaw quadrant that are divided by a tooth-less region, the diastema. Although mice lost teeth in the diastema during evolution, the remnants of the evolutionary lost teeth are observed as transient epithelial buds in the wild-type diastema during early stages of development. Shh and Fgf signaling pathways that are essential for tooth development have been shown to be repressed in the diastema. It remains unclear however how Wnt signaling, that is also required for tooth development, is regulated in the diastema. In this study we found that in the embryonic diastema, Wnt5a expression was observed in mesenchyme, whereas Wnt4 and Wnt10b were expressed in epithelium. The expression of Wnt6 and Wnt11 was found in both tissues. The Wnt co-receptor, Lrp6, was weakly expressed in the diastema overlapping with weak Lrp4 expression, a co-receptor that inhibits Wnt signaling. Secreted Wnt inihibitors Dkk1, Dkk2, and Dkk3 were also expressed in the diastema. Lrp4 mutant mice develop supernumerary teeth in the diastema that is accompanied by upregulation of Wnt signaling and Lrp6 expression. Wnt signaling is thus usually attenuated in the diastema by these secreted and membrane bound Wnt inhibitors. PMID:21531785

  15. Tay Bridge Is a Negative Regulator of EGFR Signalling and Interacts with Erk and Mkp3 in the Drosophila melanogaster Wing

    PubMed Central

    Molnar, Cristina; de Celis, Jose F.

    2013-01-01

    The regulation of Extracellular regulated kinase (Erk) activity is a key aspect of signalling by pathways activated by extracellular ligands acting through tyrosine kinase transmembrane receptors. In this process, participate proteins with kinase activity that phosphorylate and activate Erk, as well as different phosphatases that inactivate Erk by de-phosphorylation. The state of Erk phosphorylation affects not only its activity, but also its subcellular localization, defining the repertoire of Erk target proteins, and consequently, the cellular response to Erk. In this work, we characterise Tay bridge as a novel component of the EGFR/Erk signalling pathway. Tay bridge is a large nuclear protein with a domain of homology with human AUTS2, and was previously identified due to the neuronal phenotypes displayed by loss-of-function mutations. We show that Tay bridge antagonizes EGFR signalling in the Drosophila melanogaster wing disc and other tissues, and that the protein interacts with both Erk and Mkp3. We suggest that Tay bridge constitutes a novel element involved in the regulation of Erk activity, acting as a nuclear docking for Erk that retains this protein in an inactive form in the nucleus. PMID:24348264

  16. Tay bridge is a negative regulator of EGFR signalling and interacts with Erk and Mkp3 in the Drosophila melanogaster wing.

    PubMed

    Molnar, Cristina; de Celis, Jose F

    2013-01-01

    The regulation of Extracellular regulated kinase (Erk) activity is a key aspect of signalling by pathways activated by extracellular ligands acting through tyrosine kinase transmembrane receptors. In this process, participate proteins with kinase activity that phosphorylate and activate Erk, as well as different phosphatases that inactivate Erk by de-phosphorylation. The state of Erk phosphorylation affects not only its activity, but also its subcellular localization, defining the repertoire of Erk target proteins, and consequently, the cellular response to Erk. In this work, we characterise Tay bridge as a novel component of the EGFR/Erk signalling pathway. Tay bridge is a large nuclear protein with a domain of homology with human AUTS2, and was previously identified due to the neuronal phenotypes displayed by loss-of-function mutations. We show that Tay bridge antagonizes EGFR signalling in the Drosophila melanogaster wing disc and other tissues, and that the protein interacts with both Erk and Mkp3. We suggest that Tay bridge constitutes a novel element involved in the regulation of Erk activity, acting as a nuclear docking for Erk that retains this protein in an inactive form in the nucleus.

  17. Norovirus Mechanisms of Immune Antagonism

    PubMed Central

    Roth, Alexa N.; Karst, Stephanie M.

    2015-01-01

    Noroviruses are a leading cause of gastroenteritis outbreaks globally. Several lines of evidence indicate that noroviruses can antagonize or evade host immune responses, including the absence of long-lasting immunity elicited during a primary norovirus exposure and the ability of noroviruses to establish prolonged infections that are associated with protracted viral shedding. Specific norovirus proteins possessing immune antagonist activity have been described in recent years although mechanistic insight in most cases is limited. In this review, we discuss these emerging strategies used by noroviruses to subvert the immune response, including the actions of two nonstructural proteins (p48 and p22) to impair cellular protein trafficking and secretory pathways; the ability of the VF1 protein to inhibit cytokine induction; and the ability of the minor structural protein VP2 to regulate antigen presentation. We also discuss the current state of the understanding of host and viral factors regulating the establishment of persistent norovirus infections along the gastrointestinal tract. A more detailed understanding of immune antagonism by pathogenic viruses will inform prevention and treatment of disease. PMID:26673810

  18. Activation and Inhibition of The Wnt3A Signaling Pathway in Buffalo (Bubalus bubalis) Embryonic Stem Cells: Effects of WNT3A, Bio and Dkk1.

    PubMed

    Zandi, Mohammad; Shah, Syed Mohamad; Muzaffar, Musharifa; Kumar Singh, Manoj; Palta, Prabhat; Kumar Singla, Suresh; Sham Manik, Radhey; Chauhan, Manmohan Singh

    2015-01-01

    maintain the pluripotency of ES cell-like cells both as an exogenous growth factor as well as an endogenously expressed gene. It complements the absence of FGF-2 and LIF, otherwise propounded essential for buffalo ES cell culture. WNT3A antagonizes the inhibitory effects of Dkk1 and acts in combination with its activator, Bio, to activate the Wnt signaling pathway.

  19. Developmental ERK Signaling Goes Digital.

    PubMed

    Dessauges, Coralie; Pertz, Olivier

    2017-09-11

    Reporting in Developmental Cell, de la Cova et al. (2017) present a biosensor to measure ERK activity dynamics in C. elegans larvae. They find that fate decision signaling involves frequency-modulated, digital ERK activity pulses. These findings may explain how graded morphogen signals are converted into precise and robust cell fate patterns. Copyright © 2017 Elsevier Inc. All rights reserved.

  20. Wnt Signaling in Cancer Stem Cell Biology

    PubMed Central

    de Sousa e Melo, Felipe; Vermeulen, Louis

    2016-01-01

    Aberrant regulation of Wnt signaling is a common theme seen across many tumor types. Decades of research have unraveled the epigenetic and genetic alterations that result in elevated Wnt pathway activity. More recently, it has become apparent that Wnt signaling levels identify stem-like tumor cells that are responsible for fueling tumor growth. As therapeutic targeting of these tumor stem cells is an intense area of investigation, a concise understanding on how Wnt activity relates to cancer stem cell traits is needed. This review attempts at summarizing the intricacies between Wnt signaling and cancer stem cell biology with a special emphasis on colorectal cancer. PMID:27355964

  1. Loss Of Klotho During Melanoma Progression Leads To Increased Filamin Cleavage, Increased Wnt5A Expression and Enhanced Melanoma Cell Motility

    PubMed Central

    Camilli, Tura C.; Xu, Mai; O'Connell, Michael P.; Chien, Bonnie; Frank, Brittany P.; Subaran, Sarah; Indig, Fred E.; Morin, Patrice J.; Hewitt, Stephen M.; Weeraratna, Ashani T.

    2010-01-01

    Summary We have previously shown that Wnt5A-mediated signaling can promote melanoma metastasis. It has been shown that Wnt signaling is antagonized by the protein Klotho, which has been implicated in aging. We show here that in melanoma cells, expressions of Wnt5A and Klotho are inversely correlated. In the presence of recombinant Klotho (rKlotho) we show that Wnt5A internalization and signaling is decreased in high Wnt5A expressing cells. Moreover, in the presence of rKlotho, we observe an increase in Wnt5A remaining in the medium, coincident with an increase in sialidase activity and decrease in syndecan expression. These effects can be inhibited using a sialidase inhibitor. In addition to its effects on Wnt5A internalization, we also demonstrate that Klotho decreases melanoma cell invasive potential by a second mechanism, that involves the inhibition of calpain and a resultant decrease in filamin cleavage, which we demonstrate is critical for melanoma cell motility. PMID:20955350

  2. Fresh WNT into the regulation of mitosis.

    PubMed

    Stolz, Ailine; Bastians, Holger

    2015-01-01

    Canonical Wnt signaling triggering β-catenin-dependent gene expression contributes to cell cycle progression, in particular at the G1/S transition. Recently, however, it became clear that the cell cycle can also feed back on Wnt signaling at the G2/M transition. This is illustrated by the fact that mitosis-specific cyclin-dependent kinases can phosphorylate the Wnt co-receptor LRP6 to prime the pathway for incoming Wnt signals when cells enter mitosis. In addition, there is accumulating evidence that various Wnt pathway components might exert additional, Wnt-independent functions that are important for proper regulation of mitosis. The importance of Wnt pathways during mitosis was most recently enforced by the discovery of Wnt signaling contributing to the stabilization of proteins other than β-catenin, specifically at G2/M and during mitosis. This Wnt-mediated stabilization of proteins, now referred to as Wnt/STOP, might on one hand contribute to maintaining a critical cell size required for cell division and, on the other hand, for the faithful execution of mitosis itself. In fact, most recently we have shown that Wnt/STOP is required for ensuring proper microtubule dynamics within mitotic spindles, which is pivotal for accurate chromosome segregation and for the maintenance of euploidy.

  3. Wnt5a Signaling in Cancer

    PubMed Central

    Asem, Marwa S.; Buechler, Steven; Wates, Rebecca Burkhalter; Miller, Daniel L.; Stack, M. Sharon

    2016-01-01

    Wnt5a is involved in activating several non-canonical WNT signaling pathways, through binding to different members of the Frizzled- and Ror-family receptors. Wnt5a signaling is critical for regulating normal developmental processes, including proliferation, differentiation, migration, adhesion and polarity. However, the aberrant activation or inhibition of Wnt5a signaling is emerging as an important event in cancer progression, exerting both oncogenic and tumor suppressive effects. Recent studies show the involvement of Wnt5a in regulating cancer cell invasion, metastasis, metabolism and inflammation. In this article, we review findings regarding the molecular mechanisms and roles of Wnt5a signaling in various cancer types, and highlight Wnt5a in ovarian cancer. PMID:27571105

  4. Disulfide Bond Requirements for Active Wnt Ligands*

    PubMed Central

    MacDonald, Bryan T.; Hien, Annie; Zhang, Xinjun; Iranloye, Oladoyin; Virshup, David M.; Waterman, Marian L.; He, Xi

    2014-01-01

    Secreted Wnt lipoproteins are cysteine-rich and lipid-modified morphogens that bind to the Frizzled (FZD) receptor and LDL receptor-related protein 6 (LRP6). Wnt engages FZD through protruding thumb and index finger domains, which are each assembled from paired β strands secured by disulfide bonds and grasp two sides of the FZD ectodomain. The importance of Wnt disulfide bonds has been assumed but uncharacterized. We systematically analyzed cysteines and associated disulfide bonds in the prototypic Wnt3a. Our data show that mutation of any individual cysteine of Wnt3a results in covalent Wnt oligomers through ectopic intermolecular disulfide bond formation and diminishes/abolishes Wnt signaling. Although individual cysteine mutations in the amino part of the saposin-like domain and in the base of the index finger are better tolerated and permit residual Wnt3a secretion/activity, those in the amino terminus, the thumb, and at the tip of the index finger are incompatible with secretion and/or activity. A few select double cysteine mutants based on the disulfide bond pattern restore Wnt secretion/activity. Further, a double cysteine mutation at the index finger tip results in a Wnt3a with normal secretion but minimal FZD binding and dominant negative properties. Our results experimentally validate predictions from the Wnt crystal structure and highlight critical but different roles of the saposin-like and cytokine-like domains, including the thumb and the index finger in Wnt folding/secretion and FZD binding. Finally, we modified existing expression vectors for 19 epitope-tagged human WNT proteins by removal of a tag-supplied ectopic cysteine, thereby generating tagged WNT ligands active in canonical and non-canonical signaling. PMID:24841207

  5. WNT5A-JNK regulation of vascular insulin resistance in human obesity.

    PubMed

    Farb, Melissa G; Karki, Shakun; Park, Song-Young; Saggese, Samantha M; Carmine, Brian; Hess, Donald T; Apovian, Caroline; Fetterman, Jessica L; Bretón-Romero, Rosa; Hamburg, Naomi M; Fuster, José J; Zuriaga, María A; Walsh, Kenneth; Gokce, Noyan

    2016-12-01

    Obesity is associated with the development of vascular insulin resistance; however, pathophysiological mechanisms are poorly understood. We sought to investigate the role of WNT5A-JNK in the regulation of insulin-mediated vasodilator responses in human adipose tissue arterioles prone to endothelial dysfunction. In 43 severely obese (BMI 44±11 kg/m(2)) and five metabolically normal non-obese (BMI 26±2 kg/m(2)) subjects, we isolated arterioles from subcutaneous and visceral fat during planned surgeries. Using videomicroscopy, we examined insulin-mediated, endothelium-dependent vasodilator responses and characterized adipose tissue gene and protein expression using real-time polymerase chain reaction and Western blot analyses. Immunofluorescence was used to quantify endothelial nitric oxide synthase (eNOS) phosphorylation. Insulin-mediated vasodilation was markedly impaired in visceral compared to subcutaneous vessels from obese subjects (p<0.001), but preserved in non-obese individuals. Visceral adiposity was associated with increased JNK activation and elevated expression of WNT5A and its non-canonical receptors, which correlated negatively with insulin signaling. Pharmacological JNK antagonism with SP600125 markedly improved insulin-mediated vasodilation by sixfold (p<0.001), while endothelial cells exposed to recombinant WNT5A developed insulin resistance and impaired eNOS phosphorylation (p<0.05). We observed profound vascular insulin resistance in the visceral adipose tissue arterioles of obese subjects that was associated with up-regulated WNT5A-JNK signaling and impaired endothelial eNOS activation. Pharmacological JNK antagonism markedly improved vascular endothelial function, and may represent a potential therapeutic target in obesity-related vascular disease. © The Author(s) 2016.

  6. Matrix Contraction by Dermal Fibroblasts Requires Transforming Growth Factor-β/Activin-Linked Kinase 5, Heparan Sulfate-Containing Proteoglycans, and MEK/ERK

    PubMed Central

    Chen, Yunliang; Shi-wen, Xu; van Beek, Jonathan; Kennedy, Laura; McLeod, Marilyn; Renzoni, Elisabetta A.; Bou-Gharios, George; Wilcox-Adelman, Sarah; Goetinck, Paul F.; Eastwood, Mark; Black, Carol M.; Abraham, David J.; Leask, Andrew

    2005-01-01

    Scarring is characterized by excessive synthesis and contraction of extracellular matrix. Here, we show that fibroblasts from scarred (lesional) areas of patients with the chronic fibrotic disorder diffuse scleroderma [diffuse systemic sclerosis (dSSc)] show an enhanced ability to adhere to and contract extracellular matrix, relative to fibroblasts from unscarred (nonlesional) areas of dSSc patients and dermal fibroblasts from normal, healthy individuals. The contractile abilities of normal and dSSc dermal fibroblasts were suppressed by blocking heparin sulfate-containing proteoglycan biosynthesis or antagonizing transforming growth factor-β receptor type I [activin-linked kinase (ALK5)] or ras/mitogen-activated protein kinase (MEK)/extracellular signal-regulated kinase (ERK). Compared with both normal and nonlesional fibroblasts, lesional dSSc fibroblasts overexpressed the heparin sulfate-containing proteoglycan syndecan 4. We also found that the procontractile signals from transforming growth factor (TGF)-β were integrated through syndecan 4 and MEK/ERK because the ability of TGFβ to induce contraction of dermal fibroblasts was prevented by MEK antagonism. TGFβ could not induce a contractile phenotype or phosphorylate ERK in syndecan 4−/− dermal fibroblasts. These results suggest that integrating TGFβ and ERK signals via syndecan 4 is essential for the contractile ability of dermal fibroblasts. We conclude that antagonizing MEK/ERK, TGFβ1/ALK5, or syndecan 4 may alleviate scarring in chronic fibrotic disease. PMID:16314481

  7. Phenotypic spandrel: absolute discrimination and ligand antagonism

    NASA Astrophysics Data System (ADS)

    François, Paul; Hemery, Mathieu; Johnson, Kyle A.; Saunders, Laura N.

    2016-12-01

    We consider the general problem of sensitive and specific discrimination between biochemical species. An important instance is immune discrimination between self and not-self, where it is also observed experimentally that ligands just below the discrimination threshold negatively impact response, a phenomenon called antagonism. We characterize mathematically the generic properties of such discrimination, first relating it to biochemical adaptation. Then, based on basic biochemical rules, we establish that, surprisingly, antagonism is a generic consequence of any strictly specific discrimination made independently from ligand concentration. Thus antagonism constitutes a ‘phenotypic spandrel’: a phenotype existing as a necessary by-product of another phenotype. We exhibit a simple analytic model of discrimination displaying antagonism, where antagonism strength is linear in distance from the detection threshold. This contrasts with traditional proofreading based models where antagonism vanishes far from threshold and thus displays an inverted hierarchy of antagonism compared to simpler models. The phenotypic spandrel studied here is expected to structure many decision pathways such as immune detection mediated by TCRs and FCɛRIs, as well as endocrine signalling/disruption.

  8. Coordination of kidney organogenesis by Wnt signaling.

    PubMed

    Halt, Kimmo; Vainio, Seppo

    2014-04-01

    Several Wnt proteins are expressed in the embryonic kidney during various stages of development. Gene knockout models and ex vivo studies have provided strong evidence that Wnt-mediated signals are essential in renal ontogeny. Perhaps the most critical factors, Wnt9b and Wnt4, function during the early phase when the cap mesenchyme is induced to undergo morphogenesis into a nephron. Wnt11 controls early ureteric bud branching and contributes to the final kidney size. In addition to its inductive role, later on Wnt9b plays a significant role in the convergent extension of the tubular epithelial cells, while Wnt4 signaling controls smooth muscle cell fates in the medulla. Wnt7b has a specific function together with its likely antagonist Dkk1 in controlling the morphogenesis of the renal medulla. The signal-transduction mechanisms of the Wnts in kidney ontogeny have not been resolved, but studies characterizing the downstream signaling pathways are emerging. Aberrant Wnt signaling may lead to kidney diseases ranging from fatal kidney agenesis to more benign phenotypes. Wnt-mediated signaling regulates several critical aspects of kidney development from the early inductive stages to later steps of tubular epithelial maturation.

  9. miR-135b Stimulates Osteosarcoma Recurrence and Lung Metastasis via Notch and Wnt/β-Catenin Signaling.

    PubMed

    Jin, Hua; Luo, Song; Wang, Yun; Liu, Chang; Piao, Zhenghao; Xu, Meng; Guan, Wei; Li, Qing; Zou, Hua; Tan, Qun-You; Yang, Zhen-Zhou; Wang, Yan; Wang, Dong; Xu, Cheng-Xiong

    2017-09-15

    Cancer stem cells (CSCs) play an important role in osteosarcoma (OS) metastasis and recurrence, and both Wnt/β-catenin and Notch signaling are essential for the development of the biological traits of CSCs. However, the mechanism that underlies the simultaneous hyperactivation of both Wnt/β-catenin and Notch signaling in OS remains unclear. Here, we report that expression of miR-135b correlates with the overall and recurrence-free survival of OS patients, and that miR-135b has an activating effect on both Wnt/β-catenin and Notch signaling. The overexpression of miR-135b simultaneously targets multiple negative regulators of the Wnt/β-catenin and Notch signaling pathways, including glycogen synthase kinase-3 beta (GSK3β), casein kinase 1a (CK1α), and ten-eleven translocation 3 (TET3). Therefore, upregulated miR-135b promotes CSC traits, lung metastasis, and tumor recurrence in OS. Notably, antagonizing miR-135b potently inhibits OS lung metastasis, cancer cell stemness, CSC-induced tumor formation, and recurrence in xenograft animal models. These findings suggest that miR-135b mediates the constitutive activation of Wnt/β-catenin and Notch signaling, and that the inhibition of miR-135b is a novel strategy to inhibit tumor metastasis and prevent CSC-induced recurrence in OS. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

  10. The Wnt Blows: On the Functional Role of Wnt Signaling in Mycobacterium tuberculosis Infection and Beyond.

    PubMed

    Brandenburg, Julius; Reiling, Norbert

    2016-01-01

    In recent years, it has become apparent that the Wnt signaling pathway, known for its essential functions in embryonic development and tissue homeostasis, exerts immunomodulatory functions during inflammation and infection. Most functional studies indicate that Wnt5a exerts pro-inflammatory functions on its cellular targets, which include various types of immune and non-immune cells. Wnt5a expression has also been linked to the pathogenesis of chronic inflammatory diseases. Activation of beta-catenin-dependent Wnt signaling, e.g., by Wnt3a, has however been shown to limit inflammation by interfering with the nuclear factor kappa-light chain-enhancer of activated B-cells (NF-kappaB) pathway. This review focuses on the regulation of Wnt5a, Wnt3a, and the recently identified Wnt6 and their functional role in bacterial infections with a primary focus on pulmonary tuberculosis, a leading infectious cause of morbidity and mortality worldwide.

  11. The Wnt Blows: On the Functional Role of Wnt Signaling in Mycobacterium tuberculosis Infection and Beyond

    PubMed Central

    Brandenburg, Julius; Reiling, Norbert

    2016-01-01

    In recent years, it has become apparent that the Wnt signaling pathway, known for its essential functions in embryonic development and tissue homeostasis, exerts immunomodulatory functions during inflammation and infection. Most functional studies indicate that Wnt5a exerts pro-inflammatory functions on its cellular targets, which include various types of immune and non-immune cells. Wnt5a expression has also been linked to the pathogenesis of chronic inflammatory diseases. Activation of beta-catenin-dependent Wnt signaling, e.g., by Wnt3a, has however been shown to limit inflammation by interfering with the nuclear factor kappa-light chain-enhancer of activated B-cells (NF-kappaB) pathway. This review focuses on the regulation of Wnt5a, Wnt3a, and the recently identified Wnt6 and their functional role in bacterial infections with a primary focus on pulmonary tuberculosis, a leading infectious cause of morbidity and mortality worldwide. PMID:28082976

  12. Decorin Antagonizes the Angiogenic Network

    PubMed Central

    Neill, Thomas; Painter, Hannah; Buraschi, Simone; Owens, Rick T.; Lisanti, Michael P.; Schaefer, Liliana; Iozzo, Renato V.

    2012-01-01

    Decorin, a small leucine-rich proteoglycan, inhibits tumor growth by antagonizing multiple receptor tyrosine kinases including EGFR and Met. Here, we investigated decorin during normoxic angiogenic signaling. An angiogenic PCR array revealed a profound decorin-evoked transcriptional inhibition of pro-angiogenic genes, such as HIF1A. Decorin evoked a reduction of hypoxia inducible factor (HIF)-1α and vascular endothelial growth factor A (VEGFA) in MDA-231 breast carcinoma cells expressing constitutively-active HIF-1α. Suppression of Met with decorin or siRNA evoked a similar reduction of VEGFA by attenuating downstream β-catenin signaling. These data establish a noncanonical role for β-catenin in regulating VEGFA expression. We found that exogenous decorin induced expression of thrombospondin-1 and TIMP3, two powerful angiostatic agents. In contrast, decorin suppressed both the expression and enzymatic activity of matrix metalloprotease (MMP)-9 and MMP-2, two pro-angiogenic proteases. Our data establish a novel duality for decorin as a suppressor of tumor angiogenesis under normoxia by simultaneously down-regulating potent pro-angiogenic factors and inducing endogenous anti-angiogenic agents. PMID:22194599

  13. The canonical Wnt signaling pathway in autism.

    PubMed

    Zhang, Yinghua; Yuan, Xiangshan; Wang, Zhongping; Li, Ruixi

    2014-01-01

    Mounting attention is being focused on the canonical Wnt signaling pathway which has been implicated in the pathogenesis of autism in some our and other recent studies. The canonical Wnt pathway is involved in cell proliferation, differentiation and migration, especially during nervous system development. Given its various functions, dysfunction of the canonical Wnt pathway may exert adverse effects on neurodevelopment and therefore leads to the pathogenesis of autism. Here, we review human and animal studies that implicate the canonical Wnt signal transduction pathway in the pathogenesis of autism. We also describe the crosstalk between the canonical Wnt pathway and the Notch signaling pathway in several types of autism spectrum disorders, including Asperger syndrome and Fragile X. Further research on the crosstalk between the canonical Wnt signaling pathway and other signaling cascades in autism may be an efficient avenue to understand the etiology of autism and ultimately lead to alternative medications for autism-like phenotypes.

  14. Role of Wnt signaling in fracture healing.

    PubMed

    Xu, Huiyun; Duan, Jing; Ning, Dandan; Li, Jingbao; Liu, Ruofei; Yang, Ruixin; Jiang, Jean X; Shang, Peng

    2014-12-01

    The Wnt signaling pathway is well known to play major roles in skeletal development and homeostasis. In certain aspects, fracture repair mimics the process of bone embryonic development. Thus, the importance of Wnt signaling in fracture healing has become more apparent in recent years. Here, we summarize recent research progress in the area, which may be conducive to the development of Wnt-based therapeutic strategies for bone repair.

  15. Wnt signaling in kidney tubulointerstitium during disease.

    PubMed

    Maarouf, Omar H; Ikeda, Yoichiro; Humphreys, Benjamin D

    2015-02-01

    The evolutionary conserved Wnt signaling transduction pathway plays essential roles in a wide array of biologic processes including embryonic development, branching morphogenesis, proliferation and carcinogenesis. Over the past ten years it has become increasingly clear that Wnt signaling also regulates the response of adult organs to disease processes, including kidney disease. This review will focus on the growing literature implicating important roles for Wnt signaling during disease in two separate kidney compartments: the tubular epithelium and the interstitium.

  16. WNT10A and isolated hypodontia.

    PubMed

    Kantaputra, Piranit; Sripathomsawat, Warissara

    2011-05-01

    WNT10A has been associated with various syndromes with ectodermal dysplasia from severe autosomal recessive SchO?pf-Schulz-Passarge syndrome to odonto-onycho-dermal dysplasia and autosomal dominant hypodontia. We report WNT10A mutations in an American family of which four members are affected with isolated hypodontia or microdontia. Here we demonstrate that in addition to MSX1, PAX9, AXIN2, and EDA, mutations in WNT10A can cause isolated hypodontia.

  17. The Evolution of the Wnt Pathway

    PubMed Central

    Holstein, Thomas W.

    2012-01-01

    Wnt genes are important regulators of embryogenesis and cell differentiation in vertebrates and insects. New data revealed by comparative genomics have now shown that members of the Wnt signaling pathway can be found in all clades of metazoans, but not in fungi, plants, or unicellular eukaryotes. This article focuses on new data from recent genomic analyses of several basal metazoan organisms, providing evidence that the Wnt pathway was a primordial signaling pathway during evolution. The formation of a Wnt signaling center at the site of gastrulation was instrumental for the formation of a primary, anterior–posterior body axis, which can be traced throughout animal evolution. PMID:22751150

  18. Wnt signaling in development and disease.

    PubMed

    Freese, Jennifer L; Pino, Darya; Pleasure, Samuel J

    2010-05-01

    The Wnt signaling pathway is one of the central morphogenic signaling pathways regulating early vertebrate development. In recent years, it has become clear that the Wnt pathway also regulates many aspects of nervous system development from the patterning stage through the regulation of neural plasticity. In this review, we first present an overview of the components of the Wnt signaling pathway and then go on to discuss the literature describing the multitude of roles of Wnts in nervous system. In the latter portion of the review, we turn to the ways that defects in Wnt signaling lead to neurologic disease.

  19. Rescuing failed oral implants via Wnt activation

    PubMed Central

    Yin, Xing; Li, Jingtao; Chen, Tao; Mouraret, Sylvain; Dhamdhere, Girija; Brunski, John B.; Zou, Shujuan; Helms, Jill A.

    2016-01-01

    Aim Implant osseointegration is not always guaranteed and once fibrous encapsulation occurs clinicians have few options other than implant removal. Our goal was to test whether a WNT protein therapeutic could rescue such failed implants. Material and Methods Titanium implants were placed in over-sized murine oral osteotomies. A lack of primary stability was verified by mechanical testing. Interfacial strains were estimated by finite element modelling and histology coupled with histomorphometry confirmed the lack of peri-implant bone. After fibrous encapsulation was established peri-implant injections of a liposomal formulation of WNT3A protein (L-WNT3A) or liposomal PBS (L-PBS) were then initiated. Quantitative assays were employed to analyse the effects of L-WNT3A treatment. Results Implants in gap-type interfaces exhibited high interfacial strains and no primary stability. After verification of implant failure, L-WNT3A or L-PBS injections were initiated. L-WNT3A induced a rapid, significant increase in Wnt responsiveness in the peri-implant environment, cell proliferation and osteogenic protein expression. The amount of peri-implant bone and bone in contact with the implant were significantly higher in L-WNT3A cases. Conclusions These data demonstrate L-WNT3A can induce peri-implant bone formation even in cases where fibrous encapsulation predominates. PMID:26718012

  20. Noncanonical Wnt11 signaling and cardiomyogenic differentiation.

    PubMed

    Flaherty, Michael P; Dawn, Buddhadeb

    2008-10-01

    Although the molecular details remain unclear, Wnt signaling via both canonical and noncanonical pathways is integral to cardiac specification and morphogenesis. A growing body of evidence also suggests substantial overlap between these supposedly discrete pathways in cell- and context-dependent manners. The ability of Wnt11 to induce cardiomyogenesis both during embryonic development and in adult cells makes it an attractive candidate for preprogramming cells for cardiac repair. This review primarily discusses various aspects of noncanonical Wnt signaling in cardiogenesis with particular emphasis on Wnt11.

  1. Both ERK1 and ERK2 are required for enterovirus 71 (EV71) efficient replication.

    PubMed

    Zhu, Meng; Duan, Hao; Gao, Meng; Zhang, Hao; Peng, Yihong

    2015-03-20

    It has been demonstrated that MEK1, one of the two MEK isoforms in Raf-MEK-ERK1/2 pathway, is essential for successful EV71 propagation. However, the distinct function of ERK1 and ERK2 isoforms, the downstream kinases of MEKs, remains unclear in EV71 replication. In this study, specific ERK siRNAs and selective inhibitor U0126 were applied. Silencing specific ERK did not significantly impact on the EV71-caused biphasic activation of the other ERK isoform, suggesting the EV71-induced activations of ERK1 and ERK2 were non-discriminative and independent to one another. Knockdown of either ERK1 or ERK2 markedly impaired progeny EV71 propagation (both by more than 90%), progeny viral RNA amplification (either by about 30% to 40%) and protein synthesis (both by around 70%), indicating both ERK1 and ERK2 were critical and not interchangeable to EV71 propagation. Moreover, suppression of EV71 replication by inhibiting both early and late phases of ERK1/2 activation showed no significant difference from that of only blocking the late phase, supporting the late phase activation was more importantly responsible for EV71 life cycle. Taken together, this study for the first time identified both ERK1 and ERK2 were required for EV71 efficient replication and further verified the important role of MEK1-ERK1/2 in EV71 replication.

  2. The antidepressant roles of Wnt2 and Wnt3 in stress-induced depression-like behaviors

    PubMed Central

    Zhou, W-J; Xu, N; Kong, L; Sun, S-C; Xu, X-F; Jia, M-Z; Wang, Y; Chen, Z-Y

    2016-01-01

    Wnts-related signaling pathways have been reported to play roles in the pathogenesis of stress-induced depression-like behaviors. However, there is relatively few direct evidence to indicate the effect of Wnt ligands on this process. Here, we investigated the role of Wnts in mediating chronic restraint stress (CRS)-induced depression-like behaviors. We found that CRS induced a significant decrease in the expression of Wnt2 and Wnt3 in the ventral hippocampus (VH) but not in the dorsal hippocampus. Knocking down Wnt2 or Wnt3 in the VH led to impaired Wnt/β-catenin signaling, neurogenesis deficits and depression-like behaviors. In contrast, overexpression of Wnt2 or Wnt3 reversed CRS-induced depression-like behaviors. Moreover, Wnt2 and Wnt3 activated cAMP response element-binding protein (CREB) and there was CREB-dependent positive feedback between Wnt2 and Wnt3. Finally, fluoxetine treatment increased Wnt2 and Wnt3 levels in the VH and knocking down Wnt2 or Wnt3 abolished the antidepressant effect of fluoxetine. Taken together, our study indicates essential roles for Wnt2 and Wnt3 in CRS-induced depression-like behaviors and antidepressant. PMID:27622936

  3. Probing Wnt Receptor Turnover: A Critical Regulatory Point of Wnt Pathway.

    PubMed

    Jiang, Xiaomo; Cong, Feng

    2016-01-01

    Wnt pathways are critical for embryonic development and adult tissue homeostasis in all multicellular animals. Many regulatory mechanisms exist to control proper signaling output. Recent studies suggest that cell surface Wnt receptor level is controlled by ubiquitination, and serve as a critical regulatory point of Wnt pathway activity as it determines the responsiveness of cells to Wnt signal. Here, we describe flow cytometry, cell surface protein biotinylation, and immunofluorescence pulse-chase methods to probe the surface expression, ubiquitination, and internalization of the Wnt receptors FZD and LRP6.

  4. Regulation of atypical MAP kinases ERK3 and ERK4 by the phosphatase DUSP2

    PubMed Central

    Perander, Maria; Al-Mahdi, Rania; Jensen, Thomas C.; Nunn, Jennifer A. L.; Kildalsen, Hanne; Johansen, Bjarne; Gabrielsen, Mads; Keyse, Stephen M.; Seternes, Ole-Morten

    2017-01-01

    The atypical MAP kinases ERK3 and ERK4 are activated by phosphorylation of a serine residue lying within the activation loop signature sequence S-E-G. However, the regulation of ERK3 and ERK4 phosphorylation and activity is poorly understood. Here we report that the inducible nuclear dual-specificity MAP kinase phosphatase (MKP) DUSP2, a known regulator of the ERK and p38 MAPKs, is unique amongst the MKP family in being able to bind to both ERK3 and ERK4. This interaction is mediated by a conserved common docking (CD) domain within the carboxyl-terminal domains of ERK3 and ERK4 and the conserved kinase interaction motif (KIM) located within the non-catalytic amino terminus of DUSP2. This interaction is direct and results in the dephosphorylation of ERK3 and ERK4 and the stabilization of DUSP2. In the case of ERK4 its ability to stabilize DUSP2 requires its kinase activity. Finally, we demonstrate that expression of DUSP2 inhibits ERK3 and ERK4-mediated activation of its downstream substrate MK5. We conclude that the activity of DUSP2 is not restricted to the classical MAPK pathways and that DUSP2 can also regulate the atypical ERK3/4-MK5 signalling pathway in mammalian cells. PMID:28252035

  5. A WNT of things to come: evolution of Wnt signaling and polarity in cnidarians.

    PubMed

    Lee, Patricia N; Pang, Kevin; Matus, David Q; Martindale, Mark Q

    2006-04-01

    The conserved family of Wnt signaling molecules mediates various developmental processes including governing cell fate, proliferation, and polarity. The diverse developmental functions of the Wnt genes in bilaterians have obscured the evolutionary origin of this important signaling pathway. Recent work in the Cnidaria has shown the diversity of Wnt genes, and regulatory components of Wnt signaling, evolved early in metazoan evolution, prior to the divergence of cnidarians and bilaterians. Evidence from Hydra and the sea anemone, Nematostella, demonstrates a role for Wnt signaling in axis formation and patterning, as well as gastrulation and germ-layer specification. In this review, we examine what is currently known about Wnt signaling in cnidarians, and discuss what this group of "simple" animals may reveal about the evolution of Wnt signaling and polarity.

  6. Hypermethylation of Wnt antagonist gene promoters and activation of Wnt pathway in myelodysplastic marrow cells.

    PubMed

    Masala, Erico; Valencia, Ana; Buchi, Francesca; Nosi, Daniele; Spinelli, Elena; Gozzini, Antonella; Sassolini, Francesca; Sanna, Alessandro; Zecchi, Sandra; Bosi, Alberto; Santini, Valeria

    2012-10-01

    We observed aberrant gene methylation of Wnt antagonists: sFRP1, sFRP2, sFRP4, sFRP5 and DKK1 in marrow cells of 55 MDS cases. Methylation of Wnt antagonist genes was associated with activation of the Wnt signaling pathway, consistent with the up-regulation of the Wnt downstream genes TCF1 and LEF1. Azacitidine exposure induced demethylation of Wnt-antagonist gene promoters and reduction of the non-phosphorylated β-catenin (NPBC) which is prevalent during Wnt pathway inactivation. Presence of ≥5% of bone marrow blasts was associated with methylation of sFRP1 and DKK1 and with methylation of more than two of the five Wnt antagonist genes.

  7. In the Wnt-er of life: Wnt signalling in melanoma and ageing

    PubMed Central

    Kaur, Amanpreet; Webster, Marie R; Weeraratna, Ashani T

    2016-01-01

    Although the clinical landscape of melanoma is improving rapidly, metastatic melanoma remains a deadly disease. Age remains one of the greatest risk factors for melanoma, and patients older than 55 have a much poorer prognosis than younger individuals, even when the data are controlled for grade and stage. The reasons for this disparity have not been fully uncovered, but there is some recent evidence that Wnt signalling may have a role. Wnt signalling is known to have roles both in cancer progression as well as in organismal ageing. In melanoma, the interplay of Wnt signalling pathways is complex, with different members of the Wnt family guiding different aspects of invasion and proliferation. Here, we will briefly review the current literature addressing the roles of different Wnt pathways in melanoma pathogenesis, provide an overview of Wnt signalling during ageing, and discuss the intersection between melanoma and ageing in terms of Wnt signalling. PMID:27764844

  8. The Wnt code: cnidarians signal the way.

    PubMed

    Guder, C; Philipp, I; Lengfeld, T; Watanabe, H; Hobmayer, B; Holstein, T W

    2006-12-04

    Cnidarians are the simplest metazoans with a nervous system. They are well known for their regeneration capacity, which is based on the restoration of a signalling centre (organizer). Recent work has identified the canonical Wnt pathway in the freshwater polyp Hydra, where it acts in organizer formation and regeneration. Wnt signalling is also essential for cnidarian embryogenesis. In the sea anemone Nematostella vectensis 11 of the 12 known wnt gene subfamilies were identified. Different wnt genes exhibit serial and overlapping expression domains along the oral-aboral axis of the embryo (the 'wnt code'). This is reminiscent of the hox code (cluster) in bilaterian embryogenesis that is, however, absent in cnidarians. It is proposed that the common ancestor of cnidarians and bilaterians invented a set of wnt genes that patterned the ancient main body axis. Major antagonists of Wnt ligands (e.g. Dkk 1/2/4) that were previously known only from chordates, are also present in cnidarians and exhibit a similar conserved function. The unexpectedly high level of genetic complexity of wnt genes evolved in early multi-cellular animals about 650 Myr ago and suggests a radical expansion of the genetic repertoire, concurrent with the evolution of multi-cellularity and the diversification of eumetazoan body plans.

  9. WNT Signaling in Bone Development and Homeostasis

    PubMed Central

    Zhong, Zhendong; Ethen, Nicole J.; Williams, Bart O.

    2014-01-01

    The balance between bone formation and bone resorption controls postnatal bone homeostasis. Research over the last decade has provided a vast amount of evidence that WNT signaling plays a pivotal role in regulating this balance. Therefore, understanding how the WNT signaling pathway regulates skeletal development and homeostasis is of great value for human skeletal health and disease. PMID:25270716

  10. The Wnt signaling pathway in cancer.

    PubMed

    Duchartre, Yann; Kim, Yong-Mi; Kahn, Michael

    2016-03-01

    The Wnt signaling pathway is critically involved in both the development and homeostasis of tissues via regulation of their endogenous stem cells. Aberrant Wnt signaling has been described as a key player in the initiation of and/or maintenance and development of many cancers, via affecting the behavior of Cancer Stem Cells (CSCs). CSCs are considered by most to be responsible for establishment of the tumor and also for disease relapse, as they possess inherent drug-resistance properties. The development of new therapeutic compounds targeting the Wnt signaling pathway promises new hope to eliminate CSCs and achieve cancer eradication. However, a major challenge resides in developing a strategy efficient enough to target the dysregulated Wnt pathway in CSCs, while being safe enough to not damage the normal somatic stem cell population required for tissue homeostasis and repair. Here we review recent therapeutic approaches to target the Wnt pathway and their clinical applications.

  11. Harmine Induces Adipocyte Thermogenesis through RAC1-MEK-ERK-CHD4 Axis

    PubMed Central

    Nie, Tao; Hui, Xiaoyan; Mao, Liufeng; Nie, Baoming; Li, Kuai; Sun, Wei; Gao, Xuefei; Tang, Xiaofeng; Xu, Yong; Jiang, Baishan; Tu, Zhengcao; Li, Peng; Ding, Ke; Han, Weiping; Zhang, Shaoping; Xu, Aimin; Ding, Sheng; Liu, Pentao; Patterson, Adam; Cooper, Garth; Wu, Donghai

    2016-01-01

    Harmine is a natural compound possessing insulin-sensitizing effect in db/db diabetic mice. However its effect on adipose tissue browning is unknown. Here we reveal that harmine antagonizes high fat diet-induced adiposity. Harmine-treated mice gained less weight on a high fat diet and displayed increased energy expenditure and adipose tissue thermogenesis. In vitro, harmine potently induced the expression of thermogenic genes in both brown and white adipocytes, which was largely abolished by inhibition of RAC1/MEK/ERK pathway. Post-transcriptional modification analysis revealed that chromodomain helicase DNA binding protein 4 (CHD4) is a potential downstream target of harmine-mediated ERK activation. CHD4 directly binds the proximal promoter region of Ucp1, which is displaced upon treatment of harmine, thereby serving as a negative modulator of Ucp1. Thus, here we reveal a new application of harmine in combating obesity via this off-target effect in adipocytes. PMID:27805061

  12. Wnt modulating agents inhibit human cytomegalovirus replication.

    PubMed

    Kapoor, Arun; He, Ran; Venkatadri, Rajkumar; Forman, Michael; Arav-Boger, Ravit

    2013-06-01

    Infection with human cytomegalovirus (HCMV) continues to be a threat for pregnant women and immunocompromised hosts. Although limited anti-HCMV therapies are available, development of new agents is desired. The Wnt signaling pathway plays a critical role in embryonic and cancer stem cell development and is targeted by gammaherpesviruses, Epstein-Barr virus (EBV), and Kaposi's sarcoma-associated herpesvirus (KSHV). HCMV infects stem cells, including neural progenitor cells, during embryogenesis. To investigate the role of Wnt in HCMV replication in vitro, we tested monensin, nigericin, and salinomycin, compounds that inhibit cancer stem cell growth by modulating the Wnt pathway. These compounds inhibited the replication of HCMV Towne and a clinical isolate. Inhibition occurred prior to DNA replication but persisted throughout the full replication cycle. There was a significant decrease in expression of IE2, UL44, and pp65 proteins. HCMV infection resulted in a significant and sustained decrease in expression of phosphorylated and total lipoprotein receptor-related protein 6 (pLRP6 and LRP6, respectively), Wnt 5a/b, and β-catenin and a modest decrease in Dvl2/3, while levels of the negative regulator axin 1 were increased. Nigericin decreased the expression of pLRP6, LRP6, axin 1, and Wnt 5a/b in noninfected and HCMV-infected cells. For all three compounds, a correlation was found between expression levels of Wnt 5a/b and axin 1 and HCMV inhibition. The decrease in Wnt 5a/b and axin 1 expression was more significant in HCMV-infected cells than noninfected cells. These data illustrate the complex effects of HCMV on the Wnt pathway and the fine balance between Wnt and HCMV, resulting in abrogation of HCMV replication. Additional studies are required to elucidate how HCMV targets Wnt for its benefit.

  13. Wnt Modulating Agents Inhibit Human Cytomegalovirus Replication

    PubMed Central

    Kapoor, Arun; He, Ran; Venkatadri, Rajkumar; Forman, Michael

    2013-01-01

    Infection with human cytomegalovirus (HCMV) continues to be a threat for pregnant women and immunocompromised hosts. Although limited anti-HCMV therapies are available, development of new agents is desired. The Wnt signaling pathway plays a critical role in embryonic and cancer stem cell development and is targeted by gammaherpesviruses, Epstein-Barr virus (EBV), and Kaposi's sarcoma-associated herpesvirus (KSHV). HCMV infects stem cells, including neural progenitor cells, during embryogenesis. To investigate the role of Wnt in HCMV replication in vitro, we tested monensin, nigericin, and salinomycin, compounds that inhibit cancer stem cell growth by modulating the Wnt pathway. These compounds inhibited the replication of HCMV Towne and a clinical isolate. Inhibition occurred prior to DNA replication but persisted throughout the full replication cycle. There was a significant decrease in expression of IE2, UL44, and pp65 proteins. HCMV infection resulted in a significant and sustained decrease in expression of phosphorylated and total lipoprotein receptor-related protein 6 (pLRP6 and LRP6, respectively), Wnt 5a/b, and β-catenin and a modest decrease in Dvl2/3, while levels of the negative regulator axin 1 were increased. Nigericin decreased the expression of pLRP6, LRP6, axin 1, and Wnt 5a/b in noninfected and HCMV-infected cells. For all three compounds, a correlation was found between expression levels of Wnt 5a/b and axin 1 and HCMV inhibition. The decrease in Wnt 5a/b and axin 1 expression was more significant in HCMV-infected cells than noninfected cells. These data illustrate the complex effects of HCMV on the Wnt pathway and the fine balance between Wnt and HCMV, resulting in abrogation of HCMV replication. Additional studies are required to elucidate how HCMV targets Wnt for its benefit. PMID:23571549

  14. Wnt genes in the mouse uterus: potential regulation of implantation.

    PubMed

    Hayashi, Kanako; Erikson, David W; Tilford, Sarah A; Bany, Brent M; Maclean, James A; Rucker, Edmund B; Johnson, Greg A; Spencer, Thomas E

    2009-05-01

    Wnt genes are involved in critical developmental and growth processes. The present study comprehensively analyzed temporal and spatial alterations in Wnt and Fzd gene expression in the mouse uterus during peri-implantation of pregnancy. Expression of Wnt4, Wnt5a, Wnt7a, Wnt7b, Wnt11, Wnt16, Fzd2, Fzd4, and Fzd6 was detected in the uterus during implantation. Wnt4 mRNA was most abundant in the decidua, whereas Wnt5a mRNA was restricted to the mesometrial decidua during decidualization. Wnt7a, Wnt7b, and Wnt11 mRNAs were abundantly detected in the endometrial epithelia. The expression of Wnt7b was robust in the luminal epithelium (LE) at the implantation site on Gestational Day 5, whereas Wnt11 mRNA disappeared in the LE adjacent to the embryo in the antimesometrial implantation chamber but remained abundant in the LE. Wnt16 mRNA was localized to the stroma surrounding the LE on Day 4 and remained in the stroma adjacent to the LE but not in areas undergoing the decidual reaction. Fzd2 mRNA was detected in the decidua, Fzd4 mRNA was in the vessels and stroma surrounding the embryo, and Fzd6 mRNA was observed in the endometrial epithelia, stroma, and some blood vessels during implantation. Ovarian steroid hormone treatment was found to regulate Wnt genes and Fzd receptors in ovariectomized mice. Especially, single injections of progesterone stimulated Wnt11 mRNA, and estrogen stimulated Wnt4 and Wnt7b. The temporal and spatial alterations in Wnt genes likely play a critical role during implantation and decidualization in mice.

  15. Nonmyocyte ERK1/2 signaling contributes to load-induced cardiomyopathy in Marfan mice

    PubMed Central

    MacFarlane, Elena Gallo; Takimoto, Eiki; Chaudhary, Rahul; Nagpal, Varun; Rainer, Peter P.; Bindman, Julia G.; Gerber, Elizabeth E.; Bedja, Djahida; Schiefer, Christopher; Miller, Karen L.; Zhu, Guangshuo; Myers, Loretha; Amat-Alarcon, Nuria; Lee, Dong I.; Koitabashi, Norimichi; Judge, Daniel P.; Dietz, Harry C.

    2017-01-01

    Among children with the most severe presentation of Marfan syndrome (MFS), an inherited disorder of connective tissue caused by a deficiency of extracellular fibrillin-1, heart failure is the leading cause of death. Here, we show that, while MFS mice (Fbn1C1039G/+ mice) typically have normal cardiac function, pressure overload (PO) induces an acute and severe dilated cardiomyopathy in association with fibrosis and myocyte enlargement. Failing MFS hearts show high expression of TGF-β ligands, with increased TGF-β signaling in both nonmyocytes and myocytes; pathologic ERK activation is restricted to the nonmyocyte compartment. Informatively, TGF-β, angiotensin II type 1 receptor (AT1R), or ERK antagonism (with neutralizing antibody, losartan, or MEK inhibitor, respectively) prevents load-induced cardiac decompensation in MFS mice, despite persistent PO. In situ analyses revealed an unanticipated axis of activation in nonmyocytes, with AT1R-dependent ERK activation driving TGF-β ligand expression that culminates in both autocrine and paracrine overdrive of TGF-β signaling. The full compensation seen in wild-type mice exposed to mild PO correlates with enhanced deposition of extracellular fibrillin-1. Taken together, these data suggest that fibrillin-1 contributes to cardiac reserve in the face of hemodynamic stress, critically implicate nonmyocytes in disease pathogenesis, and validate ERK as a therapeutic target in MFS-related cardiac decompensation. PMID:28768908

  16. Wnt signaling blockage inhibits cell proliferation and migration, and induces apoptosis in triple-negative breast cancer cells

    PubMed Central

    2013-01-01

    Background Triple-negative breast cancer (TNBC) is an aggressive clinical subtype of breast cancer that is characterized by the lack of estrogen receptor (ER) and progesterone receptor (PR) expression as well as human epidermal growth factor receptor 2 (HER2) overexpression. The TNBC subtype constitutes approximately 10%–20% of all breast cancers, but has no effective molecular targeted therapies. Previous meta-analysis of gene expression profiles of 587 TNBC cases from 21 studies demonstrated high expression of Wnt signaling pathway-associated genes in basal-like 2 and mesenchymal subtypes of TNBC. In this study, we investigated the potential of Wnt pathway inhibitors in effective treatment of TNBC. Methods Activation of Wnt pathway was assessed in four TNBC cell lines (BT-549, MDA-MB-231, HCC-1143 and HCC-1937), and the ER+ cell line MCF-7 using confocal microscopy and Western blot analysis of pathway components. Effectiveness of five different Wnt pathway inhibitors (iCRT-3, iCRT-5, iCRT-14, IWP-4 and XAV-939) on cell proliferation and apoptosis were tested in vitro. The inhibitory effects of iCRT-3 on canonical Wnt signaling in TNBC was evaluated by quantitative real-time RT-PCR analysis of Axin2 and dual-luciferase reporter assays. The effects of shRNA knockdown of SOX4 in combination with iCRT-3 and/or genistein treatments on cell proliferation, migration and invasion on BT-549 cells were also evaluated. Results Immunofluorescence staining of β-catenin in TNBC cell lines showed both nuclear and cytoplasmic localization, indicating activation of Wnt pathway in TNBC cells. iCRT-3 was the most effective compound for inhibiting proliferation and antagonizing Wnt signaling in TNBC cells. In addition, treatment with iCRT-3 resulted in increased apoptosis in vitro. Knockdown of the Wnt pathway transcription factor, SOX4 in triple negative BT-549 cells resulted in decreased cell proliferation and migration, and combination treatment of iCRT-3 with SOX4 knockdown

  17. Gangliosides, or sialic acid, antagonize ethanol intoxication

    SciTech Connect

    Klemm, W.R.; Boyles, R.; Matthew, J.; Cherian, L.

    1988-01-01

    Because ethanol elicits a dose-dependent hydrolysis of brain sialogangliosides, the authors tested the possibility that injected gangliosides might antagonize intoxicating doses of ethanol. Clear anti-intoxication effects were seen at 24 hr post-injection of mixed mouse-brain gangliosides at 125-130 mg/kg, but not at lower or higher doses. Sleep time was reduced on the order of 50%, and roto-rod agility was significantly enhanced. Sialic acid (SA) similarly antagonized ethanol; however, the precursor of SA, N-acetyl-D-mannosamine, as well as ceramide and asialoganglioside did not.

  18. C. elegans EOR-1/PLZF and EOR-2 positively regulate Ras and Wnt signaling and function redundantly with LIN-25 and the SUR-2 Mediator component.

    PubMed

    Howard, Robyn M; Sundaram, Meera V

    2002-07-15

    In Caenorhabditis elegans, Ras/ERK and Wnt/beta-catenin signaling pathways cooperate to induce P12 and vulval cell fates in a Hox-dependent manner. Here we describe eor-1 and eor-2, two new positively acting nuclear components of the Ras and Wnt pathways. eor-1 and eor-2 act downstream or in parallel to ERK and function redundantly with the Mediator complex gene sur-2 and the functionally related gene lin-25, such that removal of both eor-1/eor-2 and sur-2/lin-25 mimics the removal of a main Ras pathway component. Furthermore, the eor-1 and eor-2 mutant backgrounds reveal an essential role for the Elk1-related gene lin-1. eor-1 and eor-2 also act downstream or in parallel to pry-1 Axin and therefore act at the convergence of the Ras and Wnt pathways. eor-1 encodes the ortholog of human PLZF, a BTB/zinc-finger transcription factor that is fused to RARalpha in acute promyelocytic leukemia. eor-2 encodes a novel protein. EOR-1/PLZF and EOR-2 appear to function closely together and cooperate with Hox genes to promote the expression of Ras- and Wnt-responsive genes. Further studies of eor-1 and eor-2 may provide insight into the roles of PLZF in normal development and leukemogenesis.

  19. WNT signaling in neuronal maturation and synaptogenesis

    PubMed Central

    Rosso, Silvana B.; Inestrosa, Nibaldo C.

    2013-01-01

    The Wnt signaling pathway plays a role in the development of the central nervous system and growing evidence indicates that Wnts also regulates the structure and function of the adult nervous system. Wnt components are key regulators of a variety of developmental processes, including embryonic patterning, cell specification, and cell polarity. In the nervous system, Wnt signaling also regulates the formation and function of neuronal circuits by controlling neuronal differentiation, axon outgrowth and guidance, dendrite development, synaptic function, and neuronal plasticity. Wnt factors can signal through three very well characterized cascades: canonical or β-catenin pathway, planar cell polarity pathway and calcium pathway that control different processes. However, divergent downstream cascades have been identified to control neuronal morphogenesis. In the nervous system, the expression of Wnt proteins is a highly controlled process. In addition, deregulation of Wnt signaling has been associated with neurodegenerative diseases. Here, we will review different aspects of neuronal and dendrite maturation, including spinogenesis and synaptogenesis. Finally, the role of Wnt pathway components on Alzheimer’s disease will be revised. PMID:23847469

  20. Wnt signaling inhibits CTL memory programming.

    PubMed

    Xiao, Zhengguo; Sun, Zhifeng; Smyth, Kendra; Li, Lei

    2013-12-01

    Induction of functional CTLs is one of the major goals for vaccine development and cancer therapy. Inflammatory cytokines are critical for memory CTL generation. Wnt signaling is important for CTL priming and memory formation, but its role in cytokine-driven memory CTL programming is unclear. We found that wnt signaling inhibited IL-12-driven CTL activation and memory programming. This impaired memory CTL programming was attributed to up-regulation of eomes and down-regulation of T-bet. Wnt signaling suppressed the mTOR pathway during CTL activation, which was different to its effects on other cell types. Interestingly, the impaired memory CTL programming by wnt was partially rescued by mTOR inhibitor rapamycin. In conclusion, we found that crosstalk between wnt and the IL-12 signaling inhibits T-bet and mTOR pathways and impairs memory programming which can be recovered in part by rapamycin. In addition, direct inhibition of wnt signaling during CTL activation does not affect CTL memory programming. Therefore, wnt signaling may serve as a new tool for CTL manipulation in autoimmune diseases and immune therapy for certain cancers. Copyright © 2013 Elsevier Ltd. All rights reserved.

  1. Heparin activates Wnt signaling for neuronal morphogenesis.

    PubMed

    Colombres, Marcela; Henríquez, Juan Pablo; Reig, Germán F; Scheu, Jessica; Calderón, Rosario; Alvarez, Alejandra; Brandan, Enrique; Inestrosa, Nibaldo C

    2008-09-01

    Wnt factors are secreted ligands that affect different aspects of the nervous system behavior like neurodevelopment, synaptogenesis and neurodegeneration. In different model systems, Wnt signaling has been demonstrated to be regulated by heparan sulfate proteoglycans (HSPGs). Whether HSPGs modulate Wnt signaling in the context of neuronal behavior is currently unknown. Here we demonstrate that activation of Wnt signaling with the endogenous ligand Wnt-7a results in an increased of neurite outgrowth in the neuroblastoma N2a cell line. Interestingly, heparin induces glycogen synthase kinase-3beta (GSK-3beta) inhibition, beta-catenin stabilization and morphological differentiation in both N2a cells and in rat primary hippocampal neuronal cultures. We also show that heparin modulates Wnt-3a-induced stabilization of beta-catenin. Several extracellular matrix and membrane-attached HSPGs were found to be expressed in both in vitro neuronal models. Changes in the expression of specific HSPGs were observed upon differentiation of N2a cells. Taken together, our findings suggest that HSPGs may modulate canonical Wnt signaling for neuronal morphogenesis.

  2. Mutual Antagonism of Wilms’ Tumor 1 and β-Catenin Dictates Podocyte Health and Disease

    PubMed Central

    Zhou, Lili; Li, Yingjian; He, Weichun; Zhou, Dong; Tan, Roderick J.; Nie, Jing; Hou, Fan Fan

    2015-01-01

    Activation of β-catenin, the intracellular mediator of canonical Wnt signaling, has a critical role in mediating podocyte injury and proteinuria. However, the underlying mechanisms remain poorly understood. Here, we show that β-catenin triggers ubiquitin-mediated protein degradation of Wilms’ tumor 1 (WT1) and functionally antagonizes its action. In mice injected with adriamycin, WT1 protein was progressively lost in glomerular podocytes at 1, 3, and 5 weeks after injection. Notably, loss of WT1 apparently did not result from podocyte depletion but was closely associated with upregulation of β-catenin. This change in WT1/β-catenin ratio was accompanied by loss of podocyte-specific nephrin, podocalyxin, and synaptopodin and acquisition of mesenchymal markers Snail1, α-smooth muscle actin, and fibroblast-specific protein 1. In vitro, overexpression of β-catenin induced WT1 protein degradation through the ubiquitin proteasomal pathway, which was blocked by MG-132. WT1 and β-catenin also competed for binding to common transcriptional coactivator CREB-binding protein and mutually repressed the expression of their respective target genes. In glomerular miniorgan culture, activation of β-catenin by Wnt3a repressed WT1 and its target gene expression. In vivo, blockade of Wnt/β-catenin signaling by endogenous antagonist Klotho induced WT1 and restored podocyte integrity in adriamycin nephropathy. These results show that β-catenin specifically targets WT1 for ubiquitin-mediated degradation, leading to podocyte dedifferentiation and mesenchymal transition. Our data also suggest that WT1 and β-catenin have opposing roles in podocyte biology, and that the ratio of their expression levels dictates the state of podocyte health and disease in vivo. PMID:25071087

  3. Expression of MIG-6, WNT-9A, and WNT-7B during osteoarthritis.

    PubMed

    Velasquillo, Cristina; Garciadiego-Cázares, David; Almonte, Maylin; Bustamante, Marcia; Ibarra, Clemente; Kouri, Juan B; Chimal-Monroy, Jesús

    2007-11-01

    Although the molecular mechanisms for initiation of cartilage destruction in osteoarthritis (OA) are unknown, it has been demonstrated that disruption of mitogen-inducible gene 6 (Mig-6) in mice leads to the onset of a degenerative joint disease like OA. On this basis, we correlated gene expression of Mig-6 with Wnt-9a and Wnt-7b genes; we showed downregulation of Mig-6, Wnt-7b, and Wnt-9a during OA, while Wnt-7b was expressed also in osteoblast-like cells. Here we suggest that Aggrecan degradation occurs before the downregulation of Mig-6. It remains to be proven whether there is any relation between Wnt signaling and Aggrecan degradation.

  4. Pleiotrophin antagonizes Brd2 during neuronal differentiation

    PubMed Central

    Garcia-Gutierrez, Pablo; Juarez-Vicente, Francisco; Wolgemuth, Debra J.; Garcia-Dominguez, Mario

    2014-01-01

    ABSTRACT Bromodomain-containing protein 2 (Brd2) is a BET family chromatin adaptor required for expression of cell-cycle-associated genes and therefore involved in cell cycle progression. Brd2 is expressed in proliferating neuronal progenitors, displays cell-cycle-stimulating activity and, when overexpressed, impairs neuronal differentiation. Paradoxically, Brd2 is also detected in differentiating neurons. To shed light on the role of Brd2 in the transition from cell proliferation to differentiation, we had previously looked for proteins that interacted with Brd2 upon induction of neuronal differentiation. Surprisingly, we identified the growth factor pleiotrophin (Ptn). Here, we show that Ptn antagonized the cell-cycle-stimulating activity associated with Brd2, thus enhancing induced neuronal differentiation. Moreover, Ptn knockdown reduced neuronal differentiation. We analyzed Ptn-mediated antagonism of Brd2 in a cell differentiation model and in two embryonic processes associated with the neural tube: spinal cord neurogenesis and neural crest migration. Finally, we investigated the mechanisms of Ptn-mediated antagonism and determined that Ptn destabilizes the association of Brd2 with chromatin. Thus, Ptn-mediated Brd2 antagonism emerges as a modulation system accounting for the balance between cell proliferation and differentiation in the vertebrate nervous system. PMID:24695857

  5. Pleiotrophin antagonizes Brd2 during neuronal differentiation.

    PubMed

    Garcia-Gutierrez, Pablo; Juarez-Vicente, Francisco; Wolgemuth, Debra J; Garcia-Dominguez, Mario

    2014-06-01

    Bromodomain-containing protein 2 (Brd2) is a BET family chromatin adaptor required for expression of cell-cycle-associated genes and therefore involved in cell cycle progression. Brd2 is expressed in proliferating neuronal progenitors, displays cell-cycle-stimulating activity and, when overexpressed, impairs neuronal differentiation. Paradoxically, Brd2 is also detected in differentiating neurons. To shed light on the role of Brd2 in the transition from cell proliferation to differentiation, we had previously looked for proteins that interacted with Brd2 upon induction of neuronal differentiation. Surprisingly, we identified the growth factor pleiotrophin (Ptn). Here, we show that Ptn antagonized the cell-cycle-stimulating activity associated with Brd2, thus enhancing induced neuronal differentiation. Moreover, Ptn knockdown reduced neuronal differentiation. We analyzed Ptn-mediated antagonism of Brd2 in a cell differentiation model and in two embryonic processes associated with the neural tube: spinal cord neurogenesis and neural crest migration. Finally, we investigated the mechanisms of Ptn-mediated antagonism and determined that Ptn destabilizes the association of Brd2 with chromatin. Thus, Ptn-mediated Brd2 antagonism emerges as a modulation system accounting for the balance between cell proliferation and differentiation in the vertebrate nervous system. © 2014. Published by The Company of Biologists Ltd.

  6. Wnt3-frizzled 1 chimera as a model to study canonical Wnt signaling.

    PubMed

    Bhat, Ramesh A; Stauffer, Barbara; Della Pietra, Anthony; Bodine, Peter V N

    2010-04-01

    Wnt proteins initiate signaling by binding to seven transmembrane spanning receptors of the frizzled (Fz) family together with the members of the low-density lipoprotein receptor-related protein (LRP) 5 and 6. A chimera of human Wnt3 and Fz1 receptor was developed that efficiently activated the TCF-luciferase reporter. Deletion of the cytoplasmic tail and point mutations in the PDZ binding region in the chimera resulted in the loss of Wnt signaling, suggesting a critical role for the Fz cytoplasmic region in Wnt signaling. The Fz CRD is also critical for Wnt signaling, as a deletion of 29 amino acids in the 2nd cysteine loop resulted in the total loss of TCF-luciferase activation. DKK-1 protein blocks upregulation of the TCF-luciferase reporter by the Wnt3-Fz1 chimera, suggesting involvement of LRP in Wnt3-Fz1 signaling. Expression of a Wnt3-Fz1 chimera in C3H10T1/2 cells resulted in the upregulation of alkaline phosphatase activity and inhibition of adipocyte formation, demonstrating that the Wnt3-Fz1 chimera is a potent activator of differentiation of C3H10T1/2 cells into osteoblasts and an inhibitor of their differentiation into the adipocyte lineage. In summary, the Wnt-Fz chimera approach has the potential to better our understanding of the mechanism of Wnt action and its role, particularly in stem cell differentiation. In addition, this methodology can be utilized to identify inhibitors of either Wnt, Fz or interactors of the canonical pathway, which may have potential therapeutic value in the treatment of cancers and other diseases.

  7. The N-Terminal Domain of ERK1 Accounts for the Functional Differences with ERK2

    PubMed Central

    Marchi, Matilde; D'Antoni, Angela; Formentini, Ivan; Parra, Riccardo; Brambilla, Riccardo

    2008-01-01

    The Extracellular Regulated Kinase 1 and 2 transduce a variety of extracellular stimuli regulating processes as diverse as proliferation, differentiation and synaptic plasticity. Once activated in the cytoplasm, ERK1 and ERK2 translocate into the nucleus and interact with nuclear substrates to induce specific programs of gene expression. ERK1/2 share 85% of aminoacid identity and all known functional domains and thence they have been considered functionally equivalent until recent studies found that the ablation of either ERK1 or ERK2 causes dramatically different phenotypes. To search a molecular justification of this dichotomy we investigated whether the different functions of ERK1 and 2 might depend on the properties of their cytoplasmic-nuclear trafficking. Since in the nucleus ERK1/2 is predominantly inactivated, the maintenance of a constant level of nuclear activity requires continuous shuttling of activated protein from the cytoplasm. For this reason, different nuclear-cytoplasmic trafficking of ERK1 and 2 would cause a differential signalling capability. We have characterised the trafficking of fluorescently tagged ERK1 and ERK2 by means of time-lapse imaging in living cells. Surprisingly, we found that ERK1 shuttles between the nucleus and cytoplasm at a much slower rate than ERK2. This difference is caused by a domain of ERK1 located at its N-terminus since the progressive deletion of these residues converted the shuttling features of ERK1 into those of ERK2. Conversely, the fusion of this ERK1 sequence at the N-terminus of ERK2 slowed down its shuttling to a similar value found for ERK1. Finally, computational, biochemical and cellular studies indicated that the reduced nuclear shuttling of ERK1 causes a strong reduction of its nuclear phosphorylation compared to ERK2, leading to a reduced capability of ERK1 to carry proliferative signals to the nucleus. This mechanism significantly contributes to the differential ability of ERK1 and 2 to generate an

  8. Activation of the Wnt Pathway by Mycobacterium tuberculosis: A Wnt-Wnt Situation.

    PubMed

    Villaseñor, Tomás; Madrid-Paulino, Edgardo; Maldonado-Bravo, Rafael; Urbán-Aragón, Antonio; Pérez-Martínez, Leonor; Pedraza-Alva, Gustavo

    2017-01-01

    Mycobacterium tuberculosis (M. tuberculosis), an intracellular pathogenic Gram-positive bacterium, is the cause of tuberculosis (TB), a major worldwide human infectious disease. The innate immune system is the first host defense against M. tuberculosis. The recognition of this pathogen is mediated by several classes of pattern recognition receptors expressed on the host innate immune cells, including Toll-like receptors, Nod-like receptors, and C-type lectin receptors like Dectin-1, the Mannose receptor, and DC-SIGN. M. tuberculosis interaction with any of these receptors activates multiple signaling pathways among which the protein kinase C, the MAPK, and the NFκB pathways have been widely studied. These pathways have been implicated in macrophage invasion, M. tuberculosis survival, and impaired immune response, thus promoting a successful infection and disease. Interestingly, the Wnt signaling pathway, classically regarded as a pathway involved in the control of cell proliferation, migration, and differentiation in embryonic development, has recently been involved in immunoregulatory mechanisms in infectious and inflammatory diseases, such as TB, sepsis, psoriasis, rheumatoid arthritis, and atherosclerosis. In this review, we present the current knowledge supporting a role for the Wnt signaling pathway during macrophage infection by M. tuberculosis and the regulation of the immune response against M. tuberculosis. Understanding the cross talk between different signaling pathways activated by M. tuberculosis will impact on the search for new therapeutic targets to fuel the rational design of drugs aimed to restore the immunological response against M. tuberculosis.

  9. Polymorphisms in WNT6 and WNT10A and Colorectal Adenoma Risk

    PubMed Central

    Galbraith, Rachel L.; Poole, Elizabeth M.; Duggan, David; Muehling, Jill; Hsu, Li; Makar, Karen; Xiao, Liren; Potter, John D.; Ulrich, Cornelia M.

    2015-01-01

    The WNT/β-catenin signaling pathway upregulates transcription of genes involved in cell proliferation and cancer progression; it has been implicated in colorectal adenoma formation. To date, no studies have examined polymorphisms in WNT genes or WNT gene–environment interactions in relation to adenoma risk. Within a colonoscopy-based case-control study of 628 adenoma cases and 516 polyp-free controls, we analyzed two tagSNPs in WNT6 (rs6747776 G > C, rs6754599 G > C) and WNT10A (rs7349332 G > A, rs10177996 A > G). The WNT6 rs6747776 homozygous minor allele (CC) was associated with increased risk of colorectal adenoma (OR = 2.75, 95% CI: 1.03–7.31). We observed a statistically significant interaction between WNT6 rs6747776 and the proportion of calories from total fat (P-int = 0.02), where the highest risk was observed among those with minor alleles and lowest fat intake. We also detected a marginally significant (0.05 < P ≤ 0.10) interaction with fish intake (P-int = 0.09). Additionally, a marginally significant interaction was observed between proportion of calories from saturated fat and the WNT10A rs7349332 polymorphism. Our results suggest that genetic variability in the WNT pathway may play a role in colorectal adenoma formation or may partly mediate the increased risk of colorectal cancer associated with fat intake. PMID:21547848

  10. Polymorphisms in WNT6 and WNT10A and colorectal adenoma risk.

    PubMed

    Galbraith, Rachel L; Poole, Elizabeth M; Duggan, David; Muehling, Jill; Hsu, Li; Makar, Karen; Xiao, Liren; Potter, John D; Ulrich, Cornelia M

    2011-01-01

    The WNT/β-catenin signaling pathway upregulates transcription of genes involved in cell proliferation and cancer progression; it has been implicated in colorectal adenoma formation. To date, no studies have examined polymorphisms in WNT genes or WNT gene-environment interactions in relation to adenoma risk. Within a colonoscopy-based case-control study of 628 adenoma cases and 516 polyp-free controls, we analyzed two tagSNPs in WNT6 (rs6747776 G > C, rs6754599 G > C) and WNT10A (rs7349332 G > A, rs10177996 A > G). The WNT6 rs6747776 homozygous minor allele (CC) was associated with increased risk of colorectal adenoma (OR = 2.75, 95% CI: 1.03-7.31). We observed a statistically significant interaction between WNT6 rs6747776 and the proportion of calories from total fat (P-int = 0.02), where the highest risk was observed among those with minor alleles and lowest fat intake. We also detected a marginally significant (0.05 < P ≤ 0.10) interaction with fish intake (P-int = 0.09). Additionally, a marginally significant interaction was observed between proportion of calories from saturated fat and the WNT10A rs7349332 polymorphism. Our results suggest that genetic variability in the WNT pathway may play a role in colorectal adenoma formation or may partly mediate the increased risk of colorectal cancer associated with fat intake.

  11. Altered Expression of PRKX, WNT3 and WNT16 in Human Nodular Basal Cell Carcinoma.

    PubMed

    DO Carmo, Natalia Gurgel; Sakamoto, Luis Henrique Toshihiro; Pogue, Robert; DO Couto Mascarenhas, Cintia; Passos, Simone Karst; Felipe, Maria Sueli Soares; DE Andrade, Rosângela Vieira

    2016-09-01

    Nodular and superficial are the most common subtypes of basal cell carcinoma (BCC). Signaling pathways such as Hedgehog (HH) and Wingless (WNT) signaling are associated with BCC phenotypic variation. The aim of the study was to evaluate of the expression profiles of 84 genes related to the WNT and HH signaling pathways in patients with nodular and superficial BCC. A total of 58 BCCs and 13 samples of normal skin were evaluated by quantitative real-time polymerase chain reaction (qPCR) to detect the gene-expression profile. qPCR array showed segregation in BCC subtypes compared to healthy skin. PRKX, WNT3 and WNT16 were significantly (p<0.05) altered: PRKX was up-regulated, and WNT3 and WNT16 were down-regulated in nodular BCC. PRKX, WNT3 and WNT16 genes, belonging to the WNT signaling pathway, are involved in the tumorigenic process of nodular BCC. Copyright© 2016 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

  12. Quetiapine and aripiprazole signal differently to ERK, p90RSK and c-Fos in mouse frontal cortex and striatum: role of the EGF receptor

    PubMed Central

    2014-01-01

    Background Signaling pathways outside dopamine D2 receptor antagonism may govern the variable clinical profile of antipsychotic drugs (APD) in schizophrenia. One postulated mechanism causal to APD action may regulate synaptic plasticity and neuronal connectivity via the extracellular signal-regulated kinase (ERK) cascade that links G-protein coupled receptors (GPCR) and ErbB growth factor signaling, systems disturbed in schizophrenia. This was based upon our finding that the low D2 receptor affinity APD clozapine induced initial down-regulation and delayed epidermal growth factor receptor (EGFR or ErbB1) mediated activation of the cortical and striatal ERK response in vivo distinct from olanzapine or haloperidol. Here we map whether the second generation atypical APDs aripiprazole and quetiapine affect the EGFR-ERK pathway and its substrates p90RSK and c-Fos in mouse brain, given their divergent agonist and antagonist properties on dopaminergic transmission, respectively. Results In prefrontal cortex, aripiprazole triggered triphasic ERK phosphorylation that was EGFR-independent but had no significant effect in striatum. Conversely quetiapine did not alter cortical ERK signaling but elevated striatal ERK levels in an EGFR-dependent manner. Induction of ERK by aripiprazole did not affect p90RSK signaling but quetiapine decreased RSK phosphorylation within 1-hour of administration. The transcription factor c-Fos by comparison was a direct target of ERK phosphorylation induced by aripiprazole in cortex and quetiapine in striatum with protein levels in temporal alignment with that of ERK. Conclusions These data indicate that aripiprazole and quetiapine signal to specific nuclear targets of ERK, which for quetiapine occurs via an EGFR-linked mechanism, possibly indicating involvement of this system in its action. PMID:24552586

  13. Aberrantly expressed miR-582-3p maintains lung cancer stem cell-like traits by activating Wnt/β-catenin signalling

    PubMed Central

    Fang, Lishan; Cai, Junchao; Chen, Baixue; Wu, Shanshan; Li, Rong; Xu, Xiaonan; Yang, Yi; Guan, Hongyu; Zhu, Xun; Zhang, Le; Yuan, Jie; Wu, Jueheng; Li, Mengfeng

    2015-01-01

    Cancer stem cells (CSCs) are involved in tumorigenesis, tumour recurrence and therapy resistance and Wnt signalling is essential for the development of the biological traits of CSCs. In non-small cell lung carcinoma (NSCLC), unlike in colon cancer, mutations in β-catenin and APC genes are uncommon; thus, the mechanism underlying the constitutive activation of Wnt signalling in NSCLC remains unclear. Here we report that miR-582-3p expression correlates with the overall- and recurrence-free-survival of NSCLC patients, and miR-582-3p has an activating effect on Wnt/β-catenin signalling. miR-582-3p overexpression simultaneously targets multiple negative regulators of the Wnt/β-catenin pathway, namely, AXIN2, DKK3 and SFRP1. Consequently, miR-582-3p promotes CSC traits of NSCLC cells in vitro and tumorigenesis and tumour recurrence in vivo. Antagonizing miR-582-3p potently inhibits tumour initiation and progression in xenografted animal models. These findings suggest that miR-582-3p mediates the constitutive activation of Wnt/β-catenin signalling, likely serving as a potential therapeutic target for NSCLC. PMID:26468775

  14. Augmenting Endogenous Wnt Signaling Improves Skin Wound Healing

    PubMed Central

    Liu, Bo; Manzano, Wilfred R.; Evans, Nick D.; Dhamdhere, Girija R.; Fang, Mark Y.; Chang, Howard Y.

    2013-01-01

    Wnt signaling is required for both the development and homeostasis of the skin, yet its contribution to skin wound repair remains controversial. By employing Axin2LacZ/+ reporter mice we evaluated the spatial and temporal distribution patterns of Wnt responsive cells, and found that the pattern of Wnt responsiveness varies with the hair cycle, and correlates with wound healing potential. Using Axin2LacZ/LacZ mice and an ear wound model, we demonstrate that amplified Wnt signaling leads to improved healing. Utilizing a biochemical approach that mimics the amplified Wnt response of Axin2LacZ/LacZ mice, we show that topical application of liposomal Wnt3a to a non-healing wound enhances endogenous Wnt signaling, and results in better skin wound healing. Given the importance of Wnt signaling in the maintenance and repair of skin, liposomal Wnt3a may have widespread application in clinical practice. PMID:24204695

  15. Antagonism by bioactive polyphenols against inflammation: a systematic view.

    PubMed

    Chu, Arthur J

    2014-02-01

    Through pattern recognition receptors, infections and tissue injuries drive innate immune cells to trigger inflammation with elevated cytokines, chemokines, growth factors, and other mediators. Inflammation resolves upon removal of pathogenic signals and the presence of pro-resolving conditions including combating adaptive immunity. Failure of resolution progresses into chronic inflammation, manifesting as detrimental disease development known as inflammatory diseases including cardiovascular diseases, diabetes, obesity, cancers, etc. Inflammation typically involves activations of many intracellular signaling pathways such as PI3K/AkT/mTORC1, PI3K/AkT/IKK(JNK), Ras/Raf/MEK/ERK, JAK/STAT, etc.; these pathways could in turn mediate the upregulations of proinflammatory transcription factors (e.g., NFκB, activator protein 1 (AP-1), HIF, signal transducer and activator of transcription (STAT), etc.). Furthermore, the resulting FOXO inactivation ensures inflammatory proceeding. This review provides a systematic view that polyphenols target multiple inflammatory components and reinforce anti-inflammatory mechanisms by antioxidant potentials, AMPK activation, PI3K/AkT inhibition, IKK/JNK inhibition, mTORC1 inhibition, JAK/STAT inhibition, TLR suppression, and ACE inhibition. As a result, polyphenols readily lead to NFκB, AP-1, HIF, and STAT inactivations with reduced proinflammatory mediator generation. In conclusion, polyphenols sustain resolution of inflammation and antagonize against proinflammation, which is readily consistent with diverse anti-inflammatory actions. The promoted, restored, and maintained tissue homeostasis beyond its anti-inflammatory effects also extends to diverse health benefits for disease preventions and interventions.

  16. Wnt signaling: why is everything so negative?

    PubMed

    Brown, J D; Moon, R T

    1998-04-01

    The Wnt proteins constitute a family of secreted glycoproteins the members of which have essential signaling roles during embryogenesis. The recent identification of several new regulators of this signal transduction pathway have revealed unexpectedly intricate levels of constraint on Wnt-dependent gene activation, and studies in developing embryos and in cell culture systems have allowed a more complete understanding of the functional and biochemical interactions between components of this evolutionarily conserved pathway.

  17. Wnt Modulators in the Biotech Pipeline

    PubMed Central

    Rey, Jean-Philippe; Ellies, Debra L.

    2011-01-01

    The purpose of this review is to provide a better understanding for the LRP co-receptor mediated Wnt pathway signaling. Using proteomics we have also subdivided the LRP receptor family into six subfamilies, encompassing the twelve family members. This review includes a discussion of proteins containing a cystine-knot protein motif (i.e. Sclerostin, Dan, Sostdc1, Vwf, Norrin, Pdgf, Mucin) and discusses how this motif plays a role in mediating Wnt signaling through interactions with LRP. PMID:20014100

  18. ERK1 and ERK2 activation modulates diet-induced obesity in mice.

    PubMed

    Khan, Amira Sayed; Subramaniam, Selvakumar; Dramane, Gado; Khelifi, Douadi; Khan, Naim Akhtar

    2017-06-01

    Obesity is a worldwide problem, and dietary lipids play an important role in its pathogenesis. Recently, Erk1 knock-out (ERK1(-/-)) mice have been shown to exhibit low preference for dietary fatty acids. Hence, we maintained Erk1(-/-) mice on a high-fat diet (HFD) to assess the implication of this mitogen-activated protein (MAP) kinase in obesity. The Erk1(-/-) mice, fed the HFD, were more obese than wild-type (WT) animals, fed the same diet. Erk1(-/-) obese mice gained more fat and liver mass than WT obese animals. No difference was observed in daily food and energy intake in HFD-fed both group of animals. However, feed efficiency was higher in Erk1(-/-) than WT animals. Blood cholesterol, triglyceride and insulin concentrations were higher in Erk1(-/-) obese mice compared to WT obese animals. Accordingly, homeostatic model assessment of insulin resistance (HOMA-IR) value was higher in Erk1(-/-) obese mice compared to WT obese animals. Interestingly, only Erk1(-/-) obese mice, but not WT-obese animals, exhibited high degree of phosphorylation of liver MEK, the upstream regulator of ERK1/2. This phenomenon was associated with high liver ERK2 phosphorylation in Erk1(-/-) obese mice which also had high liver acetyl-CoA carboxylase 1 (ACC1) and fatty acid synthase (FAS) mRNA expression, suggesting high lipogenesis in these animals. The Erk1(-/-) obese mice also had low PPAR-α and CPT1β mRNA, indicating low fatty acid oxidation. Our observations suggest that ERK1 and ERK2 might play key roles in the regulation of obesity. Copyright © 2017 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.

  19. ERK1 and ERK2 Map Kinases: Specific Roles or Functional Redundancy?

    PubMed Central

    Buscà, Roser; Pouysségur, Jacques; Lenormand, Philippe

    2016-01-01

    The MAP kinase signaling cascade Ras/Raf/MEK/ERK has been involved in a large variety of cellular and physiological processes that are crucial for life. Many pathological situations have been associated to this pathway. More than one isoform has been described at each level of the cascade. In this review we devoted our attention to ERK1 and ERK2, which are the effector kinases of the pathway. Whether ERK1 and ERK2 specify functional differences or are in contrast functionally redundant, constitutes an ongoing debate despite the huge amount of studies performed to date. In this review we compiled data on ERK1 vs. ERK2 gene structures, protein sequences, expression levels, structural and molecular mechanisms of activation and substrate recognition. We have also attempted to perform a rigorous analysis of studies regarding the individual roles of ERK1 and ERK2 by the means of morpholinos, siRNA, and shRNA silencing as well as gene disruption or gene replacement in mice. Finally, we comment on a recent study of gene and protein evolution of ERK isoforms as a distinct approach to address the same question. Our review permits the evaluation of the relevance of published studies in the field especially when measurements of global ERK activation are taken into account. Our analysis favors the hypothesis of ERK1 and ERK2 exhibiting functional redundancy and points to the concept of the global ERK quantity, and not isoform specificity, as being the essential determinant to achieve ERK function. PMID:27376062

  20. Differential transformation of mammary epithelial cells by Wnt genes.

    PubMed Central

    Wong, G T; Gavin, B J; McMahon, A P

    1994-01-01

    The mouse Wnt family includes at least 10 genes that encode structurally related secreted glycoproteins. Wnt-1 and Wnt-3 were originally identified as oncogenes activated by the insertion of mouse mammary tumor virus in virus-induced mammary adenocarcinomas, although they are not expressed in the normal mammary gland. However, five other Wnt genes are differentially expressed during development of adult mammary tissue, suggesting that they may play distinct roles in various phases of mammary gland growth and development. Induction of transformation by Wnt-1 and Wnt-3 may be due to interference with these normal regulatory events; however, there is no direct evidence for this hypothesis. We have tested Wnt family members for the ability to induce transformation of cultured mammary cells. The results demonstrate that the Wnt gene family can be divided into three groups depending on their ability to induce morphological transformation and altered growth characteristics of the C57MG mammary epithelial cell line. Wnt-1, Wnt-3A, and Wnt-7A were highly transforming and induced colonies which formed and shed balls of cells. Wnt-2, Wnt-5B, and Wnt-7B also induced transformation but with a lower frequency and an apparent decrease in saturation density. In contrast, Wnt-6 and two other family members which are normally expressed in C57MG cells, Wnt-4 and Wnt-5A, failed to induce transformation. These data demonstrate that the Wnt genes have distinct effects on cell growth and should not be regarded as functionally equivalent. Images PMID:8065359

  1. TIAM1 Antagonizes TAZ/YAP Both in the Destruction Complex in the Cytoplasm and in the Nucleus to Inhibit Invasion of Intestinal Epithelial Cells.

    PubMed

    Diamantopoulou, Zoi; White, Gavin; Fadlullah, Muhammad Z H; Dreger, Marcel; Pickering, Karen; Maltas, Joe; Ashton, Garry; MacLeod, Ruth; Baillie, George S; Kouskoff, Valerie; Lacaud, Georges; Murray, Graeme I; Sansom, Owen J; Hurlstone, Adam F L; Malliri, Angeliki

    2017-05-08

    Aberrant WNT signaling drives colorectal cancer (CRC). Here, we identify TIAM1 as a critical antagonist of CRC progression through inhibiting TAZ and YAP, effectors of WNT signaling. We demonstrate that TIAM1 shuttles between the cytoplasm and nucleus antagonizing TAZ/YAP by distinct mechanisms in the two compartments. In the cytoplasm, TIAM1 localizes to the destruction complex and promotes TAZ degradation by enhancing its interaction with βTrCP. Nuclear TIAM1 suppresses TAZ/YAP interaction with TEADs, inhibiting expression of TAZ/YAP target genes implicated in epithelial-mesenchymal transition, cell migration, and invasion, and consequently suppresses CRC cell migration and invasion. Importantly, high nuclear TIAM1 in clinical specimens associates with increased CRC patient survival. Together, our findings suggest that in CRC TIAM1 suppresses tumor progression by regulating YAP/TAZ activity. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

  2. The mTOR and canonical Wnt signaling pathways mediate the mnemonic effects of progesterone in the dorsal hippocampus.

    PubMed

    Fortress, Ashley M; Heisler, John D; Frick, Karyn M

    2015-05-01

    Although much is known about the neural mechanisms responsible for the mnemonic effects of 17β-estradiol (E2 ), very little is understood about the mechanisms through which progesterone (P4 ) regulates memory. We previously showed that intrahippocampal infusion of P4 in ovariectomized female mice enhances object recognition (OR) memory consolidation in a manner dependent on activation of dorsal hippocampal ERK and mTOR signaling. However, the role of specific progesterone receptors (PRs) in mediating the effects of progesterone on memory consolidation and hippocampal cell signaling are unknown. Therefore, the goals of this study were to investigate the roles of membrane-associated and intracellular PRs in mediating hippocampal memory consolidation, and identify downstream cell signaling pathways activated by PRs. Membrane-associated PRs were targeted using bovine serum albumin-conjugated progesterone (BSA-P), and intracellular PRs (PR-A, PR-B) were targeted using the intracellular PR agonist R5020. Immediately after OR training, ovariectomized mice received bilateral dorsal hippocampal infusion of vehicle, P4 , BSA-P, or R5020. OR memory consolidation was enhanced by P4 , BSA-P, and R5020. However, only P4 and BSA-P activated ERK and mTOR signaling. Furthermore, dorsal hippocampal infusion of the ERK inhibitor U0126 blocked the memory-enhancing effects of BSA-P, but not R5020. The intracellular PR antagonist RU486 blocked the memory-enhancing effects of R5020, but not BSA-P. Interestingly, P4 robustly activated canonical Wnt signaling in the dorsal hippocampus, which is consistent with our recent findings that canonical Wnt signaling is necessary for OR memory consolidation. R5020, but not BSA-P, also elicited a modest increase in canonical Wnt signaling. Collectively, these data suggest that activation of ERK signaling is necessary for membrane-associated PRs to enhance OR, and indicate a role for canonical Wnt signaling in the memory-enhancing effects of

  3. Wnt8a and Wnt3a Cooperate in the Axial Stem Cell Niche to Promote Mammalian Body Axis Extension

    PubMed Central

    Cunningham, Thomas J.; Kumar, Sandeep; Yamaguchi, Terry P.; Duester, Gregg

    2015-01-01

    Background Vertebrate body axis extension occurs in a head-to-tail direction from a caudal progenitor zone that responds to interacting signals. Wnt/β-catenin signaling is critical for generation of paraxial mesoderm, somite formation, and maintenance of the axial stem cell pool. Body axis extension requires Wnt8a in lower vertebrates, but in mammals Wnt3a is required, although the anterior trunk develops in the absence of Wnt3a. Results We examined mouse Wnt8a−/− and Wnt3a−/− single and double mutants to explore whether mammalian Wnt8a contributes to body axis extension and to determine whether a posterior growth function for Wnt8a is conserved throughout the vertebrate lineage. We find that caudal Wnt8a is expressed only during early somite stages and is required for normal development of the anterior trunk in the absence of Wnt3a. During this time, we show that Wnt8a and Wnt3a cooperate to maintain Fgf8 expression and prevent premature Sox2 upregulation in the axial stem cell niche, critical for posterior growth. Similar to Fgf8, Wnt8a requires retinoic acid (RA) signaling to establish its expression boundaries and possesses an upstream RA response element that binds RA receptors. Conclusions These findings provide new insight into interaction of caudal Wnt-FGF-RA signals required for body axis extension. PMID:25809880

  4. Identification and mechanism of ABA receptor antagonism

    SciTech Connect

    Melcher, Karsten; Xu, Yong; Ng, Ley-Moy; Zhou, X. Edward; Soon, Fen-Fen; Chinnusamy, Viswanathan; Suino-Powell, Kelly M; Kovach, Amanda; Tham, Fook S.; Cutler, Sean R.; Li, Jun; Yong, Eu-Leong; Zhu, Jian-Kang; Xu, H. Eric

    2010-11-11

    The phytohormone abscisic acid (ABA) functions through a family of fourteen PYR/PYL receptors, which were identified by resistance to pyrabactin, a synthetic inhibitor of seed germination. ABA activates these receptors to inhibit type 2C protein phosphatases, such as ABI1, yet it remains unclear whether these receptors can be antagonized. Here we demonstrate that pyrabactin is an agonist of PYR1 and PYL1 but is unexpectedly an antagonist of PYL2. Crystal structures of the PYL2-pyrabactin and PYL1-pyrabactin-ABI1 complexes reveal the mechanism responsible for receptor-selective activation and inhibition, which enables us to design mutations that convert PYL1 to a pyrabactin-inhibited receptor and PYL2 to a pyrabactin-activated receptor and to identify new pyrabactin-based ABA receptor agonists. Together, our results establish a new concept of ABA receptor antagonism, illustrate its underlying mechanisms and provide a rational framework for discovering novel ABA receptor ligands.

  5. Wnt some lose some: transcriptional governance of stem cells by Wnt/β-catenin signaling

    PubMed Central

    Lien, Wen-Hui; Fuchs, Elaine

    2014-01-01

    In mammals, Wnt/β-catenin signaling features prominently in stem cells and cancers, but how and for what purposes have been matters of much debate. In this review, we summarize our current knowledge of Wnt/β-catenin signaling and its downstream transcriptional regulators in normal and malignant stem cells. We centered this review largely on three types of stem cells—embryonic stem cells, hair follicle stem cells, and intestinal epithelial stem cells—in which the roles of Wnt/β-catenin have been extensively studied. Using these models, we unravel how many controversial issues surrounding Wnt signaling have been resolved by dissecting the diversity of its downstream circuitry and effectors, often leading to opposite outcomes of Wnt/β-catenin-mediated regulation and differences rooted in stage- and context-dependent effects. PMID:25030692

  6. Wnt some lose some: transcriptional governance of stem cells by Wnt/β-catenin signaling.

    PubMed

    Lien, Wen-Hui; Fuchs, Elaine

    2014-07-15

    In mammals, Wnt/β-catenin signaling features prominently in stem cells and cancers, but how and for what purposes have been matters of much debate. In this review, we summarize our current knowledge of Wnt/β-catenin signaling and its downstream transcriptional regulators in normal and malignant stem cells. We centered this review largely on three types of stem cells--embryonic stem cells, hair follicle stem cells, and intestinal epithelial stem cells--in which the roles of Wnt/β-catenin have been extensively studied. Using these models, we unravel how many controversial issues surrounding Wnt signaling have been resolved by dissecting the diversity of its downstream circuitry and effectors, often leading to opposite outcomes of Wnt/β-catenin-mediated regulation and differences rooted in stage- and context-dependent effects. © 2014 Lien and Fuchs; Published by Cold Spring Harbor Laboratory Press.

  7. Triclosan Antagonizes Fluconazole Activity against Candida albicans

    PubMed Central

    Higgins, J.; Pinjon, E.; Oltean, H.N.; White, T.C.; Kelly, S.L.; Martel, C.M.; Sullivan, D.J.; Coleman, D.C.; Moran, G.P.

    2012-01-01

    Triclosan is a broad-spectrum antimicrobial compound commonly used in oral hygiene products. Investigation of its activity against Candida albicans showed that triclosan was fungicidal at concentrations of 16 mg/L. However, at subinhibitory concentrations (0.5-2 mg/L), triclosan antagonized the activity of fluconazole. Although triclosan induced CDR1 expression in C. albicans, antagonism was still observed in cdr1Δ and cdr2Δ strains. Triclosan did not affect fluconazole uptake or alter total membrane sterol content, but did induce the expression of FAS1 and FAS2, indicating that its mode of action may involve inhibition of fatty acid synthesis, as it does in prokaryotes. However, FAS2 mutants did not exhibit increased susceptibility to triclosan, and overexpression of both FAS1 and FAS2 alleles did not alter triclosan susceptibility. Unexpectedly, the antagonistic effect was specific for C. albicans under hypha-inducing conditions and was absent in the non-filamentous efg1Δ strain. This antagonism may be due to the membranotropic activity of triclosan and the unique composition of hyphal membranes. PMID:21972257

  8. wnt3a but not wnt11 supports self-renewal of embryonic stem cells

    SciTech Connect

    Singla, Dinender K. . E-mail: Dinender.Kumar@uvm.edu; Schneider, David J.; LeWinter, Martin M.; Sobel, Burton E.

    2006-06-30

    wnt proteins (wnts) promote both differentiation of midbrain dopaminergic cells and self-renewal of haematopoietic stem cells. Mouse embryonic stem (ES) cells can be maintained and self-renew on mouse feeder cell layers or in media containing leukemia inhibitory factor (LIF). However, the effects of wnts on ES cells self-renewal and differentiation are not clearly understood. In the present study, we found that conditioned medium prepared from L cells expressing wnt3a can replace feeder cell layers and medium containing LIF in maintaining ES cells in the proliferation without differentiation (self-renewal) state. By contrast, conditioned medium from NIH3T3 cells expressing wnt11 did not. Alkaline phosphatase staining and compact colony formation were used as criteria of cells being in the undifferentiated state. ES cells maintained in medium conditioned by Wnt3a expressing cells underwent freezing and thawing while maintaining properties seen with LIF maintained ES cells. Purified wnt3a did not maintain self-renewal of ES cells for prolonged intervals. Thus, other factors in the medium conditioned by wnt3a expressing cells may have contributed to maintenance of ES cells in a self-renewal state. Pluripotency of ES cells was determined with the use of embryoid bodies in vitro. PD98059, a MEK specific inhibitor, promoted the growth of undifferentiated ES cells maintained in conditioned medium from wnt3a expressing cells. By contrast, the P38 MAPK inhibitor SB230580 did not, suggesting a role for the MEK pathway in self-renewal and differentiation of ES cells maintained in the wnt3a cell conditioned medium. Thus, our results show that conditioned medium from wnt3a but not wnt11 expressing cells can maintain ES cells in self-renewal and in a pluripotent state.

  9. WNT10B mutations in human obesity

    PubMed Central

    Christodoulides, C.; Scarda, A.; Granzotto, M.; Milan, G.; Dalla Nora, E.; Keogh, J.; De Pergola, G.; Stirling, H.; Pannacciulli, N.; Sethi, J. K.; Federspil, G.; Vidal-Puig, A.; Farooqi, I. S.; O’Rahilly, S.; Vettor, R.

    2015-01-01

    Aims/hypothesis Recent studies suggest that wingless-type MMTV integration site family, member 10B (WNT10B) may play a role in the negative regulation of adipocyte differentiation in vitro and in vivo. In order to determine whether mutations in WNT10B contribute to human obesity, we screened two independent populations of obese subjects for mutations in this gene. Subjects and methods We studied 96 subjects with severe obesity of early onset (less than 10 years of age) from the UK Genetics of Obesity Study and 115 obese Italian subjects of European origin. Results One proband with early-onset obesity was found to be heterozygous for a C256Y mutation, which abrogated the ability of WNT10B to activate canonical WNT signalling and block adipogenesis and was not found in 600 control alleles. All relatives of the proband who carried this allele were either overweight or obese. Three other rare missense variants were found in obese probands, but these did not clearly cosegregate with obesity in family studies and one (P301S), which was found in three unrelated subjects with early-onset obesity, had normal functional properties. Conclusions/interpretation These mutations represent the first naturally occurring missense variants of WNT10B. While the pedigree analysis in the case of C256Y WNT10B does not provide definitive proof of a causal link of this variant with obesity, the finding of a non-functioning WNT10B allele in a human family affected by obesity should encourage further study of this gene in other obese populations. PMID:16477437

  10. Modulating effects of acyl-CoA synthetase 5-derived mitochondrial Wnt2B palmitoylation on intestinal Wnt activity

    PubMed Central

    Klaus, Christina; Schneider, Ursula; Hedberg, Christian; Schütz, Anke K; Bernhagen, Jürgen; Waldmann, Herbert; Gassler, Nikolaus; Kaemmerer, Elke

    2014-01-01

    AIM: To investigate the role of acyl-CoA synthetase 5 (ACSL5) activity in Wnt signaling in intestinal surface epithelia. METHODS: Several cell lines were used to investigate the ACSL5-dependent expression and synthesis of Wnt2B, a mitochondrially expressed protein of the Wnt signaling family. Wnt activity was functionally assessed with a luciferase reporter assay. ACSL5-related biochemical Wnt2B modifications were investigated with a modified acyl-exchange assay. The findings from the cell culture models were verified using an Apcmin/+ mouse model as well as normal and neoplastic diseased human intestinal tissues. RESULTS: In the presence of ACSL5, Wnt2B was unable to translocate into the nucleus and was enriched in mitochondria, which was paralleled by a significant decrease in Wnt activity. ACSL5-dependent S-palmitoylation of Wnt2B was identified as a molecular reason for mitochondrial Wnt2B accumulation. In cell culture systems, a strong relation of ACSL5 expression, Wnt2B palmitoylation, and degree of malignancy were found. Using normal mucosa, the association of ACSL5 and Wnt2B was seen, but in intestinal neoplasias the mechanism was only rudimentarily observed. CONCLUSION: ACSL5 mediates antiproliferative activities via Wnt2B palmitoylation with diminished Wnt activity. The molecular pathway is probably relevant for intestinal homeostasis, overwhelmed by other pathways in carcinogenesis. PMID:25356045

  11. Regulation of NMDA-receptor synaptic transmission by Wnt signaling

    PubMed Central

    Cerpa, Waldo; Gambrill, Abigail; Inestrosa, Nibaldo C.; Barria, Andres

    2011-01-01

    Wnt ligands are secreted glycoproteins controlling gene expression and cytoskeleton reorganization involved in embryonic development of the nervous system. However, their role in later stages of brain development, particularly in the regulation of established synaptic connections is not known. We found that Wnt-5a acutely and specifically up-regulates synaptic NMDAR currents in rat hippocampal slices facilitating induction of LTP, a cellular model of learning and memory. This effect requires an increase in postsynaptic Ca2+ and activation of non-canonical downstream effectors of the Wnt signaling pathway. In contrast, Wnt-7a, an activator of the canonical Wnt signaling pathway, has no effect on NMDAR mediated synaptic transmission. Moreover, endogenous Wnt ligands are necessary to maintain basal NMDAR synaptic transmission adjusting the threshold for synaptic potentiation. This novel role for Wnt ligands provides a mechanism for Wnt signaling to acutely modulate synaptic plasticity and brain function in later stages of development and in the mature organism. PMID:21715611

  12. Sialidase NEU3 contributes neoplastic potential on colon cancer cells as a key modulator of gangliosides by regulating Wnt signaling.

    PubMed

    Takahashi, Kohta; Hosono, Masahiro; Sato, Ikuro; Hata, Keiko; Wada, Tadashi; Yamaguchi, Kazunori; Nitta, Kazuo; Shima, Hiroshi; Miyagi, Taeko

    2015-10-01

    The plasma membrane-associated sialidase NEU3 is a key enzyme for ganglioside degradation. We previously demonstrated remarkable up-regulation of NEU3 in various human cancers, with augmented malignant properties. Here, we provide evidence of a close link between NEU3 expression and Wnt/β-catenin signaling in colon cancer cells by analyzing tumorigenic potential and cancer stem-like characteristics. NEU3 silencing in HT-29 and HCT116 colon cancer cells resulted in significant decrease in clonogenicity on soft agar and in vivo tumor growth, along with down-regulation of stemness and Wnt-related genes. Analyses further revealed that NEU3 enhanced phosphorylation of the Wnt receptor LRP6 and consequently β-catenin activation by accelerating complex formation with LRP6 and recruitment of GSK3β and Axin, whereas its silencing exerted the opposite effects. NEU3 activity-null mutants failed to demonstrate the activation, indicating the requirement of ganglioside modulation by the sialidase for the effects. Under sphere-forming conditions, when stemness genes are up-regulated, endogenous NEU3 expression was found to be significantly increased, whereas NEU3 silencing suppressed sphere-formation and in vivo tumor incidence in NOD-SCID mice. Increased ability of clonogenicity on soft agar and sphere formation by Wnt stimulation was abrogated by NEU3 silencing. Furthermore, NEU3 was found to regulate phosphorylation of ERK and Akt via EGF receptor and Ras cascades, thought to be additionally required for tumor progression. The results indicate an essential contribution of NEU3 to tumorigenic potential through maintenance of stem-like characteristics of colon cancer cells by regulating Wnt signaling at the receptor level, in addition to tumor progression via Ras/MAPK signaling.

  13. Exosomes go with the Wnt.

    PubMed

    Koles, Kate; Budnik, Vivian

    2012-07-01

    Exosomes, small secreted microvesicles, are implicated in intercellular communication in diverse cell types, transporting protein, lipid and nucleic acid cargo that impact the physiology of recipient cells. Besides the signaling function of exosomes they also serve as a mechanism to dispose obsolete cellular material.(1) Particularly exciting is the involvement of exosomal communication in the nervous system, as this has important implications for brain development and function. The properties of exosomes are also beginning to entice the biomedical community since they represent potentially novel avenues for the targeted delivery of customized exosome cargo, such as miRNAs, during disease. Our findings implicating exosomes in trans-synaptic communication emerged from the serendipitous observation that at the Drosophila larval neuromuscular junction (NMJ) the release of a signaling molecule, Wnt1/Wingless (Wg) and its binding partner Evenness Interrupted (Evi)/Wntless (Wls)/Sprint (Srt), were released by motorneurons in association with vesicles, which we postulated to be exosomes.(2) In our most recent paper(3) using in vivo analysis at the Drosophila NMJ as well as in cultured insect cells we formally demonstrate that Evi rides in exosomes that are released to the extracellular space and identify some of the players involved in their release. In addition, a proteomic analysis of exosomes highlights novel potential function of exosomes.

  14. Delivery of the Porcupine Inhibitor WNT974 in Mice

    PubMed Central

    Zhang, Li-shu; Lum, Lawrence

    2016-01-01

    We describe here a technique for delivering the porcupine inhibitor WNT974 (formerly LGK974) in mice. The protocol entails once-a-day oral delivery of WNT974 for up to 3 months at a concentration sufficient to achieve systemic Wnt pathway inhibition with limited toxicity as measured by weight change. This route of delivery enables extended durations of Wnt signaling inhibition in a mammalian model organism. PMID:27590157

  15. ERKed by LRRK2: A cell biological perspective on hereditary and sporadic Parkinson’s disease

    PubMed Central

    Verma, Manish; Steer, Erin K.; Chu, Charleen T.

    2014-01-01

    The leucine rich repeat kinase 2 (LRRK2/dardarin) is implicated in autosomal dominant familial and sporadic Parkinson’s disease (PD); mutations in LRRK2 account for up to 40% of PD cases in some populations. LRRK2 is a large protein with a kinase domain, a GTPase domain, and multiple potential protein interaction domains. As such, delineating the functional pathways for LRRK2 and mechanisms by which PD-linked variants contribute to age-related neurodegeneration could result in pharmaceutically tractable therapies. A growing number of recent studies implicate dysregulation of mitogen activated protein kinases 3 and 1 (also known as ERK1/2) as possible downstream mediators of mutant LRRK2 effects. As these master regulators of growth, differentiation, neuronal plasticity and cell survival have also been implicated in other PD models, a set of common cell biological pathways may contribute to neuronal susceptibility in PD. Here, we review the literature on several major cellular pathways impacted by LRRK2 mutations – autophagy, microtubule/cytoskeletal dynamics, and protein synthesis – in context of potential signaling crosstalk involving the ERK1/2 and Wnt signaling pathways. Emerging implications for calcium homeostasis, mitochondrial biology and synaptic dysregulation are discussed in relation to LRRK2 interactions with other PD gene products. It has been shown that substantia nigra neurons in human PD and Lewy body dementia patients exhibit cytoplasmic accumulations of ERK1/2 in mitochondria, autophagosomes and bundles of intracellular fibrils. Both experimental and human tissue data implicate pathogenic changes in ERK1/2 signaling in sporadic, toxin-based and mutant LRRK2 settings, suggesting engagement of common cell biological pathways by divergent PD etiologies. PMID:24225420

  16. A Second WNT for Old Drugs: Drug Repositioning against WNT-Dependent Cancers

    PubMed Central

    Ahmed, Kamal; Shaw, Holly V.; Koval, Alexey; Katanaev, Vladimir L.

    2016-01-01

    Aberrant WNT signaling underlies cancerous transformation and growth in many tissues, such as the colon, breast, liver, and others. Downregulation of the WNT pathway is a desired mode of development of targeted therapies against these cancers. Despite the urgent need, no WNT signaling-directed drugs currently exist, and only very few candidates have reached early phase clinical trials. Among different strategies to develop WNT-targeting anti-cancer therapies, repositioning of existing drugs previously approved for other diseases is a promising approach. Nonsteroidal anti-inflammatory drugs like aspirin, the anti-leprotic clofazimine, and the anti-trypanosomal suramin are among examples of drugs having recently revealed WNT-targeting activities. In total, 16 human-use drug compounds have been found to be working through the WNT pathway and show promise for their prospective repositioning against various cancers. Advances, hurdles, and prospects of developing these molecules as potential drugs against WNT-dependent cancers, as well as approaches for discovering new ones for repositioning, are the foci of the current review. PMID:27429001

  17. Wnt signaling: role in LTP, neural networks and memory.

    PubMed

    Oliva, Carolina A; Vargas, Jessica Y; Inestrosa, Nibaldo C

    2013-06-01

    Wnt components are key regulators of a variety of developmental processes, including embryonic patterning, cell specification, and cell polarity. The Wnt signaling pathway participates in the development of the central nervous system and growing evidence indicates that Wnts also regulates the function of the adult nervous system. In fact, most of the key components including Wnts and Frizzled receptors are expressed in the adult brain. Wnt ligands have been implicated in the regulation of synaptic assembly as well as in neurotransmission and synaptic plasticity. Deregulation of Wnt signaling has been associated with several pathologies, and more recently has been related to neurodegenerative diseases and to mental and mood disorders. In this review, we focus our attention on the Wnt signaling cascade in postnatal life and we review in detail the presence of Wnt signaling components in pre- and postsynaptic regions. Due to the important role of Wnt proteins in wiring neural circuits, we discuss recent findings about the role of Wnt pathways both in basal spontaneous activities as well as in activity-dependent processes that underlie synaptic plasticity. Finally, we review the role of Wnt in vivo and we finish with the most recent data in literature that involves the effect of components of the Wnt signaling pathway in neurological and mental disorders, including a special emphasis on in vivo studies that relate behavioral abnormalities to deficiencies in Wnt signaling, as well as the data that support a neuroprotective role of Wnt proteins in relation to the pathogenesis of Alzheimer's disease.

  18. [Wnt signalling pathway and cervical cancer].

    PubMed

    Ramos-Solano, Moisés; Álvarez-Zavala, Monserrat; García-Castro, Beatriz; Jave-Suárez, Luis Felipe; Aguilar-Lemarroy, Adriana

    2015-01-01

    Cervical cancer (CC) is a pathology that arises in the cervical epithelium, whose major cause of risk is human papillomavirus (HPV) infection. Due to the fact that HPV infection per se is not enough to generate a carcinogenic process, it has been proposed that alterations in the Wnt signaling pathway are involved in cervical carcinogenesis. The Wnt family consists of 13 receptors and 19 ligands, and it is highly conserved phylogenetically due to its contribution in different biological processes, such as embryogenesis and tissue regeneration. Additionally, this signaling pathway modulates various cellular functions, for instance: cell proliferation, differentiation, migration and cell polarity. This paper describes the Wnt signaling pathways and alterations that have been found in members of this family in different cancer types and, especially, in CC.

  19. Perfluorooctanoic acid induces human Ishikawa endometrial cancer cell migration and invasion through activation of ERK/mTOR signaling.

    PubMed

    Ma, Zhinan; Liu, Xiaoqiu; Li, Fujun; Wang, Yixong; Xu, Yang; Zhang, Mei; Zhang, Xiaoqian; Ying, Xiaoyan; Zhang, Xuesen

    2016-10-11

    Perfluorooctanoic acid (PFOA) is a common environmental pollutant that has been associated with various diseases, including cancer. We explored the molecular mechanisms underlying PFOA-induced endometrial cancer cell invasion and migration. PFOA treatment enhanced migration and invasion by human Ishikawa endometrial cancer cells, which correlated with decreased E-cadherin expression, a marker of epithelial-mesenchymal transition. PFOA also induced activation of ERK1/2/mTOR signaling. Treatment with rapamycin, an mTOR inhibitor, antagonized the effects of PFOA and reversed the effects of PFOA activation in a xenograft mouse model of endometrial cancer. Consistent with these results, pre-treatment with rapamycin abolished PFOA-induced down-regulation of E-cadherin expression. These results indicate that PFOA is a carcinogen that promotes endometrial cancer cell migration and invasion through activation of ERK/mTOR signaling.

  20. Perfluorooctanoic acid induces human Ishikawa endometrial cancer cell migration and invasion through activation of ERK/mTOR signaling

    PubMed Central

    Li, Fujun; Wang, Yixong; Xu, Yang; Zhang, Mei; Zhang, Xiaoqian; Ying, Xiaoyan; Zhang, Xuesen

    2016-01-01

    Perfluorooctanoic acid (PFOA) is a common environmental pollutant that has been associated with various diseases, including cancer. We explored the molecular mechanisms underlying PFOA-induced endometrial cancer cell invasion and migration. PFOA treatment enhanced migration and invasion by human Ishikawa endometrial cancer cells, which correlated with decreased E-cadherin expression, a marker of epithelial-mesenchymal transition. PFOA also induced activation of ERK1/2/mTOR signaling. Treatment with rapamycin, an mTOR inhibitor, antagonized the effects of PFOA and reversed the effects of PFOA activation in a xenograft mouse model of endometrial cancer. Consistent with these results, pre-treatment with rapamycin abolished PFOA-induced down-regulation of E-cadherin expression. These results indicate that PFOA is a carcinogen that promotes endometrial cancer cell migration and invasion through activation of ERK/mTOR signaling. PMID:27589685

  1. Molecular dissection of Wnt3a-Frizzled8 interaction reveals essential and modulatory determinants of Wnt signaling activity

    PubMed Central

    2014-01-01

    Background Wnt proteins are a family of secreted signaling molecules that regulate key developmental processes in metazoans. The molecular basis of Wnt binding to Frizzled and LRP5/6 co-receptors has long been unknown due to the lack of structural data on Wnt ligands. Only recently, the crystal structure of the Wnt8-Frizzled8-cysteine-rich-domain (CRD) complex was solved, but the significance of interaction sites that influence Wnt signaling has not been assessed. Results Here, we present an extensive structure-function analysis of mouse Wnt3a in vitro and in vivo. We provide evidence for the essential role of serine 209, glycine 210 (site 1) and tryptophan 333 (site 2) in Fz binding. Importantly, we discovered that valine 337 in the site 2 binding loop is critical for signaling without contributing to binding. Mutations in the presumptive second CRD binding site (site 3) partly abolished Wnt binding. Intriguingly, most site 3 mutations increased Wnt signaling, probably by inhibiting Wnt-CRD oligomerization. In accordance, increasing amounts of soluble Frizzled8-CRD protein modulated Wnt3a signaling in a biphasic manner. Conclusions We propose a concentration-dependent switch in Wnt-CRD complex formation from an inactive aggregation state to an activated high mobility state as a possible modulatory mechanism in Wnt signaling gradients. PMID:24885675

  2. A systems biology approach to model neural stem cell regulation by notch, shh, wnt, and EGF signaling pathways.

    PubMed

    Sivakumar, Krishnankutty Chandrika; Dhanesh, Sivadasan Bindu; Shobana, Sekar; James, Jackson; Mundayoor, Sathish

    2011-10-01

    The Notch, Sonic Hedgehog (Shh), Wnt, and EGF pathways have long been known to influence cell fate specification in the developing nervous system. Here we attempted to evaluate the contemporary knowledge about neural stem cell differentiation promoted by various drug-based regulations through a systems biology approach. Our model showed the phenomenon of DAPT-mediated antagonism of Enhancer of split [E(spl)] genes and enhancement of Shh target genes by a SAG agonist that were effectively demonstrated computationally and were consistent with experimental studies. However, in the case of model simulation of Wnt and EGF pathways, the model network did not supply any concurrent results with experimental data despite the fact that drugs were added at the appropriate positions. This paves insight into the potential of crosstalks between pathways considered in our study. Therefore, we manually developed a map of signaling crosstalk, which included the species connected by representatives from Notch, Shh, Wnt, and EGF pathways and highlighted the regulation of a single target gene, Hes-1, based on drug-induced simulations. These simulations provided results that matched with experimental studies. Therefore, these signaling crosstalk models complement as a tool toward the discovery of novel regulatory processes involved in neural stem cell maintenance, proliferation, and differentiation during mammalian central nervous system development. To our knowledge, this is the first report of a simple crosstalk map that highlights the differential regulation of neural stem cell differentiation and underscores the flow of positive and negative regulatory signals modulated by drugs.

  3. Integrated omics-analysis reveals Wnt-mediated NAD+ metabolic reprogramming in cancer stem-like cells

    PubMed Central

    Min, Soonki; Park, Ki Cheong; Park, Sunho; Hwang, Tae Hyun; Ryu, Do Hyun; Hwang, Geum-Sook; Cheong, Jae-Ho

    2016-01-01

    Abnormal tumor cell metabolism is a consequence of alterations in signaling pathways that provide critical selective advantage to cancer cells. However, a systematic characterization of the metabolic and signaling pathways altered in cancer stem-like cells (CSCs) is currently lacking. Using nuclear magnetic resonance and mass spectrometry, we profiled the whole-cell metabolites of a pair of parental (P-231) and stem-like cancer cells (S-231), and then integrated with whole transcriptome profiles. We identified elevated NAAD+ in S-231 along with a coordinated increased expression of genes in Wnt/calcium signaling pathway, reflecting the correlation between metabolic reprogramming and altered signaling pathways. The expression of CD38 and ALP, upstream NAAD+ regulatory enzymes, was oppositely regulated between P- and S-231; high CD38 strongly correlated with NAADP in P-231 while high ALP with NAAD+ levels in S-231. Antagonizing Wnt activity by dnTCF4 transfection reversed the levels of NAAD+ and ALP expression in S-231. Of note, elevated NAAD+ caused a decrease of cytosolic Ca2+ levels preventing calcium-induced apoptosis in nutrient-deprived conditions. Reprograming of NAD+ metabolic pathway instigated by Wnt signaling prevented cytosolic Ca2+ overload thereby inhibiting calcium-induced apoptosis in S-231. These results suggest that “oncometabolites” resulting from cross talk between the deranged core cancer signaling pathway and metabolic network provide a selective advantage to CSCs. PMID:27391070

  4. Murrayafoline A attenuates the Wnt/{beta}-catenin pathway by promoting the degradation of intracellular {beta}-catenin proteins

    SciTech Connect

    Choi, Hyuk; Gwak, Jungsug; Cho, Munju; Ryu, Min-Jung; Lee, Jee-Hyun; Kim, Sang Kyum; Kim, Young Ho; Lee, Gye Won; Yun, Mi-Young; Cuong, Nguyen Manh; Shin, Jae-Gook; Song, Gyu-Yong; Oh, Sangtaek

    2010-01-01

    Molecular lesions in Wnt/{beta}-catenin signaling and subsequent up-regulation of {beta}-catenin response transcription (CRT) occur frequently during the development of colon cancer. To identify small molecules that suppress CRT, we screened natural compounds in a cell-based assay for detection of TOPFalsh reporter activity. Murrayafoline A, a carbazole alkaloid isolated from Glycosmis stenocarpa, antagonized CRT that was stimulated by Wnt3a-conditioned medium (Wnt3a-CM) or LiCl, an inhibitor of glycogen synthase kinase-3{beta} (GSK-3{beta}), and promoted the degradation of intracellular {beta}-catenin without altering its N-terminal phosphorylation at the Ser33/37 residues, marking it for proteasomal degradation, or the expression of Siah-1, an E3 ubiquitin ligase. Murrayafoline A repressed the expression of cyclin D1 and c-myc, which is known {beta}-catenin/T cell factor (TCF)-dependent genes and thus inhibited the proliferation of various colon cancer cells. These findings indicate that murrayafoline A may be a potential chemotherapeutic agent for use in the treatment of colon cancer.

  5. Stress antagonizes morphine-induced analgesia in rats

    NASA Technical Reports Server (NTRS)

    Vernikos, J.; Shannon, L.; Heybach, J. P.

    1981-01-01

    Exposure to restraint stress resulted in antagonism of the analgesic effect of administered morphine in adult male rats. This antagonism of morphine-induced analgesia by restraint stress was not affected by adrenalectomy one day prior to testing, suggesting that stress-induced secretion of corticosteroids is not critical to this antagonism. In addition, parenteral administration of exogenous adrenocorticotropin (ACTH) mimicked the effect of stress in antagonizing morphine's analgesic efficacy. The hypothesis that ACTH is an endogenous opiate antagonist involved in modulating pain sensitivity is supported.

  6. Wnt signalling in neuronal differentiation and development.

    PubMed

    Inestrosa, Nibaldo C; Varela-Nallar, Lorena

    2015-01-01

    Wnts are secreted glycoproteins that play multiple roles in early development, including the differentiation of precursor cells. During this period, gradients of Wnts and other morphogens are formed and regulate the differentiation and migration of neural progenitor cells. Afterwards, Wnt signalling cascades participate in the formation of neuronal circuits, playing roles in dendrite and axon development, dendritic spine formation and synaptogenesis. Finally, in the adult brain, Wnts control hippocampal plasticity, regulating synaptic transmission and neurogenesis. In this review, we summarize the reported roles of Wnt signalling cascades in these processes with a particular emphasis on the role of Wnts in neuronal differentiation and development.

  7. Genome-wide gene expression profiling reveals aberrant MAPK and Wnt signaling pathways associated with early parthenogenesis.

    PubMed

    Liu, Na; Enkemann, Steven A; Liang, Ping; Hersmus, Remko; Zanazzi, Claudia; Huang, Junjiu; Wu, Chao; Chen, Zhisheng; Looijenga, Leendert H J; Keefe, David L; Liu, Lin

    2010-12-01

    Mammalian parthenogenesis could not survive but aborted during mid-gestation, presumably because of lack of paternal gene expression. To understand the molecular mechanisms underlying the failure of parthenogenesis at early stages of development, we performed global gene expression profiling and functional analysis of parthenogenetic blastocysts in comparison with those of blastocysts from normally fertilized embryos. Parthenogenetic blastocysts exhibited changes in the expression of 749 genes, of which 214 had lower expression and 535 showed higher expressions than fertilized embryos using a minimal 1.8-fold change as a cutoff. Genes important for placenta development were decreased in their expression in parthenote blastocysts. Some maternally expressed genes were up-regulated and paternal-related genes were down-regulated. Moreover, aberrantly increased Wnt signaling and reduced mitogen-activated protein kinase (MAPK) signaling were associated with early parthenogenesis. The protein level of extracellular signal-regulated kinase 2 (ERK2) was low in parthenogenetic blastocysts compared with that of fertilized blastocysts 120 h after fertilization. 6-Bromoindirubin-3'-oxime, a specific glycogen synthase kinase-3 (GSK-3) inhibitor, significantly decreased embryo hatching. The expression of several imprinted genes was altered in parthenote blastocysts. Gene expression also linked reduced expression of Xist to activation of X chromosome. Our findings suggest that failed X inactivation, aberrant imprinting, decreased ERK/MAPK signaling and possibly elevated Wnt signaling, and reduced expression of genes for placental development collectively may contribute to abnormal placenta formation and failed fetal development in parthenogenetic embryos.

  8. Wnt signaling in axial patterning and regeneration: lessons from planaria.

    PubMed

    De Robertis, Edward M

    2010-06-22

    Wnt signal transduction plays a crucial role in stem cell proliferation and regeneration. When canonical Wnt signaling is low, heads develop, and when it is high, tails are formed. In planarians, Wnt transcription is activated by wounding in a beta-catenin-independent way. Hedgehog is one of the signals involved, because it induces regeneration of tails (instead of heads) through the activation of Wnt transcription. Depletion of Smad4 blocks regeneration entirely, which suggests that the bone morphogenetic protein signaling pathway and the Wnt pathway are required for regeneration and body patterning.

  9. Wnt Signaling in Axial Patterning and Regeneration: Lessons from Planaria

    PubMed Central

    De Robertis, Edward M.

    2011-01-01

    Wnt signal transduction plays a crucial role in stem cell proliferation and regeneration. When canonical Wnt signaling is low, heads develop, and when it is high, tails are formed. In planarians, Wnt transcription is activated by wounding in a β-catenin–independent way. Hedgehog is one of the signals involved, because it induces regeneration of tails (instead of heads) through the activation of Wnt transcription. Depletion of Smad4 blocks regeneration entirely, which suggests that the bone morphogenetic protein signaling pathway and the Wnt pathway are required for regeneration and body patterning. PMID:20571126

  10. Revisiting the ERK/Src cortactin switch

    PubMed Central

    Kelley, Laura C; Hayes, Karen E; Ammer, Amanda Gatesman; Martin, Karen H

    2011-01-01

    The filamentous (F)-actin regulatory protein cortactin plays an important role in tumor cell movement and invasion by promoting and stabilizing actin related protein (Arp)2/3-mediated actin networks necessary for plasma membrane protrusion. Cortactin is a substrate for ERK1/2 and Src family kinases, with previous in vitro findings demonstrating ERK1/2 phosphorylation of cortactin as a positive and Src phosphorylation as a negative regulatory event in promoting Arp2/3 activation through neuronal Wiskott Aldrich Syndrome protein (N-WASp). Evidence for this regulatory cortactin “switch” in cells has been hampered due to the lack of phosphorylation-specific antibodies that recognize ERK1/2-phosphorylated cortactin. Our findings with phosphorylation-specific antibodies against these ERK1/2 sites (pS405 and pS418) indicate that cortactin can be co-phosphorylated at 405/418 and tyrosine residues targeted by Src family tyrosine kinases. These results indicate that the ERK/Src cortactin switch is not the sole mechanism by which ERK1/2 and tyrosine phosphorylation events regulate cortactin function in cell systems. PMID:21655441

  11. 6-HYDROXYDOPAMINE INDUCES MITOCHONDRIAL ERK ACTIVATION

    PubMed Central

    Kulich, Scott M.; Horbinski, Craig; Patel, Manisha; Chu, Charleen T.

    2007-01-01

    Reactive oxygen species (ROS) are implicated in 6-hydroxydopamine (6-OHDA) injury to catecholaminergic neurons; however, the mechanism(s) are unclear. In addition to ROS generated during autoxidation, 6-OHDA may initiate secondary cellular sources of ROS that contribute to toxicity. Using a neuronal cell line, we found that catalytic metalloporphyrin antioxidants conferred protection if added 1 hour after exposure to 6-OHDA, whereas the hydrogen peroxide scavenger catalase failed to protect if added more than 15 min after 6-OHDA. There was a temporal correspondence between loss of protection and loss of the ability of the antioxidant to inhibit 6-OHDA-induced ERK phosphorylation. Time course studies of aconitase inactivation, as an indicator of intracellular superoxide, and MitoSOX red, a mitochondria targeted ROS indicator, demonstrate early intracellular ROS followed by a delayed phase of mitochondrial ROS production, associated with phosphorylation of a mitochondrial pool of ERK. Furthermore, upon initiation of mitochondrial ROS and ERK activation, 6-OHDA-injured cells became refractory to rescue by metalloporphyrin antioxidants. Together with previous studies showing that inhibition of the ERK pathway confers protection from 6-OHDA toxicity, and that phosphorylated ERK accumulates in mitochondria of degenerating human Parkinson’s disease neurons, these studies implicate mitochondrial ERK activation in Parkinsonian oxidative neuronal injury. PMID:17602953

  12. Identification and Mechanism of ABA Receptor Antagonism

    PubMed Central

    Melcher, Karsten; Xu, Yong; Ng, Ley-Moy; Zhou, X. Edward; Soon, Fen-Fen; Chinnusamy, Viswanathan; Suino-Powell, Kelly M.; Kovach, Amanda; Tham, Fook S.; Cutler, Sean R.; Li, Jun; Yong, Eu-Leong; Zhu, Jian-Kang; Xu, H. Eric

    2010-01-01

    The phytohormone abscisic acid (ABA) functions through a family of fourteen PYR/PYL receptors, which were identified by resistance to pyrabactin, a synthetic inhibitor of seed germination. ABA activates these receptors to inhibit type 2C protein phosphatases, such as ABI1, yet it remains unclear whether these receptors can be antagonized. Here we demonstrate that pyrabactin is an agonist of PYR1 and PYL1, but unexpectedly an antagonist of PYL2. Crystal structures of the PYL2–pyrabactin and PYL1–pyrabactin–ABI1 complexes reveal the mechanism responsible for receptor-selective activation and inhibition, which enables us to design mutations that convert PYL1 to a pyrabactin-inhibited receptor and PYL2 to a pyrabactin-activated receptor, and to identify new pyrabactin-based ABA receptor agonists. Together, our results establish a new concept of ABA receptor antagonism, illustrate its underlying mechanisms, and provide a rational framework for discovering novel ABA receptor ligands. PMID:20729862

  13. Does zaltoprofen antagonize the bradykinin receptors?

    PubMed

    Bawolak, Marie-Thérèse; Marceau, François

    2007-05-03

    Zaltoprofen is a nonsteroidal antiinflammatory drug that has been proposed to inhibit with some selectivity the nociception mediated by the bradykinin (BK) B(2) receptor. In order to test the predictive power of this claim, we applied the drug to vascular smooth muscle assays previously found useful to characterize B(2) receptor antagonists (contractility, human isolated umbilical vein) or B(1) receptor antagonists (contraction, rabbit aorta; relaxation, rabbit mesenteric artery). Zaltoprofen (up to 30 microM) failed to antagonize BK or des-Arg(9)-BK-induced contraction in the umbilical vein and aorta, respectively. The drug (1 microM) abated des-Arg(9)-BK-induced, prostaglandin-mediated relaxation of the precontracted mesenteric artery, consistent with its known activity as a cyclooxygenase (COX) inhibitor. However, zaltoprofen (10 microM) did not inhibit kinin-stimulated phospholipase A(2) activity in HEK 293 cells expressing recombinant forms of the rabbit B(1) or B(2) receptors. Nonpeptide antagonists of either receptor subtype were active in this respect. The results do not support that zaltoprofen, a COX inhibitor, antagonizes kinin receptors or influences their signaling with selectivity in the tested systems.

  14. Targeting the Wnt/β-catenin pathway in primary ovarian cancer with the porcupine inhibitor WNT974.

    PubMed

    Boone, Jonathan D; Arend, Rebecca C; Johnston, Bobbi E; Cooper, Sara J; Gilchrist, Scott A; Oelschlager, Denise K; Grizzle, William E; McGwin, Gerald; Gangrade, Abhishek; Straughn, J Michael; Buchsbaum, Donald J

    2016-02-01

    Preclinical studies in ovarian cancer have demonstrated upregulation of the Wnt/β-catenin pathway promoting tumor proliferation and chemoresistance. Our objective was to evaluate the effect of the Wnt/β-catenin pathway inhibitor, WNT974, in primary ovarian cancer ascites cells. Ascites cells from patients with papillary serous ovarian cancer were isolated and treated with 1 μM WNT974±100 μM carboplatin. Viability was evaluated with the ATPlite assay. The IC50 was calculated using a dose-response analysis. Immunohistochemistry (IHC) was performed on ascites cells and tumor. Expression of R-spondin 2 (RSPO2), RSPO3, PORCN, WLS, AXIN2, and three previously characterized RSPO fusion transcripts were assessed using Taqman assays. Sixty ascites samples were analyzed for response to WNT974. The ascites samples that showed a decrease in ATP concentration after treatment demonstrated no difference from the untreated cells in percent viability with trypan blue staining. Flow cytometry demonstrated fewer cells in the G2 phase and more in the G1 and S phases after treatment with WNT974. Combination therapy with WNT974 and carboplatin resulted in a higher percentage of samples that showed ≥30% reduction in ATP concentration than either single drug treatment. IHC analysis of Wnt pathway proteins suggests cell cycle arrest rather than cytotoxicity after WNT974 treatment. QPCR indicated that RSPO fusions are not prevalent in ovarian cancer tissues or ascites. However, higher PORCN expression correlated to sensitivity to WNT974 (P=0.0073). In conclusion, WNT974 produces cytostatic effects in patient ascites cells with primary ovarian cancer through inhibition of the Wnt/β-catenin pathway. The combination of WNT974 and carboplatin induces cytotoxicity plus cell cycle arrest in a higher percentage of ascites samples than with single drug treatment. RSPO fusions do not contribute to WNT974 sensitivity; however, higher PORCN expression indicates increased WNT974 sensitivity.

  15. Transcriptional regulation of WNT2B based on the balance of Hedgehog, Notch, BMP and WNT signals.

    PubMed

    Katoh, Masuko; Katoh, Masaru

    2009-05-01

    We cloned and characterized human WNT2B in 1996, and then others cloned and characterized mouse, chicken, and zebrafish WNT2B orthologs. WNT2B is expressed in several types of human cancer, such as basal cell carcinoma, gastric cancer, breast cancer, head/neck squamous cell carcinoma, cervical cancer and leukemia. WNT2B is one of canonical WNTs transducing signals through Frizzled (FZD) and LRP5/LRP6 receptors to beta-catenin-TCF/LEF signaling cascade. Here, refined integrative genomic analyses on WNT2B orthologs were carried out to elucidate its transcriptional mechanisms. GLI-, double FOX-, HES/HEY-, bHLH-, and Sp1-binding sites within mammalian WNT2B promoter were well conserved. Because GLI1, FOXA2, FOXC2, FOXE1, FOXF1 and FOXL1 are direct target genes of Hedgehog-GLI2 signaling cascade, Hedgehog signals should induce WNT2B upregulation through GLI family members as well as FOX family members. Notch, BMP and Hedgehog signals inhibit WNT2B expression via HES/HEY-binding to N-box, whereas BMP and WNT signals inhibit bHLH transcription factor-induced WNT2B expression via ID1, ID2, ID3, MSX1 or MSX2. Together these facts indicate that Hedgehog signals and bHLH transcription factors are involved in WNT2B upregulation, which is counteracted by BMP, WNT and Notch signals. Mesenchymal BMP induces IHH expression in gastrointestinal epithelial cells, and then epithelial Hedgehog induces WNT2B and BMP4 expression in mesenchymal cells. NF-kappaB signals induce SHH upregulation, and WNT2B is upregulated in inflammatory bowel disease (IBD). BMP-IHH and inflammation-SHH signaling loops are involved in WNT2B up-regulation during embryogenesis, adult tissue homeostasis, and carcinogenesis.

  16. The WNT-less wonder: WNT-independent β-catenin signaling.

    PubMed

    Aktary, Zackie; Bertrand, Juliette U; Larue, Lionel

    2016-09-01

    β-catenin is known as an Armadillo protein that regulates gene expression following WNT pathway activation. However, WNT-independent pathways also activate β-catenin. During the establishment of the melanocyte lineage, β-catenin plays an important role. In the context of physiopathology, β-catenin is activated genetically or transiently in various cancers, including melanoma, where it can be found in the nucleus of tumors. In this review, we discuss alternative pathways that activate β-catenin independent of WNTs and highlight what is known regarding these pathways in melanoma. We also discuss the role of β-catenin as a transcriptional regulator in various cell types, with emphasis on the different transcription factors it associates with independent of WNT induction. Finally, the role of WNT-independent β-catenin in melanocyte development and melanomagenesis is also discussed. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  17. USP6 oncogene promotes Wnt signaling by deubiquitylating Frizzleds

    PubMed Central

    Madan, Babita; Walker, Matthew P.; Young, Robert; Quick, Laura; Orgel, Kelly A.; Ryan, Meagan; Gupta, Priti; Henrich, Ian C.; Ferrer, Marc; Marine, Shane; Roberts, Brian S.; Arthur, William T.; Berndt, Jason D.; Oliveira, Andre M.; Moon, Randall T.; Chou, Margaret M.; Major, Michael B.

    2016-01-01

    The Wnt signaling pathways play pivotal roles in carcinogenesis. Modulation of the cell-surface abundance of Wnt receptors is emerging as an important mechanism for regulating sensitivity to Wnt ligands. Endocytosis and degradation of the Wnt receptors Frizzled (Fzd) and lipoprotein-related protein 6 (LRP6) are regulated by the E3 ubiquitin ligases zinc and ring finger 3 (ZNRF3) and ring finger protein 43 (RNF43), which are disrupted in cancer. In a genome-wide small interfering RNA screen, we identified the deubiquitylase ubiquitin-specific protease 6 (USP6) as a potent activator of Wnt signaling. USP6 enhances Wnt signaling by deubiquitylating Fzds, thereby increasing their cell-surface abundance. Chromosomal translocations in nodular fasciitis result in USP6 overexpression, leading to transcriptional activation of the Wnt/β-catenin pathway. Inhibition of Wnt signaling using Dickkopf-1 (DKK1) or a Porcupine (PORCN) inhibitor significantly decreased the growth of USP6-driven xenograft tumors, indicating that Wnt signaling is a key target of USP6 during tumorigenesis. Our study defines an additional route to ectopic Wnt pathway activation in human disease, and identifies a potential approach to modulate Wnt signaling for therapeutic benefit. PMID:27162353

  18. Distinct Wnt signaling pathways have opposing roles in appendage regeneration.

    PubMed

    Stoick-Cooper, Cristi L; Weidinger, Gilbert; Riehle, Kimberly J; Hubbert, Charlotte; Major, Michael B; Fausto, Nelson; Moon, Randall T

    2007-02-01

    In contrast to mammals, lower vertebrates have a remarkable capacity to regenerate complex structures damaged by injury or disease. This process, termed epimorphic regeneration, involves progenitor cells created through the reprogramming of differentiated cells or through the activation of resident stem cells. Wnt/beta-catenin signaling regulates progenitor cell fate and proliferation during embryonic development and stem cell function in adults, but its functional involvement in epimorphic regeneration has not been addressed. Using transgenic fish lines, we show that Wnt/beta-catenin signaling is activated in the regenerating zebrafish tail fin and is required for formation and subsequent proliferation of the progenitor cells of the blastema. Wnt/beta-catenin signaling appears to act upstream of FGF signaling, which has recently been found to be essential for fin regeneration. Intriguingly, increased Wnt/beta-catenin signaling is sufficient to augment regeneration, as tail fins regenerate faster in fish heterozygous for a loss-of-function mutation in axin1, a negative regulator of the pathway. Likewise, activation of Wnt/beta-catenin signaling by overexpression of wnt8 increases proliferation of progenitor cells in the regenerating fin. By contrast, overexpression of wnt5b (pipetail) reduces expression of Wnt/beta-catenin target genes, impairs proliferation of progenitors and inhibits fin regeneration. Importantly, fin regeneration is accelerated in wnt5b mutant fish. These data suggest that Wnt/beta-catenin signaling promotes regeneration, whereas a distinct pathway activated by wnt5b acts in a negative-feedback loop to limit regeneration.

  19. Phosphatase resistance of ERK2 brain kinase PK40erk2.

    PubMed

    Roder, H M; Hoffman, F J; Schröder, W

    1995-05-01

    We have previously shown that a brain protein kinase, termed PK40, catalyzes the multiple phosphorylation of the KSP-repeat site of neurofilaments (NFs) and also can transform tau proteins into the paired helical filament-like state as found in Alzheimer's disease (AD) brains. Protein sequence analysis suggests that PK40 is a form of the extracellular signal-regulated kinase ERK2. A subpopulation of ERK2 species in soluble brain fractions can be efficiently phosphorylated and activated in cell-free systems, simply by adding Mg(2+)-ATP. Two phosphoisoforms of PK40erk2 are formed in this process, which have a reduced gel mobility, very much like the ERK2 form obtained in cell culture by stimulation with growth factors. One of these low-mobility forms cannot be inactivated with protein phosphatase 2A (PP2A) or with tyrosine phosphatases. The second form can be slowly inactivated by PP2A. In this case two Ser/Thr phosphates are removed at different rates during inactivation: One phosphate is very quickly removed to result in the formation of a high-mobility 39-kDa ERK2 species without consequence for activity; the other, slowly removed Ser/Thr phosphate controls the activity but has no effect on the gel mobility of ERK2. These results show that forms of ERK2 exist with properties different from the previously characterized ERK2 (p42mapk) from stimulated cell cultures. The active ERK2 forms produced in the presence of Mg(2+)-ATP alone could provide an explanation for the existence of constitutive ERK2-like NF phosphorylation in vivo. Excessive formation of an ERK2 species resistant to inactivation by PP2A might be relevant to the persistent pathological tau hyperphosphorylation in AD.

  20. The nuclear receptor NHR-25 cooperates with the Wnt/β-catenin asymmetry pathway to control differentiation of the T seam cell in C. elegans

    PubMed Central

    Hajduskova, Martina; Jindra, Marek; Herman, Michael A.; Asahina, Masako

    2009-01-01

    Summary Asymmetric cell divisions produce new cell types during animal development. Studies in Caenorhabditis elegans have identified major signal-transduction pathways that determine the polarity of cell divisions. How these relatively few conserved pathways interact and what modulates them to ensure the diversity of multiple tissue types is an open question. The Wnt/β-catenin asymmetry pathway governs polarity of the epidermal T seam cell in the C. elegans tail. Here, we show that the asymmetry of T-seam-cell division and morphogenesis of the male sensory rays require NHR-25, an evolutionarily conserved nuclear receptor. NHR-25 ensures the neural fate of the T-seam-cell descendants in cooperation with the Wnt/β-catenin asymmetry pathway. Loss of NHR-25 enhances the impact of mutated nuclear effectors of this pathway, POP-1 (TCF) and SYS-1 (β-catenin), on T-seam-cell polarity, whereas it suppresses the effect of the same mutations on asymmetric division of the somatic gonad precursor cells. Therefore, NHR-25 can either synergize with or antagonize the Wnt/β-catenin asymmetry pathway depending on the tissue context. Our findings define NHR-25 as a versatile modulator of Wnt/β-catenin-dependent cell-fate decisions. PMID:19654209

  1. Wnt2 regulates progenitor proliferation in the developing ventral midbrain.

    PubMed

    Sousa, Kyle M; Villaescusa, J Carlos; Cajanek, Lukas; Ondr, Jennifer K; Castelo-Branco, Goncalo; Hofstra, Wytske; Bryja, Vitezslav; Palmberg, Carina; Bergman, Tomas; Wainwright, Brandon; Lang, Richard A; Arenas, Ernest

    2010-03-05

    Wnts are secreted, lipidated proteins that regulate multiple aspects of brain development, including dopaminergic neuron development. In this study, we perform the first purification and signaling analysis of Wnt2 and define the function of Wnt2 in ventral midbrain precursor cultures, as well as in Wnt2-null mice in vivo. We found that purified Wnt2 induces the phosphorylation of both Lrp5/6 and Dvl-2/3, and activates beta-catenin in SN4741 dopaminergic cells. Moreover, purified Wnt2 increases progenitor proliferation, and the number of dopaminergic neurons in ventral midbrain precursor cultures. In agreement with these findings, analysis of the ventral midbrain of developing Wnt2-null mice revealed a decrease in progenitor proliferation and neurogenesis that lead to a decrease in the number of postmitotic precursors and dopaminergic neurons. Collectively, our observations identify Wnt2 as a novel regulator of dopaminergic progenitors and dopaminergic neuron development.

  2. Wnt2 Regulates Progenitor Proliferation in the Developing Ventral Midbrain*

    PubMed Central

    Sousa, Kyle M.; Villaescusa, J. Carlos; Cajanek, Lukas; Ondr, Jennifer K.; Castelo-Branco, Goncalo; Hofstra, Wytske; Bryja, Vitezslav; Palmberg, Carina; Bergman, Tomas; Wainwright, Brandon; Lang, Richard A.; Arenas, Ernest

    2010-01-01

    Wnts are secreted, lipidated proteins that regulate multiple aspects of brain development, including dopaminergic neuron development. In this study, we perform the first purification and signaling analysis of Wnt2 and define the function of Wnt2 in ventral midbrain precursor cultures, as well as in Wnt2-null mice in vivo. We found that purified Wnt2 induces the phosphorylation of both Lrp5/6 and Dvl-2/3, and activates β-catenin in SN4741 dopaminergic cells. Moreover, purified Wnt2 increases progenitor proliferation, and the number of dopaminergic neurons in ventral midbrain precursor cultures. In agreement with these findings, analysis of the ventral midbrain of developing Wnt2-null mice revealed a decrease in progenitor proliferation and neurogenesis that lead to a decrease in the number of postmitotic precursors and dopaminergic neurons. Collectively, our observations identify Wnt2 as a novel regulator of dopaminergic progenitors and dopaminergic neuron development. PMID:20018874

  3. Antagonizing Retinoic Acid and FGF/MAPK Pathways Control Posterior Body Patterning in the Invertebrate Chordate Ciona intestinalis

    PubMed Central

    Pasini, Andrea; Manenti, Raoul; Rothbächer, Ute; Lemaire, Patrick

    2012-01-01

    Vertebrate embryos exploit the mutual inhibition between the RA and FGF signalling pathways to coordinate the proliferative elongation of the main body axis with the progressive patterning and differentiation of its neuroectodermal and paraxial mesodermal structures. The evolutionary history of this patterning system is still poorly understood. Here, we investigate the role played by the RA and FGF/MAPK signals during the development of the tail structures in the tunicate Ciona intestinalis, an invertebrate chordate belonging to the sister clade of vertebrates, in which the prototypical chordate body plan is established through very derived morphogenetic processes. Ciona embryos are constituted of few cells and develop according to a fixed lineage; elongation of the tail occurs largely by rearrangement of postmitotic cells; mesoderm segmentation and somitogenesis are absent. We show that in the Ciona embryo, the antagonism of the RA and FGF/MAPK signals is required to control the anteroposterior patterning of the tail epidermis. We also demonstrate that the RA, FGF/MAPK and canonical Wnt pathways control the anteroposterior patterning of the tail peripheral nervous system, and reveal the existence of distinct subpopulations of caudal epidermal neurons with different responsiveness to the RA, FGF/MAPK and canonical Wnt signals. Our data provide the first demonstration that the use of the antagonism between the RA and FGF signals to pattern the main body axis predates the emergence of vertebrates and highlight the evolutionary plasticity of this patterning strategy, showing that in different chordates it can be used to pattern different tissues within the same homologous body region. PMID:23049976

  4. Antagonizing retinoic acid and FGF/MAPK pathways control posterior body patterning in the invertebrate chordate Ciona intestinalis.

    PubMed

    Pasini, Andrea; Manenti, Raoul; Rothbächer, Ute; Lemaire, Patrick

    2012-01-01

    Vertebrate embryos exploit the mutual inhibition between the RA and FGF signalling pathways to coordinate the proliferative elongation of the main body axis with the progressive patterning and differentiation of its neuroectodermal and paraxial mesodermal structures. The evolutionary history of this patterning system is still poorly understood. Here, we investigate the role played by the RA and FGF/MAPK signals during the development of the tail structures in the tunicate Ciona intestinalis, an invertebrate chordate belonging to the sister clade of vertebrates, in which the prototypical chordate body plan is established through very derived morphogenetic processes. Ciona embryos are constituted of few cells and develop according to a fixed lineage; elongation of the tail occurs largely by rearrangement of postmitotic cells; mesoderm segmentation and somitogenesis are absent. We show that in the Ciona embryo, the antagonism of the RA and FGF/MAPK signals is required to control the anteroposterior patterning of the tail epidermis. We also demonstrate that the RA, FGF/MAPK and canonical Wnt pathways control the anteroposterior patterning of the tail peripheral nervous system, and reveal the existence of distinct subpopulations of caudal epidermal neurons with different responsiveness to the RA, FGF/MAPK and canonical Wnt signals. Our data provide the first demonstration that the use of the antagonism between the RA and FGF signals to pattern the main body axis predates the emergence of vertebrates and highlight the evolutionary plasticity of this patterning strategy, showing that in different chordates it can be used to pattern different tissues within the same homologous body region.

  5. The natural product 4,10-aromadendranediol induces neuritogenesis in neuronal cells in vitro through activation of the ERK pathway

    PubMed Central

    Chang, Sai; Ruan, Wen-chen; Xu, Ya-zhou; Wang, Yun-jie; Pang, Jie; Zhang, Lu-yong; Liao, Hong; Pang, Tao

    2017-01-01

    Recent studies focus on promoting neurite outgrowth to remodel the central nervous network after brain injury. Currently, however, there are few drugs treating brain diseases in the clinic by enhancing neurite outgrowth. In this study, we established an NGF-induced PC12 differentiation model to screen novel compounds that have the potential to induce neuronal differentiation, and further characterized 4,10-Aromadendranediol (ARDD) isolated from the dried twigs of the Baccharis gaudichaudiana plant, which exhibited the capability of promoting neurite outgrowth in neuronal cells in vitro. ARDD (1, 10 μmol/L) significantly enhanced neurite outgrowth in NGF-treated PC12 cells and N1E115 cells in a time-dependent manner. In cultured primary cortical neurons, ARDD (5, 10 μmol/L) not only significantly increased neurite outgrowth but also increased the number of neurites on the soma and the number of bifurcations. Further analyses showed that ARDD (10 μmol/L) significantly increased the phosphorylation of ERK1/2 and the downstream GSK-3β, subsequently induced β-catenin expression and up-regulated the gene expression of the Wnt ligands Fzd1 and Wnt3a in neuronal cells. The neurite outgrowth-promoting effect of ARDD in neuronal cells was abolished by pretreatment with the specific ERK1/2 inhibitor PD98059, but was partially reversed by XAV939, an inhibitor of the Wnt/β-catenin pathway. ARDD also increased the expression of BDNF, CREB and GAP-43 in N1E115 cells, which was reversed by pretreatment with PD98059. In N1E115 cells subjected to oxygen and glucose deprivation (OGD), pretreatment with ARDD (1–10 μmol/L) significantly enhanced the phosphorylation of ERK1/2 and induced neurite outgrowth. These results demonstrated that the natural product ARDD exhibits neurite outgrowth-inducing activity in neurons via activation of the ERK signaling pathway, which may be beneficial to the treatment of brain diseases. PMID:27840407

  6. The natural product 4,10-aromadendranediol induces neuritogenesis in neuronal cells in vitro through activation of the ERK pathway.

    PubMed

    Chang, Sai; Ruan, Wen-Chen; Xu, Ya-Zhou; Wang, Yun-Jie; Pang, Jie; Zhang, Lu-Yong; Liao, Hong; Pang, Tao

    2017-01-01

    Recent studies focus on promoting neurite outgrowth to remodel the central nervous network after brain injury. Currently, however, there are few drugs treating brain diseases in the clinic by enhancing neurite outgrowth. In this study, we established an NGF-induced PC12 differentiation model to screen novel compounds that have the potential to induce neuronal differentiation, and further characterized 4,10-Aromadendranediol (ARDD) isolated from the dried twigs of the Baccharis gaudichaudiana plant, which exhibited the capability of promoting neurite outgrowth in neuronal cells in vitro. ARDD (1, 10 μmol/L) significantly enhanced neurite outgrowth in NGF-treated PC12 cells and N1E115 cells in a time-dependent manner. In cultured primary cortical neurons, ARDD (5, 10 μmol/L) not only significantly increased neurite outgrowth but also increased the number of neurites on the soma and the number of bifurcations. Further analyses showed that ARDD (10 μmol/L) significantly increased the phosphorylation of ERK1/2 and the downstream GSK-3β, subsequently induced β-catenin expression and up-regulated the gene expression of the Wnt ligands Fzd1 and Wnt3a in neuronal cells. The neurite outgrowth-promoting effect of ARDD in neuronal cells was abolished by pretreatment with the specific ERK1/2 inhibitor PD98059, but was partially reversed by XAV939, an inhibitor of the Wnt/β-catenin pathway. ARDD also increased the expression of BDNF, CREB and GAP-43 in N1E115 cells, which was reversed by pretreatment with PD98059. In N1E115 cells subjected to oxygen and glucose deprivation (OGD), pretreatment with ARDD (1-10 μmol/L) significantly enhanced the phosphorylation of ERK1/2 and induced neurite outgrowth. These results demonstrated that the natural product ARDD exhibits neurite outgrowth-inducing activity in neurons via activation of the ERK signaling pathway, which may be beneficial to the treatment of brain diseases.

  7. Altered Expression of Wnt Signaling Pathway Components in Osteogenesis of Mesenchymal Stem Cells in Osteoarthritis Patients

    PubMed Central

    Herranz, Eva; Rodríguez-Rodríguez, Luis; Mucientes, Arkaitz; Abásolo, Lydia; Marco, Fernando; Fernández-Gutiérrez, Benjamín; Lamas, José Ramón

    2015-01-01

    Introduction Osteoarthritis (OA) is characterized by altered homeostasis of joint cartilage and bone, whose functional properties rely on chondrocytes and osteoblasts, belonging to mesenchymal stem cells (MSCs). WNT signaling acts as a hub integrating and crosstalking with other signaling pathways leading to the regulation of MSC functions. The aim of this study was to evaluate the existence of a differential signaling between Healthy and OA-MSCs during osteogenesis. Methods MSCs of seven OA patients and six healthy controls were isolated, characterised and expanded. During in vitro osteogenesis, cells were recovered at days 1, 10 and 21. RNA and protein content was obtained. Expression of WNT pathway genes was evaluated using RT-qPCR. Functional studies were also performed to study the MSC osteogenic commitment and functional and post-traslational status of β-catenin and several receptor tyrosine kinases. Results Several genes were downregulated in OA-MSCs during osteogenesis in vitro. These included soluble Wnts, inhibitors, receptors, co-receptors, several kinases and transcription factors. Basal levels of β-catenin were higher in OA-MSCs, but calcium deposition and expression of osteogenic genes was similar between Healthy and OA-MSCs. Interestingly an increased phosphorylation of p44/42 MAPK (ERK1/2) signaling node was present in OA-MSCs. Conclusion Our results point to the existence in OA-MSCs of alterations in expression of Wnt pathway components during in vitro osteogenesis that are partially compensated by post-translational mechanisms modulating the function of other pathways. We also point the relevance of other signaling pathways in OA pathophysiology suggesting their role in the maintenance of joint homeostasis through modulation of MSC osteogenic potential. PMID:26352263

  8. Expression of Wnt Signaling Components during Xenopus Pronephros Development

    PubMed Central

    Zhang, Bo; Tran, Uyen; Wessely, Oliver

    2011-01-01

    Background The formation of the vertebrate kidney is tightly regulated and relies on multiple evolutionarily conserved inductive events. These are present in the complex metanephric kidney of higher vertebrates, but also in the more primitive pronephric kidney functional in the larval stages of amphibians and fish. Wnts have long been viewed as central in this process. Canonical β-Catenin-dependent Wnt signaling establishes kidney progenitors and non-canonical β-Catenin-independent Wnt signaling participate in the morphogenetic processes that form the highly sophisticated nephron structure. While some individual Wnt signaling components have been studied extensively in the kidney, the overall pathway has not yet been analyzed in depth. Methodology/Principal Findings Here we report a detailed expression analysis of all Wnt ligands, receptors and several downstream Wnt effectors during pronephros development in Xenopus laevis using in situ hybridization. Out of 19 Wnt ligands, only three, Wnt4, Wnt9a and Wnt11, are specifically expressed in the pronephros. Others such as Wnt8a are present, but in a broader domain comprising adjacent tissues in addition to the kidney. The same paradigm is observed for the Wnt receptors and its downstream signaling components. Fzd1, Fzd4, Fzd6, Fzd7, Fzd8 as well as Celsr1 and Prickle1 show distinct expression domains in the pronephric kidney, whereas the non-traditional Wnt receptors, Ror2 and Ryk, as well as the majority of the effector molecules are rather ubiquitous. In addition to this spatial regulation, the timing of expression is also tightly regulated. In particular, non-canonical Wnt signaling seems to be restricted to later stages of pronephros development. Conclusion/Significance Together these data suggest a complex cross talk between canonical and non-canonical Wnt signaling is required to establish a functional pronephric kidney. PMID:22028899

  9. p120-catenin in canonical Wnt signaling.

    PubMed

    Duñach, Mireia; Del Valle-Pérez, Beatriz; García de Herreros, Antonio

    2017-03-03

    Canonical Wnt signaling controls β-catenin protein stabilization, its translocation to the nucleus and the activation of β-catenin/Tcf-4-dependent transcription. In this review, we revise and discuss the recent results describing actions of p120-catenin in different phases of this pathway. More specifically, we comment its involvement in four different steps: (i) the very early activation of CK1ɛ, essential for Dvl-2 binding to the Wnt receptor complex; (ii) the internalization of GSK3 and Axin into multivesicular bodies, necessary for a complete stabilization of β-catenin; (iii) the activation of Rac1 small GTPase, required for β-catenin translocation to the nucleus; and (iv) the release of the inhibitory action caused by Kaiso transcriptional repressor. We integrate these new results with the previously known action of other elements in this pathway, giving a particular relevance to the responses of the Wnt pathway not required for β-catenin stabilization but for β-catenin transcriptional activity. Moreover, we discuss the possible future implications, suggesting that the two cellular compartments where β-catenin is localized, thus, the adherens junction complex and the Wnt signalosome, are more physically connected that previously thought.

  10. Wnt5a-mediated non-canonical Wnt signalling regulates human endothelial cell proliferation and migration

    SciTech Connect

    Cheng Chingwen Yeh Juching; Fan Taiping; Smith, Stephen K.; Charnock-Jones, D. Stephen

    2008-01-11

    Cell to cell interaction is one of the key processes effecting angiogenesis and endothelial cell function. Wnt signalling is mediated through cell-cell interaction and is involved in many developmental processes and cellular functions. In this study, we investigated the possible function of Wnt5a and the non-canonical Wnt pathway in human endothelial cells. We found that Wnt5a-mediated non-canonical Wnt signalling regulated endothelial cell proliferation. Blocking this pathway using antibody, siRNA or a down-stream inhibitor led to suppression of endothelial cell proliferation, migration, and monolayer wound closure. We also found that the mRNA level of Wnt5a is up-regulated when endothelial cells are treated with a cocktail of inflammatory cytokines. Our findings suggest non-canonical Wnt signalling plays a role in regulating endothelial cell growth and possibly in angiogenesis.

  11. Gut Commensal Bacteria and Regional Wnt Gene Expression in the Proximal Versus Distal Colon

    PubMed Central

    Neumann, Philipp-Alexander; Koch, Stefan; Hilgarth, Roland S.; Perez-Chanona, Ernesto; Denning, Patricia; Jobin, Christian; Nusrat, Asma

    2015-01-01

    Regional expression of Wingless/Int (Wnt) genes plays a central role in regulating intestinal development and homeostasis. However, our knowledge of such regional Wnt proteins in the colon remains limited. To understand further the effect of Wnt signaling components in controlling intestinal epithelial homeostasis, we investigated whether the physiological heterogeneity of the proximal and distal colon can be explained by differential Wnt signaling. With the use of a Wnt signaling-specific PCR array, expression of 84 Wnt-mediated signal transduction genes was analyzed, and a differential signature of Wnt-related genes in the proximal versus distal murine colon was identified. Several Wnt agonists (Wnt5a, Wnt8b, and Wnt11), the Wnt receptor frizzled family receptor 3, and the Wnt inhibitory factor 1 were differentially expressed along the colon length. These Wnt signatures were associated with differential epithelial cell proliferation and migration in the proximal versus distal colon. Furthermore, reduced Wnt/β-catenin activity and decreased Wnt5a and Wnt11 expression were observed in mice lacking commensal bacteria, an effect that was reversed by conventionalization of germ-free mice. Interestingly, myeloid differentiation primary response gene 88 knockout mice showed decreased Wnt5a levels, indicating a role for Toll-like receptor signaling in regulating Wnt5a expression. Our results suggest that the morphological and physiological heterogeneity within the colon is in part facilitated by the differential expression of Wnt signaling components and influenced by colonization with bacteria. PMID:24418259

  12. Gut commensal bacteria and regional Wnt gene expression in the proximal versus distal colon.

    PubMed

    Neumann, Philipp-Alexander; Koch, Stefan; Hilgarth, Roland S; Perez-Chanona, Ernesto; Denning, Patricia; Jobin, Christian; Nusrat, Asma

    2014-03-01

    Regional expression of Wingless/Int (Wnt) genes plays a central role in regulating intestinal development and homeostasis. However, our knowledge of such regional Wnt proteins in the colon remains limited. To understand further the effect of Wnt signaling components in controlling intestinal epithelial homeostasis, we investigated whether the physiological heterogeneity of the proximal and distal colon can be explained by differential Wnt signaling. With the use of a Wnt signaling-specific PCR array, expression of 84 Wnt-mediated signal transduction genes was analyzed, and a differential signature of Wnt-related genes in the proximal versus distal murine colon was identified. Several Wnt agonists (Wnt5a, Wnt8b, and Wnt11), the Wnt receptor frizzled family receptor 3, and the Wnt inhibitory factor 1 were differentially expressed along the colon length. These Wnt signatures were associated with differential epithelial cell proliferation and migration in the proximal versus distal colon. Furthermore, reduced Wnt/β-catenin activity and decreased Wnt5a and Wnt11 expression were observed in mice lacking commensal bacteria, an effect that was reversed by conventionalization of germ-free mice. Interestingly, myeloid differentiation primary response gene 88 knockout mice showed decreased Wnt5a levels, indicating a role for Toll-like receptor signaling in regulating Wnt5a expression. Our results suggest that the morphological and physiological heterogeneity within the colon is in part facilitated by the differential expression of Wnt signaling components and influenced by colonization with bacteria.

  13. Endothelin ETA receptor antagonism in cardiovascular disease.

    PubMed

    Nasser, Suzanne A; El-Mas, Mahmoud M

    2014-08-15

    Since the discovery of the endothelin system in 1988, it has been implicated in numerous physiological and pathological phenomena. In the cardiovascular system, endothelin-1 (ET-1) acts through intracellular pathways of two endothelin receptors (ETA and ETB) located mainly on smooth muscle and endothelial cells to regulate vascular tone and provoke mitogenic and proinflammatory reactions. The endothelin ETA receptor is believed to play a pivotal role in the pathogenesis of several cardiovascular disease including systemic hypertension, pulmonary arterial hypertension (PAH), dilated cardiomyopathy, and diabetic microvascular dysfunction. Growing evidence from recent experimental and clinical studies indicates that the blockade of endothelin receptors, particularly the ETA subtype, grasps promise in the treatment of major cardiovascular pathologies. The simultaneous blockade of endothelin ETB receptors might not be advantageous, leading possibly to vasoconstriction and salt and water retentions. This review summarizes the role of ET-1 in cardiovascular modulation and the therapeutic potential of endothelin receptor antagonism.

  14. Wnt signaling and the control of human stem cell fate.

    PubMed

    Van Camp, J K; Beckers, S; Zegers, D; Van Hul, W

    2014-04-01

    Wnt signaling determines major developmental processes in the embryonic state and regulates maintenance, self-renewal and differentiation of adult mammalian tissue stem cells. Both β-catenin dependent and independent Wnt pathways exist, and both affect stem cell fate in developing and adult tissues. In this review, we debate the response to Wnt signal activation in embryonic stem cells and human, adult stem cells of mesenchymal, hematopoetic, intestinal, gastric, epidermal, mammary and neural lineages, and discuss the need for Wnt signaling in these cell types. Due to the vital actions of Wnt signaling in developmental and maintenance processes, deregulation of the pathway can culminate into a broad spectrum of developmental and genetic diseases, including cancer. The way in which Wnt signals can feed tumors and maintain cancer stem stells is discussed as well. Manipulation of Wnt signals both in vivo and in vitro thus carries potential for therapeutic approaches such as tissue engineering for regenerative medicine and anti-cancer treatment. Although many questions remain regarding the complete Wnt signal cell-type specific response and interplay of Wnt signaling with pathways such as BMP, Hedgehog and Notch, we hereby provide an overview of current knowledge on Wnt signaling and its control over human stem cell fate.

  15. Wnt signaling in heart valve development and osteogenic gene induction

    PubMed Central

    Alfieri, Christina M.; Cheek, Jonathan; Chakraborty, Santanu; Yutzey, Katherine E.

    2009-01-01

    Wnt signaling mediated by beta-catenin has been implicated in early endocardial cushion development, but its roles in later stages of heart valve maturation and homeostasis have not been identified. Multiple Wnt ligands and pathway genes are differentially expressed during heart valve development. At E12.5, Wnt2 is expressed in cushion mesenchyme, whereas Wnt4 and Wnt9b are predominant in overlying endothelial cells. At E17.5, both Wnt3a and Wnt7b are expressed in the remodeling atrioventricular (AV) and semilunar valves. In addition, the TOPGAL Wnt reporter transgene is active throughout the developing AV and semilunar valves at E16.5, with more localized expression in the stratified valve leaflets after birth. In chicken embryo aortic valves, genes characteristic of osteogenic cell lineages including periostin, osteonectin, and Id2 are expressed specifically in the collagen-rich fibrosa layer at E14. Treatment of E14 aortic valve interstitial cells (VIC) in culture with osteogenic media results in increased expression of multiple genes associated with bone formation. Treatment of VIC with Wnt3a leads to nuclear localization of beta-catenin and induction of periostin and matrix gla-protein, but does not induce genes associated with later stages of osteogenesis. Together, these studies provide evidence for Wnt signaling as a regulator of endocardial cushion maturation as well as valve leaflet stratification, homeostasis and pathogenesis. PMID:19961844

  16. Wnt-3a is critical for caudal embryonic development

    SciTech Connect

    Camper, S.A.; Greco, T.L.; Newhouse, M.M.

    1994-09-01

    Skeletal and neural tube defects represent an important class of birth defects. The majority of mouse mutants with neural tube defects also have malformations of the tail. Vestigial tail (vt) is an autosomal recessive mouse mutation characterized by reduction or absence of the tail, vertebral abnormalities, and reduced fertility. The phenotype has been described as the result of failure of cell migration through the primitive streak, causing abnormalities in the development of the neural tube and a reduction in the ventral ectodermal ridge. Wnt3a is an excellent candidate gene for vt because Wnt3a is expressed in the primitive streak and in the embryonic mesoderm, and it is thought to be involved in cell-to-cell communication and formation of the dorsal-ventral axis in the CNS. A lack of Wnt3a might be expected to result in overdorsalization of the neural tube and reduction of the ventral ectodermal ridge characteristic of vt/vt embryos. In a high resolution backcross segregating vt, we observed no recombination between vt and Wnt3a in 363 individuals analyzed. In vt/vt mice, Southern blot analysis revealed no abnormalities in the Wnt3a gene, and the Wnt3a cDNA sequence does not encode any amino acid changes. Whole mount in situ hybridization analysis demonstrated that Wnt3a expression is severely reduced in the developing tailbud of day 9.5 vt/vt embryos, suggestive of a lesion in the regulation on Wnt3a expression. An alleleism test, carried out by mating vt/vt males with Wnt3a +/Wnt3a- females, demonstrated that vt and Wnt3a are noncomplementing alleles. All of the compound heterozygotes exhibited severe tail defects, including occasional examples of hind limb parlaysis and spina bifida. The vertebral defects are intermediate between those of vt and Wnt3a homozygotes, suggesting that the concentration of Wnt3a correlates with the severity of the defect.

  17. Wnt-11 overexpression promoting the invasion of cervical cancer cells.

    PubMed

    Wei, Heng; Wang, Ning; Zhang, Yao; Wang, Shizhuo; Pang, Xiaoao; Zhang, Shulan

    2016-09-01

    Wnt-11 is a positive regulator of the Wnt signaling pathway, which plays a crucial role in carcinogenesis. However, Wnt-11 expression in cervical cancer has not been well investigated. The aim of this study was to investigate the role of Wnt-11 in cervical tumor proliferation and invasion. This study examined 24 normal cervical squamous epithelia, 29 cervical intraepithelial neoplasia (CIN), and 78 cervical cancer samples. The expression of Wnt-11 was investigated by immunohistochemistry and quantitative reverse transcription-polymerase chain reaction analysis. The expression of the high-risk human papilloma virus (HR-HPV) E6 oncoprotein was also investigated by immunohistochemistry. In addition, the expression of Wnt-11, HR-HPV E6, JNK-1, phosphorylated JNK-1(P-JNK1), and β-catenin was examined by western blot analysis following Wnt-11 knockdown or overexpression in HeLa or SiHa cells, respectively. The promotion of cervical cancer cell proliferation and invasion was investigated using the cell counting kit-8 and Matrigel invasion assay, respectively. Wnt-11 and HR-HPV E6 expression increased in a manner that corresponded with the progression of cervical cancer and was significantly correlated with the International Federation of Gynecology and Obstetrics cancer stage, lymph node metastasis, tumor size, and HPV infection. Wnt-11 protein expression was positively associated with HR-HPV E6 protein expression in all 78 cervical cancer samples (P < 0.001). Furthermore, Wnt-11 was positively associated with P-JNK1 expression and promoted cervical cancer cell proliferation and invasion. These observations suggest that the increased Wnt-11 expression observed in cervical cancer cells may lead to the phosphorylation and activation of JNK-1 and significantly promote tumor cell proliferation and cell migration/invasion through activation of the Wnt/JNK pathway. Consequently, Wnt-11 may serve as a novel target for cervical cancer therapy.

  18. FGFR Inhibitor Ameliorates Hypophosphatemia and Impaired Engrailed-1/Wnt Signaling in FGF2 High Molecular Weight Isoform Transgenic Mice.

    PubMed

    Du, Erxia; Xiao, Liping; Hurley, Marja M

    2016-09-01

    High molecular weight FGF2 transgenic (HMWTg) mouse phenocopies the Hyp mouse, homolog of human X-linked hypophosphatemic rickets with hypophosphatemis, and abnormal FGF23, FGFR, Klotho signaling in kidney. Since abnormal Wnt signaling was reported in Hyp mice we assessed whether Wnt signaling was impaired in HMWTg kidneys and the effect of blocking FGF receptor (FGFR) signaling. Bone mineral density and bone mineral content in female HMWTg mice were significantly reduced. HMWTg mice were gavaged with FGFR inhibitor NVP-BGJ398, or vehicle and were euthanized 24 h post treatment. Serum phosphate was significantly reduced and urine phosphate was significantly increased in HMWTg and was rescued by NVP-BGJ398. Analysis of kidneys revealed a significant reduction in Npt2a mRNA in HMWTg that was significantly increased by NVP-BGJ398. Increased FGFR1, KLOTHO, P-ERK1/2, and decreased NPT2a protein in HMWTg were rescued by NVP-BGJ398. Wnt inhibitor Engrailed-1 mRNA and protein was increased in HMWTg and was decreased by BGJ398. Akt mRNA and protein was decreased in HMWTg and was increased by NVP-BGJ398. The active form of glycogen synthase 3 beta (pGSK3-β) and phosphor-β-catenin were increased in HMWTg and were both decreased by NVP-BGJ398 while decreased active-β-catenin in HMWTg was increased by NVP-BGJ398. We conclude that FGFR blockade rescued hypophosphatemia by regulating FGF and WNT signaling in HMWTg kidneys. J. Cell. Biochem. 117: 1991-2000, 2016. © 2016 Wiley Periodicals, Inc.

  19. Itraconazole exerts its anti-melanoma effect by suppressing Hedgehog, Wnt, and PI3K/mTOR signaling pathways

    PubMed Central

    Liang, Guanzhao; Liu, Musang; Wang, Qiong; Shen, Yongnian; Mei, Huan; Li, Dongmei; Liu, Weida

    2017-01-01

    Malignant melanoma is the deadliest form of all skin cancers. Itraconazole, a commonly used systemic antifungal drug, has been tested for its anti-tumor effects on basal cell carcinoma, prostate cancer, and non-small cell lung cancer. Whether itraconazole has any specific anti-tumor effect on melanoma remains unknown. However, the goal of this study is to investigate the effect of itraconazole on melanoma and to reveal some details of its underlying mechanism. In the in vivo xenograft mouse model, we find that itraconazole can inhibit melanoma growth and extend the survival of melanoma xenograft mice, compared to non-itraconazole-treated mice. Also, itraconazole can significantly inhibit cell proliferation, as demonstrated by Ki-67 staining in itraconazole-treated tumor tissues. In in vitro, we show that itraconazole inhibits the proliferation and colony formation of both SK-MEL-28 and A375 human melanoma cells. Moreover, we demonstrate that itraconazole significantly down-regulates Gli-1, Gli-2, Wnt3A, β-catenin and cyclin D1, while it up-regulates Gli-3 and Axin-1, indicating potent inhibitory effects of itraconazole on Hedgehog (Hh) and Wnt signaling pathways. Furthermore, itraconazole significantly suppresses the PI3K/mTOR signaling pathway – indicated by the down-regulated phosphorylation of p70S6K, 4E-BP1 and AKT – but has no effect on the phosphorylation of MEK or ERK. Our data suggest that itraconazole inhibits melanoma growth through an interacting regulatory network that includes Hh, Wnt, and PI3K/mTOR signaling pathways. These results suggest that this agent has several potent anti-melanoma features and may be useful in the synergesis of other anti-cancer drugs via blockage of the Hh, Wnt and PI3K/mTOR signaling pathways. PMID:28212537

  20. Genetic Screening of WNT4 and WNT5B in Two Populations with Deviating Bone Mineral Densities.

    PubMed

    Hendrickx, Gretl; Boudin, Eveline; Steenackers, Ellen; Nielsen, Torben Leo; Andersen, Marianne; Brixen, Kim; Van Hul, Wim

    2017-03-01

    A role for WNT4 and WNT5B in bone metabolism was indicated by genome-wide association studies (GWAS) and a Wnt4 knockout mouse model. The aim of this study was therefore to replicate and further investigate the causality between genetic variation in WNT4 and WNT5B and deviating bone mineral density (BMD) values. A WNT4 and WNT5B mutation screening was performed in patients with craniotubular hyperostosis using Sanger sequencing. Here, no putative causal mutations were detected. Moreover, a high and low BMD cohort was selected from the Odense Androgen Study population for re-sequencing. In WNT4 we detected four variants (three rare, one common), while in WNT5B we detected five variants (two rare, three common). For the common variants, no significant difference in genotype frequencies between the high and low BMD cohorts was observed. The SNPs associated with the GWAS were genotyped in these cohorts, but again no significant difference in genotype frequencies was observed. Despite the findings of the GWAS, we were not able to replicate or further verify the genetic association of polymorphisms in WNT4 and WNT5B with BMD. In order to do so, the intronic regions of both genes could be investigated more thoroughly in more extended populations (or extremes) with greater power. Future genetic and functional studies toward adjacent genes of WNT4 and WNT5B can also be interesting to figure out whether the signal from GWAS could possibly be attributed to genetic variation in these genes.

  1. Expression of WNT genes in cervical cancer-derived cells: Implication of WNT7A in cell proliferation and migration

    SciTech Connect

    Ramos-Solano, Moisés; Meza-Canales, Ivan D.; Torres-Reyes, Luis A.; Alvarez-Zavala, Monserrat; and others

    2015-07-01

    According to the multifactorial model of cervical cancer (CC) causation, it is now recognized that other modifications, in addition to Human papillomavirus (HPV) infection, are necessary for the development of this neoplasia. Among these, it has been proposed that a dysregulation of the WNT pathway might favor malignant progression of HPV-immortalized keratinocytes. The aim of this study was to identify components of the WNT pathway differentially expressed in CC vs. non-tumorigenic, but immortalized human keratinocytes. Interestingly, WNT7A expression was found strongly downregulated in cell lines and biopsies derived from CC. Restoration of WNT7A in CC-derived cell lines using a lentiviral gene delivery system or after adding a recombinant human protein decreases cell proliferation. Likewise, WNT7A silencing in non-tumorigenic cells markedly accelerates proliferation. Decreased WNT7A expression was due to hypermethylation at particular CpG sites. To our knowledge, this is the first study reporting reduced WNT7A levels in CC-derived cells and that ectopic WNT7A restoration negatively affects cell proliferation and migration. - Highlights: • WNT7A is expressed in normal keratinocytes or cervical cells without lesion. • WNT7A is significantly reduced in cervical cancer-derived cells. • Restoration of WNT7A expression in HeLa decreases proliferation and cell migration. • Silencing of WNT7A in HaCaT induces an increased proliferation and migration rate. • Decreased WNT7A expression in this model is due to hypermethylation.

  2. Estrogen Receptor α Mediates Proliferation of Osteoblastic Cells Stimulated by Estrogen and Mechanical Strain, but Their Acute Down-regulation of the Wnt Antagonist Sost Is Mediated by Estrogen Receptor β*

    PubMed Central

    Galea, Gabriel L.; Meakin, Lee B.; Sugiyama, Toshihiro; Zebda, Noureddine; Sunters, Andrew; Taipaleenmaki, Hanna; Stein, Gary S.; van Wijnen, Andre J.; Lanyon, Lance E.; Price, Joanna S.

    2013-01-01

    Mechanical strain and estrogens both stimulate osteoblast proliferation through estrogen receptor (ER)-mediated effects, and both down-regulate the Wnt antagonist Sost/sclerostin. Here, we investigate the differential effects of ERα and -β in these processes in mouse long bone-derived osteoblastic cells and human Saos-2 cells. Recruitment to the cell cycle following strain or 17β-estradiol occurs within 30 min, as determined by Ki-67 staining, and is prevented by the ERα antagonist 1,3-bis(4-hydroxyphenyl)-4-methyl-5-[4-(2-piperidinylethoxy)phenol]-1H-pyrazole dihydrochloride. ERβ inhibition with 4-[2-phenyl-5,7-bis(trifluoromethyl)pyrazolo[1,5-β]pyrimidin-3-yl] phenol (PTHPP) increases basal proliferation similarly to strain or estradiol. Both strain and estradiol down-regulate Sost expression, as does in vitro inhibition or in vivo deletion of ERα. The ERβ agonists 2,3-bis(4-hydroxyphenyl)-propionitrile and ERB041 also down-regulated Sost expression in vitro, whereas the ERα agonist 4,4′,4″-[4-propyl-(1H)-pyrazol-1,3,5-triyl]tris-phenol or the ERβ antagonist PTHPP has no effect. Tamoxifen, a nongenomic ERβ agonist, down-regulates Sost expression in vitro and in bones in vivo. Inhibition of both ERs with fulvestrant or selective antagonism of ERβ, but not ERα, prevents Sost down-regulation by strain or estradiol. Sost down-regulation by strain or ERβ activation is prevented by MEK/ERK blockade. Exogenous sclerostin has no effect on estradiol-induced proliferation but prevents that following strain. Thus, in osteoblastic cells the acute proliferative effects of both estradiol and strain are ERα-mediated. Basal Sost down-regulation follows decreased activity of ERα and increased activity of ERβ. Sost down-regulation by strain or increased estrogens is mediated by ERβ, not ERα. ER-targeting therapy may facilitate structurally appropriate bone formation by enhancing the distinct ligand-independent, strain-related contributions to proliferation

  3. Divergent Wnt8a gene expression in teleosts.

    PubMed

    Mwafi, Nesrin; Beretta, Carlo A; Paolini, Alessio; Carl, Matthias

    2014-01-01

    The analysis of genes in evolutionarily distant but morphologically similar species is of major importance to unravel the changes in genomes over millions of years, which led to gene silencing and functional diversification. We report the analysis of Wnt8a gene expression in the medakafish and provide a detailed comparison to other vertebrates. In all teleosts analyzed there are two paralogous Wnt8a copies. These show largely overlapping expression in the early developing zebrafish embryo, an evolutionarily distant relative of medaka. In contrast to zebrafish, we find that both maternal and zygotic expression of particularly one Wnt8a paralog has diverged in medaka. While Wnt8a1 expression is mostly conserved at early embryonic stages, the expression of Wnt8a2 differs markedly. In addition, both genes are distinctly expressed during organogenesis unlike the zebrafish homologs, which may hint at the emergence of functional diversification of Wnt8a ligands during evolution.

  4. Probing the canonicity of the Wnt/Wingless signaling pathway

    PubMed Central

    Brunner, Erich; Stark, Alexander; Basler, Konrad

    2017-01-01

    The hallmark of canonical Wnt signaling is the transcriptional induction of Wnt target genes by the beta-catenin/TCF complex. Several studies have proposed alternative interaction partners for beta-catenin or TCF, but the relevance of potential bifurcations in the distal Wnt pathway remains unclear. Here we study on a genome-wide scale the requirement for Armadillo (Arm, Drosophila beta-catenin) and Pangolin (Pan, Drosophila TCF) in the Wnt/Wingless(Wg)-induced transcriptional response of Drosophila Kc cells. Using somatic genetics, we demonstrate that both Arm and Pan are absolutely required for mediating activation and repression of target genes. Furthermore, by means of STARR-sequencing we identified Wnt/Wg-responsive enhancer elements and found that all responsive enhancers depend on Pan. Together, our results confirm the dogma of canonical Wnt/Wg signaling and argue against the existence of distal pathway branches in this system. PMID:28369070

  5. Probing the canonicity of the Wnt/Wingless signaling pathway.

    PubMed

    Franz, Alexandra; Shlyueva, Daria; Brunner, Erich; Stark, Alexander; Basler, Konrad

    2017-04-01

    The hallmark of canonical Wnt signaling is the transcriptional induction of Wnt target genes by the beta-catenin/TCF complex. Several studies have proposed alternative interaction partners for beta-catenin or TCF, but the relevance of potential bifurcations in the distal Wnt pathway remains unclear. Here we study on a genome-wide scale the requirement for Armadillo (Arm, Drosophila beta-catenin) and Pangolin (Pan, Drosophila TCF) in the Wnt/Wingless(Wg)-induced transcriptional response of Drosophila Kc cells. Using somatic genetics, we demonstrate that both Arm and Pan are absolutely required for mediating activation and repression of target genes. Furthermore, by means of STARR-sequencing we identified Wnt/Wg-responsive enhancer elements and found that all responsive enhancers depend on Pan. Together, our results confirm the dogma of canonical Wnt/Wg signaling and argue against the existence of distal pathway branches in this system.

  6. Tankyrase is necessary for canonical Wnt signaling during kidney development

    PubMed Central

    Karner, Courtney M.; Merkel, Calli E; Dodge, Michael; Ma, Zhiqiang; Lu, Jianming; Chen, Chuo; Lum, Lawrence; Carroll, Thomas J.

    2010-01-01

    Recent studies utilizing small molecule antagonists have revealed that the poly(ADP-ribose) polymerases (PARPs) Tankyrase 1 and 2 are critical regulators of canonical Wnt signaling in some cellular contexts. However, the absence of any activity during zebrafish embryogenesis suggested that the tankyrases may not be general/core components of the Wnt pathway. Here we show that Tnks1 and 2 are broadly expressed during mouse development and are essential during kidney and lung development. In the kidney, blockage of tankyrase activity phenocopies the effect of blocking production of all Wnt ligands. Tankyrase inhibition can be rescued by activation of β-catenin demonstrating its specificity for the Wnt pathway. In addition, treatment with tankyrase inhibitors appears to be completely reversible in some cell types. These studies suggest that the tankyrases are core components of the canonical Wnt pathway and their inhibitors should enjoy broad usage as antagonists of Wnt signaling. PMID:20549720

  7. Secretion and extracellular space travel of Wnt proteins.

    PubMed

    Gross, Julia Christina; Boutros, Michael

    2013-08-01

    Wnt signaling pathways control many processes during development, stem cell maintenance and homeostasis, and their aberrant regulation has been linked to diseases in man including diabetes, neurodegeneration and cancer. Wnts are hydrophobic proteins, however, quite paradoxically, they can travel over distances to induce cell-type specific responses. While there has been an initial focus on elucidating the intracellular signaling cascade, discoveries in the past few years have shed light on a highly complex, and regulated secretory process that guides Wnt proteins through the exocytic pathway. Wnt proteins are at least in portion packaged onto extracellular carriers such as exosomes. Similar to dysregulation of components in the Wnt receiving cell, failure to regulate Wnt secretion has been linked to cancer. Here, we review recent discoveries on factors and processes implicated in Wnt secretion.

  8. Wnt secretion is required to maintain high levels of Wnt activity in colon cancer cells

    PubMed Central

    Voloshanenko, Oksana; Erdmann, Gerrit; Dubash, Taronish D.; Augustin, Iris; Metzig, Marie; Moffa, Giusi; Hundsrucker, Christian; Kerr, Grainne; Sandmann, Thomas; Anchang, Benedikt; Demir, Kubilay; Boehm, Christina; Leible, Svenja; Ball, Claudia R.; Glimm, Hanno; Spang, Rainer; Boutros, Michael

    2013-01-01

    Aberrant regulation of the Wnt/β-catenin pathway has an important role during the onset and progression of colorectal cancer, with over 90% of cases of sporadic colon cancer featuring mutations in APC or β-catenin. However, it has remained a point of controversy whether these mutations are sufficient to activate the pathway or require additional upstream signals. Here we show that colorectal tumours express elevated levels of Wnt3 and Evi/Wls/GPR177. We found that in colon cancer cells, even in the presence of mutations in APC or β-catenin, downstream signalling remains responsive to Wnt ligands and receptor proximal signalling. Furthermore, we demonstrate that truncated APC proteins bind β-catenin and key components of the destruction complex. These results indicate that cells with mutations in APC or β-catenin depend on Wnt ligands and their secretion for a sufficient level of β-catenin signalling, which potentially opens new avenues for therapeutic interventions by targeting Wnt secretion via Evi/Wls. PMID:24162018

  9. Wnt secretion is required to maintain high levels of Wnt activity in colon cancer cells.

    PubMed

    Voloshanenko, Oksana; Erdmann, Gerrit; Dubash, Taronish D; Augustin, Iris; Metzig, Marie; Moffa, Giusi; Hundsrucker, Christian; Kerr, Grainne; Sandmann, Thomas; Anchang, Benedikt; Demir, Kubilay; Boehm, Christina; Leible, Svenja; Ball, Claudia R; Glimm, Hanno; Spang, Rainer; Boutros, Michael

    2013-01-01

    Aberrant regulation of the Wnt/β-catenin pathway has an important role during the onset and progression of colorectal cancer, with over 90% of cases of sporadic colon cancer featuring mutations in APC or β-catenin. However, it has remained a point of controversy whether these mutations are sufficient to activate the pathway or require additional upstream signals. Here we show that colorectal tumours express elevated levels of Wnt3 and Evi/Wls/GPR177. We found that in colon cancer cells, even in the presence of mutations in APC or β-catenin, downstream signalling remains responsive to Wnt ligands and receptor proximal signalling. Furthermore, we demonstrate that truncated APC proteins bind β-catenin and key components of the destruction complex. These results indicate that cells with mutations in APC or β-catenin depend on Wnt ligands and their secretion for a sufficient level of β-catenin signalling, which potentially opens new avenues for therapeutic interventions by targeting Wnt secretion via Evi/Wls.

  10. Omega-3 polyunsaturated fatty acids selectively inhibit growth in neoplastic oral keratinocytes by differentially activating ERK1/2

    PubMed Central

    Parkinson, Eric Kenneth

    2013-01-01

    The long-chain omega-3 polyunsaturated fatty acids (n-3 PUFAs)—eicosapentaenoic acid (EPA) and its metabolite docosahexaenoic acid (DHA)—inhibit cancer formation in vivo, but their mechanism of action is unclear. Extracellular signal-regulated kinase 1/2 (ERK1/2) activation and inhibition have both been associated with the induction of tumour cell apoptosis by n-3 PUFAs. We show here that low doses of EPA, in particular, inhibited the growth of premalignant and malignant keratinocytes more than the growth of normal counterparts by a combination of cell cycle arrest and apoptosis. The growth inhibition of the oral squamous cell carcinoma (SCC) lines, but not normal keratinocytes, by both n-3 PUFAs was associated with epidermal growth factor receptor (EGFR) autophosphorylation, a sustained phosphorylation of ERK1/2 and its downstream target p90RSK but not with phosphorylation of the PI3 kinase target Akt. Inhibition of EGFR with either the EGFR kinase inhibitor AG1478 or an EGFR-blocking antibody inhibited ERK1/2 phosphorylation, and the blocking antibody partially antagonized growth inhibition by EPA but not by DHA. DHA generated more reactive oxygen species and activated more c-jun N-terminal kinase than EPA, potentially explaining its increased toxicity to normal keratinocytes. Our results show that, in part, EPA specifically inhibits SCC growth and development by creating a sustained signalling imbalance to amplify the EGFR/ERK/p90RSK pathway in neoplastic keratinocytes to a supraoptimal level, supporting the chemopreventive potential of EPA, whose toxicity to normal cells might be reduced further by blocking its metabolism to DHA. Furthermore, ERK1/2 phosphorylation may have potential as a biomarker of n-3 PUFA function in vivo. PMID:23892603

  11. Canonical Wnt signaling is necessary for object recognition memory consolidation.

    PubMed

    Fortress, Ashley M; Schram, Sarah L; Tuscher, Jennifer J; Frick, Karyn M

    2013-07-31

    Wnt signaling has emerged as a potent regulator of hippocampal synaptic function, although no evidence yet supports a critical role for Wnt signaling in hippocampal memory. Here, we sought to determine whether canonical β-catenin-dependent Wnt signaling is necessary for hippocampal memory consolidation. Immediately after training in a hippocampal-dependent object recognition task, mice received a dorsal hippocampal (DH) infusion of vehicle or the canonical Wnt antagonist Dickkopf-1 (Dkk-1; 50, 100, or 200 ng/hemisphere). Twenty-four hours later, mice receiving vehicle remembered the familiar object explored during training. However, mice receiving Dkk-1 exhibited no memory for the training object, indicating that object recognition memory consolidation is dependent on canonical Wnt signaling. To determine how Dkk-1 affects canonical Wnt signaling, mice were infused with vehicle or 50 ng/hemisphere Dkk-1 and protein levels of Wnt-related proteins (Dkk-1, GSK3β, β-catenin, TCF1, LEF1, Cyclin D1, c-myc, Wnt7a, Wnt1, and PSD95) were measured in the dorsal hippocampus 5 min or 4 h later. Dkk-1 produced a rapid increase in Dkk-1 protein levels and a decrease in phosphorylated GSK3β levels, followed by a decrease in β-catenin, TCF1, LEF1, Cyclin D1, c-myc, Wnt7a, and PSD95 protein levels 4 h later. These data suggest that alterations in Wnt/GSK3β/β-catenin signaling may underlie the memory impairments induced by Dkk-1. In a subsequent experiment, object training alone rapidly increased DH GSK3β phosphorylation and levels of β-catenin and Cyclin D1. These data suggest that canonical Wnt signaling is regulated by object learning and is necessary for hippocampal memory consolidation.

  12. Wnt signaling regulates homeostasis of the periodontal ligament

    PubMed Central

    Lim, W.H.; Liu, B.; Cheng, D.; Williams, B.O.; Mah, S.J.; Helms, J.A.

    2014-01-01

    Background and Objective In health, the periodontal ligament maintains a constant width throughout an organism’s lifetime. The molecular signals responsible for maintaining homeostatic control over the periodontal ligament are unknown. The purpose of this study was to investigate the role of Wnt signaling in this process by removing an essential chaperone protein, Wntless (Wls) from odontoblasts and cementoblasts, and observing the effects of Wnt depletion on cells of the periodontal complex. Material and Methods The Wnt responsive status of the periodontal complex was assessed using two strains of Wnt reporter mice, Axin2LacZ/+ mice and Lgr5LacZ/+. The function of this endogenous Wnt signal was evaluated by conditionally eliminating the Wntless (Wls) gene using an Osteocalcin Cre driver. The resulting OCN-Cre;Wlsfl/fl mice were examined using micro-CT and histology, immunohistochemical analyses for Osteopontin, Runx2 and Fibromodulin, in situ hybridization for Osterix, and alkaline phosphatase activity. Results The adult periodontal ligament is Wnt responsive. Elimination of Wnt signaling in the periodontal complex of OCN-Cre;Wlsfl/fl mice results in a wider periodontal ligament space. This pathologically increased periodontal width is due to a reduction in the expression of osteogenic genes and proteins, which results in thinner alveolar bone. A concomitant increase in fibrous tissue occupying the periodontal space was observed along with a disruption in the orientation of the periodontal ligament. Conclusion The periodontal ligament is a Wnt dependent tissue. Cells in the periodontal complex are Wnt responsive and eliminating an essential component of the Wnt signaling network leads to a pathological widening of the periodontal ligament space. Osteogenic stimuli are reduced and a disorganized fibrillary matrix results from depletion of Wnt signaling. Collectively, these data underscore the importance of Wnt signaling in homeostasis of the periodontal ligament

  13. Targeting the WNT Signaling Pathway in Cancer Therapeutics.

    PubMed

    Tai, David; Wells, Keith; Arcaroli, John; Vanderbilt, Chad; Aisner, Dara L; Messersmith, Wells A; Lieu, Christopher H

    2015-10-01

    The WNT signaling cascade is integral in numerous biological processes including embryonic development, cell cycle regulation, inflammation, and cancer. Hyperactivation of WNT signaling secondary to alterations to varying nodes of the pathway have been identified in multiple tumor types. These alterations converge into increased tumorigenicity, sustained proliferation, and enhanced metastatic potential. This review seeks to evaluate the evidence supporting the WNT pathway in cancer, the therapeutic strategies in modulating this pathway, and potential challenges in drug development.

  14. Epilepsy and the Wnt Signaling Pathway

    DTIC Science & Technology

    2015-06-01

    molecular mechanisms and uncovered new possible insights for therapeutics with a drug combination that we had been developing for breast cancer treatment ...interventions into the epileptogenic period. Practically, a patient would seek treatment after the first seizure to prevent a second seizure, thus... treatment at day 0 and then the early epileptogenic period from day 1 to 7 was analyzed. In Figure 2, we found that Wnt signaling occurred in the

  15. Epilepsy and the Wnt Signaling Pathway

    DTIC Science & Technology

    2016-09-01

    therapeutic drug combination that we had been developing for breast cancer treatment aimed at Wnt signaling. Surprisingly, the combination attenuates...treating some forms of drug -intractable epilepsy already suggest a role for glucose metabolism in the etiology of certain epilepsies. The basis of the...Furthermore, the HBP1-/- mouse has exciting potential for future anti-epileptogenic drug development, based on the human genetics. In principle

  16. [Cytokines in bone diseases. Wnt signal and excessive bone formation].

    PubMed

    Hosoi, Takayuki

    2010-10-01

    Wnt signal has been known to play various roles in many organ from the beginning of embryogensis. Its role in bone metabolism has also been investigated and established. Lipoprotein receptor-related protein 5 (LRP5) is one of the important molecules in wnt signal pathway whose point mutations are related to both bone loss and excessive bone formation. Wnt signal is involved in the action of sclerostin which was found as a gene for osteosclerosis, one of the diseases of excessive bone formation. Wnt signal is keeping the position as an important research target for normal and pathological bone formation.

  17. Wnt proteins can direct planar cell polarity in vertebrate ectoderm

    PubMed Central

    Chu, Chih-Wen; Sokol, Sergei Y

    2016-01-01

    The coordinated orientation of cells across the tissue plane, known as planar cell polarity (PCP), is manifested by the segregation of core PCP proteins to different sides of the cell. Secreted Wnt ligands are involved in many PCP-dependent processes, yet whether they act as polarity cues has been controversial. We show that in Xenopus early ectoderm, the Prickle3/Vangl2 complex was polarized to anterior cell edges and this polarity was disrupted by several Wnt antagonists. In midgastrula embryos, Wnt5a, Wnt11, and Wnt11b, but not Wnt3a, acted across many cell diameters to orient Prickle3/Vangl2 complexes away from their sources regardless of their positions relative to the body axis. The planar polarity of endogenous Vangl2 in the neuroectoderm was similarly redirected by an ectopic Wnt source and disrupted after depletion of Wnt11b in the presumptive posterior region of the embryo. These observations provide evidence for the instructive role of Wnt ligands in vertebrate PCP. DOI: http://dx.doi.org/10.7554/eLife.16463.001 PMID:27658614

  18. Wnt/β-catenin signaling enables developmental transitions during valvulogenesis.

    PubMed

    Bosada, Fernanda M; Devasthali, Vidusha; Jones, Kimberly A; Stankunas, Kryn

    2016-03-15

    Heart valve development proceeds through coordinated steps by which endocardial cushions (ECs) form thin, elongated and stratified valves. Wnt signaling and its canonical effector β-catenin are proposed to contribute to endocardial-to-mesenchymal transformation (EMT) through postnatal steps of valvulogenesis. However, genetic redundancy and lethality have made it challenging to define specific roles of the canonical Wnt pathway at different stages of valve formation. We developed a transgenic mouse system that provides spatiotemporal inhibition of Wnt/β-catenin signaling by chemically inducible overexpression of Dkk1. Unexpectedly, this approach indicates canonical Wnt signaling is required for EMT in the proximal outflow tract (pOFT) but not atrioventricular canal (AVC) cushions. Furthermore, Wnt indirectly promotes pOFT EMT through its earlier activity in neighboring myocardial cells or their progenitors. Subsequently, Wnt/β-catenin signaling is activated in cushion mesenchymal cells where it supports FGF-driven expansion of ECs and then AVC valve extracellular matrix patterning. Mice lacking Axin2, a negative Wnt regulator, have larger valves, suggesting that accumulating Axin2 in maturing valves represents negative feedback that restrains tissue overgrowth rather than simply reporting Wnt activity. Disruption of these Wnt/β-catenin signaling roles that enable developmental transitions during valvulogenesis could account for common congenital valve defects.

  19. Wnt/β-catenin signaling enables developmental transitions during valvulogenesis

    PubMed Central

    Bosada, Fernanda M.; Devasthali, Vidusha; Jones, Kimberly A.; Stankunas, Kryn

    2016-01-01

    Heart valve development proceeds through coordinated steps by which endocardial cushions (ECs) form thin, elongated and stratified valves. Wnt signaling and its canonical effector β-catenin are proposed to contribute to endocardial-to-mesenchymal transformation (EMT) through postnatal steps of valvulogenesis. However, genetic redundancy and lethality have made it challenging to define specific roles of the canonical Wnt pathway at different stages of valve formation. We developed a transgenic mouse system that provides spatiotemporal inhibition of Wnt/β-catenin signaling by chemically inducible overexpression of Dkk1. Unexpectedly, this approach indicates canonical Wnt signaling is required for EMT in the proximal outflow tract (pOFT) but not atrioventricular canal (AVC) cushions. Furthermore, Wnt indirectly promotes pOFT EMT through its earlier activity in neighboring myocardial cells or their progenitors. Subsequently, Wnt/β-catenin signaling is activated in cushion mesenchymal cells where it supports FGF-driven expansion of ECs and then AVC valve extracellular matrix patterning. Mice lacking Axin2, a negative Wnt regulator, have larger valves, suggesting that accumulating Axin2 in maturing valves represents negative feedback that restrains tissue overgrowth rather than simply reporting Wnt activity. Disruption of these Wnt/β-catenin signaling roles that enable developmental transitions during valvulogenesis could account for common congenital valve defects. PMID:26893350

  20. Wnt-regulated dynamics of positional information in zebrafish somitogenesis

    PubMed Central

    Bajard, Lola; Morelli, Luis G.; Ares, Saúl; Pécréaux, Jacques; Jülicher, Frank; Oates, Andrew C.

    2014-01-01

    How signaling gradients supply positional information in a field of moving cells is an unsolved question in patterning and morphogenesis. Here, we ask how a Wnt signaling gradient regulates the dynamics of a wavefront of cellular change in a flow of cells during somitogenesis. Using time-controlled perturbations of Wnt signaling in the zebrafish embryo, we changed segment length without altering the rate of somite formation or embryonic elongation. This result implies specific Wnt regulation of the wavefront velocity. The observed Wnt signaling gradient dynamics and timing of downstream events support a model for wavefront regulation in which cell flow plays a dominant role in transporting positional information. PMID:24595291

  1. Wnt to Build a Tube: Contributions of Wnt signaling to epithelial tubulogenesis

    PubMed Central

    Miller, Rachel K.; McCrea, Pierre D.

    2012-01-01

    Epithelial tubes are crucial to the function of organ systems including the cardiovascular system, pulmonary system, gastrointestinal tract, reproductive organ systems, excretory system, and auditory system. Using a variety of animal model systems, recent studies have substantiated the role of Wnt signaling via the canonical/ β-catenin mediated trajectory, the non-canonical Wnt trajectories, or both, in forming epithelial tubular tissues. This review focuses on the involvement of the Wnt pathways in the induction, specification, proliferation and morphogenesis involved in tubulogenesis within tissues including the lungs, kidneys, ears, mammary glands, gut and heart. The ultimate goal is to describe the developmental processes forming the various tubulogenic organ systems to determine the relationships between these processes. PMID:19681164

  2. Wnt Signaling in Renal Cell Carcinoma

    PubMed Central

    Xu, Qi; Krause, Mirja; Samoylenko, Anatoly; Vainio, Seppo

    2016-01-01

    Renal cell carcinoma (RCC) accounts for 90% of all kidney cancers. Due to poor diagnosis, high resistance to the systemic therapies and the fact that most RCC cases occur sporadically, current research switched its focus on studying the molecular mechanisms underlying RCC. The aim is the discovery of new effective and less toxic anti-cancer drugs and novel diagnostic markers. Besides the PI3K/Akt/mTOR, HGF/Met and VHL/hypoxia cellular signaling pathways, the involvement of the Wnt/β-catenin pathway in RCC is commonly studied. Wnt signaling and its targeted genes are known to actively participate in different biological processes during embryonic development and renal cancer. Recently, studies have shown that targeting this pathway by alternating/inhibiting its intracellular signal transduction can reduce cancer cells viability and inhibit their growth. The targets and drugs identified show promising potential to serve as novel RCC therapeutics and prognostic markers. This review aims to summarize the current status quo regarding recent research on RCC focusing on the involvement of the Wnt/β-catenin pathway and how its understanding could facilitate the identification of potential therapeutic targets, new drugs and diagnostic biomarkers. PMID:27322325

  3. Anti-Melanogenic Activities of Heracleum moellendorffii via ERK1/2-Mediated MITF Downregulation

    PubMed Central

    Alam, Md Badrul; Seo, Bum-Ju; Zhao, Peijun; Lee, Sang-Han

    2016-01-01

    In this study, the anti-melanogenic effects of Heracleum moellendorffii Hance extract (HmHe) and the mechanisms through which it inhibits melanogenesis in melan-a cells were investigated. Mushroom tyrosinase (TYR) activity and melanin content as well as cellular tyrosinase activity were measured in the cells. mRNA and protein expression of microphthalmia-associated transcription factor (MITF), tyrosinase (TYR), TYR-related protein-1 (TYRP-1) and -2 were also examined. The results demonstrate that treatment with HmHe significantly inhibits mushroom tyrosinase activity. Furthermore, HmHe also markedly inhibits melanin production and intracellular tyrosinase activity. By suppressing the expression of TYR, TYRP-1, TYRP-2, and MITF, HmHe treatment antagonized melanin production in melan-a cells. Additionally, HmHe interfered with the phosphorylation of extracellular signal-regulated kinase (ERK) 1/2, with reversal of HmHe-induced melanogenesis inhibition after treatment with specific inhibitor U0126. In summary, HmHe can be said to stimulate ERK1/2 phosphorylation and subsequent degradation of MITF, resulting in suppression of melanogenic enzymes and melanin production, possibly due to the presence of polyphenolic compounds. PMID:27827938

  4. Agonism, Antagonism, and Inverse Agonism Bias at the Ghrelin Receptor Signaling*

    PubMed Central

    M'Kadmi, Céline; Leyris, Jean-Philippe; Onfroy, Lauriane; Galés, Céline; Saulière, Aude; Gagne, Didier; Damian, Marjorie; Mary, Sophie; Maingot, Mathieu; Denoyelle, Séverine; Verdié, Pascal; Fehrentz, Jean-Alain; Martinez, Jean; Banères, Jean-Louis; Marie, Jacky

    2015-01-01

    The G protein-coupled receptor GHS-R1a mediates ghrelin-induced growth hormone secretion, food intake, and reward-seeking behaviors. GHS-R1a signals through Gq, Gi/o, G13, and arrestin. Biasing GHS-R1a signaling with specific ligands may lead to the development of more selective drugs to treat obesity or addiction with minimal side effects. To delineate ligand selectivity at GHS-R1a signaling, we analyzed in detail the efficacy of a panel of synthetic ligands activating the different pathways associated with GHS-R1a in HEK293T cells. Besides β-arrestin2 recruitment and ERK1/2 phosphorylation, we monitored activation of a large panel of G protein subtypes using a bioluminescence resonance energy transfer-based assay with G protein-activation biosensors. We first found that unlike full agonists, Gq partial agonists were unable to trigger β-arrestin2 recruitment and ERK1/2 phosphorylation. Using G protein-activation biosensors, we then demonstrated that ghrelin promoted activation of Gq, Gi1, Gi2, Gi3, Goa, Gob, and G13 but not Gs and G12. Besides, we identified some GHS-R1a ligands that preferentially activated Gq and antagonized ghrelin-mediated Gi/Go activation. Finally, we unambiguously demonstrated that in addition to Gq, GHS-R1a also promoted constitutive activation of G13. Importantly, we identified some ligands that were selective inverse agonists toward Gq but not of G13. This demonstrates that bias at GHS-R1a signaling can occur not only with regard to agonism but also to inverse agonism. Our data, combined with other in vivo studies, may facilitate the design of drugs selectively targeting individual signaling pathways to treat only the therapeutically relevant function. PMID:26363071

  5. Systematic Mapping of WNT-FZD Protein Interactions Reveals Functional Selectivity by Distinct WNT-FZD Pairs*

    PubMed Central

    Dijksterhuis, Jacomijn P.; Baljinnyam, Bolormaa; Stanger, Karen; Sercan, Hakki O.; Ji, Yun; Andres, Osler; Rubin, Jeffrey S.; Hannoush, Rami N.; Schulte, Gunnar

    2015-01-01

    The seven-transmembrane-spanning receptors of the FZD1–10 class are bound and activated by the WNT family of lipoglycoproteins, thereby inducing a complex network of signaling pathways. However, the specificity of the interaction between mammalian WNT and FZD proteins and the subsequent signaling cascade downstream of the different WNT-FZD pairs have not been systematically addressed to date. In this study, we determined the binding affinities of various WNTs for different members of the FZD family by using bio-layer interferometry and characterized their functional selectivity in a cell system. Using purified WNTs, we show that different FZD cysteine-rich domains prefer to bind to distinct WNTs with fast on-rates and slow off-rates. In a 32D cell-based system engineered to overexpress FZD2, FZD4, or FZD5, we found that WNT-3A (but not WNT-4, -5A, or -9B) activated the WNT-β-catenin pathway through FZD2/4/5 as measured by phosphorylation of LRP6 and β-catenin stabilization. Surprisingly, different WNT-FZD pairs showed differential effects on phosphorylation of DVL2 and DVL3, revealing a previously unappreciated DVL isoform selectivity by different WNT-FZD pairs in 32D cells. In summary, we present extensive mapping of WNT-FZD cysteine-rich domain interactions complemented by analysis of WNT-FZD pair functionality in a unique cell system expressing individual FZD isoforms. Differential WNT-FZD binding and selective functional readouts suggest that endogenous WNT ligands evolved with an intrinsic natural bias toward different downstream signaling pathways, a phenomenon that could be of great importance in the design of FZD-targeting drugs. PMID:25605717

  6. Smooth muscle archvillin is an ERK scaffolding protein.

    PubMed

    Gangopadhyay, Samudra S; Kengni, Edouard; Appel, Sarah; Gallant, Cynthia; Kim, Hak Rim; Leavis, Paul; DeGnore, Jon; Morgan, Kathleen G

    2009-06-26

    ERK influences a number of pathways in all cells, but how ERK activities are segregated between different pathways has not been entirely clear. Using immunoprecipitation and pulldown experiments with domain-specific recombinant fragments, we show that smooth muscle archvillin (SmAV) binds ERK and members of the ERK signaling cascade in a domain-specific, stimulus-dependent, and pathway-specific manner. MEK binds specifically to the first 445 residues of SmAV. B-Raf, an upstream regulator of MEK, constitutively interacts with residues 1-445 and 446-1250. Both ERK and 14-3-3 bind to both fragments, but in a stimulus-specific manner. Phosphorylated ERK is associated only with residues 1-445. An ERK phosphorylation site was determined by mass spectrometry to reside at Ser132. A phospho-antibody raised to this site shows that the site is phosphorylated during alpha-agonist-mediated ERK activation in smooth muscle tissue. Phosphorylation of SmAV by ERK decreases the association of phospho-ERK with SmAV. These results, combined with previous observations, indicate that SmAV serves as a new ERK scaffolding protein and provide a mechanism for regulation of ERK binding, activation, and release from the signaling complex.

  7. A Strategy for Antagonizing Quorum Sensing

    SciTech Connect

    G Chen; L Swem; D Swem; D Stauff; C OLoughlin; P Jeffrey; B Bassler; F Hughson

    2011-12-31

    Quorum-sensing bacteria communicate via small molecules called autoinducers to coordinate collective behaviors. Because quorum sensing controls virulence factor expression in many clinically relevant pathogens, membrane-permeable quorum sensing antagonists that prevent population-wide expression of virulence genes offer a potential route to novel antibacterial therapeutics. Here, we report a strategy for inhibiting quorum-sensing receptors of the widespread LuxR family. Structure-function studies with natural and synthetic ligands demonstrate that the dimeric LuxR-type transcription factor CviR from Chromobacterium violaceum is potently antagonized by molecules that bind in place of the native acylated homoserine lactone autoinducer, provided that they stabilize a closed conformation. In such conformations, each of the two DNA-binding domains interacts with the ligand-binding domain of the opposing monomer. Consequently, the DNA-binding helices are held apart by {approx}60 {angstrom}, twice the {approx}30 {angstrom} separation required for operator binding. This approach may represent a general strategy for the inhibition of multidomain proteins.

  8. Antagonism of cyanide poisoning by dihydroxyacetone.

    PubMed

    Niknahad, Hossein; Ghelichkhani, Esmaeel

    2002-06-14

    Dihydroxyacetone (DHA) effectively antagonized the lethal effect of cyanide in mice and rabbits, particularly if administered in combination with thiosulfate. Oral DHA (2 and 4 g/kg) given to mice 10 min before injection (i.p.) of cyanide increased the LD50 values of cyanide from 5.7 mg/kg to 12 and 17.6 mg/kg, respectively. DHA prevented cyanide-induced lethality most effectively, if given orally 10-15 min before injection of cyanide. A combination of pretreatment with oral DHA (4 g/kg) and post-treatment with sodium thiosulfate (1 g/kg) increased the LD50 of cyanide by a factor of 9.9. Furthermore, DHA given intravenously to rabbits 5 min after subcutaneous injection of cyanide increased the LD50 of cyanide from 6 mg/kg to more than 11 mg/kg, while thiosulfate (1 g/kg) given intravenously 5 min after cyanide injection increased the LD50 of cyanide only to 8.5 mg/kg. DHA also prevented the convulsions that occurred after cyanide intoxication.

  9. Lung epithelial branching program antagonizes alveolar differentiation.

    PubMed

    Chang, Daniel R; Martinez Alanis, Denise; Miller, Rachel K; Ji, Hong; Akiyama, Haruhiko; McCrea, Pierre D; Chen, Jichao

    2013-11-05

    Mammalian organs, including the lung and kidney, often adopt a branched structure to achieve high efficiency and capacity of their physiological functions. Formation of a functional lung requires two developmental processes: branching morphogenesis, which builds a tree-like tubular network, and alveolar differentiation, which generates specialized epithelial cells for gas exchange. Much progress has been made to understand each of the two processes individually; however, it is not clear whether the two processes are coordinated and how they are deployed at the correct time and location. Here we show that an epithelial branching morphogenesis program antagonizes alveolar differentiation in the mouse lung. We find a negative correlation between branching morphogenesis and alveolar differentiation temporally, spatially, and evolutionarily. Gain-of-function experiments show that hyperactive small GTPase Kras expands the branching program and also suppresses molecular and cellular differentiation of alveolar cells. Loss-of-function experiments show that SRY-box containing gene 9 (Sox9) functions downstream of Fibroblast growth factor (Fgf)/Kras to promote branching and also suppresses premature initiation of alveolar differentiation. We thus propose that lung epithelial progenitors continuously balance between branching morphogenesis and alveolar differentiation, and such a balance is mediated by dual-function regulators, including Kras and Sox9. The resulting temporal delay of differentiation by the branching program may provide new insights to lung immaturity in preterm neonates and the increase in organ complexity during evolution.

  10. Wnt5b-associated exosomes promote cancer cell migration and proliferation.

    PubMed

    Harada, Takeshi; Yamamoto, Hideki; Kishida, Shosei; Kishida, Michiko; Awada, Chihiro; Takao, Toshifumi; Kikuchi, Akira

    2017-01-01

    Wnt5b is a member of the same family of proteins as Wnt5a, the overexpression of which is associated with cancer aggressiveness. Wnt5b is also suggested to be involved in cancer progression, however, details remain unclarified. We analyzed the biochemical properties of purified Wnt5b and the mode of secretion of Wnt5b by cancer cells. Wnt5b was glycosylated at three asparagine residues and lipidated at one serine residue, and these post-translational modifications of Wnt5b were essential for secretion. Purified Wnt5b showed Dvl2 phosphorylation and Rac activation abilities to a similar extent as Wnt5a. In cultured-cell conditioned medium, Wnt5b was detected in supernatant or precipitation fractions that were separated by centrifugation at 100 000 g. In PANC-1 pancreatic cancer cells, 55% of secreted endogenous Wnt5b was associated with exosomes. Exosomes from wild-type PANC-1 cells, but not those from Wnt5b-knockout PANC-1 cells, activated Wnt5b signaling in CHO cells and stimulated migration and proliferation of A549 lung adenocarcinoma cells, suggesting that endogenous, Wnt5b-associated exosomes are active. The exosomes were taken up by CHO cells and immunoelectron microscopy revealed that Wnt5b is indeed associated with exosomes. In Caco-2 colon cancer cells, most Wnt5b was recovered in precipitation fractions when Wnt5b was ectopically expressed (Caco-2/Wnt5b cells). Knockdown of TSG101, an exosome marker, decreased the secretion of Wnt5b-associated exosomes from Caco-2/Wnt5b cells and inhibited Wnt5b-dependent cell proliferation. Exosomes secreted from Caco-2/Wnt5b cells stimulated migration and proliferation of A549 cells. These results suggest that Wnt5b-associated exosomes promote cancer cell migration and proliferation in a paracrine manner.

  11. The MAPK ERK5, but not ERK1/2, inhibits the progression of monocytic phenotype to the functioning macrophage

    SciTech Connect

    Wang, Xuening; Pesakhov, Stella; Harrison, Jonathan S; Kafka, Michael; Danilenko, Michael; Studzinski, George P

    2015-01-01

    Intracellular signaling pathways present targets for pharmacological agents with potential for treatment of neoplastic diseases, with some disease remissions already recorded. However, cellular compensatory mechanisms usually negate the initial success. For instance, attempts to interrupt aberrant signaling downstream of the frequently mutated ras by inhibiting ERK1/2 has shown only limited usefulness for cancer therapy. Here, we examined how ERK5, that overlaps the functions of ERK1/2 in cell proliferation and survival, functions in a manner distinct from ERK1/2 in human AML cells induced to differentiate by 1,25D-dihydroxyvitamin D{sub 3} (1,25D). Using inhibitors of ERK1/2 and of MEK5/ERK5 at concentrations specific for each kinase in HL60 and U937 cells, we observed that selective inhibition of the kinase activity of ERK5, but not of ERK1/2, in the presence of 1,25D resulted in macrophage-like cell morphology and enhancement of phagocytic activity. Importantly, this was associated with increased expression of the macrophage colony stimulating factor receptor (M-CSFR), but was not seen when M-CSFR expression was knocked down. Interestingly, inhibition of ERK1/2 led to activation of ERK5 in these cells. Our results support the hypothesis that ERK5 negatively regulates the expression of M-CSFR, and thus has a restraining function on macrophage differentiation. The addition of pharmacological inhibitors of ERK5 may influence trials of differentiation therapy of AML. - Highlights: • ERK5 has at least some functions in AML cells which are distinct from those of ERK1/2. • ERK5 activity negatively controls the expression of M-CSFR. • ERK5 retards the progression of differentiation from monocyte to functional macrophage.

  12. Antagonism and synergism between lead and zinc in amphibian larvae.

    PubMed

    Herkovits, J; Pérez-Coll, C S

    1991-01-01

    Lead and zinc effects on Bufo arenarum larval survival were studied in single and combined treatments. On a weight basis, lead is about twice as toxic as zinc. The antagonism or synergism between these heavy metals is dose-dependent.

  13. Regulation of Adipogenesis by Quinine through the ERK/S6 Pathway

    PubMed Central

    Ning, Xiaomin; He, Jingjing; Shi, Xin’e; Yang, Gongshe

    2016-01-01

    Quinine is a bitter tasting compound that is involved in the regulation of body weight as demonstrated in in vivo animal models and in vitro models of the adipogenic system. Arguments exist over the positive or negative roles of quinine in both in vivo animal models and in vitro cell models, which motivates us to further investigate the functions of quinine in the in vitro adipogenic system. To clarify the regulatory functions of quinine in adipogenesis, mouse primary preadipocytes were induced for differentiation with quinine supplementation. The results showed that quinine enhanced adipogenesis in a dose dependent manner without affecting lipolysis. The pro-adipogenic effect of quinine was specific, as other bitter tasting agonists had no effect on adipogenesis. Moreover, the pro-adipogenic effect of quinine was mediated by activation of ERK/S6 (extracellular-signal-regulated kinase/Ribosomal protein S6) signaling. Knockdown of bitter taste receptor T2R106 (taste receptor, type 2, member 106) impaired the pro-adipogenic effect of quinine and suppressed the activation of ERK/S6 signaling. Taken together, quinine stimulates adipogenesis through ERK/S6 signaling, which at least partly functions via T2R106. PMID:27089323

  14. Insulin antagonizes the phagocytosis stimulating action of histamine in Tetrahymena.

    PubMed

    Csaba, G; Darvas, Z

    1992-02-01

    Histamine increased specifically the phagocytic activity of the unicellular Tetrahymena, whereas insulin had no influence on it. Insulin antagonized the phagocytosis stimulating action of histamine after simultaneous exposure and after preexposure two days earlier as well, although in the latter case to a lesser degree. Double exposure to a combination of histamine+insulin didn't influence the phagocytic activity at all, demonstrating the histamine antagonizing effect of insulin in this model.

  15. Wnt4 Participates in the Formation of Vertebrate Neuromuscular Junction

    PubMed Central

    Strochlic, Laure; Falk, Julien; Goillot, Evelyne; Sigoillot, Séverine; Bourgeois, Francine; Delers, Perrine; Rouvière, Jérôme; Swain, Amanda; Castellani, Valérie; Schaeffer, Laurent; Legay, Claire

    2012-01-01

    Neuromuscular junction (NMJ) formation requires the highly coordinated communication of several reciprocal signaling processes between motoneurons and their muscle targets. Identification of the early, spatially restricted cues in target recognition at the NMJ is still poorly documented, especially in mammals. Wnt signaling is one of the key pathways regulating synaptic connectivity. Here, we report that Wnt4 contributes to the formation of vertebrate NMJ in vivo. Results from a microarray screen and quantitative RT-PCR demonstrate that Wnt4 expression is regulated during muscle cell differentiation in vitro and muscle development in vivo, being highly expressed when the first synaptic contacts are formed and subsequently downregulated. Analysis of the mouse Wnt4−/− NMJ phenotype reveals profound innervation defects including motor axons overgrowing and bypassing AChR aggregates with 30% of AChR clusters being unapposed by nerve terminals. In addition, loss of Wnt4 function results in a 35% decrease of the number of prepatterned AChR clusters while Wnt4 overexpression in cultured myotubes increases the number of AChR clusters demonstrating that Wnt4 directly affects postsynaptic differentiation. In contrast, muscle structure and the localization of several synaptic proteins including acetylcholinesterase, MuSK and rapsyn are not perturbed in the Wnt4 mutant. Finally, we identify MuSK as a Wnt4 receptor. Wnt4 not only interacts with MuSK ectodomain but also mediates MuSK activation. Taken together our data reveal a new role for Wnt4 in mammalian NMJ formation that could be mediated by MuSK, a key receptor in synaptogenesis. PMID:22253844

  16. Wnt pathway in Dupuytren disease: connecting profibrotic signals.

    PubMed

    van Beuge, Marike M; Ten Dam, Evert-Jan P M; Werker, Paul M N; Bank, Ruud A

    2015-12-01

    A role of Wnt signaling in Dupuytren disease, a fibroproliferative disease of the hand and fingers, has not been fully elucidated. We examined a large set of Wnt pathway components and signaling targets and found significant dysregulation of 41 Wnt-related genes in tissue from the Dupuytren nodules compared with patient-matched control tissue. A large proportion of genes coding for Wnt proteins themselves was downregulated. However, both canonical Wnt targets and components of the noncanonical signaling pathway were upregulated. Immunohistochemical analysis revealed that protein expression of Wnt1-inducible secreted protein 1 (WISP1), a known Wnt target, was increased in nodules compared with control tissue, but knockdown of WISP1 using small interfering RNA (siRNA) in the Dupuytren myofibroblasts did not confirm a functional role. The protein expression of noncanonical pathway components Wnt5A and VANGL2 as well as noncanonical coreceptors Ror2 and Ryk was increased in nodules. On the contrary, the strongest downregulated genes in this study were 4 antagonists of Wnt signaling (DKK1, FRZB, SFRP1, and WIF1). Downregulation of these genes in the Dupuytren tissue was mimicked in vitro by treating normal fibroblasts with transforming growth factor β1 (TGF-β1), suggesting cross talk between different profibrotic pathways. Furthermore, siRNA-mediated knockdown of these antagonists in normal fibroblasts led to increased nuclear translocation of Wnt target β-catenin in response to TGF-β1 treatment. In conclusion, we have shown extensive dysregulation of Wnt signaling in affected tissue from Dupuytren disease patients. Components of both the canonical and the noncanonical pathways are upregulated, whereas endogenous antagonists are downregulated, possibly via interaction with other profibrotic pathways. Copyright © 2015 Elsevier Inc. All rights reserved.

  17. Stearoyl CoA desaturase is required to produce active, lipid-modified Wnt proteins.

    PubMed

    Rios-Esteves, Jessica; Resh, Marilyn D

    2013-09-26

    Wnt proteins contain palmitoleic acid, an unusual lipid modification. Production of an active Wnt signal requires the acyltransferase Porcupine and depends on the attachment of palmitoleic acid to Wnt. The source of this monounsaturated fatty acid has not been identified, and it is not known how Porcupine recognizes its substrate and whether desaturation occurs before or after fatty acid transfer to Wnt. Here, we show that stearoyl desaturase (SCD) generates a monounsaturated fatty acid substrate that is then transferred by Porcupine to Wnt. Treatment of cells with SCD inhibitors blocked incorporation of palmitate analogs into Wnt3a and Wnt5a and reduced Wnt secretion as well as autocrine and paracrine Wnt signaling. The SCD inhibitor effects were rescued by exogenous addition of monounsaturated fatty acids. We propose that SCD is a key molecular player responsible for Wnt biogenesis and processing and that SCD inhibition provides an alternative mechanism for blocking Wnt pathway activation.

  18. WNT signaling affects gene expression in the ventral diencephalon and pituitary gland growth

    PubMed Central

    Potok, Mary Anne; Cha, Kelly B.; Hunt, Andrea; Brinkmeier, Michelle L.; Leitges, Michael; Kispert, Andreas; Camper, Sally A.

    2009-01-01

    We examined the role of WNT signaling in pituitary development by characterizing the pituitary phenotype of three WNT knockout mice and assessing the expression of WNT pathway components. Wnt5a mutants have expanded domains of Fgf10 and BMP expression in the ventral diencephalon and a reduced domain of LHX3 expression in Rathke's pouch. Wnt4 mutants have mildly reduced cell differentiation, reduced POU1F1 expression, and mild anterior lobe hypoplasia. Wnt4, Wnt5a double mutants exhibit an additive pituitary phenotype of dysmorphology and mild hypoplasia. Wnt6 mutants have no obvious pituitary phenotype. We surveyed WNT expression and identified transcripts for numerous Wnts, Frizzleds and downstream pathway members in the pituitary and ventral diencephalon. These findings support the emerging model that WNT signaling affects the pituitary gland via effects on ventral diencephalon signaling, and suggest additional Wnt genes that are worthy of functional studies. PMID:18351662

  19. Agonism and Antagonism at the Insulin Receptor

    PubMed Central

    Knudsen, Louise; Hansen, Bo Falck; Jensen, Pia; Pedersen, Thomas Åskov; Vestergaard, Kirsten; Schäffer, Lauge; Blagoev, Blagoy; Oleksiewicz, Martin B.; Kiselyov, Vladislav V.; De Meyts, Pierre

    2012-01-01

    Insulin can trigger metabolic as well as mitogenic effects, the latter being pharmaceutically undesirable. An understanding of the structure/function relationships between insulin receptor (IR) binding and mitogenic/metabolic signalling would greatly facilitate the preclinical development of new insulin analogues. The occurrence of ligand agonism and antagonism is well described for G protein-coupled receptors (GPCRs) and other receptors but in general, with the exception of antibodies, not for receptor tyrosine kinases (RTKs). In the case of the IR, no natural ligand or insulin analogue has been shown to exhibit antagonistic properties, with the exception of a crosslinked insulin dimer (B29-B’29). However, synthetic monomeric or dimeric peptides targeting sites 1 or 2 of the IR were shown to be either agonists or antagonists. We found here that the S961 peptide, previously described to be an IR antagonist, exhibited partial agonistic effects in the 1–10 nM range, showing altogether a bell-shaped dose-response curve. Intriguingly, the agonistic effects of S961 were seen only on mitogenic endpoints (3H-thymidine incorporation), and not on metabolic endpoints (14C-glucose incorporation in adipocytes and muscle cells). The agonistic effects of S961 were observed in 3 independent cell lines, with complete concordance between mitogenicity (3H-thymidine incorporation) and phosphorylation of the IR and Akt. Together with the B29-B’29 crosslinked dimer, S961 is a rare example of a mixed agonist/antagonist for the human IR. A plausible mechanistic explanation based on the bivalent crosslinking model of IR activation is proposed. PMID:23300584

  20. ERK Phosphorylation Regulates Sleep and Plasticity in Drosophila

    PubMed Central

    Vanderheyden, William M.; Gerstner, Jason R.; Tanenhaus, Anne; Yin, Jerry C.; Shaw, Paul J.

    2013-01-01

    Given the relationship between sleep and plasticity, we examined the role of Extracellular signal-regulated kinase (ERK) in regulating baseline sleep, and modulating the response to waking experience. Both sleep deprivation and social enrichment increase ERK phosphorylation in wild-type flies. The effects of both sleep deprivation and social enrichment on structural plasticity in the LNvs can be recapitulated by expressing an active version of ERK (UAS-ERKSEM) pan-neuronally in the adult fly using GeneSwitch (Gsw) Gsw-elav-GAL4. Conversely, disrupting ERK reduces sleep and prevents both the behavioral and structural plasticity normally induced by social enrichment. Finally, using transgenic flies carrying a cAMP response Element (CRE)-luciferase reporter we show that activating ERK enhances CRE-Luc activity while disrupting ERK reduces it. These data suggest that ERK phosphorylation is an important mediator in transducing waking experience into sleep. PMID:24244744

  1. Role of the Wnt Pathway in Thyroid Cancer

    PubMed Central

    Sastre-Perona, Ana; Santisteban, Pilar

    2012-01-01

    Aberrant activation of Wnt signaling is involved in the development of several epithelial tumors. Wnt signaling includes two major types of pathways: (i) the canonical or Wnt/β-catenin pathway; and (ii) the non-canonical pathways, which do not involve β-catenin stabilization. Among these pathways, the Wnt/β-catenin pathway has received most attention during the past years for its critical role in cancer. A number of publications emphasize the role of the Wnt/β-catenin pathway in thyroid cancer. This pathway plays a crucial role in development and epithelial renewal, and components such as β-catenin and Axin are often mutated in thyroid cancer. Although it is accepted that altered Wnt signaling is a late event in thyroid cell transformation that affects anaplastic thyroid tumors, recent data suggest that it is also altered in papillary thyroid carcinoma (PTC) with RET/PTC mutations. Therefore, the purpose of this review is to summarize the main relevant data of Wnt signaling in thyroid cancer, with special emphasis on the Wnt/β-catenin pathway. PMID:22645520

  2. Mesodermal Wnt signaling organizes the neural plate via Meis3.

    PubMed

    Elkouby, Yaniv M; Elias, Sarah; Casey, Elena S; Blythe, Shelby A; Tsabar, Nir; Klein, Peter S; Root, Heather; Liu, Karen J; Frank, Dale

    2010-05-01

    In vertebrates, canonical Wnt signaling controls posterior neural cell lineage specification. Although Wnt signaling to the neural plate is sufficient for posterior identity, the source and timing of this activity remain uncertain. Furthermore, crucial molecular targets of this activity have not been defined. Here, we identify the endogenous Wnt activity and its role in controlling an essential downstream transcription factor, Meis3. Wnt3a is expressed in a specialized mesodermal domain, the paraxial dorsolateral mesoderm, which signals to overlying neuroectoderm. Loss of zygotic Wnt3a in this region does not alter mesoderm cell fates, but blocks Meis3 expression in the neuroectoderm, triggering the loss of posterior neural fates. Ectopic Meis3 protein expression is sufficient to rescue this phenotype. Moreover, Wnt3a induction of the posterior nervous system requires functional Meis3 in the neural plate. Using ChIP and promoter analysis, we show that Meis3 is a direct target of Wnt/beta-catenin signaling. This suggests a new model for neural anteroposterior patterning, in which Wnt3a from the paraxial mesoderm induces posterior cell fates via direct activation of a crucial transcription factor in the overlying neural plate.

  3. Mutations in WNT1 Cause Different Forms of Bone Fragility

    PubMed Central

    Keupp, Katharina; Beleggia, Filippo; Kayserili, Hülya; Barnes, Aileen M.; Steiner, Magdalena; Semler, Oliver; Fischer, Björn; Yigit, Gökhan; Janda, Claudia Y.; Becker, Jutta; Breer, Stefan; Altunoglu, Umut; Grünhagen, Johannes; Krawitz, Peter; Hecht, Jochen; Schinke, Thorsten; Makareeva, Elena; Lausch, Ekkehart; Cankaya, Tufan; Caparrós-Martín, José A.; Lapunzina, Pablo; Temtamy, Samia; Aglan, Mona; Zabel, Bernhard; Eysel, Peer; Koerber, Friederike; Leikin, Sergey; Garcia, K. Christopher; Netzer, Christian; Schönau, Eckhard; Ruiz-Perez, Victor L.; Mundlos, Stefan; Amling, Michael; Kornak, Uwe; Marini, Joan; Wollnik, Bernd

    2013-01-01

    We report that hypofunctional alleles of WNT1 cause autosomal-recessive osteogenesis imperfecta, a congenital disorder characterized by reduced bone mass and recurrent fractures. In consanguineous families, we identified five homozygous mutations in WNT1: one frameshift mutation, two missense mutations, one splice-site mutation, and one nonsense mutation. In addition, in a family affected by dominantly inherited early-onset osteoporosis, a heterozygous WNT1 missense mutation was identified in affected individuals. Initial functional analysis revealed that altered WNT1 proteins fail to activate canonical LRP5-mediated WNT-regulated β-catenin signaling. Furthermore, osteoblasts cultured in vitro showed enhanced Wnt1 expression with advancing differentiation, indicating a role of WNT1 in osteoblast function and bone development. Our finding that homozygous and heterozygous variants in WNT1 predispose to low-bone-mass phenotypes might advance the development of more effective therapeutic strategies for congenital forms of bone fragility, as well as for common forms of age-related osteoporosis. PMID:23499309

  4. Non-canonical WNT signalling in the lung

    PubMed Central

    Li, Changgong; Bellusci, Saverio; Borok, Zea; Minoo, Parviz

    2015-01-01

    The role of WNT signalling in metazoan organogenesis has been a topic of widespread interest. In the lung, while the role of canonical WNT signalling has been examined in some detail by multiple studies, the non-canonical WNT signalling has received limited attention. Reliable evidence shows that this important signalling mechanism constitutes a major regulatory pathway in lung development. In addition, accumulating evidence has also shown that the non-canonical WNT pathway is critical for maintaining lung homeostasis and that aberrant activation of this pathway may underlie several debilitating lung diseases. Functional analyses have further revealed that the non-canonical WNT pathway regulates multiple cellular activities in the lung that are dependent on the specific cellular context. In most cell types, non-canonical WNT signalling regulates canonical WNT activity, which is also critical for many aspects of lung biology. This review will summarize what is currently known about the role of non-canonical WNT signalling in lung development, homeostasis and pathogenesis of disease. PMID:26261051

  5. Non-canonical WNT signalling in the lung.

    PubMed

    Li, Changgong; Bellusci, Saverio; Borok, Zea; Minoo, Parviz

    2015-11-01

    The role of WNT signalling in metazoan organogenesis has been a topic of widespread interest. In the lung, while the role of canonical WNT signalling has been examined in some detail by multiple studies, the non-canonical WNT signalling has received limited attention. Reliable evidence shows that this important signalling mechanism constitutes a major regulatory pathway in lung development. In addition, accumulating evidence has also shown that the non-canonical WNT pathway is critical for maintaining lung homeostasis and that aberrant activation of this pathway may underlie several debilitating lung diseases. Functional analyses have further revealed that the non-canonical WNT pathway regulates multiple cellular activities in the lung that are dependent on the specific cellular context. In most cell types, non-canonical WNT signalling regulates canonical WNT activity, which is also critical for many aspects of lung biology. This review will summarize what is currently known about the role of non-canonical WNT signalling in lung development, homeostasis and pathogenesis of disease.

  6. The role of the Wnt canonical signaling in neurodegenerative diseases.

    PubMed

    Libro, Rosaliana; Bramanti, Placido; Mazzon, Emanuela

    2016-08-01

    The Wnt/β-catenin or Wnt canonical pathway controls multiple biological processes throughout development and adult life. Growing evidences have suggested that deregulation of the Wnt canonical pathway could be involved in the pathogenesis of neurodegenerative diseases. The Wnt canonical signaling is a pathway tightly regulated, which activation results in the inhibition of the Glycogen Synthase Kinase 3β (GSK-3β) function and in increased β-catenin activity, that migrates into the nucleus, activating the transcription of the Wnt target genes. Conversely, when the Wnt canonical pathway is turned off, increased levels of GSK-3β promote β-catenin degradation. Hence, GSK-3β could be considered as a key regulator of the Wnt canonical pathway. Of note, GSK-3β has also been involved in the modulation of inflammation and apoptosis, determining the delicate balance between immune tolerance/inflammation and neuronal survival/neurodegeneration. In this review, we have summarized the current acknowledgements about the role of the Wnt canonical pathway in the pathogenesis of some neurodegenerative diseases including Alzheimer's disease, cerebral ischemia, Parkinson's disease, Huntington's disease, multiple sclerosis and amyotrophic lateral sclerosis, with particular regard to the main in vitro and in vivo studies in this field, by reviewing 85 research articles about. Copyright © 2016 Elsevier Inc. All rights reserved.

  7. Testosterone/bicalutamide antagonism at the predicted extracellular androgen binding site of ZIP9.

    PubMed

    Bulldan, Ahmed; Malviya, Viveka Nand; Upmanyu, Neha; Konrad, Lutz; Scheiner-Bobis, Georgios

    2017-09-21

    ZIP9 is a Zn(2+) transporter, testosterone receptor, and mediator of signaling events through G-proteins. Despite these pivotal properties, however, its physiological and pathophysiological significance has not yet been comprehensively addressed. Using a cell line that lacks the classical androgen receptor we show that ZIP9-mediated phosphorylation of Erk1/2, CREB, or ATF-1 and expression of claudin-5 and zonula occludens-1 by testosterone can be completely antagonized by bicalutamide (Casodex), an anti-androgen of significant clinical impact. Computational modeling and docking experiments with ZIP9 reveal typical characteristics of ZIP transporters and an extracellular binding site for testosterone capable of accommodating bicalutamide. The presence of this site is verified by our demonstration that the membrane-impermeable testosterone analogue T-BSA-FITC labels the membrane only when ZIP9 is expressed and that this labeling is completely prevented by bicalutamide. The study connects structural features of ZIP9 to its functions and indicates a possible relevance of ZIP9 as a pharmacological target. Copyright © 2017. Published by Elsevier B.V.

  8. MAP kinase signaling antagonizes PAR-1 function during polarization of the early Caenorhabditis elegans embryo.

    PubMed

    Spilker, Annina C; Rabilotta, Alexia; Zbinden, Caroline; Labbé, Jean-Claude; Gotta, Monica

    2009-11-01

    PAR proteins (partitioning defective) are major regulators of cell polarity and asymmetric cell division. One of the par genes, par-1, encodes a Ser/Thr kinase that is conserved from yeast to mammals. In Caenorhabditis elegans, par-1 governs asymmetric cell division by ensuring the polar distribution of cell fate determinants. However the precise mechanisms by which PAR-1 regulates asymmetric cell division in C. elegans remain to be elucidated. We performed a genomewide RNAi screen and identified six genes that specifically suppress the embryonic lethal phenotype associated with mutations in par-1. One of these suppressors is mpk-1, the C. elegans homolog of the conserved mitogen activated protein (MAP) kinase ERK. Loss of function of mpk-1 restored embryonic viability, asynchronous cell divisions, the asymmetric distribution of cell fate specification markers, and the distribution of PAR-1 protein in par-1 mutant embryos, indicating that this genetic interaction is functionally relevant for embryonic development. Furthermore, disrupting the function of other components of the MAPK signaling pathway resulted in suppression of par-1 embryonic lethality. Our data therefore indicates that MAP kinase signaling antagonizes PAR-1 signaling during early C. elegans embryonic polarization.

  9. The acceleration of implant osseointegration by liposomal Wnt3a.

    PubMed

    Popelut, Antoine; Rooker, Scott M; Leucht, Philipp; Medio, Marie; Brunski, John B; Helms, Jill A

    2010-12-01

    The strength of a Wnt-based strategy for tissue regeneration lies in the central role that Wnts play in healing. Tissue injury triggers local Wnt activation at the site of damage, and this Wnt signal is required for the repair and/or regeneration of almost all tissues including bone, neural tissues, myocardium, and epidermis. We developed a biologically based approach to create a transient elevation in Wnt signaling in peri-implant tissues, and in doing so, accelerated bone formation around the implant. Our subsequent molecular and cellular analyses provide mechanistic insights into the basis for this pro-osteogenic effect. Given the essential role of Wnt signaling in bone formation, this protein-based approach may have widespread application in implant osseointegration. Copyright © 2010 Elsevier Ltd. All rights reserved.

  10. Keeping Wnt Signalosome in Check by Vesicular Traffic

    PubMed Central

    FENG, QIANG; GAO, NAN

    2015-01-01

    Wg/Wnts are paracrine and autocrine ligands that activate distinct signaling pathways while being internalized through surface receptors. Converging and contrasting views are shaping our understanding of whether, where, and how endocytosis may modulate Wnt signaling. We gather considerable amount of evidences to elaborate the point that signal-receiving cells utilize distinct, flexible, and sophisticated vesicular trafficking mechanisms to keep Wnt signaling activity in check. Same molecules in a highly context-dependent fashion serve as regulatory hub for various signaling purposes: amplification, maintenance, inhibition, and termination. Updates are provided for the regulatory mechanisms related to the three critical cell surface complexes, Wnt-Fzd-LRP6, Dkk1-Kremen-LRP6, and R-spondin-LGR5-RNF43, which potently influence Wnt signaling. We pay particular attentions to how cells achieve sustained and delicate control of Wnt signaling strength by employing comprehensive aspects of vesicular trafficking. PMID:25336320

  11. Wnt pathway regulation of embryonic stem cell self-renewal.

    PubMed

    Merrill, Bradley J

    2012-09-01

    Embryonic stem cells (ESCs) can generate all of the cell types found in the adult organism. Remarkably, they retain this ability even after many cell divisions in vitro, as long as the culture conditions prevent differentiation of the cells. Wnt signaling and β-catenin have been shown to cause strong effects on ESCs both in terms of stimulating the expansion of stem cells and stimulating differentiation toward lineage committed cell types. The varied effects of Wnt signaling in ESCs, alongside the sometimes unconventional mechanisms underlying the effects, have generated a fair amount of controversy and intrigue regarding the role of Wnt signaling in pluripotent stem cells. Insights into the mechanisms of Wnt function in stem cells can be gained by examination of the causes for seemingly opposing effects of Wnt signaling on self-renewal versus differentiation.

  12. Regulation of Wnt/β-catenin signaling by herpesviruses

    PubMed Central

    Zwezdaryk, Kevin J; Combs, Joseph A; Morris, Cindy A; Sullivan, Deborah E

    2016-01-01

    The Wnt/β-catenin signaling pathway is instrumental in successful differentiation and proliferation of mammalian cells. It is therefore not surprising that the herpesvirus family has developed mechanisms to interact with and manipulate this pathway. Successful coexistence with the host requires that herpesviruses establish a lifelong infection that includes periods of latency and reactivation or persistence. Many herpesviruses establish latency in progenitor cells and viral reactivation is linked to host-cell proliferation and differentiation status. Importantly, Wnt/β-catenin is tightly connected to stem/progenitor cell maintenance and differentiation. Numerous studies have linked Wnt/β-catenin signaling to a variety of cancers, emphasizing the importance of Wnt/β-catenin pathways in development, tissue homeostasis and disease. This review details how the alpha-, beta-, and gammaherpesviruses interact and manipulate the Wnt/β-catenin pathway to promote a virus-centric agenda. PMID:27878101

  13. Frat is dispensable for canonical Wnt signaling in mammals

    PubMed Central

    van Amerongen, Renée; Nawijn, Martijn; Franca-Koh, Jonathan; Zevenhoven, John; van der Gulden, Hanneke; Jonkers, Jos; Berns, Anton

    2005-01-01

    Wnt-signal transduction through β-catenin is thought to require the inhibition of GSK3 by Frat/GBP. To investigate the role of Frat in mammalian development, we have generated mice with targeted mutations in all three murine Frat homologs. We show that Frat is normally expressed at sites of active Wnt signaling. Surprisingly, Frat-deficient mice do not display gross abnormalities. Moreover, canonical Wnt signaling in primary cells is unaffected by the loss of Frat. These studies show that Frat is not an essential component of the canonical Wnt pathway in higher organisms, despite the strict requirement of Frat/GBP for maternal Wnt signaling in Xenopus. PMID:15681612

  14. Wntless in Wnt secretion: molecular, cellular and genetic aspects.

    PubMed

    Das, Soumyashree; Yu, Shiyan; Sakamori, Ryotaro; Stypulkowski, Ewa; Gao, Nan

    2012-12-01

    Throughout the animal kingdom, Wnt-triggered signal transduction pathways play fundamental roles in embryonic development and tissue homeostasis. Wnt proteins are modified as glycolipoproteins and are secreted into the extracellular environment as morphogens. Recent studies on the intracellular trafficking of Wnt proteins demonstrate multiple layers of regulation along its secretory pathway. These findings have propelled a great deal of interest among researchers to further investigate the molecular mechanisms that control the release of Wnts and hence the level of Wnt signaling. This review is dedicated to Wntless, a putative G-protein coupled receptor that transports Wnts intracellularly for secretion. Here, we highlight the conclusions drawn from the most recent cellular, molecular and genetic studies that affirm the role of Wntless in the secretion of Wnt proteins.

  15. Follistatin Operates Downstream of Wnt4 in Mammalian Ovary Organogenesis

    PubMed Central

    Yao, Humphrey H.C.; Matzuk, Martin M.; Jorgez, Carolina J.; Menke, Douglas B.; Page, David C.; Swain, Amanda; Capel, Blanche

    2014-01-01

    Wnt4−/− XX gonads display features normally associated with testis differentiation, suggesting that WNT4 actively represses elements of the male pathway during ovarian development. Here, we show that follistatin (Fst), which encodes a TGFβ superfamily binding protein, is a downstream component of Wnt4 signaling. Fst inhibits formation of the XY-specific coelomic vessel in XX gonads. In addition, germ cells in the ovarian cortex are almost completely lost in both Wnt4 and Fst null gonads before birth. Thus, we propose that WNT4 acts through FST to regulate vascular boundaries and maintain germ cell survival in the ovary. Developmental Dynamics 230:210–215, 2004. PMID:15162500

  16. Ursodeoxycholic acid (UDCA) can inhibit deoxycholic acid (DCA)-induced apoptosis via modulation of EGFR/Raf-1/ERK signaling in human colon cancer cells.

    PubMed

    Im, Eunok; Martinez, Jesse D

    2004-02-01

    Ursodeoxycholic acid (UDCA), a hydrophilic bile acid, is known as a cytoprotective agent. UDCA prevents apoptosis induced by a variety of stress stimuli including cytotoxic bile acids such as deoxycholic acid (DCA). Here we examined the molecular mechanism by which UDCA can antagonize DCA-induced apoptosis in human colon cancer cells. UDCA pretreatment decreases the number of apoptotic cells caused by exposure to DCA and UDCA. Further studies of the signaling pathway showed that UDCA pretreatment suppressed DNA binding activity of activator protein-1 and this was accompanied by downregulation of both extracellular signal-regulated kinase (ERK) and Raf-1 kinase activities stimulated by exposure to DCA. DCA was also found to activate epidermal growth factor receptor (EGFR) activity and UDCA inhibited this. Collectively, these findings suggest that the inhibitory effect of UDCA in DCA-induced apoptosis is partly mediated by modulation of EGFR/Raf-1/ERK signaling.

  17. IGF-1R inhibition in mammary epithelia promotes canonical Wnt signaling and Wnt1-driven tumors

    PubMed Central

    Rota, Lauren M.; Albanito, Lidia; Shin, Marcus E.; Goyeneche, Corey L.; Shushanov, Sain; Gallagher, Emily J.; LeRoith, Derek; Lazzarino, Deborah A.; Wood, Teresa L.

    2014-01-01

    Triple-negative breast cancers (TNBC) are an aggressive disease subtype which unlike other subtypes lack an effective targeted therapy. Inhibitors of the insullin-like growth factor receptor (IGF-1R) have been considered for use in treating TNBC. Here we provide genetic evidence that IGF-1R inhibition promotes development of Wnt1-mediated murine mammary tumors that offer a model of TNBC. We found that in a double transgenic mouse model carrying activated Wnt-1 and mutant IGF-1R, a reduction in IGF-1R signaling reduced tumor latency and promoted more aggressive phenotypes. These tumors displayed a squamal cell phenotype with increased expression of keratins 5/6 and β-catenin. Notably, cell lineage analyses revealed an increase in basal (CD29hi/CD24+) and luminal (CD24+/CD61+/CD29lo) progenitor cell populations, along with increased Nanog expression and decreased Elf5 expression. In these doubly transgenic mice, lung metastases developed with characteristics of the primary tumors, unlike MMTV-Wnt1 mice. Mechanistic investigations showed that pharmacological inhibition of the IGF-1R in vitro was sufficient to increase the tumorsphere-forming efficiency of MMTV-Wnt1 tumor cells. Tumors from doubly transgenic mice also exhibited an increase in the expression ratio of the IGF-II-sensitive, A isoform of the insulin receptor vs the IR-B isoform, which in vitro resulted in enhanced expression of β-catenin. Overall, our results revealed that in Wnt-driven tumors an attenuation of IGF-1R signaling accelerates tumorigenesis and promotes more aggressive phenotypes, with potential implications for understanding TNBC pathobiology and treatment. PMID:25092896

  18. Down-regulation of ERK1 and ERK2 activity during differentiation of the intestinal cell line HT-29.

    PubMed

    Luongo, Diomira; Mazzarella, Giuseppe; Della, Ragione Fulvio; Maurano, Francesco; Rossi, Mauro

    2002-02-01

    The role and regulation of signal transduction pathways in proliferation and differentiation of intestinal epithelial cells are still poorly understood. However, growing evidences have been recently accumulated demonstrating that mitogen-activated protein kinases (MAPKs) play a pivotal function in the normal development of intestine. We have investigated, in the intestinal cell line HT-29, the regulation (namely activity and phosphorylation degree) of MAP kinases ERK 1 (p44) and ERK 2 (p42) during differentiation. Addition of fetal calf serum to HT-29 undifferentiated resting cells caused a rapid phosphorylation of both ERKs and an increase of their specific kinase activity. Moreover, nuclear translocation of ERK 1 and ERK 2 occurred concurrently to their activation, leading to the conclusion that ERK 1 and ERK 2 are classically regulated when quiescent HT-29 cells are induced to proliferate. Butyrate addition to the intestinal cell line resulted in terminal differentiation and in a selective down-regulation of ERK 2 activity (and phosphorylation degree) without any effect on ERK 1. Conversely, when HT-29 cells were differentiated by repeated passages in a glucose-free medium, we observed a progressive dephosphorylation and inactivation of p42 and p44 kinases along with the failure of serum to activate both the enzymes. Our findings suggest that, during the differentiation of intestinal cells, remarkable changes occur in ERK 1 and ERK 2 control mechanisms leading to an unresponsiveness of MAP kinase pathway.

  19. Wnt antagonist gene polymorphisms and renal cancer

    PubMed Central

    Hirata, Hiroshi; Hinoda, Yuji; Nakajima, Koichi; Kikuno, Nobuyuki; Yamamura, Soichiro; Kawakami, Kazumori; Suehiro, Yutaka; Tabatabai, Z. Laura; Ishii, Nobuhisa; Dahiya, Rajvir

    2014-01-01

    Purpose Epigenetic silencing of several Wnt pathway related genes has been reported in renal cancer. Except for the TCF4 gene, there are no reports regarding Wnt pathway gene polymorphisms in renal cancer. Therefore, we hypothesized that the polymorphisms in Wnt signaling genes may be risk factors for renal cancer. Experimental Design A total of 210 patients (145 male and 65 female) with pathologically confirmed renal cell carcinoma (RCC), and 200 age- and sex-matched control individuals were enrolled in this study. We genotyped 14 SNPs in six genes including DKK2 (rs17037102, rs419558, rs447372), DKK3 (rs3206824, rs11022095, rs1472189, rs7396187, rs2291599), DKK4 (rs2073664), sFRP4 (rs1802073, rs1802074), SMAD7 (rs12953717), DAAM2 (rs6937133, rs2504106) using PCR-RFLP and direct sequencing in RCC and age-matched healthy subjects. We also tested the relationship between these polymorphisms and clinicopathologic data including gender, grade, tumor stage, lymph-node involvement, distant metastasis, and overall survival. Results A significant decrease in the frequency of the G/A+A/A genotypes in the DKK3 codon335 rs3206824 was observed in RCC patients compared with controls. The frequency of the rs3206824 (G/A) A- rs7396187 (G/C) C haplotype was significantly lower in RCC compared with other haplotypes. We also found that DKK3 rs1472189 C/T is associated with distant metastasis and furthermore, DKK2 rs17037102 G homozygous patients had a decreased risk for death by multivariate Cox regression analysis. Conclusions This is the first report documenting that DKK3 polymorphisms are associated with RCC and that the DKK2 rs17037102 polymorphism may be a predictor for survival in RCC patients after radical nephrectomy. PMID:19562778

  20. Wnt signaling: role in Alzheimer disease and schizophrenia.

    PubMed

    Inestrosa, Nibaldo C; Montecinos-Oliva, Carla; Fuenzalida, Marco

    2012-12-01

    Wnt signaling function starts during the development of the nervous system and is crucial for synaptic plasticity in the adult brain. Clearly Wnt effects in synaptic and plastic processes are relevant, however the implication of this pathway in the prevention of neurodegenerative diseases that produce synaptic impairment, is even more interesting. Several years ago our laboratory found a relationship between the loss of Wnt signaling and the neurotoxicity of the amyloid-β-peptide (Aβ), one of the main players in Alzheimer's disease (AD). Moreover, the activation of the Wnt signaling cascade prevents Aβ-dependent cytotoxic effects. In fact, disrupted Wnt signaling may be a direct link between Aβ-toxicity and tau hyperphosphorylation, ultimately leading to impaired synaptic plasticity and/or neuronal degeneration, indicating that a single pathway can account for both neuro-pathological lesions and altered synaptic function. These observations, suggest that a sustained loss of Wnt signaling function may be a key relevant factor in the pathology of AD. On the other hand, Schizophrenia remains one of the most debilitating and intractable illness in psychiatry. Since Wnt signaling is important in organizing the developing brain, it is reasonable to propose that defects in Wnt signaling could contribute to Schizophrenia, particularly since the neuro-developmental hypothesis of the disease implies subtle dys-regulation of brain development, including some core components of the Wnt signaling pathways such as GSK-3β or Disrupted in Schizophrenia-1 (DISC-1). This review focuses on the relationship between Wnt signaling and its potential relevance for the treatment of neurodegenerative and neuropsychiatric diseases including AD and Schizophrenia.

  1. Skeletal characteristics associated with homozygous and heterozygous WNT1 mutations.

    PubMed

    Palomo, Telma; Al-Jallad, Hadil; Moffatt, Pierre; Glorieux, Francis H; Lentle, Brian; Roschger, Paul; Klaushofer, Klaus; Rauch, Frank

    2014-10-01

    Recent reports have shown that homozygous or compound heterozygous mutations in WNT1 can give rise to severe bone fragility resembling osteogenesis imperfecta, whereas heterozygous WNT1 mutations have been found in adults with dominant early-onset osteoporosis. Here we assessed the effects of WNT1 mutations in four children with recessive severe bone fragility and in heterozygous family members. In vitro studies using the Topflash luciferase reporter system showed that two WNT1 missense mutations that were observed in these families, p.Cys143Phe and p.Val355Phe, decreased the ability of WNT1 to stimulate WNT signaling by >90%. Analyses of iliac bone samples revealed no major abnormalities in bone mineralization density distribution, an indicator of material bone properties, whereas a shift towards higher bone mineralization density is characteristic of classical osteogenesis imperfecta caused by mutations in COL1A1/COL1A2. Intravenous bisphosphonate treatment of four children with homozygous or compound heterozygous WNT1 mutations was associated with increasing lumbar spine areal bone mineral density z-scores, as measured by dual energy X-ray absorptiometry, but the effect was smaller than what had previously been reported for children with classical osteogenesis imperfecta. Family members with heterozygous WNT1 mutation tended to have low bone mass. Three of these heterozygous individuals had radiographic signs of vertebral fractures. These observations suggest that more effective treatment approaches are needed for children with recessive WNT1-related bone fragility and that a systematic work-up for osteoporosis is warranted for WNT1 mutation carriers in these families. Copyright © 2014. Published by Elsevier Inc.

  2. Regulation of Wnt4 in chronic obstructive pulmonary disease

    PubMed Central

    Durham, Andrew L.; McLaren, Alistair; Hayes, Brian P.; Caramori, Gaetano; Clayton, Chris L.; Barnes, Peter J.; Chung, K. Fan; Adcock, Ian M.

    2013-01-01

    Chronic obstructive pulmonary disease (COPD) is associated with persistent inflammation and oxidative stress in susceptible individuals. Using microarray analysis of bronchial biopsy samples from patients with COPD and controls, we identified Wnt4 as being up-regulated in COPD. Analysis of bronchial biopsy samples showed a very strong correlation between Wnt4 and IL8 gene expression, suggesting that Wnt4 plays a role in chronic lung inflammation. In vitro, Wnt4 induced proliferation and inflammation in human epithelial cells (BEAS-2B) and normal primary human bronchial epithelial cells in a concentration-dependent manner. This effect was enhanced in the presence of interleukin-1β (IL-1β) as a result of activation of the p38 and c-Jun NH2-terminal kinase mitogen-activated protein kinase pathways. Hydrogen peroxide, but not proinflammatory stimuli, up-regulated Wnt4 expression in epithelial cells. In monocytic THP-1 and primary airway smooth muscle cells, Wnt4 induced inflammation and enhanced the inflammatory response to lipopolysaccharide and IL-1β but did not induce proliferation. In addition, these other cell types did not have enhanced Wnt4 expression in response to hydrogen peroxide. Our results indicate that airway epithelial activation, due to oxidative stress, may lead to Wnt4 induction. Wnt4, in turn, acts through the noncanonical pathway to activate epithelial cell remodeling and IL8 gene expression, leading to neutrophil infiltration and inflammation.—Durham, A. L., McLaren, A., Hayes, B. P., Caramori, G., Clayton, C. L., Barnes, P. J., Chung, K. F., Adcock, I. M. Regulation of Wnt4 in chronic obstructive pulmonary disease. PMID:23463699

  3. Interplay between Wnt2 and Wnt2bb controls multiple steps of early foregut-derived organ development

    PubMed Central

    Poulain, Morgane; Ober, Elke A.

    2011-01-01

    The vertebrate liver, pancreas and lung arise in close proximity from the multipotent foregut endoderm. Tissue-explant experiments uncovered instructive signals emanating from the neighbouring lateral plate mesoderm, directing the endoderm towards specific organ fates. This suggested that an intricate network of signals is required to control the specification and differentiation of each organ. Here, we show that sequential functions of Wnt2bb and Wnt2 control liver specification and proliferation in zebrafish. Their combined specific activities are essential for liver specification, as their loss of function causes liver agenesis. Conversely, excess wnt2bb or wnt2 induces ectopic liver tissue at the expense of pancreatic and anterior intestinal tissues, revealing the competence of intestinal endoderm to respond to hepatogenic signals. Epistasis experiments revealed that the receptor frizzled homolog 5 (fzd5) mediates part of the broader hepatic competence of the alimentary canal. fzd5 is required for early liver formation and interacts genetically with wnt2 as well as wnt2bb. In addition, lack of both ligands causes agenesis of the swim bladder, the structural homolog of the mammalian lung. Thus, tightly regulated spatiotemporal expression of wnt2bb, wnt2 and fzd5 is central to coordinating early liver, pancreas and swim bladder development from a multipotent foregut endoderm. PMID:21771809

  4. Wnt3a Promotes the Vasculogenic Mimicry Formation of Colon Cancer via Wnt/β-Catenin Signaling.

    PubMed

    Qi, Lisha; Song, Wangzhao; Liu, Zhiyong; Zhao, Xiulan; Cao, Wenfeng; Sun, Baocun

    2015-08-10

    Our previous study provided evidence that non-canonical Wnt signaling is involved in regulating vasculogenic mimicry (VM) formation. However, the functions of canonical Wnt signaling in VM formation have not yet been explored. In this study, we found the presence of VM was related to colon cancer histological differentiation (p < 0.001), the clinical stage (p < 0.001), and presence of metastasis and recurrence (p < 0.001). VM-positive colon cancer samples showed increased Wnt3a expression (p < 0.001) and β-catenin nuclear expression (p < 0.001) compared with the VM-negative samples. In vitro, over-regulated Wnt3a expression in HT29 colon cancer cells promoted the capacity to form tube-like structures in the three-dimensional (3-D) culture together with increased expression of endothelial phenotype-associated proteins such as VEGFR2 and VE-cadherin. The mouse xenograft model showed that Wnt3a-overexpressing cells grew into larger tumor masses and formed more VM than the control cells. In addition, the Wnt/β-catenin signaling antagonist Dickkopf-1(Dkk1) can reverse the capacity to form tube-like structures and can decrease the expressions of VEGFR2 and VE-cadherin in Wnt3a-overexpressing cells. Taken together, our results suggest that Wnt/β-catenin signaling is involved in VM formation in colon cancer and might contribute to the development of more accurate treatment modalities aimed at VM.

  5. Jade-1 inhibits Wnt signalling by ubiquitylating beta-catenin and mediates Wnt pathway inhibition by pVHL.

    PubMed

    Chitalia, Vipul C; Foy, Rebecca L; Bachschmid, Markus M; Zeng, Liling; Panchenko, Maria V; Zhou, Mina I; Bharti, Ajit; Seldin, David C; Lecker, Stewart H; Dominguez, Isabel; Cohen, Herbert T

    2008-10-01

    The von Hippel-Lindau protein pVHL suppresses renal tumorigenesis in part by promoting the degradation of hypoxia-inducible HIF-alpha transcription factors; additional mechanisms have been proposed. pVHL also stabilizes the plant homeodomain protein Jade-1, which is a candidate renal tumour suppressor that may correlate with renal cancer risk. Here we show that Jade-1 binds the oncoprotein beta-catenin in Wnt-responsive fashion. Moreover, Jade-1 destabilizes wild-type beta-catenin but not a cancer-causing form of beta-catenin. Whereas the well-established beta-catenin E3 ubiquitin ligase component beta-TrCP ubiquitylates only phosphorylated beta-catenin, Jade-1 ubiquitylates both phosphorylated and non-phosphorylated beta-catenin and therefore regulates canonical Wnt signalling in both Wnt-off and Wnt-on phases. Thus, the different characteristics of beta-TrCP and Jade-1 may ensure optimal Wnt pathway regulation. Furthermore, pVHL downregulates beta-catenin in a Jade-1-dependent manner and inhibits Wnt signalling, supporting a role for Jade-1 and Wnt signalling in renal tumorigenesis. The pVHL tumour suppressor and the Wnt tumorigenesis pathway are therefore directly linked through Jade-1.

  6. Jade-1 inhibits Wnt signaling by ubiquitinating β-catenin and mediates Wnt pathway inhibition by pVHL

    PubMed Central

    Chitalia, Vipul C.; Foy, Rebecca L.; Bachschmid, Markus M.; Zeng, Liling; Panchenko, Maria V.; Zhou, Mina I.; Bharti, Ajit; Seldin, David C.; Lecker, Stewart H.; Dominguez, Isabel; Cohen, Herbert T.

    2009-01-01

    The von Hippel-Lindau protein pVHL suppresses renal tumorigenesis in part by promoting degradation of hypoxia-inducible HIF-alpha transcription factors1, and additional mechanisms have been proposed2. pVHL also stabilizes plant homeodomain (PHD) protein Jade-1, which is a candidate renal tumor suppressor that may correlate with renal cancer risk3-5. We show here that Jade-1 binds the oncoprotein β-catenin in Wnt-responsive fashion. Moreover, Jade-1 destabilizes wild-type β-catenin, but not a cancer-causing form of β-catenin. While β-TrCP ubiquitinates only phosphorylated β-catenin6, Jade-1 ubiquitinates both phosphorylated and non-phosphorylated β-catenin and therefore regulates canonical Wnt signaling in both Wnt-off and Wnt-on phases. Thus, the different characteristics of β-TrCP and Jade-1 may ensure optimal Wnt pathway regulation. Furthermore, pVHL down-regulates β-catenin in a Jade-1-dependent manner and inhibits Wnt signaling, supporting a role for Jade-1 and Wnt signaling in renal tumorigenesis. The pVHL tumor suppressor and the Wnt tumorigenesis pathway are therefore directly linked through Jade-1. PMID:18806787

  7. Initiation of Wnt signaling: control of Wnt coreceptor Lrp6 phosphorylation/activation via frizzled, dishevelled and axin functions

    PubMed Central

    Zeng, Xin; Huang, He; Tamai, Keiko; Zhang, Xinjun; Harada, Yuko; Yokota, Chika; Almeida, Karla; Wang, Jianbo; Doble, Brad; Woodgett, Jim; Wynshaw-Boris, Anthony; Hsieh, Jen-Chieh; He, Xi

    2016-01-01

    Canonical Wnt/β-catenin signaling has central roles in development and diseases, and is initiated by the action of the frizzled (Fz) receptor, its coreceptor LDL receptor-related protein 6 (Lrp6), and the cytoplasmic dishevelled (Dvl) protein. The functional relationships among Fz, Lrp6 and Dvl have long been enigmatic. We demonstrated previously that Wnt-induced Lrp6 phosphorylation via glycogen synthase kinase 3 (Gsk3) initiates Wnt/β-catenin signaling. Here we show that both Fz and Dvl functions are critical for Wnt-induced Lrp6 phosphorylation through Fz-Lrp6 interaction. We also show that axin, a key scaffolding protein in the Wnt pathway, is required for Lrp6 phosphorylation via its ability to recruit Gsk3, and inhibition of Gsk3 at the plasma membrane blocks Wnt/β-catenin signaling. Our results suggest a model that upon Wnt-induced Fz-Lrp6 complex formation, Fz recruitment of Dvl in turn recruits the axin-Gsk3 complex, thereby promoting Lrp6 phosphorylation to initiate β-catenin signaling. We discuss the dual roles of the axin-Gsk3 complex and signal amplification by Lrp6-axin interaction during Wnt/β-catenin signaling. PMID:18077588

  8. Expression of WNT genes in cervical cancer-derived cells: Implication of WNT7A in cell proliferation and migration.

    PubMed

    Ramos-Solano, Moisés; Meza-Canales, Ivan D; Torres-Reyes, Luis A; Alvarez-Zavala, Monserrat; Alvarado-Ruíz, Liliana; Rincon-Orozco, Bladimiro; Garcia-Chagollan, Mariel; Ochoa-Hernández, Alejandra B; Ortiz-Lazareno, Pablo C; Rösl, Frank; Gariglio, Patricio; Jave-Suárez, Luis F; Aguilar-Lemarroy, Adriana

    2015-07-01

    According to the multifactorial model of cervical cancer (CC) causation, it is now recognized that other modifications, in addition to Human papillomavirus (HPV) infection, are necessary for the development of this neoplasia. Among these, it has been proposed that a dysregulation of the WNT pathway might favor malignant progression of HPV-immortalized keratinocytes. The aim of this study was to identify components of the WNT pathway differentially expressed in CC vs. non-tumorigenic, but immortalized human keratinocytes. Interestingly, WNT7A expression was found strongly downregulated in cell lines and biopsies derived from CC. Restoration of WNT7A in CC-derived cell lines using a lentiviral gene delivery system or after adding a recombinant human protein decreases cell proliferation. Likewise, WNT7A silencing in non-tumorigenic cells markedly accelerates proliferation. Decreased WNT7A expression was due to hypermethylation at particular CpG sites. To our knowledge, this is the first study reporting reduced WNT7A levels in CC-derived cells and that ectopic WNT7A restoration negatively affects cell proliferation and migration. Copyright © 2015 Elsevier Inc. All rights reserved.

  9. Wnt3a regulates proliferation and migration of HUVEC via canonical and non-canonical Wnt signaling pathways

    SciTech Connect

    Samarzija, Ivana; Sini, Patrizia; Schlange, Thomas; MacDonald, Gwen; Hynes, Nancy E.

    2009-08-28

    Untangling the signaling pathways involved in endothelial cell biology is of central interest for the development of antiangiogenesis based therapies. Here we report that Wnt3a induces the proliferation and migration of HUVECs, but does not affect their survival. Wnt3a-induced proliferation was VEGFR signaling independent, but reduced upon CamKII inhibition. In a search for the downstream mediators of Wnt3a's effects on HUVEC biology, we found that Wnt3a treatment leads to phosphorylation of DVL3 and stabilization of {beta}-catenin. Moreover, under the same conditions we observed an upregulation in c-MYC, TIE-2 and GJA1 mRNA transcripts. Although treatment of HUVECs with Wnt5a induced DVL3 phosphorylation, we did not observe any of the other effects seen upon Wnt3a stimulation. Taken together, our data indicate that Wnt3a induces canonical and non-canonical Wnt signaling in HUVECs, and stimulates their proliferation and migration.

  10. Wnt signalling requires MTM-6 and MTM-9 myotubularin lipid-phosphatase function in Wnt-producing cells

    PubMed Central

    Silhankova, Marie; Port, Fillip; Harterink, Martin; Basler, Konrad; Korswagen, Hendrik C

    2010-01-01

    Wnt proteins are lipid-modified glycoproteins that have important roles in development, adult tissue homeostasis and disease. Secretion of Wnt proteins from producing cells is mediated by the Wnt-binding protein MIG-14/Wls, which binds Wnt in the Golgi network and transports it to the cell surface for release. It has recently been shown that recycling of MIG-14/Wls from the plasma membrane to the trans-Golgi network is required for efficient Wnt secretion, but the mechanism of this retrograde transport pathway is still poorly understood. In this study, we report the identification of MTM-6 and MTM-9 as novel regulators of MIG-14/Wls trafficking in Caenorhabditis elegans. MTM-6 and MTM-9 are myotubularin lipid phosphatases that function as a complex to dephosphorylate phosphatidylinositol-3-phosphate, a central regulator of endosomal trafficking. We show that mutation of mtm-6 or mtm-9 leads to defects in several Wnt-dependent processes and demonstrate that MTM-6 is required in Wnt-producing cells as part of the MIG-14/Wls-recycling pathway. This function is evolutionarily conserved, as the MTM-6 orthologue DMtm6 is required for Wls stability and Wg secretion in Drosophila. We conclude that regulation of endosomal trafficking by the MTM-6/MTM-9 myotubularin complex is required for the retromer-dependent recycling of MIG-14/Wls and Wnt secretion. PMID:21076391

  11. Wnt signalling requires MTM-6 and MTM-9 myotubularin lipid-phosphatase function in Wnt-producing cells.

    PubMed

    Silhankova, Marie; Port, Fillip; Harterink, Martin; Basler, Konrad; Korswagen, Hendrik C

    2010-12-15

    Wnt proteins are lipid-modified glycoproteins that have important roles in development, adult tissue homeostasis and disease. Secretion of Wnt proteins from producing cells is mediated by the Wnt-binding protein MIG-14/Wls, which binds Wnt in the Golgi network and transports it to the cell surface for release. It has recently been shown that recycling of MIG-14/Wls from the plasma membrane to the trans-Golgi network is required for efficient Wnt secretion, but the mechanism of this retrograde transport pathway is still poorly understood. In this study, we report the identification of MTM-6 and MTM-9 as novel regulators of MIG-14/Wls trafficking in Caenorhabditis elegans. MTM-6 and MTM-9 are myotubularin lipid phosphatases that function as a complex to dephosphorylate phosphatidylinositol-3-phosphate, a central regulator of endosomal trafficking. We show that mutation of mtm-6 or mtm-9 leads to defects in several Wnt-dependent processes and demonstrate that MTM-6 is required in Wnt-producing cells as part of the MIG-14/Wls-recycling pathway. This function is evolutionarily conserved, as the MTM-6 orthologue DMtm6 is required for Wls stability and Wg secretion in Drosophila. We conclude that regulation of endosomal trafficking by the MTM-6/MTM-9 myotubularin complex is required for the retromer-dependent recycling of MIG-14/Wls and Wnt secretion.

  12. Wnt Genes in the Mouse Uterus: Potential Regulation of Implantation1

    PubMed Central

    Hayashi, Kanako; Erikson, David W.; Tilford, Sarah A.; Bany, Brent M.; Maclean, James A.; Rucker, Edmund B.; Johnson, Greg A.; Spencer, Thomas E.

    2009-01-01

    Wnt genes are involved in critical developmental and growth processes. The present study comprehensively analyzed temporal and spatial alterations in Wnt and Fzd gene expression in the mouse uterus during peri-implantation of pregnancy. Expression of Wnt4, Wnt5a, Wnt7a, Wnt7b, Wnt11, Wnt16, Fzd2, Fzd4, and Fzd6 was detected in the uterus during implantation. Wnt4 mRNA was most abundant in the decidua, whereas Wnt5a mRNA was restricted to the mesometrial decidua during decidualization. Wnt7a, Wnt7b, and Wnt11 mRNAs were abundantly detected in the endometrial epithelia. The expression of Wnt7b was robust in the luminal epithelium (LE) at the implantation site on Gestational Day 5, whereas Wnt11 mRNA disappeared in the LE adjacent to the embryo in the antimesometrial implantation chamber but remained abundant in the LE. Wnt16 mRNA was localized to the stroma surrounding the LE on Day 4 and remained in the stroma adjacent to the LE but not in areas undergoing the decidual reaction. Fzd2 mRNA was detected in the decidua, Fzd4 mRNA was in the vessels and stroma surrounding the embryo, and Fzd6 mRNA was observed in the endometrial epithelia, stroma, and some blood vessels during implantation. Ovarian steroid hormone treatment was found to regulate Wnt genes and Fzd receptors in ovariectomized mice. Especially, single injections of progesterone stimulated Wnt11 mRNA, and estrogen stimulated Wnt4 and Wnt7b. The temporal and spatial alterations in Wnt genes likely play a critical role during implantation and decidualization in mice. PMID:19164167

  13. Can we safely target the WNT pathway?

    PubMed Central

    Kahn, Michael

    2015-01-01

    WNT–β-catenin signalling is involved in a multitude of developmental processes and the maintenance of adult tissue homeostasis by regulating cell proliferation, differentiation, migration, genetic stability and apoptosis, as well as by maintaining adult stem cells in a pluripotent state. Not surprisingly, aberrant regulation of this pathway is therefore associated with a variety of diseases, including cancer, fibrosis and neurodegeneration. Despite this knowledge, therapeutic agents specifically targeting the WNT pathway have only recently entered clinical trials and none has yet been approved. This Review examines the problems and potential solutions to this vexing situation and attempts to bring them into perspective. PMID:24981364

  14. Isorhamnetin inhibits H₂O₂-induced activation of the intrinsic apoptotic pathway in H9c2 cardiomyocytes through scavenging reactive oxygen species and ERK inactivation.

    PubMed

    Sun, Bing; Sun, Gui-Bo; Xiao, Jing; Chen, Rong-Chang; Wang, Xin; Wu, Ying; Cao, Li; Yang, Zhi-Hong; Sun, Xiao-Bo

    2012-02-01

    As a traditional Chinese medicine, the sea buckthorn (Hippophae rhamnoides L.) has a long history in the treatment of ischemic heart disease and circulatory disorders. However, the active compounds responsible for and the underlying mechanisms of these effects are not fully understood. In this article, isorhamnetin pretreatment counteracted H(2)O(2)-induced apoptotic damage in H9c2 cardiomyocytes. Isorhamnetin did not inhibit the death receptor-dependent or extrinsic apoptotic pathways, as characterized by its absence in both caspase-8 inactivation and tBid downregulation along with unchanged Fas and TNFR1 mRNA levels. Instead, isorhamnetin specifically suppressed the mitochondria-dependent or intrinsic apoptotic pathways, as characterized by inactivation of caspase-9 and -3, maintenance of the mitochondrial membrane potential (ΔΨm), and regulation of a series of Bcl-2 family genes upstream of ΔΨm. The anti-apoptotic effects of isorhamnetin were linked to decreased ROS generation. H(2)O(2) activated ERK and p53, whereas isorhamnetin inhibited their activation. ERK overexpression overrode the isorhamnetin-induced inhibition of the intrinsic apoptotic pathway in H9c2 cardiomyocytes, which indicated that an ERK-dependent pathway was involved. Furthermore, N-acetyl cysteine (a potent ROS scavenger) could attenuate the H(2)O(2)-induced apoptosis. However, PD98059 (an ERK-specific inhibitor) could not effectively antagonize ROS generation, which indicates that ROS may be an upstream inducer of ERK. In conclusion, isorhamnetin inhibits the H(2)O(2)-induced activation of the intrinsic apoptotic pathway via ROS scavenging and ERK inactivation. Therefore, isorhamnetin is a promising reagent for the treatment of ROS-induced cardiomyopathy.

  15. Purified Wnt-5a increases differentiation of midbrain dopaminergic cells and dishevelled phosphorylation.

    PubMed

    Schulte, Gunnar; Bryja, Vítezslav; Rawal, Nina; Castelo-Branco, Goncalo; Sousa, Kyle M; Arenas, Ernest

    2005-03-01

    The Wnt family of lipoproteins regulates several aspects of the development of the nervous system. Recently, we reported that Wnt-3a enhances the proliferation of midbrain dopaminergic precursors and that Wnt-5a promotes their differentiation into dopaminergic neurones. Here we report the purification of hemagglutinin-tagged Wnt-5a using a three-step purification method similar to that previously described for Wnt-3a. Haemagglutinin-tagged Wnt-5a was biologically active and induced the differentiation of immature primary midbrain precursors into tyrosine hydroxylase-positive dopaminergic neurones. Using a substantia nigra-derived dopaminergic cell line (SN4741), we found that Wnt-5a, unlike Wnt-3a, did not promote beta-catenin phosphorylation or stabilization. However, both Wnt-5a and Wnt-3a activated dishevelled, as assessed by a phosphorylation-dependent mobility shift. Moreover, the activity of Wnt-5a on dishevelled was blocked by pre-treatment with acyl protein thioesterase-1, indicating that palmitoylation of Wnt-5a is necessary for its function. Thus, our results suggest that Wnt-3a and Wnt-5a, respectively, activate canonical and non-canonical Wnt signalling pathways in ventral midbrain dopaminergic cells. Furthermore, we identify dishevelled as a key player in transducing both Wnt canonical and non-canonical signals in dopaminergic cells.

  16. Sprouty 2: a novel attenuator of B-cell receptor and MAPK-Erk signaling in CLL.

    PubMed

    Shukla, Ashima; Rai, Karan; Shukla, Vipul; Chaturvedi, Nagendra K; Bociek, R Gregory; Pirruccello, Samuel J; Band, Hamid; Lu, Runqing; Joshi, Shantaram S

    2016-05-12

    Clinical heterogeneity is a major barrier to effective treatment of chronic lymphocytic leukemia (CLL). Emerging evidence suggests that constitutive activation of various signaling pathways like mitogen-activated protein kinase-extracellular signal-regulated kinase (MAPK-Erk) signaling plays a role in the heterogeneous clinical outcome of CLL patients. In this study, we have investigated the role of Sprouty (SPRY)2 as a negative regulator of receptor and nonreceptor tyrosine kinase signaling in the pathogenesis of CLL. We show that SPRY2 expression is significantly decreased in CLL cells, particularly from poor-prognosis patients compared with those from good-prognosis patients. Overexpression of SPRY2 in CLL cells from poor-prognosis patients increased their apoptosis. Conversely, downregulation of SPRY2 in CLL cells from good-prognosis patients resulted in increased proliferation. Furthermore, CLL cells with low SPRY2 expression grew more rapidly in a xenograft model of CLL. Strikingly, B-cell-specific transgenic overexpression of spry2 in mice led to a decrease in the frequency of B1 cells, the precursors of CLL cells in rodents. Mechanistically, we show that SPRY2 attenuates the B-cell receptor (BCR) and MAPK-Erk signaling by binding to and antagonizing the activities of RAF1, BRAF, and spleen tyrosine kinase (SYK) in normal B cells and CLL cells. We also show that SPRY2 is targeted by microRNA-21, which in turn leads to increased activity of Syk and Erk in CLL cells. Taken together, these results establish SPRY2 as a critical negative regulator of BCR-mediated MAPK-Erk signaling in CLL, thereby providing one of the molecular mechanisms to explain the clinical heterogeneity of CLL. © 2016 by The American Society of Hematology.

  17. Hematopoietic stem cell fate decisions are regulated by Wnt antagonists: comparisons and current controversies.

    PubMed

    Cain, Corey J; Manilay, Jennifer O

    2013-01-01

    Wingless and int (Wnt) proteins are secreted proteins that are important for regulating hematopoietic stem cell self-renewal and differentiation in the bone marrow microenvironment in mice. The mechanisms by which Wnt signaling regulates these hematopoietic cell fate decisions are not fully understood. Secreted Wnt antagonists, which are expressed in bone and bone marrow stromal cells, either bind to Wnt ligands directly or block Wnt receptors and co-receptors to halt Wnt-mediated signal transduction in both osteolineage and hematopoietic cell types. Secreted frizzled related proteins-1 and -2, Wnt inhibitory factor-1, Dickkopf-1, and Sclerostin are Wnt antagonists that influence hematopoietic cell fate decisions in the bone marrow niche. In this review, we compare and contrast the roles of these Wnt antagonists and their effects on hematopoietic development in mice, and also discuss the clinical significance of targeting Wnt antagonists within the context of hematopoietic disease.

  18. Exploring Leptin Antagonism in Ophthalmic Cell Models

    PubMed Central

    Coroniti, Roberta; Otvos, Laszlo; Surmacz, Eva

    2013-01-01

    Background Emerging evidence suggests that angiogenic and pro-inflammatory cytokine leptin might be implicated in ocular neovascularization. However, the potential of inhibiting leptin function in ophthalmic cells has never been explored. Here we assessed mitogenic, angiogenic, and signaling leptin activities in retinal and corneal endothelial cells and examined the capability of a specific leptin receptor (ObR) antagonist, Allo-aca, to inhibit these functions. Methods and Results The experiments were carried out in monkey retinal (RF/6A) and bovine corneal (BCE) endothelial cells. Leptin at 50-250 ng/mL stimulated the growth of both cell lines in a dose-dependent manner. The maximal mitogenic response (35±7 and 27±3% in RF6A and BCE cells, respectively) was noted at 24 h of 250 ng/mL leptin treatments. Leptin-dependent proliferation was reduced to base levels with 10 and 100 nM Allo-aca in BCE and RF6A cells, respectively. In both cell lines, leptin promoted angiogenic responses, with the maximal increase in tube formation (163±10 and 133±8% in RF6A and BCE cultures, respectively) observed under a 250 ng/mL leptin treatment for 3 h. Furthermore, in both cell lines 250 ng/mL leptin modulated the activity or expression of several signaling molecules involved in proliferation, inflammatory activity and angiogenesis, such as STAT3, Akt, and ERK1/2, COX2, and NFκB. In both cell lines, leptin-induced angiogenic and signaling responses were significantly inhibited with 100 nM Allo-aca. We also found that leptin increased its own mRNA and protein expression in both cell lines, and this autocrine effect was abolished by 100-250 nM Allo-aca. Conclusions Our data provide new insights into the role of leptin in ocular endothelial cells and represent the first original report on targeting ObR in ophthalmic cell models. PMID:24098500

  19. Evolutionary Dynamics of the wnt Gene Family: A Lophotrochozoan Perspective

    PubMed Central

    Cho, Sung-Jin; Vallès, Yvonne; Giani, Vincent C.; Seaver, Elaine C.; Weisblat, David A.

    2010-01-01

    The wnt gene family encodes a set of secreted glycoproteins involved in key developmental processes, including cell fate specification and regulation of posterior growth (Cadigan KM, Nusse R. 1997. Wnt signaling: a common theme in animal development. Genes Dev. 11:3286–3305.; Martin BL, Kimelman D. 2009. Wnt signaling and the evolution of embryonic posterior development. Curr Biol. 19:R215–R219.). As for many other gene families, evidence for expansion and/or contraction of the wnt family is available from deuterostomes (e.g., echinoderms and vertebrates [Nusse R, Varmus HE. 1992. Wnt genes. Cell. 69:1073–1087.; Schubert M, Holland LZ, Holland ND, Jacobs DK. 2000. A phylogenetic tree of the Wnt genes based on all available full-length sequences, including five from the cephalochordate amphioxus. Mol Biol Evol. 17:1896–1903.; Croce JC, Wu SY, Byrum C, Xu R, Duloquin L, Wikramanayake AH, Gache C, McClay DR. 2006. A genome-wide survey of the evolutionarily conserved Wnt pathways in the sea urchin Strongylocentrotus purpuratus. Dev Biol. 300:121–131.]) and ecdysozoans (e.g., arthropods and nematodes [Eisenmann DM. 2005. Wnt signaling. WormBook. 1–17.; Bolognesi R, Farzana L, Fischer TD, Brown SJ. 2008. Multiple Wnt genes are required for segmentation in the short-germ embryo of Tribolium castaneum. Curr Biol. 18:1624–1629.]), but little is known from the third major bilaterian group, the lophotrochozoans (e.g., mollusks and annelids [Prud'homme B, Lartillot N, Balavoine G, Adoutte A, Vervoort M. 2002. Phylogenetic analysis of the Wnt gene family. Insights from lophotrochozoan members. Curr Biol. 12:1395.]). To obtain a more comprehensive scenario of the evolutionary dynamics of this gene family, we exhaustively mined wnt gene sequences from the whole genome assemblies of a mollusk (Lottia gigantea) and two annelids (Capitella teleta and Helobdella robusta) and examined them by phylogenetic, genetic linkage, intron–exon structure, and embryonic

  20. Wnt Signaling and Its Contribution to Craniofacial Tissue Homeostasis.

    PubMed

    Yin, X; Li, J; Salmon, B; Huang, L; Lim, W H; Liu, B; Hunter, D J; Ransom, R C; Singh, G; Gillette, M; Zou, S; Helms, J A

    2015-11-01

    A new field of dental medicine seeks to exploit nature's solution for repairing damaged tissues, through the process of regeneration. Most adult mammalian tissues have limited regenerative capacities, but in lower vertebrates, the molecular machinery for regeneration is an elemental part of their genetic makeup. Accumulating data suggest that the molecular pathways responsible for the regenerative capacity of teleosts, amphibians, and reptiles have fallen into disuse in mammals but that they can be "jumpstarted" by the selective activation of key molecules. The Wnt family of secreted proteins constitutes one such critical pathway: Wnt proteins rank among the most potent and ubiquitous stem cell self-renewing factors, with tremendous potential for promoting human tissue regeneration. Wnt reporter and lineage-tracing strains of mice have been employed to create molecular maps of Wnt responsiveness in the craniofacial tissues, and these patterns of Wnt signaling colocalize with stem/progenitor populations in the rodent incisor apex, the dental pulp, the alveolar bone, the periodontal ligament, the cementum, and oral mucosa. The importance of Wnt signaling in both the maintenance and healing of these craniofacial tissues is summarized, and the therapeutic potential of Wnt-based strategies to accelerate healing through activation of endogenous stem cells is highlighted.

  1. Canonical Wnt Signaling Regulates Atrioventricular Junction Programming and Electrophysiological Properties

    PubMed Central

    Gillers, Benjamin S; Chiplunkar, Aditi; Aly, Haytham; Valenta, Tomas; Basler, Konrad; Christoffels, Vincent M.; Efimov, Igor R; Boukens, Bastiaan J; Rentschler, Stacey

    2014-01-01

    Rationale Proper patterning of the atrioventricular canal (AVC) is essential for delay of electrical impulses between atria and ventricles, and defects in AVC maturation can result in congenital heart disease. Objective To determine the role of canonical Wnt signaling in the myocardium during AVC development. Methods and Results We utilized a novel allele of β-catenin that preserves β-catenin’s cell adhesive functions but disrupts canonical Wnt signaling, allowing us to probe the effects of Wnt loss of function independently. We show that loss of canonical Wnt signaling in the myocardium results in tricuspid atresia with hypoplastic right ventricle associated with loss of AVC myocardium. In contrast, ectopic activation of Wnt signaling was sufficient to induce formation of ectopic AV junction-like tissue as assessed by morphology, gene expression, and electrophysiologic criteria. Aberrant AVC development can lead to ventricular preexcitation, a characteristic feature of Wolff-Parkinson-White syndrome. We demonstrate that postnatal activation of Notch signaling downregulates canonical Wnt targets within the AV junction. Stabilization of β-catenin protein levels can rescue Notch-mediated ventricular preexcitation and dysregulated ion channel gene expression. Conclusions Our data demonstrate that myocardial canonical Wnt signaling is an important regulator of AVC maturation and electrical programming upstream of Tbx3. Our data further suggests that ventricular preexcitation may require both morphologic patterning defects, as well as myocardial lineage reprogramming, to allow robust conduction across accessory pathway tissue. PMID:25599332

  2. The complex roles of Wnt antagonists in RCC.

    PubMed

    Saini, Sharanjot; Majid, Shahana; Dahiya, Rajvir

    2011-10-25

    Renal cell carcinoma (RCC) is the most lethal of all the genitourinary cancers, as it is generally refractory to current treatment regimens, including chemotherapy and radiation therapy. Targeted therapies against critical signaling pathways associated with RCC pathogenesis, such as vascular endothelial growth factor, von Hippel-Lindau tumor suppressor and mammalian target of rapamycin, have shown limited efficacy so far. Thus, Wnt signaling, which is known to be intricately involved in the pathogenesis of RCC, has attracted much interest. Several Wnt signaling components have been examined in RCC, and, while studies suggest that Wnt signaling is constitutively active in RCC, the molecular mechanisms differ considerably from other human carcinomas. Increasing evidence indicates that secreted Wnt antagonists have important roles in RCC pathogenesis. Considering these vital roles, it has been postulated--and supported by experimental evidence--that the functional loss of Wnt antagonists, for example by promoter hypermethylation, can contribute to constitutive activation of the Wnt pathway, resulting in carcinogenesis through dysregulation of cell proliferation and differentiation. However, subsequent functional studies of these Wnt antagonists have demonstrated the inherent complexities underlying their role in RCC pathogenesis.

  3. Canonical wnt signaling regulates atrioventricular junction programming and electrophysiological properties.

    PubMed

    Gillers, Benjamin S; Chiplunkar, Aditi; Aly, Haytham; Valenta, Tomas; Basler, Konrad; Christoffels, Vincent M; Efimov, Igor R; Boukens, Bastiaan J; Rentschler, Stacey

    2015-01-30

    Proper patterning of the atrioventricular canal (AVC) is essential for delay of electrical impulses between atria and ventricles, and defects in AVC maturation can result in congenital heart disease. To determine the role of canonical Wnt signaling in the myocardium during AVC development. We used a novel allele of β-catenin that preserves β-catenin's cell adhesive functions but disrupts canonical Wnt signaling, allowing us to probe the effects of Wnt loss of function independently. We show that the loss of canonical Wnt signaling in the myocardium results in tricuspid atresia with hypoplastic right ventricle associated with the loss of AVC myocardium. In contrast, ectopic activation of Wnt signaling was sufficient to induce formation of ectopic AV junction-like tissue as assessed by morphology, gene expression, and electrophysiological criteria. Aberrant AVC development can lead to ventricular pre-excitation, a characteristic feature of Wolff-Parkinson-White syndrome. We demonstrate that postnatal activation of Notch signaling downregulates canonical Wnt targets within the AV junction. Stabilization of β-catenin protein levels can rescue Notch-mediated ventricular pre-excitation and dysregulated ion channel gene expression. Our data demonstrate that myocardial canonical Wnt signaling is an important regulator of AVC maturation and electric programming upstream of Tbx3. Our data further suggest that ventricular pre-excitation may require both morphological patterning defects, as well as myocardial lineage reprogramming, to allow robust conduction across accessory pathway tissue. © 2014 American Heart Association, Inc.

  4. SOX9 drives WNT pathway activation in prostate cancer

    PubMed Central

    Ma, Fen; Ye, Huihui; He, Housheng Hansen; Gerrin, Sean J.; Chen, Sen; Tanenbaum, Benjamin A.; Sowalsky, Adam G.; He, Lingfeng; Wang, Hongyun; Balk, Steven P.; Yuan, Xin

    2016-01-01

    The transcription factor SOX9 is critical for prostate development, and dysregulation of SOX9 is implicated in prostate cancer (PCa). However, the SOX9-dependent genes and pathways involved in both normal and neoplastic prostate epithelium are largely unknown. Here, we performed SOX9 ChIP sequencing analysis and transcriptome profiling of PCa cells and determined that SOX9 positively regulates multiple WNT pathway genes, including those encoding WNT receptors (frizzled [FZD] and lipoprotein receptor-related protein [LRP] family members) and the downstream β-catenin effector TCF4. Analyses of PCa xenografts and clinical samples both revealed an association between the expression of SOX9 and WNT pathway components in PCa. Finally, treatment of SOX9-expressing PCa cells with a WNT synthesis inhibitor (LGK974) reduced WNT pathway signaling in vitro and tumor growth in murine xenograft models. Together, our data indicate that SOX9 expression drives PCa by reactivating the WNT/β−catenin signaling that mediates ductal morphogenesis in fetal prostate and define a subgroup of patients who would benefit from WNT-targeted therapy. PMID:27043282

  5. RhoA Controls Wnt Upregulation on Microstructured Titanium Surfaces

    PubMed Central

    Mazzotta, Silvia; Piergianni, Maddalena; Piemontese, Marilina; Passeri, Giovanni

    2014-01-01

    Rough topography enhances the activation of Wnt canonical signaling in vitro, and this mediates its effects on cell differentiation. However, the molecular mechanisms underlying topography-dependent control of Wnt signaling are still poorly understood. As the small GTPase RhoA controls cytoskeletal reorganization and actomyosin-induced tensional forces, we hypothesized that RhoA could affect the activation of Wnt signaling in cells on micropatterned titanium surfaces. G-LISA assay revealed that RhoA activation was higher in C2C12 cells on rough (SLA) surfaces under basal conditions than on smooth (Polished) titanium. Transfection with dominant negative RhoA decreased Wnt activation by normalized TCF-Luc activity on SLA, whilst transfection with constitutively active RhoA increased TCF-Luc activation on Polished titanium. One mM Myosin II inhibitor Blebbistatin increased RhoA activation but decreased Wnt activation on SLA surfaces, indicating that tension-generating structures are required for canonical Wnt modulation on titanium surfaces. Actin inhibitor Cytochalasin markedly enhanced RhoA and TCF-Luc activation on both surfaces and increased the expression of differentiation markers in murine osteoblastic MC3T3 cells. Taken together, these data show that RhoA is upregulated in cells on rough surfaces and it affects the activation of Wnt canonical signaling through Myosin II modulation. PMID:24949442

  6. Targets of Wnt/ß-catenin transcription in penile carcinoma.

    PubMed

    Arya, Manit; Thrasivoulou, Christopher; Henrique, Rui; Millar, Michael; Hamblin, Ruth; Davda, Reena; Aare, Kristina; Masters, John R; Thomson, Calum; Muneer, Asif; Patel, Hitendra R H; Ahmed, Aamir

    2015-01-01

    Penile squamous cell carcinoma (PeCa) is a rare malignancy and little is known regarding the molecular mechanisms involved in carcinogenesis of PeCa. The Wnt signaling pathway, with the transcription activator ß-catenin as a major transducer, is a key cellular pathway during development and in disease, particularly cancer. We have used PeCa tissue arrays and multi-fluorophore labelled, quantitative, immunohistochemistry to interrogate the expression of WNT4, a Wnt ligand, and three targets of Wnt-ß-catenin transcription activation, namely, MMP7, cyclinD1 (CD1) and c-MYC in 141 penile tissue cores from 101 unique samples. The expression of all Wnt signaling proteins tested was increased by 1.6 to 3 fold in PeCa samples compared to control tissue (normal or cancer adjacent) samples (p<0.01). Expression of all proteins, except CD1, showed a significant decrease in grade II compared to grade I tumors. High magnification, deconvolved confocal images were used to measure differences in co-localization between the four proteins. Significant (p<0.04-0.0001) differences were observed for various permutations of the combinations of proteins and state of the tissue (control, tumor grades I and II). Wnt signaling may play an important role in PeCa and proteins of the Wnt signaling network could be useful targets for diagnosis and prognostic stratification of disease.

  7. Wnt signaling regulates pulp volume and dentin thickness

    PubMed Central

    Lim, Won Hee; Liu, Bo; Cheng, Du; Hunter, Daniel J; Zhong, Zhendong; Ramos, Daniel M; Williams, Bart O; Sharpe, Paul T; Bardet, Claire; Mah, Su-jung; Helms, Jill A

    2015-01-01

    Odontoblasts, cementoblasts, ameloblasts and osteoblasts all form mineralized tissues in the craniofacial complex, and all these cell types exhibit active Wnt signaling during postnatal life. We set out to understand the functions of this Wnt signaling, by evaluating the phenotypes of mice in which the essential Wnt chaperone protein, Wingless was eliminated. The deletion of Wls was restricted to cells expressing Osteocalcin, which in addition to osteoblasts includes odontoblasts, cementoblasts, and ameloblasts. Dentin, cementum, enamel, and bone all formed in OCN-Cre;Wlsfl/fl mice but their homeostasis was dramatically affected. The most notable feature was a significant increase in dentin volume and density. We attribute this gain in dentin volume to a Wnt-mediated mis-regulation of Runx2. Normally, Wnt signaling stimulates Runx2, which in turn inhibits DSP; this inhibition must be relieved for odontoblasts to differentiate. In OCN-Cre;Wlsfl/fl mice, Wnt pathway activation is reduced and Runx2 levels decline. The Runx2-mediated repression of DSP is relieved and odontoblast differentiation is accordingly enhanced. This study demonstrates the importance of Wnt signaling in the homeostasis of mineralized tissues of the craniofacial complex. PMID:23996396

  8. Targets of Wnt/ß-Catenin Transcription in Penile Carcinoma

    PubMed Central

    Henrique, Rui; Millar, Michael; Hamblin, Ruth; Davda, Reena; Aare, Kristina; Masters, John R.; Thomson, Calum; Muneer, Asif; Patel, Hitendra R. H.; Ahmed, Aamir

    2015-01-01

    Penile squamous cell carcinoma (PeCa) is a rare malignancy and little is known regarding the molecular mechanisms involved in carcinogenesis of PeCa. The Wnt signaling pathway, with the transcription activator ß-catenin as a major transducer, is a key cellular pathway during development and in disease, particularly cancer. We have used PeCa tissue arrays and multi-fluorophore labelled, quantitative, immunohistochemistry to interrogate the expression of WNT4, a Wnt ligand, and three targets of Wnt-ß-catenin transcription activation, namely, MMP7, cyclinD1 (CD1) and c-MYC in 141 penile tissue cores from 101 unique samples. The expression of all Wnt signaling proteins tested was increased by 1.6 to 3 fold in PeCa samples compared to control tissue (normal or cancer adjacent) samples (p<0.01). Expression of all proteins, except CD1, showed a significant decrease in grade II compared to grade I tumors. High magnification, deconvolved confocal images were used to measure differences in co-localization between the four proteins. Significant (p<0.04-0.0001) differences were observed for various permutations of the combinations of proteins and state of the tissue (control, tumor grades I and II). Wnt signaling may play an important role in PeCa and proteins of the Wnt signaling network could be useful targets for diagnosis and prognostic stratification of disease. PMID:25901368

  9. MKP-7, a JNK phosphatase, blocks ERK-dependent gene activation by anchoring phosphorylated ERK in the cytoplasm

    SciTech Connect

    Masuda, Kouhei; Katagiri, Chiaki; Nomura, Miyuki; Sato, Masami; Kakumoto, Kyoko; Akagi, Tsuyoshi; Kikuchi, Kunimi; Tanuma, Nobuhiro; Shima, Hiroshi

    2010-03-05

    MAPK phosphatase-7 (MKP-7) was identified as a JNK-specific phosphatase. However, despite its high specificity for JNK, MKP-7 interacts also with ERK. We previously showed that as a physiological consequence of their interaction, activated ERK phosphorylates MKP-7 at Ser-446, and stabilizing MKP-7. In the present study, we analyzed MKP-7 function in activation of ERK. A time-course experiment showed that both MKP-7 and its phosphatase-dead mutant prolonged mitogen-induced ERK phosphorylation, suggesting that MKP-7 functions as a scaffold for ERK. An important immunohistological finding was that nuclear translocation of phospho-ERK following PMA stimulation was blocked by co-expressed MKP-7 and, moreover, that phospho-ERK co-localized with MKP-7 in the cytoplasm. Reporter gene analysis indicated that MKP-7 blocks ERK-mediated transcription. Overall, our data indicate that MKP-7 down-regulates ERK-dependent gene expression by blocking nuclear accumulation of phospho-ERK.

  10. Quantifying ERK2-protein interactions by fluorescence anisotropy: PEA-15 inhibits ERK2 by blocking the binding of DEJL domains.

    PubMed

    Callaway, Kari; Rainey, Mark A; Dalby, Kevin N

    2005-12-30

    While mitogen-activated protein kinase signaling pathways constitute highly regulated networks of protein-protein interactions, little quantitative information for these interactions is available. Here we highlight recent fluorescence anisotropy binding studies that focus on the interactions of ERK1 and ERK2 with PEA-15 (antiapoptotic phosphoprotein enriched in astrocytes-15 kDa), a small protein that sequesters ERK2 in the cytoplasm. The regulation of ERK2 by PEA-15 is appraised in the light of a simple equilibrium-binding model for reversible ERK2 nucleoplasmic-cytoplasmic shuttling, which elaborates on the theory of Burack and Shaw (J. Biol. Chem. 280, 3832-3837; 2005). Also highlighted is the recent observation that the peptide N-QKGKPRDLELPLSPSL-C, derived from the docking site for ERK/JNK and LEL (DEJL) in Elk-1, displaces PEA-15 from ERK2. It is proposed that the C-terminus of PEA-15 ((121)LXLXXXXKK(129)) is a reverse DEJL domain [which has a general consensus of R/K-phi(A)-X(3/4)-phi(B), where phi(A) and phi(B) are hydrophobic residues (Leu, Ile, or Val)], which mediates one arm of a bidentate PEA-15 interaction with ERK2. The notion that PEA-15 is a potent inhibitor of many ERK2-mediated phosphorylations, by virtue of its ability to block ERK2-DEJL domain interactions, is proposed.

  11. Hypothalamic Wnt Signalling and its Role in Energy Balance Regulation.

    PubMed

    Helfer, G; Tups, A

    2016-03-01

    Wnt signalling and its downstream effectors are well known for their roles in embryogenesis and tumourigenesis, including the regulation of cell proliferation, survival and differentiation. In the nervous system, Wnt signalling has been described mainly during embryonic development, although accumulating evidence suggests that it also plays a major role in adult brain morphogenesis and function. Studies have predominantly concentrated on memory formation in the hippocampus, although recent data indicate that Wnt signalling is also critical for neuroendocrine control of the developed hypothalamus, a brain centre that is key in energy balance regulation and whose dysfunction is implicated in metabolic disorders such as type 2 diabetes and obesity. Based on scattered findings that report the presence of Wnt molecules in the tanycytes and ependymal cells lining the third ventricle and arcuate nucleus neurones of the hypothalamus, their potential importance in key regions of food intake and body weight regulation has been investigated in recent studies. The present review brings together current knowledge on Wnt signalling in the hypothalamus of adult animals and discusses the evidence suggesting a key role for members of the Wnt signalling family in glucose and energy balance regulation in the hypothalamus in diet-induced and genetically obese (leptin deficient) mice. Aspects of Wnt signalling in seasonal (photoperiod sensitive) rodents are also highlighted, given the recent evidence indicating that the Wnt pathway in the hypothalamus is not only regulated by diet and leptin, but also by photoperiod in seasonal animals, which is connected to natural adaptive changes in food intake and body weight. Thus, Wnt signalling appears to be critical as a modulator for normal functioning of the physiological state in the healthy adult brain, and is also crucial for normal glucose and energy homeostasis where its dysregulation can lead to a range of metabolic disorders. © 2016

  12. Concentration-dependent effects of WNTLESS on WNT1/3A signaling

    PubMed Central

    Galli, Lisa M.; Szabo, Linda A.; Li, Lydia; Htaik, Yin Min; Onguka, Ouma; Burrus, Laura W.

    2014-01-01

    Background WNTLESS (WLS) is a multi-transmembrane protein that transports Wnt ligands from the Golgi to the cell surface. Although WLS loss-of-function experiments in the developing central nervous system reveal phenotypes consistent with defects in WNT1 and WNT3A signaling, data from complementary gain-of-function experiments have not yet been reported. Here, we report the phenotypic consequences of WLS overexpression in cultured cells and in the developing chick spinal cord. Results Overexpression of small amounts of WLS along with either WNT1 or WNT3A promotes the Wnt/β-catenin pathway in HEK293T cells, while overexpression of higher levels of WLS inhibits the Wnt/β-catenin pathway in these cells. Similarly, overexpressed WLS inhibits the Wnt/β-catenin pathway in the developing spinal cord, as assessed by cell proliferation and specification. These effects appear to be Wnt-specific as overexpression of WLS inhibits the expression of FZD10, a target of β-catenin-dependent transcription. Conclusion Our results show that overexpression of WLS inhibits Wnt/β-catenin signaling in the spinal cord. As the activation of the Wnt/β-catenin pathway in the spinal cord requires WNT1 or WNT3A, our results are consistent with a model in which the relative concentration of WLS to Wnt regulates WNT1/3A signaling in the developing spinal cord. PMID:24866848

  13. Experimental inhibition of porcupine mediated Wnt O-acylation attenuates kidney fibrosis

    PubMed Central

    Madan, Babita; Patel, Mehul; Zhang, Jiandong; Bunte, Ralph; Rudemiller, Nathan P.; Griffiths, Robert; Virshup, David M; Crowley, Steven D

    2016-01-01

    Activated Wnt signaling is critical in the pathogenesis of renal fibrosis, a final common pathway for most forms of chronic kidney disease. Therapeutic intervention by inhibition of individual Wnts or downstream Wnt/β-catenin signaling has been proposed, but these approaches do not interrupt the functions of all Wnts nor block non-canonical Wnt signaling pathways. Alternatively, an orally bioavailable small molecule, Wnt-C59, blocks the catalytic activity of the Wnt-acyl transferase porcupine, and thereby prevents secretion of all Wnt isoforms. We found that inhibiting porcupine dramatically attenuates kidney fibrosis in the murine unilateral ureteral obstruction model. Wnt-C59 treatment similarly blunts collagen mRNA expression in the obstructed kidney. Consistent with its actions to broadly arrest Wnt signaling, porcupine inhibition reduces expression of Wnt target genes and bolsters nuclear exclusion of β-catenin in the kidney following ureteral obstruction. Importantly, prevention of Wnt secretion by Wnt-C59 blunts expression of inflammatory cytokines in the obstructed kidney that otherwise provoke a positive feedback loop of Wnt expression in collagen-producing fibroblasts and epithelial cells. Thus, therapeutic targeting of porcupine abrogates kidney fibrosis not only by overcoming the redundancy of individual Wnt isoforms but also by preventing upstream cytokine-induced Wnt generation. These findings reveal a novel therapeutic maneuver to protect the kidney from fibrosis by interrupting a pathogenic crosstalk loop between locally generated inflammatory cytokines and the Wnt/β-catenin signaling pathway. PMID:27083283

  14. Experimental inhibition of porcupine-mediated Wnt O-acylation attenuates kidney fibrosis.

    PubMed

    Madan, Babita; Patel, Mehul B; Zhang, Jiandong; Bunte, Ralph M; Rudemiller, Nathan P; Griffiths, Robert; Virshup, David M; Crowley, Steven D

    2016-05-01

    Activated Wnt signaling is critical in the pathogenesis of renal fibrosis, a final common pathway for most forms of chronic kidney disease. Therapeutic intervention by inhibition of individual Wnts or downstream Wnt/β-catenin signaling has been proposed, but these approaches do not interrupt the functions of all Wnts nor block non-canonical Wnt signaling pathways. Alternatively, an orally bioavailable small molecule, Wnt-C59, blocks the catalytic activity of the Wnt-acyl transferase porcupine, and thereby prevents secretion of all Wnt isoforms. We found that inhibiting porcupine dramatically attenuates kidney fibrosis in the murine unilateral ureteral obstruction model. Wnt-C59 treatment similarly blunts collagen mRNA expression in the obstructed kidney. Consistent with its actions to broadly arrest Wnt signaling, porcupine inhibition reduces expression of Wnt target genes and bolsters nuclear exclusion of β-catenin in the kidney following ureteral obstruction. Importantly, prevention of Wnt secretion by Wnt-C59 blunts expression of inflammatory cytokines in the obstructed kidney that otherwise provoke a positive feedback loop of Wnt expression in collagen-producing fibroblasts and epithelial cells. Thus, therapeutic targeting of porcupine abrogates kidney fibrosis not only by overcoming the redundancy of individual Wnt isoforms but also by preventing upstream cytokine-induced Wnt generation. These findings reveal a novel therapeutic maneuver to protect the kidney from fibrosis by interrupting a pathogenic crosstalk loop between locally generated inflammatory cytokines and the Wnt/β-catenin signaling pathway.

  15. Wnt therapy for bone loss: golden goose or Trojan horse?

    PubMed

    Enders, Greg H

    2009-04-01

    The Wnt pathway has been found to play a role in the development of many tissues and to spur growth and differentiation of adult osteoblasts, sparking interest in its potential clinical application for bone growth. However, when deregulated, this pathway can be oncogenic in some tissues. In this issue of the JCI, Kansara and colleagues reveal that Wnt inhibitory factor 1 is epigenetically silenced in human osteosarcomas and that its absence augments osteosarcoma formation in mice (see the related article beginning on page 837). These observations suggest the need for caution in stimulating the Wnt pathway for therapeutic bone growth.

  16. PTK7 Faces the Wnt in Development and Disease

    PubMed Central

    Berger, Hanna; Wodarz, Andreas; Borchers, Annette

    2017-01-01

    PTK7 (protein tyrosine kinase 7) is an evolutionarily conserved transmembrane receptor regulating various processes in embryonic development and tissue homeostasis. On a cellular level PTK7 affects the establishment of cell polarity, the regulation of cell movement and migration as well as cell invasion. The PTK7 receptor has been shown to interact with ligands, co-receptors, and intracellular transducers of Wnt signaling pathways, pointing to a function in the fine-tuning of the Wnt signaling network. Here we will review recent findings implicating PTK7 at the crossroads of Wnt signaling pathways in development and disease. PMID:28424771

  17. Teaching resource. Canonical Wnt/beta-catenin signaling.

    PubMed

    Moon, Randall T

    2004-06-29

    This animation provides an interactive presentation of the Wnt signaling pathway as it may occur in multiple cell types. This animation would be useful in teaching developmental biology, immunology, and cell signaling courses. Activation of Wnt pathways can modulate cell proliferation, cell survival, cell behavior, and cell fate. In the basal, unstimulated state in the absence of ligand, there is a constitutively active kinase, which phosphorylates target proteins, resulting in their degradation. Thus, the presence of the ligand Wnt inactivates the kinase allowing accumulation of beta-catenin, which then translocates to the nucleus and acts as a transcriptional regulator.

  18. Wnt therapy for bone loss: golden goose or Trojan horse?

    PubMed Central

    Enders, Greg H.

    2009-01-01

    The Wnt pathway has been found to play a role in the development of many tissues and to spur growth and differentiation of adult osteoblasts, sparking interest in its potential clinical application for bone growth. However, when deregulated, this pathway can be oncogenic in some tissues. In this issue of the JCI, Kansara and colleagues reveal that Wnt inhibitory factor 1 is epigenetically silenced in human osteosarcomas and that its absence augments osteosarcoma formation in mice (see the related article beginning on page 837). These observations suggest the need for caution in stimulating the Wnt pathway for therapeutic bone growth. PMID:19348043

  19. Antagonism by d,1-propranolol of imipramine effects in mice.

    PubMed

    Souto, M; Francès, H; Lecrubier, Y; Puech, A J; Simon, P

    1979-11-23

    Three agents with known or suspected antidepressant activity, imipramine, salbutamol and dexamphetamine, were active in animal tests predictive of an antidepressant effect in man: antagonism of the hypothermia induced by reserpine, by oxotremorine or by a high dose of apomorphine, and the potentiation of the yohimbine-induced toxicity. These effects were antagonized by d,1-propranolol, suggesting that the stimulation of beta-adrenergic receptors could be a common mechanism underlying their effects. These results agree with the noradrenergic hypothesis of the pathophysiology of affective disorders.

  20. Deferoxamine alleviates chronic hydrocephalus after intraventricular hemorrhage through iron chelation and Wnt1/Wnt3a inhibition.

    PubMed

    Meng, Hui; Li, Fei; Hu, Rong; Yuan, Yikai; Gong, Guoqi; Hu, Shengli; Feng, Hua

    2015-03-30

    Post-hemorrhagic chronic hydrocephalus (PHCH) is a common complication after intraventricular hemorrhage (IVH). The mechanism of PHCH is not fully understood, and its treatment is relatively difficult. In the present study, a rat model of PHCH was used to elucidate the role of iron in the pathogenesis of PHCH. The action of deferoxamine (DFX) in IVH-induced PHCH, the expression of brain ferritin, the concentration of iron in cerebrospinal fluid (CSF), and changes in Wnt1/Wnt3a gene expression were determined. Results indicate that iron plays an important role in the occurrence of hydrocephalus after IVH. The iron chelator, DFX, can decrease the concentrations of iron and ferritin after cerebral hemorrhage and can thereby decrease the incidence of hydrocephalus. In addition, after IVH, the gene expression of Wnt1 and Wnt3a was enhanced, with protein expression also upregulated; DFX was able to suppress both gene and protein expression of Wnt1 and Wnt3a in brain tissue. This indicates that iron may be the key stimulus that activates the Wnt signaling pathway and regulates subarachnoid fibrosis after cerebral hemorrhage, and that DFX may be a candidate for preventing PHCH in patients with IVH.

  1. Isolation and characterization of WNT8B, a novel human Wnt gene that maps to 10q24

    SciTech Connect

    Lako, Majlinda; Strachan, T.; Curtis, A.R.J.; Lindsay, S.

    1996-07-15

    Wnt genes encode intercellular signalling molecules that play important roles in key processes of embryonic development such as mesoderm induction, specification of the embryonic axis, and patterning of the central nervous system, spinal cord, and limb. Multiple such genes are known to exist in each of several species that have been investigated, and they have been classified into various groups and subgroups on the basis of high sequence homology and common expression patterns. The vertebrate Wnt8 subfamily includes genes from Xenopus, zebrafish, and chicken, but, to data, no mammalian homologues have been described. We now report cloning and characterization of a novel human member of this family that we have termed WNT8B on the basis of high sequence homology and common expression patterns. The vertebrate Wnt8 subfamily includes genes from Xenopus, zebrafish, and chicken, but, to date no mammalian homologues have been described. We now report cloning and characterization of a novel human member of this family that we have termed WNT8B on the basis of the very high sequence similarity of the inferred protein to those encoded by the Xenopus and zebrafish Wnt8b genes. PCR typing of a human monochromosomal hybrid cell panel mapped the gene to chromosome 10, and FISH mapping provided a subchromosomal location at 10q24. Northern blotting and RT-PCR assays indicated that the WNT8B gene is expressed in several human tissues during fetal and adult stages. 14 refs., 2 figs.

  2. The Wnt gatekeeper SFRP4 modulates EMT, cell migration and downstream Wnt signalling in serous ovarian cancer cells.

    PubMed

    Ford, Caroline E; Jary, Eve; Ma, Sean Si Qian; Nixdorf, Sheri; Heinzelmann-Schwarz, Viola A; Ward, Robyn L

    2013-01-01

    Aberrant Wnt signalling is implicated in numerous human cancers, and understanding the effects of modulation of pathway members may lead to the development of novel therapeutics. Expression of secreted frizzled related protein 4 (SFRP4), an extracellular modulator of the Wnt signalling pathway, is progressively lost in more aggressive ovarian cancer phenotypes. Here we show that recombinant SFRP4 (rSFRP4) treatment of a serous ovarian cancer cell line results in inhibition of β-catenin dependent Wnt signalling as measured by TOP/FOP Wnt reporter assay and decreased transcription of Wnt target genes, Axin2, CyclinD1 and Myc. In addition, rSFRP4 treatment significantly increased the ability of ovarian cancer cells to adhere to collagen and fibronectin, and decreased their ability to migrate across an inflicted wound. We conclude that these changes in cell behaviour may be mediated via mesenchymal to epithelial transition (MET), as rSFRP4 treatment also resulted in increased expression of the epithelial marker E-cadherin, and reduced expression of Vimentin and Twist. Combined, these results indicate that modulation of a single upstream gatekeeper of Wnt signalling can have effects on downstream Wnt signalling and ovarian cancer cell behaviour, as mediated through epithelial to mesenchymal plasticity (EMP). This raises the possibility that SFRP4 may be used both diagnostically and therapeutically in epithelial ovarian cancer.

  3. HPV-16 E6 promotes cell growth of esophageal cancer via downregulation of miR-125b and activation of Wnt/β-catenin signaling pathway.

    PubMed

    Zang, Bao; Huang, Guojin; Wang, Xiaowei; Zheng, Shiying

    2015-01-01

    High-risk human papillomavirus (HPV) is a possible cause of esophageal cancer. However, the molecular pathogenesis of HPV-infected esophageal cancer remains unclear. The expression levels of some microRNAs including miR-125b have been negatively correlated with HPV infection, and miR-125b downregulation is associated with tumorigenesis. In addition, Wnt/β-catenin signaling pathway has been suggested to play an important role in esophageal cancer (EC). We examined miR-125b and Wnt/β-catenin signaling pathway in HPV-16 E6 promoted tumor progression in EC. HPV-16 E6 transfection decreased markedly the expression levels of miR-125b and promoted the colony formation in the Eca 109 and Kyse 150 cell lines, and restoration of miR-125b expression level antagonized the increased colony formation in HPV-16 E6 transfected cell lines. We also demonstrated that overexpression of E6 upregulated the Wnt/β-catenin signaling activity via modulating the multiple regulators including TLE1, GSK3β, and sFRP4. Overexpression of miR-125b restored the expression levels of these proteins. Expression of miR-125b was lower in HPV-16 E6 positive esophageal cancer tissues, and was negatively correlated with E6 mRNA levels. Our results indicate that HPV-16 E6 promotes tumorigenesis in EC via down-regulation of miR-125b, and this underlying mechanism may be involved in the activation of the Wnt/β-catenin signaling pathway.

  4. Honokiol abrogates leptin-induced tumor progression by inhibiting Wnt1-MTA1-β-catenin signaling axis in a microRNA-34a dependent manner.

    PubMed

    Avtanski, Dimiter B; Nagalingam, Arumugam; Kuppusamy, Panjamurthy; Bonner, Michael Y; Arbiser, Jack L; Saxena, Neeraj K; Sharma, Dipali

    2015-06-30

    Obesity greatly influences risk, progression and prognosis of breast cancer. As molecular effects of obesity are largely mediated by adipocytokine leptin, finding effective novel strategies to antagonize neoplastic effects of leptin is desirable to disrupt obesity-cancer axis. Present study is designed to test the efficacy of honokiol (HNK), a bioactive polyphenol from Magnolia grandiflora, against oncogenic actions of leptin and systematically elucidate the underlying mechanisms. Our results show that HNK significantly inhibits leptin-induced breast-cancer cell-growth, invasion, migration and leptin-induced breast-tumor-xenograft growth. Using a phospho-kinase screening array, we discover that HNK inhibits phosphorylation and activation of key molecules of leptin-signaling-network. Specifically, HNK inhibits leptin-induced Wnt1-MTA1-β-catenin signaling in vitro and in vivo. Finally, an integral role of miR-34a in HNK-mediated inhibition of Wnt1-MTA1-β-catenin axis was discovered. HNK inhibits Stat3 phosphorylation, abrogates its recruitment to miR-34a promoter and this release of repressor-Stat3 results in miR-34a activation leading to Wnt1-MTA1-β-catenin inhibition. Accordingly, HNK treatment inhibited breast tumor growth in diet-induced-obese mouse model (exhibiting high leptin levels) in a manner associated with activation of miR-34a and inhibition of MTA1-β-catenin. These data provide first in vitro and in vivo evidence for the leptin-antagonist potential of HNK revealing a crosstalk between HNK and miR34a and Wnt1-MTA1-β-catenin axis.

  5. Honokiol abrogates leptin-induced tumor progression by inhibiting Wnt1-MTA1-β-catenin signaling axis in a microRNA-34a dependent manner

    PubMed Central

    Avtanski, Dimiter B.; Nagalingam, Arumugam; Kuppusamy, Panjamurthy; Bonner, Michael Y.; Arbiser, Jack L.; Saxena, Neeraj K.; Sharma, Dipali

    2015-01-01

    Obesity greatly influences risk, progression and prognosis of breast cancer. As molecular effects of obesity are largely mediated by adipocytokine leptin, finding effective novel strategies to antagonize neoplastic effects of leptin is desirable to disrupt obesity-cancer axis. Present study is designed to test the efficacy of honokiol (HNK), a bioactive polyphenol from Magnolia grandiflora, against oncogenic actions of leptin and systematically elucidate the underlying mechanisms. Our results show that HNK significantly inhibits leptin-induced breast-cancer cell-growth, invasion, migration and leptin-induced breast-tumor-xenograft growth. Using a phospho-kinase screening array, we discover that HNK inhibits phosphorylation and activation of key molecules of leptin-signaling-network. Specifically, HNK inhibits leptin-induced Wnt1-MTA1-β-catenin signaling in vitro and in vivo. Finally, an integral role of miR-34a in HNK-mediated inhibition of Wnt1-MTA1-β-catenin axis was discovered. HNK inhibits Stat3 phosphorylation, abrogates its recruitment to miR-34a promoter and this release of repressor-Stat3 results in miR-34a activation leading to Wnt1-MTA1-β-catenin inhibition. Accordingly, HNK treatment inhibited breast tumor growth in diet-induced-obese mouse model (exhibiting high leptin levels) in a manner associated with activation of miR-34a and inhibition of MTA1-β-catenin. These data provide first in vitro and in vivo evidence for the leptin-antagonist potential of HNK revealing a crosstalk between HNK and miR34a and Wnt1-MTA1-β-catenin axis. PMID:26036628

  6. Frequent up-regulation of WNT5A mRNA in primary gastric cancer.

    PubMed

    Saitoh, Tetsuroh; Mine, Tetsuya; Katoh, Masaru

    2002-05-01

    WNT signal is transduced to the beta-catenin - TCF pathway, the JNK pathway, or the Ca2+-releasing pathway through seven-transmembrane-type WNT receptors encoded by Frizzled genes (FZD1-FZD10). We have previously cloned and characterized human WNT2B/WNT13, WNT3, WNT3A, WNT5B, WNT6, WNT7B, WNT8A, WNT8B, WNT10A, WNT10B, WNT11, WNT14, and WNT14B/WNT15 by using bioinformatics, cDNA-library screening, and cDNA-PCR. Here, we investigated expression of human WNT5A mRNA in various normal tissues, 66 primary tumors derived from various tissues, and 15 human cancer cell lines. WNT5A mRNA was relatively highly expressed in salivary gland, bladder, uterus, placenta, and fetal kidney. Up-regulation of WNT5A mRNA was detected in 5 out of 8 cases of primary gastric cancer, 5 out of 18 cases of primary colorectal tumors, and in 2 out of 7 cases of primary uterus tumors by using matched tumor/normal expression array analysis. Up-regulation of WNT5A mRNA was also detected in 7 out of 10 other cases of primary gastric cancer by using cDNA-PCR. Although low-level expression of WNT5A mRNA was detected in gastric cancer cell line MKN45, WNT5A mRNA was almost undetectable in gastric cancer cell lines OKAJIMA, TMK1, MKN7, MKN28, MKN74, and KATO-III. Compared with frequent up-regulation of WNT5A mRNA in primary gastric cancer, expression levels of WNT5A mRNA in 7 gastric cancer cell lines were significantly lower than that in normal stomach. Frequent up-regulation of WNT5A mRNA in human primary gastric cancer might be due to cancer-stromal interaction.

  7. Specific functions for ERK/MAPK signaling during PNS development

    PubMed Central

    Newbern, Jason M.; Li, Xiaoyan; Shoemaker, Sarah E.; Zhou, Jiang; Zhong, Jian; Wu, Yaohong; Bonder, Daniel; Hollenback, Steven; Coppola, Giovanni; Geschwind, Daniel H.; Landreth, Gary E.; Snider, William D.

    2011-01-01

    We have established functions of the stimulus dependent MAPKs, ERK1/2 and ERK5 in DRG, motor neuron, and Schwann cell development. Surprisingly, many aspects of early DRG and motor neuron development were found to be ERK1/2 independent and Erk5 deletion had no obvious effect on embryonic PNS. In contrast, Erk1/2 deletion in developing neural crest resulted in peripheral nerves that were devoid of Schwann cell progenitors, and deletion of Erk1/2 in Schwann cell precursors caused disrupted differentiation and marked hypomyelination of axons. The Schwann cell phenotypes are similar to those reported in neuregulin-1 and ErbB mutant mice and neuregulin effects could not be elicited in glial precursors lacking Erk1/2. ERK/MAPK regulation of myelination was specific to Schwann cells, as deletion in oligodendrocyte precursors did not impair myelin formation, but reduced precursor proliferation. Our data suggest a tight linkage between developmental functions of ERK/MAPK signaling and biological actions of specific RTK-activating factors. PMID:21220101

  8. ERK5 Activation Is Essential for Osteoclast Differentiation

    PubMed Central

    Amano, Shigeru; Chang, Yu-Tzu; Fukui, Yasuhisa

    2015-01-01

    The MEK/ERK pathways are critical for controlling cell proliferation and differentiation. In this study, we show that the MEK5/ERK5 pathway participates in osteoclast differentiation. ERK5 was activated by M-CSF, which is one of the essential factors in osteoclast differentiation. Inhibition of MEK5 by BIX02189 or inhibition of ERK5 by XMD 8-92 blocked osteoclast differentiation. MEK5 knockdown inhibited osteoclast differentiation. RAW264.7D clone cells, which are monocytic cells, differentiate into osteoclasts after stimulation with sRANKL. ERK5 was activated without any stimulation in these cells. Inhibition of the MEK5/ERK5 pathway by the inhibitors also blocked the differentiation of RAW264.7D cells into osteoclasts. Moreover, expression of the transcription factor c-Fos, which is indispensable for osteoclast differentiation, was inhibited by treatment with MEK5 or ERK5 inhibitors. Therefore, activation of ERK5 is required for the induction of c-Fos. These events were confirmed in experiments using M-CSF-dependent bone marrow macrophages. Taken together, the present results show that activation of the MEK5/ERK5 pathway with M-CSF is required for osteoclast differentiation, which may induce differentiation through the induction of c-Fos. PMID:25885811

  9. Modeling Wnt/β-Catenin Target Gene Expression in APC and Wnt Gradients Under Wild Type and Mutant Conditions

    PubMed Central

    Benary, Uwe; Kofahl, Bente; Hecht, Andreas; Wolf, Jana

    2013-01-01

    The Wnt/β-catenin pathway is involved in the regulation of a multitude of physiological processes by controlling the differential expression of target genes. In certain tissues such as the adult liver, the Wnt/β-catenin pathway can attain different levels of activity due to gradients of Wnt ligands and/or intracellular pathway components like APC. How graded pathway activity is converted into regionally distinct patterns of Wnt/β-catenin target gene expression is largely unknown. Here, we apply a mathematical modeling approach to investigate the impact of different regulatory mechanisms on target gene expression within Wnt or APC concentration gradients. We develop a minimal model of Wnt/β-catenin signal transduction and combine it with various mechanisms of target gene regulation. In particular, the effects of activation, inhibition, and an incoherent feedforward loop (iFFL) are compared. To specify activation kinetics, we analyze experimental data that quantify the response of β-catenin/TCF reporter constructs to different Wnt concentrations, and demonstrate that the induction of these constructs occurs in a cooperative manner with Hill coefficients between 2 and 5. In summary, our study shows that the combination of specific gene regulatory mechanisms with a time-independent gradient of Wnt or APC is sufficient to generate distinct target gene expression patterns as have been experimentally observed in liver. We find that cooperative gene activation in combination with a TCF feedback can establish sharp borders of target gene expression in Wnt or APC gradients. In contrast, the iFFL renders gene expression independent of gradients of the upstream signaling components. Our subsequent analysis of carcinogenic pathway mutations reveals that their impact on gene expression is determined by the gene regulatory mechanism and the APC concentration of the cell in which the mutation occurs. PMID:23508686

  10. Wnt5a mediated canonical Wnt signaling pathway activation in orthodontic tooth movement: possible role in the tension force-induced bone formation.

    PubMed

    Fu, Hai-Di; Wang, Bei-Ke; Wan, Zi-Qiu; Lin, Heng; Chang, Mao-Lin; Han, Guang-Li

    2016-10-01

    Orthodontic tooth movement (OTM) is associated with bone remodeling mediated by orthodontic mechanical loading. Increasing studies reported that Wnt signaling played crucial roles in mechanical stimuli induced bone remodeling. However, little is known about the involvement of Wnt signaling in orthodontic force-induced bone formation during OTM. In virtue of the OTM mice model as we previously reported, where new bone formation was determined by micro-CT and immunoreactivity of osteocalcin and osterix, we explored the activation of Wnt signaling pathway during OTM. Our results proved the nuclei translocation of β-catenin, suggesting the activation of canonical Wnt signaling pathway in the periodontal ligament cells (PDLCs) near the alveolar bone at the tension site (TS). Moreover, the immunoreactivity of Wnt5a, but not Wnt3a in PDLCs indicated the activation of canonical Wnt pathway might be mediated by Wnt5a, but not Wnt3a as in most cases. The co-location of Wnt5a and β-catenin that was evidenced by double labeling immunofluorescence staining further supported the hypothesis. In addition, the high expression of FZD4 and LRP5 in PDLCs at TS of periodontium suggested that the activation of Wnt signaling pathway was mediated by these receptors. The negligible expression of ROR2 also indicated that canonical but not non-canonical Wnt signaling pathway was activated by Wnt5a, since previous studies demonstrated that the activation of canonical/non-canonical Wnt signaling pathway was largely dependent on the receptors. In summary, we here reported that Wnt5a mediated activation of canonical Wnt signaling pathway might contribute to the orthodontic force induced bone remodeling.

  11. Wnt5a is required for endothelial differentiation of embryonic stem cells and vascularization via pathways involving both Wnt/beta-catenin and protein kinase Calpha.

    PubMed

    Yang, Dong-Hwa; Yoon, Ju-Young; Lee, Soung-Hoon; Bryja, Vitezslav; Andersson, Emma R; Arenas, Ernest; Kwon, Young-Guen; Choi, Kang-Yell

    2009-02-13

    In this study, we examined the signaling pathways activated by Wnt5a in endothelial differentiation of embryonic stem (ES) cells and the function of Wnt5a during vascular development. We first found that Wnt5a(-/-) mouse embryonic stem (mES) cells exhibited a defect in endothelial differentiation, which was rescued by addition of Wnt5a, suggesting that Wnt5a is required for endothelial differentiation of ES cells. Involvement of both beta-catenin and protein kinase (PK)Calpha pathways in endothelial differentiation of mES cells requiring Wnt5a was indicated by activation of both beta-catenin and PKCalpha in Wnt5a(+/-) but not in Wnt5a(-/-) mES cells. We also found that beta-catenin or PKCalpha knockdowns inhibited the Wnt5a-induced endothelial differentiation of ES cells. Moreover, the lack of endothelial differentiation of Wnt5a(-/-) mES cells was rescued only by transfection of both beta-catenin and PKCalpha, indicating that both genes are required for Wnt5a-mediated endothelial differentiation. Wnt5a was also found to be essential for the differentiation of mES cells into immature endothelial progenitor cells, which are known to play a role in repair of damaged endothelium. Furthermore, a defect in the vascularization of the neural tissue was detected at embryonic day 14.5 in Wnt5a(-/-) mice, implicating Wnt5a in vascular development in vivo. Thus, we conclude that Wnt5a is involved in the endothelial differentiation of ES cells via both Wnt/beta-catenin and PKC signaling pathways and regulates embryonic vascular development.

  12. Tripeptidyl Peptidase II Mediates Levels of Nuclear Phosphorylated ERK1 and ERK2*

    PubMed Central

    Wiemhoefer, Anne; Stargardt, Anita; van der Linden, Wouter A.; Renner, Maria C.; van Kesteren, Ronald E.; Stap, Jan; Raspe, Marcel A.; Tomkinson, Birgitta; Kessels, Helmut W.; Ovaa, Huib; Overkleeft, Herman S.; Florea, Bogdan; Reits, Eric A.

    2015-01-01

    Tripeptidyl peptidase II (TPP2) is a serine peptidase involved in various biological processes, including antigen processing, cell growth, DNA repair, and neuropeptide mediated signaling. The underlying mechanisms of how a peptidase can influence this multitude of processes still remain unknown. We identified rapid proteomic changes in neuroblastoma cells following selective TPP2 inhibition using the known reversible inhibitor butabindide, as well as a new, more potent, and irreversible peptide phosphonate inhibitor. Our data show that TPP2 inhibition indirectly but rapidly decreases the levels of active, di-phosphorylated extracellular signal-regulated kinase 1 (ERK1) and ERK2 in the nucleus, thereby down-regulating signal transduction downstream of growth factors and mitogenic stimuli. We conclude that TPP2 mediates many important cellular functions by controlling ERK1 and ERK2 phosphorylation. For instance, we show that TPP2 inhibition of neurons in the hippocampus leads to an excessive strengthening of synapses, indicating that TPP2 activity is crucial for normal brain function. PMID:26041847

  13. Mapping the dynamic expression of Wnt11 and the lineage contribution of Wnt11-expressing cells during early mouse development

    PubMed Central

    Sinha, Tanvi; Lin, Lizhu; Li, Ding; Davis, Jennifer; Evans, Sylvia; Wynshaw-Boris, Anthony; Wang, Jianbo

    2015-01-01

    Planar cell polarity (PCP) signaling is an evolutionarily conserved mechanism that coordinates polarized cell behavior to regulate tissue morphogenesis during vertebrate gastrulation, neurulation and organogenesis. In Xenopus and zebrafish, PCP signaling is activated by non-canonical Wnts such as Wnt11, and detailed understanding of Wnt11 expression has provided important clues on when, where and how PCP may be activated to regulate tissue morphogenesis. To explore the role of Wnt11 in mammalian development, we established a Wnt11 expression and lineage map with high spatial and temporal resolution by creating and analyzing a tamoxifen-inducible Wnt11-CreER BAC (bacterial artificial chromosome) transgenic mouse line. Our short- and long-term lineage tracing experiments indicated that Wnt11-CreER could faithfully recapitulate endogenous Wnt11 expression, and revealed for the first time that cells transiently expressing Wnt11 at early gastrulation were fated to become specifically the progenitors of the entire endoderm. During mid-gastrulation, Wnt11-CreER expressing cells also contribute extensively to the endothelium in both embryonic and extraembryonic compartments, and the endocardium in all chambers of the developing heart. In contrast, Wnt11-CreER expression in the myocardium starts from late-gastrulation, and occurs in three transient, sequential waves: first in the precursors of the left ventricular (LV) myocardium from E7.0 to 8.0; subsequently in the right ventricular (RV) myocardium from E8.0 to 9.0; and finally in the superior wall of the outflow tract (OFT) myocardium from E8.5 to 10.5. These results provide formal genetic proof that the majority of the endocardium and myocardium diverge by mid-gastrulation in the mouse, and suggest a tight spatial and temporal control of Wnt11 expression in the myocardial lineage to coordinate with myocardial differentiation in the first and second heart field progenitors to form the LV, RV and OFT. The insights gained

  14. Wnt10b Activates the Wnt, Notch and NFκB Pathways in U2OS Osteosarcoma Cells

    PubMed Central

    Mödder, Ulrike I.; Oursler, Merry Jo; Khosla, Sundeep; Monroe, David G.

    2011-01-01

    Although osteosarcoma represents the most common bone malignancy, the molecular and cellular mechanisms influencing its pathogenesis have remained elusive. Recent evidence has suggested that the Wnt signaling pathway may play a crucial role in osteosarcoma. This study employed a microarray approach to discover novel genes and pathways involved in Wnt signaling in osteosarcoma. We developed a Wnt10b-expressing cell line using the human U2OS osteosarcoma model (U2OS-Wnt10b) and performed microarray and pathway analyses using parental U2OS cells as control. Differential expression of 1003 genes encompassing 28 pathways was noted. The Wnt, NFκB and Notch pathways were chosen for further study based on their known importance in bone biology. Known Wnt-responsive genes Axin-2 (4.9-fold), CD44 (2.1-fold), endothelin-1 (4.2-fold) and sclerostin domain containing-1 (43-fold) were regulated by Wnt10b. The proinflammatory cytokines interleukin-1α and tumor necrosis factor-α, known inducers of NFκB, were upregulated both at the transcript and protein level, and NFκB reporter activity was stimulated 3.8-fold, confirming NFκB activation. Interestingly, genes involved in Notch signaling [Notch-1 (2.4-fold) and Jagged-1 (3.1-fold)] were upregulated, whereas the Notch inhibitor, lunatic fringe, was downregulated (8.2-fold). This resulted in the activation of the classic Notch-responsive genes, hairy and enhancer of split-1 (Hes-1; 2.2-fold) and hairy/enhancer-of-split related with YRPW motif-1 (Hey-1; 2.5-fold). A Hey-1 reporter construct was regulated 9.1-fold in U2OS-Wnt10b cells, confirming Notch activation. Interestingly, Wnt3a failed to induce the Notch and NFκB pathways, demonstrating Wnt-specificity. In conclusion, our data demonstrate that Wnt10b, but not Wnt3a, stimulates the NFκB and Notch pathways in U2OS osteosarcoma cells. PMID:21321991

  15. THE MAPK ERK5, BUT NOT ERK1/2, INHIBITS THE PROGRESSION OF MONOCYTIC PHENOTYPE TO THE FUNCTIONING MACROPHAGE

    PubMed Central

    Wang, Xuening; Pesakhov, Stella; Harrison, Jonathan S; Kafka, Michael; Danilenko, Michael; Studzinski, George P

    2014-01-01

    Intracellular signaling pathways present targets for pharmacological agents with potential for treatment of neoplastic diseases, with some disease remissions already recorded. However, cellular compensatory mechanisms usually negate the initial success. For instance, attempts to interrupt aberrant signaling downstream of the frequently mutated ras by inhibiting ERK1/2 has shown only limited usefulness for cancer therapy. Here, we examined how ERK5, that overlaps the functions of ERK1/2 in cell proliferation and survival, functions in a manner distinct from ERK1/2 in human AML cells induced to differentiate by 1,25D-dihydroxyvitamin D3 (1,25D). Using inhibitors of ERK1/2 and of MEK5/ERK5 at concentrations specific for each kinase in HL60 and U937 cells, we observed that selective inhibition of the kinase activity of ERK5, but not of ERK1/2, in the presence of 1,25D resulted in macrophage-like cell morphology and enhancement of phagocytic activity. Importantly, this was associated with increased expression of the macrophage colony stimulating factor receptor (M-CSFR), but was not seen when M-CSFR expression was knocked down. Interestingly, inhibition of ERK1/2 led to activation of ERK5 in these cells. Our results support the hypothesis that ERK5 negatively regulates the expression of M-CSFR, and thus has a restraining function on macrophage differentiation. The addition of pharmacological inhibitors of ERK5 may influence trials of differentiation therapy of AML. PMID:25447310

  16. Nucleophosmin leukemogenic mutant activates Wnt signaling during zebrafish development

    PubMed Central

    Barbieri, Elisa; Deflorian, Gianluca; Pezzimenti, Federica; Valli, Debora; Saia, Marco; Meani, Natalia

    2016-01-01

    Nucleophosmin (NPM1) is a ubiquitous multifunctional phosphoprotein with both oncogenic and tumor suppressor functions. Mutations of the NPM1 gene are the most frequent genetic alterations in acute myeloid leukemia (AML) and result in the expression of a mutant protein with aberrant cytoplasmic localization, NPMc+. Although NPMc+ causes myeloproliferation and AML in animal models, its mechanism of action remains largely unknown. Here we report that NPMc+ activates canonical Wnt signaling during the early phases of zebrafish development and determines a Wnt-dependent increase in the number of progenitor cells during primitive hematopoiesis. Coherently, the canonical Wnt pathway is active in AML blasts bearing NPMc+ and depletion of the mutant protein in the patient derived OCI-AML3 cell line leads to a decrease in the levels of active β-catenin and of Wnt target genes. Our results reveal a novel function of NPMc+ and provide insight into the molecular pathogenesis of AML bearing NPM1 mutations. PMID:27486814

  17. Different requirements for Wnt signaling in tongue myogenic subpopulations.

    PubMed

    Zhong, Z; Zhao, H; Mayo, J; Chai, Y

    2015-03-01

    The tongue is a muscular organ that is essential in vertebrates for important functions, such as food intake and communication. Little is known about regulation of myogenic progenitors during tongue development when compared with the limb or trunk region. In this study, we investigated the relationship between different myogenic subpopulations and the function of canonical Wnt signaling in regulating these subpopulations. We found that Myf5- and MyoD-expressing myogenic subpopulations exist during embryonic tongue myogenesis. In the Myf5-expressing myogenic progenitors, there is a cell-autonomous requirement for canonical Wnt signaling for cell migration and differentiation. In contrast, the MyoD-expressing subpopulation does not require canonical Wnt signaling during tongue myogenesis. Taken together, our results demonstrate that canonical Wnt signaling differentially regulates the Myf5- and MyoD-expressing subpopulations during tongue myogenesis. © International & American Associations for Dental Research 2015.

  18. Wnt-Dependent Control of Cell Polarity in Cultured Cells.

    PubMed

    Runkle, Kristin B; Witze, Eric S

    2016-01-01

    The secreted ligand Wnt5a regulates cell polarity and polarized cell movement during development by signaling through the poorly defined noncanonical Wnt pathway. Cell polarity regulates most aspects of cell behavior including the organization of apical/basolateral membrane domains of epithelial cells, polarized cell divisions along a directional plane, and front rear polarity during cell migration. These characteristics of cell polarity allow coordinated cell movements required for tissue formation and organogenesis during embryonic development. Genetic model organisms have been used to identify multiple signaling pathways including Wnt5a that are required to establish cell polarity and regulate polarized cell behavior. However, the downstream signaling events that regulate these complex cellular processes are still poorly understood. The methods below describe assays to study Wnt5a-induced cell polarity in cultured cells, which may facilitate our understanding of these complex signaling pathways.

  19. Small molecule modulators of Wnt/β-catenin signaling.

    PubMed

    Mook, Robert A; Chen, Minyong; Lu, Jiuyi; Barak, Larry S; Lyerly, H Kim; Chen, Wei

    2013-04-01

    The Wnt signal transduction pathway is dysregulated in many highly prevalent diseases, including cancer. Unfortunately, drug discovery efforts have been hampered by the paucity of targets and drug-like lead molecules amenable to drug discovery. Recently, we reported the FDA-approved anthelmintic drug Niclosamide inhibits Wnt/β-catenin signaling by a unique mechanism, though the target responsible remains unknown. We interrogated the mechanism and structure-activity relationships to understand drivers of potency and to assist target identification efforts. We found inhibition of Wnt signaling by Niclosamide appears unique among the structurally-related anthelmintic agents tested and found the potency and functional response was dependent on small changes in the chemical structure of Niclosamide. Overall, these findings support efforts to identify the target of Niclosamide inhibition of Wnt/β-catenin signaling and the discovery of potent and selective modulators to treat human disease.

  20. Wnt family proteins are secreted and associated with the cell surface.

    PubMed Central

    Smolich, B D; McMahon, J A; McMahon, A P; Papkoff, J

    1993-01-01

    Members of the Wnt gene family are proposed to function in both normal development and differentiation as well as in mammary tumorigenesis. To understand the function of Wnt proteins in these two processes, we present here a biochemical characterization of seven Wnt family members. For these studies, AtT-20 cells, a neuroendocrine cell line previously shown to efficiently process and secrete Wnt-1, was transfected with expression vectors encoding Wnt family members. All of the newly characterized Wnt proteins are glycosylated, secreted proteins that are tightly associated with the cell surface or extracellular matrix. We have also identified native Wnt proteins in retinoic acid-treated P19 embryonal carcinoma cells, and they exhibit the same biochemical characteristics as the recombinant proteins. These data suggest that Wnt family members function in cell to cell signaling in a fashion similar to Wnt-1. Images PMID:8167409

  1. Canonical WNT signaling pathway and human AREG.

    PubMed

    Katoh, Yuriko; Katoh, Masaru

    2006-06-01

    AREG (Amphiregulin), BTC (beta-cellulin), EGF, EPGN (Epigen), EREG (Epiregulin), HBEGF, NRG1, NRG2, NRG3, NRG4 and TGFA (TGFalpha) constitute EGF family ligands for ERBB family receptors. Cetuximab (Erbitux), Pertuzumab (Omnitarg) and Trastuzumab (Herceptin) are anti-cancer drugs targeted to EGF family ligands, while Gefitinib (Iressa), Erlotinib (Tarceva) and Lapatinib (GW572016) are anti-cancer drugs targeted to ERBB family receptors. AREG and TGFA are biomarkers for Gefitinib non-responders. The TCF/LEF binding sites within the promoter region of human EGF family members were searched for by using bioinformatics and human intelligence (Humint). Because three TCF/LEF-binding sites were identified within the 5'-promoter region of human AREG gene, comparative genomics analyses on AREG orthologs were further performed. The EPGN-EREG-AREG-BTC cluster at human chromosome 4q13.3 was linked to the PPBP-CXCL segmental duplicons. AREG was the paralog of HBEGF at human chromosome 5q31.2. Chimpanzee AREG gene, consisting of six exons, was located within NW_105918.1 genome sequence. Chimpanzee AREG was a type I transmembrane protein showing 98.0% and 71.4% total amino-acid identity with human AREG and mouse Areg, respectively. Three TCF/LEF-binding sites within human AREG promoter were conserved in chimpanzee AREG promoter, but not in rodent Areg promoters. Primate AREG promoters were significantly divergent from rodent Areg promoters. AREG mRNA was expressed in a variety of human tumors, such as colorectal cancer, liver cancer, gastric cancer, breast cancer, prostate cancer, esophageal cancer and myeloma. Because human AREG was characterized as potent target gene of WNT/beta-catenin signaling pathway, WNT signaling activation could lead to Gefitinib resistance through AREG upregulation. AREG is a target of systems medicine in the field of oncology.

  2. Effect of Wnt3a on Keratinocytes Utilizing in Vitro and Bioinformatics Analysis

    PubMed Central

    Nam, Ju-Suk; Chakraborty, Chiranjib; Sharma, Ashish Ranjan; Her, Young; Bae, Kee-Jeong; Sharma, Garima; Doss, George Priya; Lee, Sang-Soo; Hong, Myung-Sun; Song, Dong-Keun

    2014-01-01

    Wingless-type (Wnt) signaling proteins participate in various cell developmental processes. A suppressive role of Wnt5a on keratinocyte growth has already been observed. However, the role of other Wnt proteins in proliferation and differentiation of keratinocytes remains unknown. Here, we investigated the effects of the Wnt ligand, Wnt3a, on proliferation and differentiation of keratinocytes. Keratinocytes from normal human skin were cultured and treated with recombinant Wnt3a alone or in combination with the inflammatory cytokine, tumor necrosis factor α (TNFα). Furthermore, using bioinformatics, we analyzed the biochemical parameters, molecular evolution, and protein–protein interaction network for the Wnt family. Application of recombinant Wnt3a showed an anti-proliferative effect on keratinocytes in a dose-dependent manner. After treatment with TNFα, Wnt3a still demonstrated an anti-proliferative effect on human keratinocytes. Exogenous treatment of Wnt3a was unable to alter mRNA expression of differentiation markers of keratinocytes, whereas an altered expression was observed in TNFα-stimulated keratinocytes. In silico phylogenetic, biochemical, and protein–protein interaction analysis showed several close relationships among the family members of the Wnt family. Moreover, a close phylogenetic and biochemical similarity was observed between Wnt3a and Wnt5a. Finally, we proposed a hypothetical mechanism to illustrate how the Wnt3a protein may inhibit the process of proliferation in keratinocytes, which would be useful for future researchers. PMID:24686518

  3. WNT2 Promotes Cervical Carcinoma Metastasis and Induction of Epithelial-Mesenchymal Transition

    PubMed Central

    Xu, Jing; Zhang, Lan; Wang, Jianhua; Huang, Long; Huang, Shuting; Yuan, Linjing; Jia, Weihua; Yu, Xingjuan; Luo, Rongzhen; Zheng, Min

    2016-01-01

    Background Previously, we found an 11-gene signature could predict pelvic lymph node metastasis (PLNM), and WNT2 is one of the key genes in the signature. This study explored the expression and underlying mechanism of WNT2 in PLNM of cervical cancer. Methods WNT2 expression level in cervical cancer was detected using western blotting, quantitative PCR, and immunohistochemistry. Two WNT2-specific small interfering RNAs (siRNAs) were used to explore the effects of WNT2 on invasive and metastatic ability of cancer cells, and to reveal the possible mechanism of WNT2 affecting epithelial—mesenchymal transition (EMT). The correlation between WNT2 expression and PLNM was further investigated in clinical cervical specimens. Results Both WNT2 mRNA and protein expression was upregulated in cervical cancer. High WNT2 expression was significantly associated with tumor size, lymphovascular space involvement, positive parametrium, and most importantly, PLNM. PLNM and WNT2 expression were independent prognostic factors for overall survival and disease-free survival. WNT2 knockdown inhibited SiHa cell motility and invasion and reversed EMT by inhibiting the WNT2/β-catenin pathway. WNT2 overexpression in cervical cancer was associated with β-catenin activation and induction of EMT, which further contributed to metastasis in cervical cancer. Conclusion WNT2 might be a novel predictor of PLNM and a promising prognostic indicator in cervical cancer. PMID:27513465

  4. Hippocampal Wnt3a is Necessary and Sufficient for Contextual Fear Memory Acquisition and Consolidation

    PubMed Central

    Xu, Ning; Zhou, Wen-Juan; Wang, Yue; Huang, Shu-Hong; Li, Xian; Chen, Zhe-Yu

    2015-01-01

    The Wnt signaling pathway plays critical roles in development. However, to date, the role of Wnts in learning and memory in adults is still not well understood. Here, we aimed to investigate the roles and mechanisms of Wnts in hippocampal-dependent contextual fear conditioning (CFC) memory formation in adult mice. CFC training induced the secretion and expression of Wnt3a and the activation of its downstream Wnt/Ca2+ and Wnt/β-catenin signaling pathways in the dorsal hippocampus (DH). Intrahippocampal infusion of Wnt3a antibody impaired CFC acquisition and consolidation, but not expression. Using the Wnt antagonist sFRP1 or the canonical Wnt inhibitor Dkk1, we found that Wnt/Ca2+ and Wnt/β-catenin signaling pathways were involved in acquisition and consolidation, respectively. Moreover, we found Wnt3a signaling is not only necessary but also sufficient for CFC memory. Intrahippocampal infusion of exogenous Wnt3a could enhance acquisition and consolidation of CFC. Overexpression of constitutively active β-catenin in the DH could rescue the deficit in CFC memory consolidation, but not acquisition induced by Wnt3a antibody injection, which suggests β-catenin signaling pathway acts downstream of Wnt3a to mediate CFC memory consolidation. Our study may help further the understanding of the precise regulation of Wnt3a in differential memory phases depending on divergent signaling pathways. PMID:24904070

  5. AOP description: ER antagonism leading to reproductive dysfunction (in fish)

    EPA Science Inventory

    This adverse outcome pathway details the linkage between antagonism of estrogen receptor in females and the adverse effect of reduced cumulative fecundity in repeat-spawning fish species. Cumulative fecundity is the most apical endpoint considered in the OECD 229 Fish Short Term ...

  6. Novel pharmacological approaches for the antagonism of neuromuscular blockade.

    PubMed

    Pic, Lisa C

    2005-02-01

    Gamma cyclodextrin and purified plasma cholinesterase are 2 novel pharmacological agents being investigated as to their suitability for antagonism of neuromuscular blockade. Both of these agents are devoid of cholinergic stimulation and the accompanying side effects because their action is independent of acetylcholinesterase inhibition. Gamma cyclodextrin antagonizes the steroidal neuromuscular blocker rocuronium via the chemical encapsulation of the molecule forming a "host-guest" complex through van der Waals and hydrophobic interactions in the plasma. Encapsulation decreases plasma drug concentrations, shifting the neuromuscular blocking drug molecules from the neuromuscular junction back to the plasma compartment resulting in a rapid recovery of the neuromuscular function. Org 25969, a modified gamma cyclodextrin, will antagonize profound neuromuscular block induced by rocuronium in approximately 2 minutes. A commercial preparation of purified human plasma cholinesterase has been shown to be effective in reversing succinylcholine or mivacurium-induced block. Administration of exogenous plasma cholinesterase also has been shown to be effective in antagonizing mivacurium-induced neuromuscular block, cocaine toxicity, and organophosphate poisoning.

  7. ANDROGEN RECEPTOR ANTAGONISM BY THE ORGANOPHOSPHATE INSECTICIDE FENITROTHION

    EPA Science Inventory

    Androgen receptor antagonism by the organophosphate insecticide fenitrothion. Tamura, H., Maness, S.C., Reischmann, K. Dorman, D.C., Gray, L.E., and Gaido, K.W. (2000). Toxicol. Sci.

    Organophosphate insecticides represent one of the most widely used classes of pesticide...

  8. The mechanism of cyanide intoxication and its antagonism.

    PubMed

    Way, J L; Leung, P; Cannon, E; Morgan, R; Tamulinas, C; Leong-Way, J; Baxter, L; Nagi, A; Chui, C

    1988-01-01

    The mechanism of cyanide intoxication has been attributed to the inhibition of cytochrome oxidase, thereby decreasing the tissue utilization of oxygen. One mechanism of cyanide antagonism is by sequestering cyanide with methaemoglobin to form cyanmethaemoglobin and another mechanism is detoxifying with a sulphur donor to thiocyanate. Questions have been raised with regard to these classical mechanisms. Oxygen with nitrite-thiosulphate antagonizes the lethal effects of cyanide. Theoretically, increased oxygen should serve no useful purpose, as it is the tissue utilization of oxygen which is inhibited. In the nitrite-thiosulphate antidotal combination, the proposal is made that the predominate antidotal action of nitrite is a vasogenic action, rather than methaemoglobin formation, because when methaemoglobin formation is inhibited by methylene blue the protective action of sodium nitrite persists. This suggests that methaemoglobin formation plays only a small part, if any, in the therapeutic antagonism of the lethal effects of cyanide. The roles and implications of sodium thiosulphate and non-rhodanese substrates in the detoxification mechanism are compared. Lastly, a new approach to cyanide antagonism has been initiated which involves the erythrocyte encapsulation of thiosulphate and sulphurtransferase as an antidote and prophylaxis against cyanide.

  9. AOP description: ER antagonism leading to reproductive dysfunction (in fish)

    EPA Science Inventory

    This adverse outcome pathway details the linkage between antagonism of estrogen receptor in females and the adverse effect of reduced cumulative fecundity in repeat-spawning fish species. Cumulative fecundity is the most apical endpoint considered in the OECD 229 Fish Short Term ...

  10. ANDROGEN RECEPTOR ANTAGONISM BY THE ORGANOPHOSPHATE INSECTICIDE FENITROTHION

    EPA Science Inventory

    Androgen receptor antagonism by the organophosphate insecticide fenitrothion. Tamura, H., Maness, S.C., Reischmann, K. Dorman, D.C., Gray, L.E., and Gaido, K.W. (2000). Toxicol. Sci.

    Organophosphate insecticides represent one of the most widely used classes of pesticide...

  11. Determine the Role of Canonical Wnt Signaling in Ovarian Tumorigenesis

    DTIC Science & Technology

    2015-12-01

    unlabeled FK and GFP-labeled WT ESCs into the right or left testis of the same non -obese diabetic (NOD)/severe combined immunodeficiency (SCID) mouse...synthetic lethality 1. Introduction The Switch/Sucrose Non -Fermentable (SWI/SNF) complex regulates gene transcription through its ATP- dependent ...developing cancer therapeutics. Wnt5a is a non - canonical Wnt ligand that plays a context- dependent role in human cancers. Recently, we discovered a lower

  12. Prognostic significance of WNT signaling in pancreatic ductal adenocarcinoma.

    PubMed

    Nakamoto, Mitsuhiro; Matsuyama, Atsuji; Shiba, Eisuke; Shibuya, Ryo; Kasai, Takahiko; Yamaguchi, Koji; Hisaoka, Masanori

    2014-10-01

    Pancreatic ductal adenocarcinoma (PDA) is one of the most lethal human malignancies and is associated with a variety of molecular abnormalities. Although WNT signaling through its canonical/non-canonical pathways is one of the major factors involved in oncogenesis or progression of PDA, the prognostic significance of WNT signaling still remains poorly investigated. In this study, the status of the WNT signaling pathways was immunohistochemically analyzed in 101 PDAs, and its potential association with patient postoperative survival was assessed. Nuclear expression of beta-catenin, a hallmark of the activated canonical pathway, was identified in 59 cases, and was associated with reduced survival compared to the patients lacking nuclear beta-catenin expression (P = 0.002). In contrast, activation of the non-canonical pathway (25 cases), as indicated by co-expression of WNT2/5a and nuclear NFATc1, was not correlated with reduced survival (P = 0.268). Co-activation of both pathways (16 cases) was associated with worse prognosis in comparison with cases with an activated non-canonical pathway (P = 0.034). In addition, nuclear beta-catenin expression was an independent unfavorable prognostic factor (P = 0.006). Our data indicate that activated WNT signaling through its canonical pathway has a significantly negative effect on the clinical course of PDA, and the canonical WNT pathway should be considered as a future therapeutic target for PDA.

  13. Two FGFRL-Wnt circuits organize the planarian anteroposterior axis.

    PubMed

    Scimone, M Lucila; Cote, Lauren E; Rogers, Travis; Reddien, Peter W

    2016-04-11

    How positional information instructs adult tissue maintenance is poorly understood. Planarians undergo whole-body regeneration and tissue turnover, providing a model for adult positional information studies. Genes encoding secreted and transmembrane components of multiple developmental pathways are predominantly expressed in planarian muscle cells. Several of these genes regulate regional identity, consistent with muscle harboring positional information. Here, single-cell RNA-sequencing of 115 muscle cells from distinct anterior-posterior regions identified 44 regionally expressed genes, including multiple Wnt and ndk/FGF receptor-like (ndl/FGFRL) genes. Two distinct FGFRL-Wnt circuits, involving juxtaposed anterior FGFRL and posterior Wnt expression domains, controlled planarian head and trunk patterning. ndl-3 and wntP-2 inhibition expanded the trunk, forming ectopic mouths and secondary pharynges, which independently extended and ingested food. fz5/8-4 inhibition, like that of ndk and wntA, caused posterior brain expansion and ectopic eye formation. Our results suggest that FGFRL-Wnt circuits operate within a body-wide coordinate system to control adult axial positioning.

  14. Wnt/β-catenin Inhibits Dental Pulp Stem Cell Differentiation

    PubMed Central

    Scheller, E.L.; Chang, J.; Wang, C.Y.

    2010-01-01

    Dental pulp stem cells (DPSCs) are a unique precursor population isolated from post-natal human dental pulp and have the ability to regenerate a reparative dentin-like complex. Canonical Wnt signaling plays a critical role in tooth development and stem cell self-renewal through β-catenin. In this study, the regulation of odontoblast-like differentiation of DPSCs by canonical Wnt signaling was examined. DPSCs were stably transduced with canonical Wnt-1 or the active form of β-catenin, with retrovirus-mediated infection. Northern blot analysis found that Wnt-1 strongly induced the expression of matricellular protein osteopontin, and modestly enhanced the expression of type I collagen in DPSCs. Unexpectedly, Wnt-1 inhibited alkaline phosphatase (ALP) activity and the formation of mineralized nodules in DPSCs. Moreover, over-expression of β-catenin was also sufficient to suppress the differentiation and mineralization of DPSCs. In conclusion, our results suggest that canonical Wnt signaling negatively regulates the odontoblast-like differentiation of DPSCs. PMID:18218837

  15. WNT/β-Catenin Signaling in Vertebrate Eye Development

    PubMed Central

    Fujimura, Naoko

    2016-01-01

    The vertebrate eye is a highly specialized sensory organ, which is derived from the anterior neural plate, head surface ectoderm, and neural crest-derived mesenchyme. The single central eye field, generated from the anterior neural plate, divides to give rise to the optic vesicle, which evaginates toward the head surface ectoderm. Subsequently, the surface ectoderm, in conjunction with the optic vesicle invaginates to form the lens vesicle and double-layered optic cup, respectively. This complex process is controlled by transcription factors and several intracellular and extracellular signaling pathways including WNT/β-catenin signaling. This signaling pathway plays an essential role in multiple developmental processes and has a profound effect on cell proliferation and cell fate determination. During eye development, the activity of WNT/β-catenin signaling is tightly controlled. Faulty regulation of WNT/β-catenin signaling results in multiple ocular malformations due to defects in the process of cell fate determination and differentiation. This mini-review summarizes recent findings on the role of WNT/β-catenin signaling in eye development. Whilst this mini-review focuses on loss-of-function and gain-of-function mutants of WNT/β-catenin signaling components, it also highlights some important aspects of β-catenin-independent WNT signaling in the eye development at later stages. PMID:27965955

  16. Monitoring Wnt/β-Catenin Signaling in Skin

    PubMed Central

    Ku, Amy T.; Miao, Qi; Nguyen, Hoang

    2017-01-01

    Wnt signaling through β-catenin plays a crucial role in skin development and homeostasis. Disruption or hyperactivation of this pathway results in skin defects and diseases (Lim and Nusse, Cold Spring Harb Perspect Biol 5(2), 2013). Monitoring Wnt signaling in skin under normal and abnormal conditions is therefore critical to understand the role of this pathway in development and homeostasis. In this chapter, we provide methods to detect Wnt/β-catenin (canonical) signaling in the skin. We present a comprehensive list of Wnt reporter mice and detail the processing of skin tissue to detect reporter genes. From this list, we focus on the three most recent lines that, according to reports, are the most sensitive in skin. Additionally, we describe a protocol to detect nuclear β-catenin, a hallmark of active Wnt signaling, although this technique should be used with caution due to its limited sensitivity. The techniques outlined below will be useful for detecting active Wnt signaling in skin. PMID:27590159

  17. Two FGFRL-Wnt circuits organize the planarian anteroposterior axis

    PubMed Central

    Scimone, M Lucila; Cote, Lauren E; Rogers, Travis; Reddien, Peter W

    2016-01-01

    How positional information instructs adult tissue maintenance is poorly understood. Planarians undergo whole-body regeneration and tissue turnover, providing a model for adult positional information studies. Genes encoding secreted and transmembrane components of multiple developmental pathways are predominantly expressed in planarian muscle cells. Several of these genes regulate regional identity, consistent with muscle harboring positional information. Here, single-cell RNA-sequencing of 115 muscle cells from distinct anterior-posterior regions identified 44 regionally expressed genes, including multiple Wnt and ndk/FGF receptor-like (ndl/FGFRL) genes. Two distinct FGFRL-Wnt circuits, involving juxtaposed anterior FGFRL and posterior Wnt expression domains, controlled planarian head and trunk patterning. ndl-3 and wntP-2 inhibition expanded the trunk, forming ectopic mouths and secondary pharynges, which independently extended and ingested food. fz5/8-4 inhibition, like that of ndk and wntA, caused posterior brain expansion and ectopic eye formation. Our results suggest that FGFRL-Wnt circuits operate within a body-wide coordinate system to control adult axial positioning. DOI: http://dx.doi.org/10.7554/eLife.12845.001 PMID:27063937

  18. Endodermal Wnt signaling is required for tracheal cartilage formation.

    PubMed

    Snowball, John; Ambalavanan, Manoj; Whitsett, Jeffrey; Sinner, Debora

    2015-09-01

    Tracheobronchomalacia is a common congenital defect in which the walls of the trachea and bronchi lack of adequate cartilage required for support of the airways. Deletion of Wls, a cargo receptor mediating Wnt ligand secretion, in the embryonic endoderm using ShhCre mice inhibited formation of tracheal-bronchial cartilaginous rings. The normal dorsal-ventral patterning of tracheal mesenchyme was lost. Smooth muscle cells, identified by Acta2 staining, were aberrantly located in ventral mesenchyme of the trachea, normally the region of Sox9 expression in cartilage progenitors. Wnt/β-catenin activity, indicated by Axin2 LacZ reporter, was decreased in tracheal mesenchyme of Wls(f/f);Shh(Cre/+) embryos. Proliferation of chondroblasts was decreased and reciprocally, proliferation of smooth muscle cells was increased in Wls(f/f);Shh(Cre/+) tracheal tissue. Expression of Tbx4, Tbx5, Msx1 and Msx2, known to mediate cartilage and muscle patterning, were decreased in tracheal mesenchyme of Wls(f/f);Shh(Cre/+) embryos. Ex vivo studies demonstrated that Wnt7b and Wnt5a, expressed by the epithelium of developing trachea, and active Wnt/β-catenin signaling are required for tracheal chondrogenesis before formation of mesenchymal condensations. In conclusion, Wnt ligands produced by the tracheal epithelium pattern the tracheal mesenchyme via modulation of gene expression and cell proliferation required for proper tracheal cartilage and smooth muscle differentiation.

  19. WNT signaling drives cholangiocarcinoma growth and can be pharmacologically inhibited.

    PubMed

    Boulter, Luke; Guest, Rachel V; Kendall, Timothy J; Wilson, David H; Wojtacha, Davina; Robson, Andrew J; Ridgway, Rachel A; Samuel, Kay; Van Rooijen, Nico; Barry, Simon T; Wigmore, Stephen J; Sansom, Owen J; Forbes, Stuart J

    2015-03-02

    Cholangiocarcinoma (CC) is typically diagnosed at an advanced stage and is refractory to surgical intervention and chemotherapy. Despite a global increase in the incidence of CC, little progress has been made toward the development of treatments for this cancer. Here we utilized human tissue; CC cell xenografts; a p53-deficient transgenic mouse model; and a non-transgenic, chemically induced rat model of CC that accurately reflects both the inflammatory and regenerative background associated with human CC pathology. Using these systems, we determined that the WNT pathway is highly activated in CCs and that inflammatory macrophages are required to establish this WNT-high state in vivo. Moreover, depletion of macrophages or inhibition of WNT signaling with one of two small molecule WNT inhibitors in mouse and rat CC models markedly reduced CC proliferation and increased apoptosis, resulting in tumor regression. Together, these results demonstrate that enhanced WNT signaling is a characteristic of CC and suggest that targeting WNT signaling pathways has potential as a therapeutic strategy for CC.

  20. WNT signaling drives cholangiocarcinoma growth and can be pharmacologically inhibited

    PubMed Central

    Boulter, Luke; Guest, Rachel V.; Kendall, Timothy J.; Wilson, David H.; Wojtacha, Davina; Robson, Andrew J.; Ridgway, Rachel A.; Samuel, Kay; Van Rooijen, Nico; Barry, Simon T.; Wigmore, Stephen J.; Sansom, Owen J.; Forbes, Stuart J.

    2015-01-01

    Cholangiocarcinoma (CC) is typically diagnosed at an advanced stage and is refractory to surgical intervention and chemotherapy. Despite a global increase in the incidence of CC, little progress has been made toward the development of treatments for this cancer. Here we utilized human tissue; CC cell xenografts; a p53-deficient transgenic mouse model; and a non-transgenic, chemically induced rat model of CC that accurately reflects both the inflammatory and regenerative background associated with human CC pathology. Using these systems, we determined that the WNT pathway is highly activated in CCs and that inflammatory macrophages are required to establish this WNT-high state in vivo. Moreover, depletion of macrophages or inhibition of WNT signaling with one of two small molecule WNT inhibitors in mouse and rat CC models markedly reduced CC proliferation and increased apoptosis, resulting in tumor regression. Together, these results demonstrate that enhanced WNT signaling is a characteristic of CC and suggest that targeting WNT signaling pathways has potential as a therapeutic strategy for CC. PMID:25689248

  1. Endodermal Wnt signaling is required for tracheal cartilage formation

    PubMed Central

    Snowball, John; Ambalavanan, Manoj; Whitsett, Jeffrey; Sinner, Debora

    2015-01-01

    Tracheobronchomalacia is a common congenital defect in which the walls of the trachea and bronchi lack of adequate cartilage required for support of the airways. Deletion of Wls, a cargo receptor mediating Wnt ligand secretion, in the embryonic endoderm using ShhCre mice inhibited formation of tracheal-bronchial cartilaginous rings. The normal dorsal-ventral patterning of tracheal mesenchyme was lost. Smooth muscle cells, identified by Acta2 staining, were aberrantly located in ventral mesenchyme of the trachea, normally the region of Sox9 expression in cartilage progenitors. Wnt/β-catenin activity, indicated by Axin2 LacZ reporter, was decreased in tracheal mesenchyme of Wlsf/f;ShhCre/+ embryos. Proliferation of chondroblasts was decreased and reciprocally, proliferation of smooth muscle cells was increased in Wlsf/f;ShhCre/+ tracheal tissue. Expression of Tbx4, Tbx5, Msx1 and Msx2, known to mediate cartilage and muscle patterning, were decreased in tracheal mesenchyme of Wlsf/f;ShhCre/+ embryos. Ex vivo studies demonstrated that Wnt7b and Wnt5a, expressed by the epithelium of developing trachea, and active Wnt/β-catenin signaling are required for tracheal chondrogenesis before formation of mesenchymal condensations. In conclusion, Wnt ligands produced by the tracheal epithelium pattern the tracheal mesenchyme via modulation of gene expression and cell proliferation required for proper tracheal cartilage and smooth muscle differentiation. PMID:26093309

  2. Role of Sost in Wnt signal pathway in osteoporosis rats and regulating effect of soybean isoflavones on Wnt signal pathway.

    PubMed

    Liang, Hai Dong; Yu, Fang; Lv, Ping; Zhao, Zheng Nan; Tong, Zhi Hong

    2014-07-01

    To explore the mechanism of soybean isoflavones (SI) on OVX-induced osteoporosis, we investigated the effect of SI on Wnt signaling that emerged as a novel key pathway for promoting bone formation. Results showed that SI decreased bone mineral elements loss, improved biomechanics parameters in OVX rats. Wnt3a activation can promote the dissociation of β-catenin complexes, release of β-catenin monomer and inhibition of β-catenin monomer degradation. SI decreased sost mRNA and sclerosteosis protein expression in a dose-dependent manner, and increased β-catenin proteins expression in femur of OVX rats. These data suggest that SI suppresses the canonical Wnt signal in OVX rats, partially through the enhancement of the dickkopf-1 production. OVX results in decreased estrogen level in rats. SI act as inhibitors of Wnt-mediated activation of by competitively binding to LRP5, and subsequently downregulating β-catenin gene.

  3. Inhibition of tumorigenesis driven by different Wnt proteins requires blockade of distinct ligand-binding regions by LRP6 antibodies

    PubMed Central

    Ettenberg, Seth A.; Charlat, Olga; Daley, Michael P.; Liu, Shanming; Vincent, Karen J.; Stuart, Darrin D.; Schuller, Alwin G.; Yuan, Jing; Ospina, Beatriz; Green, John; Yu, Qunyan; Walsh, Renee; Schmitz, Rita; Heine, Holger; Bilic, Sanela; Ostrom, Lance; Mosher, Rebecca; Hartlepp, K. Felix; Zhu, Zhenping; Fawell, Stephen; Yao, Yung-Mae; Stover, David; Finan, Peter M.; Porter, Jeffery A.; Sellers, William R.; Klagge, Ingo M.; Cong, Feng

    2010-01-01

    Disregulated Wnt/β-catenin signaling has been linked to various human diseases, including cancers. Inhibitors of oncogenic Wnt signaling are likely to have a therapeutic effect in cancers. LRP5 and LRP6 are closely related membrane coreceptors for Wnt proteins. Using a phage-display library, we identified anti-LRP6 antibodies that either inhibit or enhance Wnt signaling. Two classes of LRP6 antagonistic antibodies were discovered: one class specifically inhibits Wnt proteins represented by Wnt1, whereas the second class specifically inhibits Wnt proteins represented by Wnt3a. Epitope-mapping experiments indicated that Wnt1 class-specific antibodies bind to the first propeller and Wnt3a class-specific antibodies bind to the third propeller of LRP6, suggesting that Wnt1- and Wnt3a-class proteins interact with distinct LRP6 propeller domains. This conclusion is further supported by the structural functional analysis of LRP5/6 and the finding that the Wnt antagonist Sclerostin interacts with the first propeller of LRP5/6 and preferentially inhibits the Wnt1-class proteins. We also show that Wnt1 or Wnt3a class-specific anti-LRP6 antibodies specifically block growth of MMTV-Wnt1 or MMTV-Wnt3 xenografts in vivo. Therapeutic application of these antibodies could be limited without knowing the type of Wnt proteins expressed in cancers. This is further complicated by our finding that bivalent LRP6 antibodies sensitize cells to the nonblocked class of Wnt proteins. The generation of a biparatopic LRP6 antibody blocks both Wnt1- and Wnt3a-mediated signaling without showing agonistic activity. Our studies provide insights into Wnt-induced LRP5/6 activation and show the potential utility of LRP6 antibodies in Wnt-driven cancer. PMID:20713706

  4. ERK5 and Cell Proliferation: Nuclear Localization Is What Matters.

    PubMed

    Gomez, Nestor; Erazo, Tatiana; Lizcano, Jose M

    2016-01-01

    ERK5, the last MAP kinase family member discovered, is activated by the upstream kinase MEK5 in response to growth factors and stress stimulation. MEK5-ERK5 pathway has been associated to different cellular processes, playing a crucial role in cell proliferation in normal and cancer cells by mechanisms that are both dependent and independent of its kinase activity. Thus, nuclear ERK5 activates transcription factors by either direct phosphorylation or acting as co-activator thanks to a unique transcriptional activation TAD domain located at its C-terminal tail. Consequently, ERK5 has been proposed as an interesting target to tackle different cancers, and either inhibitors of ERK5 activity or silencing the protein have shown antiproliferative activity in cancer cells and to block tumor growth in animal models. Here, we review the different mechanisms involved in ERK5 nuclear translocation and their consequences. Inactive ERK5 resides in the cytosol, forming a complex with Hsp90-Cdc37 superchaperone. In a canonical mechanism, MEK5-dependent activation results in ERK5 C-terminal autophosphorylation, Hsp90 dissociation, and nuclear translocation. This mechanism integrates signals such as growth factors and stresses that activate the MEK5-ERK5 pathway. Importantly, two other mechanisms, MEK5-independent, have been recently described. These mechanisms allow nuclear shuttling of kinase-inactive forms of ERK5. Although lacking kinase activity, these forms activate transcription by interacting with transcription factors through the TAD domain. Both mechanisms also require Hsp90 dissociation previous to nuclear translocation. One mechanism involves phosphorylation of the C-terminal tail of ERK5 by kinases that are activated during mitosis, such as Cyclin-dependent kinase-1. The second mechanism involves overexpression of chaperone Cdc37, an oncogene that is overexpressed in cancers such as prostate adenocarcinoma, where it collaborates with ERK5 to promote cell proliferation

  5. ERK5 and Cell Proliferation: Nuclear Localization Is What Matters

    PubMed Central

    Gomez, Nestor; Erazo, Tatiana; Lizcano, Jose M.

    2016-01-01

    ERK5, the last MAP kinase family member discovered, is activated by the upstream kinase MEK5 in response to growth factors and stress stimulation. MEK5-ERK5 pathway has been associated to different cellular processes, playing a crucial role in cell proliferation in normal and cancer cells by mechanisms that are both dependent and independent of its kinase activity. Thus, nuclear ERK5 activates transcription factors by either direct phosphorylation or acting as co-activator thanks to a unique transcriptional activation TAD domain located at its C-terminal tail. Consequently, ERK5 has been proposed as an interesting target to tackle different cancers, and either inhibitors of ERK5 activity or silencing the protein have shown antiproliferative activity in cancer cells and to block tumor growth in animal models. Here, we review the different mechanisms involved in ERK5 nuclear translocation and their consequences. Inactive ERK5 resides in the cytosol, forming a complex with Hsp90-Cdc37 superchaperone. In a canonical mechanism, MEK5-dependent activation results in ERK5 C-terminal autophosphorylation, Hsp90 dissociation, and nuclear translocation. This mechanism integrates signals such as growth factors and stresses that activate the MEK5-ERK5 pathway. Importantly, two other mechanisms, MEK5-independent, have been recently described. These mechanisms allow nuclear shuttling of kinase-inactive forms of ERK5. Although lacking kinase activity, these forms activate transcription by interacting with transcription factors through the TAD domain. Both mechanisms also require Hsp90 dissociation previous to nuclear translocation. One mechanism involves phosphorylation of the C-terminal tail of ERK5 by kinases that are activated during mitosis, such as Cyclin-dependent kinase-1. The second mechanism involves overexpression of chaperone Cdc37, an oncogene that is overexpressed in cancers such as prostate adenocarcinoma, where it collaborates with ERK5 to promote cell proliferation

  6. ERK5 signalling in prostate cancer promotes an invasive phenotype

    PubMed Central

    Ramsay, A K; McCracken, S R C; Soofi, M; Fleming, J; Yu, A X; Ahmad, I; Morland, R; Machesky, L; Nixon, C; Edwards, D R; Nuttall, R K; Seywright, M; Marquez, R; Keller, E; Leung, H Y

    2011-01-01

    Background: Aberrant mitogen/extracellular signal-regulated kinase 5 (MEK5)–extracellular signal-regulated protein kinase 5 (ERK5)-mediated signalling has been implicated in a number of tumour types including prostate cancer (PCa). The molecular basis of ERK5-driven carcinogenesis and its clinical relevance remain to be fully characterised. Methods: Modulation of ERK5 expression or function in human PCa PC3 and PC3–ERK5 (stably transfected with ERK5) cells was performed using siRNA-mediated knockdown or the MEK inhibitor PD18435 respectively. In vitro significance of ERK5 signalling was assessed by assays for proliferation, motility, invasion and invadopodia. Expression of matrix metalloproteinases/tissue inhibitors of metalloproteases was determined by Q-RT–PCR. Extracellular signal-regulated protein kinase 5 expression in primary and metastatic PCa was examined using immunohistochemistry. Results: Reduction of ERK5 expression or signalling significantly inhibited the motility and invasive capability of PC3 cells. Extracellular signal-regulated protein kinase 5-mediated signalling significantly promoted formation of in vivo metastasis in an orthotopic PCa model (P<0.05). Invadopodia formation was also enhanced by forced ERK5 expression in PC3 cells. Furthermore, in metastatic PCa, nuclear ERK5 immunoreactivity was significantly upregulated when compared with benign prostatic hyperplasia and primary PCa (P=0.013 and P<0.0001, respectively). Conclusion: Our in vitro, in vivo and clinical data support an important role for the MEK5–ERK5 signalling pathway in invasive PCa, which represents a potential target for therapy in primary and metastatic PCa. PMID:21266977

  7. Zinc and the ERK Kinases in the Developing Brain

    PubMed Central

    Nuttall, J. R.

    2015-01-01

    This article reviews evidence in support of the hypothesis that impaired activation of the extracellular signal-regulated kinases (ERK1/2) contributes to the disruptions in neurodevelopment associated with zinc deficiency. These kinases are implicated in major events of brain development, including proliferation of progenitor cells, neuronal migration, differentiation, and apoptotic cell death. In humans, mutations in ERK1/2 genes have been associated with neuro-cardio-facial-cutaneous syndromes. ERK1/2 deficits in mice have revealed impaired neurogenesis, altered cellularity, and behavioral abnormalities. Zinc is an important modulator of ERK1/2 signaling. Conditions of both zinc deficiency and excess affect ERK1/2 phosphorylation in fetal and adult brains. Hypophosphorylation of ERK1/2, associated with decreased zinc availability in cell cultures, is accompanied by decreased proliferation and an arrest of the cell cycle at the G0/G1 phase. Zinc and ERK1/2 have both been shown to modulate neural progenitor cell proliferation and cell death in the brain. Furthermore, behavioral deficits resulting from developmental zinc deficiency are similar to those observed in mice with decreased ERK1/2 signaling. For example, impaired performance on behavioral tests of learning and memory; such as the Morris water maze, fear conditioning, and the radial arm maze; has been reported in both animals exposed to developmental zinc deficiency and transgenic mice with decreased ERK signaling. Future study should clarify the mechanisms through which a dysregulation of ERK1/2 may contribute to altered brain development associated with dietary zinc deficiency and with conditions that limit zinc availability. PMID:22095091

  8. Expression of WNT5A in Idiopathic Pulmonary Fibrosis and Its Control by TGF-β and WNT7B in Human Lung Fibroblasts.

    PubMed

    Newman, Donna R; Sills, W Shane; Hanrahan, Katherine; Ziegler, Amanda; Tidd, Kathleen McGinnis; Cook, Elizabeth; Sannes, Philip L

    2016-02-01

    The wingless (Wnt) family of signaling ligands contributes significantly to lung development and is highly expressed in patients with usual interstitial pneumonia (UIP). We sought to define the cellular distribution of Wnt5A in the lung tissue of patients with idiopathic pulmonary fibrosis (IPF) and the signaling ligands that control its expression in human lung fibroblasts and IPF myofibroblasts. Tissue sections from 40 patients diagnosed with IPF or UIP were probed for the immunolocalization of Wnt5A. Further, isolated lung fibroblasts from normal or IPF human lungs, adenovirally transduced for the overexpression or silencing of Wnt7B or treated with TGF-β1 or its inhibitor, were analyzed for Wnt5A protein expression. Wnt5A was expressed in IPF lungs by airway and alveolar epithelium, smooth muscle cells, endothelium, and myofibroblasts of fibroblastic foci and throughout the interstitium. Forced overexpression of Wnt7B with or without TGF-β1 treatment significantly increased Wnt5A protein expression in normal human smooth muscle cells and fibroblasts but not in IPF myofibroblasts where Wnt5A was already highly expressed. The results demonstrate a wide distribution of Wnt5A expression in cells of the IPF lung and reveal that it is significantly increased by Wnt7B and TGF-β1, which, in combination, could represent key signaling pathways that modulate the pathogenesis of IPF. © 2016 The Histochemical Society.

  9. WNT5A: a motility-promoting factor in Hodgkin lymphoma.

    PubMed

    Linke, F; Zaunig, S; Nietert, M M; von Bonin, F; Lutz, S; Dullin, C; Janovská, P; Beissbarth, T; Alves, F; Klapper, W; Bryja, V; Pukrop, T; Trümper, L; Wilting, J; Kube, D

    2017-01-05

    Classical Hodgkin lymphoma (cHL) has a typical clinical manifestation, with dissemination involving functionally neighboring lymph nodes. The factors involved in the spread of lymphoma cells are poorly understood. Here we show that cHL cell lines migrate with higher rates compared with non-Hodgkin lymphoma cell lines. cHL cell migration, invasion and adhesion depend on autocrine WNT signaling as revealed by the inhibition of WNT secretion with the porcupine inhibitors Wnt-C59/IWP-2, but did not affect cell proliferation. While application of recombinant WNT5A or WNT5A overexpression stimulates HL cell migration, neither WNT10A, WNT10B nor WNT16 did so. Time-lapse studies revealed an amoeboid type of cell migration modulated by WNT5A. Reduced migration distances and velocity of cHL cells, as well as altered movement patterns, were observed using porcupine inhibitor or WNT5A antagonist. Knockdown of Frizzled5 and Dishevelled3 disrupted the WNT5A-mediated RHOA activation and cell migration. Overexpression of DVL3-K435M or inhibition of ROCK (Rho-associated protein kinase) by Y-27632/H1152P disrupted cHL cell migration. In addition to these mechanistic insights into the role of WNT5A in vitro, global gene expression data revealed an increased WNT5A expression in primary HL cells in comparison with normal B-cell subsets and other lymphomas. Furthermore, the activity of both porcupine and WNT5A in cHL cells had an impact on lymphoma development in the chick chorionallantoic membrane assay. Massive bleeding of these lymphomas was significantly reduced after inhibition of WNT secretion by Wnt-C59. Therefore, a model is proposed where WNT signaling has an important role in regulating tumor-promoting processes.

  10. A dual role of the Wnt signaling pathway during aging in Caenorhabditis elegans

    PubMed Central

    Lezzerini, Marco; Budovskaya, Yelena

    2014-01-01

    Wnt signaling is a major and highly conserved developmental pathway that guides many important events during embryonic and larval development. In adulthood, misregulation of Wnt signaling has been implicated in tumorigenesis and various age-related diseases. These effects occur through highly complicated cell-to-cell interactions mediated by multiple Wnt-secreted proteins. While they share a high degree of sequence similarity, their function is highly diversified. Although the role of Wnt ligands during development is well studied, very little is known about the possible actions of Wnt signaling in natural aging. In this study, Caenorhabditis elegans serves, for the first time, as a model system to determine the role of Wnt ligands in aging. Caenorhabditis elegans has five Wnt proteins, mom-2, egl-20, lin-44, cwn-1, and cwn-2. We show that all five Wnt ligands are expressed and active past the development stages. The ligand mom-2/Wnt plays a major detrimental role in longevity, whereas the function of lin-44/Wnt is beneficial for long life. Interestingly, no evidence was found for Wnt signaling being involved in cellular or oxidative stress responses during aging. Our results suggest that Wnt signaling regulates aging-intrinsic genetic pathways, opening a new research direction on the role of Wnt signaling in aging and age-related diseases. PMID:23879250

  11. Effects of Wnt-10b on proliferation and differentiation of murine melanoma cells

    SciTech Connect

    Misu, Masayasu; Ouji, Yukiteru; Kawai, Norikazu; Nishimura, Fumihiko; Nakamura-Uchiyama, Fukumi; Yoshikawa, Masahide

    2015-08-07

    In spite of the strong expression of Wnt-10b in melanomas, its role in melanoma cells has not been elucidated. In the present study, the biological effects of Wnt-10b on murine B16F10 (B16) melanoma cells were investigated using conditioned medium from Wnt-10b-producing COS cells (Wnt-CM). After 2 days of culture in the presence of Wnt-CM, proliferation of B16 melanoma cells was inhibited, whereas tyrosinase activity was increased. An in vitro wound healing assay demonstrated that migration of melanoma cells to the wound area was inhibited with the addition of Wnt-CM. Furthermore, evaluation of cellular senescence revealed prominent induction of SA-β-gal-positive senescent cells in cultures with Wnt-CM. Finally, the growth of B16 melanoma cell aggregates in collagen 3D-gel cultures was markedly suppressed in the presence of Wnt-CM. These results suggest that Wnt-10b represses tumor cell properties, such as proliferation and migration of B16 melanoma cells, driving them toward a more differentiated state along a melanocyte lineage. - Highlights: • Wnt-10b inhibited proliferation and migration of melanoma cells. • Wnt-10b induced tyrosinase activity and senescence of melanoma cells. • Wnt-10b suppressed growth of cell aggregates in collagen 3D-gel cultures. • Wnt-10b represses tumor cell properties, driving them toward a more differentiated state along a melanocyte lineage.

  12. Osteocyte-specific WNT1 regulates osteoblast function during bone homeostasis.

    PubMed

    Joeng, Kyu Sang; Lee, Yi-Chien; Lim, Joohyun; Chen, Yuqing; Jiang, Ming-Ming; Munivez, Elda; Ambrose, Catherine; Lee, Brendan H

    2017-06-30

    Mutations in WNT1 cause osteogenesis imperfecta (OI) and early-onset osteoporosis, identifying it as a key Wnt ligand in human bone homeostasis. However, how and where WNT1 acts in bone are unclear. To address this mechanism, we generated late-osteoblast-specific and osteocyte-specific WNT1 loss- and gain-of-function mouse models. Deletion of Wnt1 in osteocytes resulted in low bone mass with spontaneous fractures similar to that observed in OI patients. Conversely, Wnt1 overexpression from osteocytes stimulated bone formation by increasing osteoblast number and activity, which was due in part to activation of mTORC1 signaling. While antiresorptive therapy is the mainstay of OI treatment, it has limited efficacy in WNT1-related OI. In this study, anti-sclerostin antibody (Scl-Ab) treatment effectively improved bone mass and dramatically decreased fracture rate in swaying mice, a model of global Wnt1 loss. Collectively, our data suggest that WNT1-related OI and osteoporosis are caused in part by decreased mTORC1-dependent osteoblast function resulting from loss of WNT1 signaling in osteocytes. As such, this work identifies an anabolic function of osteocytes as a source of Wnt in bone development and homoeostasis, complementing their known function as targets of Wnt signaling in regulating osteoclastogenesis. Finally, this study suggests that Scl-Ab is an effective genotype-specific treatment option for WNT1-related OI and osteoporosis.

  13. Functional conservation of Nematostella Wnts in canonical and noncanonical Wnt-signaling

    PubMed Central

    Rigo-Watermeier, T; Kraft, B; Ritthaler, M; Wallkamm, V; Holstein, T; Wedlich, D

    2012-01-01

    Summary Cnidarians surprise by the completeness of Wnt gene subfamilies (11) expressed in an overlapping pattern along the anterior-posterior axis. While the functional conservation of canonical Wnt-signaling components in cnidarian gastrulation and organizer formation is evident, a role of Nematostella Wnts in noncanonical Wnt-signaling has not been shown so far. In Xenopus, noncanonical Wnt-5a/Ror2 and Wnt-11 (PCP) signaling are distinguishable by different morphant phenotypes. They differ in PAPC regulation, cell polarization, cell protrusion formation, and the so far not reported reorientation of the microtubules. Based on these readouts, we investigated the evolutionary conservation of Wnt-11 and Wnt-5a function in rescue experiments with Nematostella orthologs and Xenopus morphants. Our results revealed that NvWnt-5 and -11 exhibited distinct noncanonical Wnt activities by disturbing convergent extension movements. However, NvWnt-5 rescued XWnt-11 and NvWnt-11 specifically XWnt-5a depleted embryos. This unexpected ‘inverse’ activity suggests that specific structures in Wnt ligands are important for receptor complex recognition in Wnt-signaling. Although we can only speculate on the identity of the underlying recognition motifs, it is likely that these crucial structural features have already been established in the common ancestor of cnidarians and vertebrates and were conserved throughout metazoan evolution. PMID:23213367

  14. Role for WNT16B in human epidermal keratinocyte proliferation and differentiation.

    PubMed

    Teh, Muy-Teck; Blaydon, Diana; Ghali, Lucy R; Briggs, Victoria; Edmunds, Scott; Pantazi, Eleni; Barnes, Michael R; Leigh, Irene M; Kelsell, David P; Philpott, Michael P

    2007-01-15

    WNT signalling regulates a variety of cell functions including cell fate, polarity, and differentiation via the canonical or beta-catenin stabilisation pathway and/or the planar cell polarity or non-canonical pathway. We have previously demonstrated that two isoforms (A and B) from the WNT16 locus have differential expression in various adult human tissues. In this study we show that WNT16B but not WNT16A isoform was upregulated in basal cell carcinomas compared with normal skin. We further investigated the cellular and molecular functions of WNT16B in primary human epidermal keratinocytes and a keratinocyte cell line. Cellular expression of WNT16B neither stabilised beta-catenin nor activated the lymphoid enhancer factor or T-cell factor transcriptional reporter in primary keratinocytes. WNT16B activated the Jun-N-terminal kinase cascade suggesting the activation of a non-canonical WNT signalling pathway. Constitutive expression of WNT16B significantly enhanced the rate of cell proliferation and prolonged clonogenicity in primary keratinocytes. Silencing WNT16B by RNA interference reduced keratinocyte proliferation. Furthermore, overexpression of WNT16B induced a hyperproliferation phenotype in an organotypical culture system. This work presents the first evidence that WNT16B activates human keratinocyte proliferation possibly via a beta-catenin-independent non-canonical WNT transduction pathway.

  15. Thymic Epithelial Cells Are a Nonredundant Source of Wnt Ligands for Thymus Development.

    PubMed

    Brunk, Fabian; Augustin, Iris; Meister, Michael; Boutros, Michael; Kyewski, Bruno

    2015-12-01

    Wnt signaling has been implicated in T cell development. However, it remained unclear which cell type is the major source of Wnt ligands and to what extent thymic epithelial cell (TEC) development is dependent on Wnt signaling. In this study, we analyzed the role of Wnt ligands provided by TECs for the development of T cells and TECs without manipulating the intracellular Wnt signaling machinery in either cell type. To this end, we used conditional knockout mice (FoxN1-Gpr177) in which TECs are unable to secrete Wnt ligands. Gpr177 (Evi/Wls) is a Wnt-specific cargo receptor that is required for the secretion of Wnt ligands. We found that TECs are the main source of Wnt ligands in the thymus, which serves a nonredundant role, and lack of TEC-provided Wnt ligands led to thymic hypotrophy, as well as a reduced peripheral T cell pool. Despite being reduced in numbers, T cells that developed in the absence of TEC-secreted Wnt ligands were functionally competent, and the subset composition of the peripheral T cell pool was not affected. Thus, our data suggest that T cell development is not directly dependent on TEC-provided Wnt ligands. Rather, TEC-secreted Wnt ligands are essential for normal thymus development and normal peripheral T cell frequencies but are dispensable for T cell function in the periphery.

  16. Wnt/β-catenin signaling is required for CNS, but not non-CNS, angiogenesis

    PubMed Central

    Daneman, Richard; Agalliu, Dritan; Zhou, Lu; Kuhnert, Frank; Kuo, Calvin J.; Barres, Ben A.

    2009-01-01

    Despite the importance of CNS blood vessels, the molecular mechanisms that regulate CNS angiogenesis and blood−brain barrier (BBB) formation are largely unknown. Here we analyze the role of Wnt/β-catenin signaling in regulating the formation of CNS blood vessels. First, through the analysis of TOP-Gal Wnt reporter mice, we identify that canonical Wnt/β-catenin signaling is specifically activated in CNS, but not non-CNS, blood vessels during development. This activation correlates with the expression of different Wnt ligands by neural progenitor cells in distinct locations throughout the CNS, including Wnt7a and Wnt7b in ventral regions and Wnt1, Wnt3, Wnt3a, and Wnt4 in dorsal regions. Blockade of Wnt/β-catenin signaling in vivo specifically disrupts CNS, but not non-CNS, angiogenesis. These defects include reduction in vessel number, loss of capillary beds, and the formation of hemorrhagic vascular malformations that remain adherent to the meninges. Furthermore, we demonstrate that Wnt/β-catenin signaling regulates the expression of the BBB-specific glucose transporter glut-1. Taken together these experiments reveal an essential role for Wnt/β-catenin signaling in driving CNS-specific angiogenesis and provide molecular evidence that angiogenesis and BBB formation are in part linked. PMID:19129494

  17. Update on the Mechanism of Action of Aripiprazole: Translational Insights into Antipsychotic Strategies Beyond Dopamine Receptor Antagonism.

    PubMed

    de Bartolomeis, Andrea; Tomasetti, Carmine; Iasevoli, Felice

    2015-09-01

    Dopamine partial agonism and functional selectivity have been innovative strategies in the pharmacological treatment of schizophrenia and mood disorders and have shifted the concept of dopamine modulation beyond the established approach of dopamine D2 receptor (D2R) antagonism. Despite the fact that aripiprazole was introduced in therapy more than 12 years ago, many questions are still unresolved regarding the complexity of the effects of this agent on signal transduction and intracellular pathways, in part linked to its pleiotropic receptor profile. The complexity of the mechanism of action has progressively shifted the conceptualization of this agent from partial agonism to functional selectivity. From the induction of early genes to modulation of scaffolding proteins and activation of transcription factors, aripiprazole has been shown to affect multiple cellular pathways and several cortical and subcortical neurotransmitter circuitries. Growing evidence shows that, beyond the consequences of D2R occupancy, aripiprazole has a unique neurobiology among available antipsychotics. The effect of chronic administration of aripiprazole on D2R affinity state and number has been especially highlighted, with relevant translational implications for long-term treatment of psychosis. The hypothesized effects of aripiprazole on cell-protective mechanisms and neurite growth, as well as the differential effects on intracellular pathways [i.e. extracellular signal-regulated kinase (ERK)] compared with full D2R antagonists, suggest further exploration of these targets by novel and future biased ligand compounds. This review aims to recapitulate the main neurobiological effects of aripiprazole and discuss the potential implications for upcoming improvements in schizophrenia therapy based on dopamine modulation beyond D2R antagonism.

  18. Characterization of Wnt/β-catenin signaling in rhabdomyosarcoma.

    PubMed

    Annavarapu, Srinivas R; Cialfi, Samantha; Dominici, Carlo; Kokai, George K; Uccini, Stefania; Ceccarelli, Simona; McDowell, Heather P; Helliwell, Timothy R

    2013-10-01

    Rhabdomyosarcoma (RMS) is the most common soft tissue sarcoma in children and accounts for about 5% of all malignant paediatric tumours. β-Catenin, a multifunctional nuclear transcription factor in the canonical Wnt signaling pathway, is active in myogenesis and embryonal somite patterning. Dysregulation of Wnt signaling facilitates tumour invasion and metastasis. This study characterizes Wnt/β-catenin signaling and functional activity in paediatric embryonal and alveolar RMS. Immunohistochemical assessment of paraffin-embedded tissues from 44 RMS showed β-catenin expression in 26 cases with cytoplasmic/membranous expression in 9/14 cases of alveolar RMS, and 15/30 cases of embryonal RMS, whereas nuclear expression was only seen in 2 cases of embryonal RMS. The potential functional significance of β-catenin expression was tested in four RMS cell lines, two derived from embryonal (RD and RD18) RMS and two from alveolar (Rh4 and Rh30) RMS. Western blot analysis demonstrated the expression of Wnt-associated proteins including β-catenin, glycogen synthase kinase-3β, disheveled, axin-1, naked, LRP-6 and cadherins in all cell lines. Cell fractionation and immunofluorescence studies of the cell lines (after stimulation by human recombinant Wnt3a) showed reduced phosphorylation of β-catenin, stabilization of the active cytosolic form and nuclear translocation of β-catenin. Reporter gene assay demonstrated a T-cell factor/lymphoid-enhancing factor-mediated transactivation in these cells. In response to human recombinant Wnt3a, the alveolar RMS cells showed a significant decrease in proliferation rate and induction of myogenic differentiation (myogenin, MyoD1 and myf5). These data indicate that the central regulatory components of canonical Wnt/β-catenin signaling are expressed and that this pathway is functionally active in a significant subset of RMS tumours and might represent a novel therapeutic target.

  19. Rapid estrogen receptor alpha signaling mediated by ERK activation regulates vascular tone in male and ovary-intact female mice.

    PubMed

    Kim, Seong Chul; Boese, Austin C; Moore, Matthew H; Cleland, Rea M; Chang, Lin; Delafontaine, Patrice; Yin, Ke-Jie; Lee, Jean-Pyo; Hamblin, Milton H

    2017-09-08

    Estrogen has been shown to affect vascular reactivity. Here, we assessed estrogen receptor-alpha (ERα) dependency of estrogenic effects on vasorelaxation via a rapid nongenomic pathway in both male and ovary-intact female mice. We compared the effect of a primary estrogen, 17 beta-estradiol (E2) or 4,4',4"-(4-propyl-[1H]pyrazole-1,3,5-triyl)tris-phenol (PPT) (selective ERα agonist). We found that E2 and PPT induced greater aortic relaxation in females than males, indicating ERα mediation, which is further validated by employing ERα antagonism. Treatment with 1,3-Bis(4-hydroxyphenyl)-4-methyl-5-[4-(2-piperidinylethoxy)phenol]-1H-pyrazole dihydrochloride (MPP dihydrochloride) (ERα antagonist) attenuated PPT-mediated vessel relaxation in both sexes. ERα-mediated vessel relaxation is further validated by the absence of significant PPT-mediated relaxation in aortas isolated from ERα knockout mice. Treatment with a specific extracellular signal-regulated kinase (ERK) inhibitor, PD98059 reduced E2-induced vessel relaxation in both sexes, but to a lesser extent in females. Further, PD98059 prevented PPT-induced vessel relaxation in both sexes. Both E2 and PPT treatment activated ERK as early as 5-10 min, which was attenuated by PD98059 in aortic tissue, cultured primary vascular smooth muscle cells (VSMCs), and endothelial cells (ECs). Aortic rings denuded of endothelium showed no differences in vessel relaxation following E2 or PPT treatment, implicating a role of ECs in the observed sex differences. Here, our results are unique to show estrogen-stimulated rapid ERα signaling mediated by ERK activation in aortic tissue, as well as VSMCs and ECs in vitro, in regulating vascular function by employing side-by-side comparisons in male and ovary-intact female mice in response to E2 or PPT. Copyright © 2017, American Journal of Physiology-Heart and Circulatory Physiology.

  20. Immunohistochemical study of hair follicle stem cells in regenerated hair follicles induced by Wnt10b

    PubMed Central

    Zhang, Yiming; Xing, Yizhan; Guo, Haiying; Ma, Xiaogen; Li, Yuhong

    2016-01-01

    The regulation of the periodic regeneration of hair follicles is complicated. Although Wnt10b has been reported to induce hair follicle regeneration, the characteristics of induced hair follicles, especially the target cells of Wnt10b, have not yet been clearly elucidated. Thus, we systematically evaluated the expression and proliferation patterns of Wnt10b-induced hair follicles. We found that Wnt10b promoted the proliferation of hair follicle stem cells from 24 hours after AdWnt10b injection. Seventy-two hours after AdWnt10b injection, cells outside of bulge area began to proliferate. When the induced hair follicle entered full anagen, although the hair follicle stem cells were normal, canonical Wnt signaling was maintained in the hair precortex cells. Our results reveal that the target cells that overexpressed Wnt10b included hair follicle stem cells, hair precortex cells, and matrix cells. PMID:27766026

  1. The evolution of reduced antagonism – a role for host-parasite coevolution

    PubMed Central

    Gibson, AK; Stoy, KS; Gelarden, IA; Penley, MJ; Lively, CM; Morran, LT

    2016-01-01

    Why do some host-parasite interactions become less antagonistic over evolutionary time? Vertical transmission can select for reduced antagonism. Vertical transmission also promotes coevolution between hosts and parasites. Therefore, we hypothesized that coevolution itself may underlie transitions to reduced antagonism. To test the coevolution hypothesis, we selected for reduced antagonism between the host Caenorhabditis elegans and its parasite Serratia marcescens. This parasite is horizontally transmitted, which allowed us to study coevolution independently of vertical transmission. After 20 generations, we observed a response to selection when coevolution was possible: reduced antagonism evolved in the co-passaged treatment. Reduced antagonism, however, did not evolve when hosts or parasites were independently selected, without coevolution. In addition, we found strong local adaptation for reduced antagonism between replicate host/parasite lines in the co-passaged treatment. Taken together, these results strongly suggest that coevolution was critical to the rapid evolution of reduced antagonism. PMID:26420682

  2. The evolution of reduced antagonism--A role for host-parasite coevolution.

    PubMed

    Gibson, A K; Stoy, K S; Gelarden, I A; Penley, M J; Lively, C M; Morran, L T

    2015-11-01

    Why do some host-parasite interactions become less antagonistic over evolutionary time? Vertical transmission can select for reduced antagonism. Vertical transmission also promotes coevolution between hosts and parasites. Therefore, we hypothesized that coevolution itself may underlie transitions to reduced antagonism. To test the coevolution hypothesis, we selected for reduced antagonism between the host Caenorhabditis elegans and its parasite Serratia marcescens. This parasite is horizontally transmitted, which allowed us to study coevolution independently of vertical transmission. After 20 generations, we observed a response to selection when coevolution was possible: reduced antagonism evolved in the copassaged treatment. Reduced antagonism, however, did not evolve when hosts or parasites were independently selected without coevolution. In addition, we found strong local adaptation for reduced antagonism between replicate host/parasite lines in the copassaged treatment. Taken together, these results strongly suggest that coevolution was critical to the rapid evolution of reduced antagonism.

  3. Basic fibroblast growth factor induces matrix metalloproteinase-13 via ERK MAP kinase-altered phosphorylation and sumoylation of Elk-1 in human adult articular chondrocytes.

    PubMed

    Im, Hee-Jeong; Sharrocks, Andrew D; Lin, Xia; Yan, Dongyao; Kim, Jaesung; van Wijnen, Andre J; Hipskind, Robert A

    2009-01-01

    Degradation of the extracellular matrix (ECM) by matrix metalloproteinases (MMPs) and release of basic fibroblast growth factor (bFGF) are principal aspects of the pathology of osteoarthritis (OA). ECM disruption leads to bFGF release, which activates the extracellular signal-regulated kinase/mitogen-activated protein kinase (ERK/MAPK) pathway and its downstream target the Ets-like transcription factor Elk-1. Previously we demonstrated that the bFGF-ERK-Elk-1 signaling axis is responsible for the potent induction of MMP-13 in human primary articular chondrocytes. Here we report that, in addition to phosphorylation of Elk-1, dynamic posttranslational modification of Elk-1 by small ubiquitin-related modifier (SUMO) serves as an important mechanism through which MMP-13 gene expression is regulated. We show that bFGF activates Elk-1 mainly through the ERK pathway and that increased phosphorylation of Elk-1 is accompanied by decreased conjugation of SUMO to Elk-1. Reporter gene assays reveal that phosphorylation renders Elk-1 competent for induction of MMP-13 gene transcription, while sumoylation has the opposite effect. Furthermore, we demonstrate that the SUMO-conjugase Ubc9 acts as a key mediator for Elk-1 sumoylation. Taken together, our results suggest that sumoylation antagonizes the phosphorylation-dependent transactivation capacity of Elk-1. This attenuates transcription of its downstream target gene MMP-13 to maintain the integrity of cartilage ECM homeostasis.

  4. Partial antagonism of tiletamine-zolazepam anesthesia in cheetah.

    PubMed

    Walzer, C; Huber, C

    2002-04-01

    This study evaluated partial antagonism of tiletamine-zolazepam (TZ) anesthesia in cheetahs (Acinonyx jubatus) and differences between two benzodiazepine antagonists, flumazenil and sarmazenil, in this species. Four cheetahs were anesthetized three times at an interval of 14 days with an average intramuscular dose of 4.2 mg/kg TZ. In trials 2 and 3 flumazenil at 0.031 mg/kg and sarmazenil at 0.1 mg/kg, respectively, were applied intramuscularly 30 min after initial TZ injection. There was a highly significant difference between the duration of TZ anesthesia with and without antagonist. Use of the antagonists significantly shortened duration and recovery and eliminated excitatory behavior during the recovery phase. No significant differences could be determined between the two antagonists. We recommend the use of sarmazenil and flumazenil to antagonize TZ anesthesia in cheetahs.

  5. Negative regulation of the Wnt signal by MM-1 through inhibiting expression of the wnt4 gene.

    PubMed

    Yoshida, Tatsuya; Kitaura, Hirotake; Hagio, Yuko; Sato, Toshiya; Iguchi-Ariga, Sanae M M; Ariga, Hiroyoshi

    2008-04-01

    We have reported that a novel c-Myc-binding protein, MM-1, repressed the E-box-dependent transcription activity of c-Myc through TIF1beta/KAP1, a transcriptional corepressor, and that the c-fms gene was a target gene involved in this pathway. We have also reported that a mutation of A157R in MM-1, which is often observed in patients with leukemia or lymphoma, abrogated all of the repressive activities of MM-1 toward c-Myc, indicating that MM-1 is a novel tumor suppressor. In this study, to further identify target genes of MM-1, DNA microarray analysis was carried out by comparing expression levels of genes in MM-1 knockdown and parental cells, and the wnt4 gene, a member of the Wnt-beta-catenin pathway, was identified as a target gene of MM-1. Increased expression level of the wnt4 gene, accumulation and translocation of beta-catenin to the cytoplasm and nucleus, and upregulation of TCF/Lef-1, a target protein of the Wnt-beta-catenin pathway, were found in MM-1 knockdown cells. Reporter assays using various deletion constructs of the wnt4 gene promoter showed that MM-1 recognized the region spanning -286 to -229 from a transcription start site, and MM-1 complex was found to bind to this region by chromatin immunoprecipitation and gel mobility shift assays. Furthermore, it was found that Egr-1 and MM-1 were bound to this region and that both proteins mutually down-regulate promoter activity of the wnt4 gene. Since the c-myc gene is the target gene of the Wnt-beta-catenin pathway, these findings suggest that MM-1 inhibits c-Myc by a dual mechanism.

  6. C3 toxin activates the stress signaling pathways, JNK and p38, but antagonizes the activation of AP-1 in rat-1 cells.

    PubMed

    Beltman, J; Erickson, J R; Martin, G A; Lyons, J F; Cook, S J

    1999-02-05

    Lysophosphatidic acid (LPA) stimulates the c-Fos serum response element (SRE) by activating two distinct signal pathways regulated by the small GTPases, Ras and RhoA. Ras activates the ERK cascade leading to phosphorylation of the transcription factors Elk-1 and Sap1a at the Ets/TCF site. RhoA regulates an undefined pathway required for the activation of the SRF/CArG site. Here we have examined the role of the Ras and RhoA pathways in activation of the SRE and c-Fos expression in Rat-1 cells. Pertussis toxin and PD98059 strongly inhibited LPA-stimulated c-Fos expression and activation of a SRE:Luc reporter. C3 toxin completely inhibited RhoA function, partially inhibited SRE:Luc activity, but had no effect on LPA-stimulated c-Fos expression. Thus, in a physiological context the Ras-Raf-MEK-ERK pathway, but not RhoA, is required for LPA-stimulated c-Fos expression in Rat-1 cells. C3 toxin stimulated the stress-activated protein kinases JNK and p38 and potentiated c-Jun expression and phosphorylation; these properties were shared by another cellular stress agonist the protein kinase C inhibitor Ro-31-8220. However, C3 toxin alone or in combination with growth factors did not stimulate AP-1:Luc activity and actually antagonized the synergistic activation of AP-1:Luc observed in response to co-stimulation with growth factors and Ro-31-8220. These data indicate that C3 toxin is a cellular stress which antagonizes activation of AP-1 at a point downstream of stress-activated kinase activation or immediate-early gene induction.

  7. Inhibition of Dengue Virus Replication by a Class of Small-Molecule Compounds That Antagonize Dopamine Receptor D4 and Downstream Mitogen-Activated Protein Kinase Signaling

    PubMed Central

    Smith, Jessica L.; Stein, David A.; Shum, David; Fischer, Matthew A.; Radu, Constantin; Bhinder, Bhavneet; Djaballah, Hakim; Nelson, Jay A.; Früh, Klaus

    2014-01-01

    ABSTRACT Dengue viruses (DENV) are endemic pathogens of tropical and subtropical regions that cause significant morbidity and mortality worldwide. To date, no vaccines or antiviral therapeutics have been approved for combating DENV-associated disease. In this paper, we describe a class of tricyclic small-molecule compounds—dihydrodibenzothiepines (DHBTs), identified through high-throughput screening—with potent inhibitory activity against DENV serotype 2. SKI-417616, a highly active representative of this class, displayed activity against all four serotypes of DENV, as well as against a related flavivirus, West Nile virus (WNV), and an alphavirus, Sindbis virus (SINV). This compound was characterized to determine its mechanism of antiviral activity. Investigation of the stage of the viral life cycle affected revealed that an early event in the life cycle is inhibited. Due to the structural similarity of the DHBTs to known antagonists of the dopamine and serotonin receptors, we explored the roles of two of these receptors, serotonin receptor 2A (5HTR2A) and the D4 dopamine receptor (DRD4), in DENV infection. Antagonism of DRD4 and subsequent downstream phosphorylation of epidermal growth factor receptor (EGFR)-related kinase (ERK) were found to impact DENV infection negatively, and blockade of signaling through this network was confirmed as the mechanism of anti-DENV activity for this class of compounds. IMPORTANCE The dengue viruses are mosquito-borne, reemerging human pathogens that are the etiological agents of a spectrum of febrile diseases. Currently, there are no approved therapeutic treatments for dengue-associated disease, nor is there a vaccine. This study identifies a small molecule, SKI-417616, with potent anti-dengue virus activity. Further analysis revealed that SKI-417616 acts through antagonism of the host cell dopamine D4 receptor and subsequent repression of the ERK phosphorylation pathway. These results suggest that SKI-417616, or other

  8. Inhibition of dengue virus replication by a class of small-molecule compounds that antagonize dopamine receptor d4 and downstream mitogen-activated protein kinase signaling.

    PubMed

    Smith, Jessica L; Stein, David A; Shum, David; Fischer, Matthew A; Radu, Constantin; Bhinder, Bhavneet; Djaballah, Hakim; Nelson, Jay A; Früh, Klaus; Hirsch, Alec J

    2014-05-01

    Dengue viruses (DENV) are endemic pathogens of tropical and subtropical regions that cause significant morbidity and mortality worldwide. To date, no vaccines or antiviral therapeutics have been approved for combating DENV-associated disease. In this paper, we describe a class of tricyclic small-molecule compounds-dihydrodibenzothiepines (DHBTs), identified through high-throughput screening-with potent inhibitory activity against DENV serotype 2. SKI-417616, a highly active representative of this class, displayed activity against all four serotypes of DENV, as well as against a related flavivirus, West Nile virus (WNV), and an alphavirus, Sindbis virus (SINV). This compound was characterized to determine its mechanism of antiviral activity. Investigation of the stage of the viral life cycle affected revealed that an early event in the life cycle is inhibited. Due to the structural similarity of the DHBTs to known antagonists of the dopamine and serotonin receptors, we explored the roles of two of these receptors, serotonin receptor 2A (5HTR2A) and the D4 dopamine receptor (DRD4), in DENV infection. Antagonism of DRD4 and subsequent downstream phosphorylation of epidermal growth factor receptor (EGFR)-related kinase (ERK) were found to impact DENV infection negatively, and blockade of signaling through this network was confirmed as the mechanism of anti-DENV activity for this class of compounds. The dengue viruses are mosquito-borne, reemerging human pathogens that are the etiological agents of a spectrum of febrile diseases. Currently, there are no approved therapeutic treatments for dengue-associated disease, nor is there a vaccine. This study identifies a small molecule, SKI-417616, with potent anti-dengue virus activity. Further analysis revealed that SKI-417616 acts through antagonism of the host cell dopamine D4 receptor and subsequent repression of the ERK phosphorylation pathway. These results suggest that SKI-417616, or other compounds targeting the same

  9. Nucleostemin Rejuvenates Cardiac Progenitor Cells and Antagonizes Myocardial Aging

    PubMed Central

    Hariharan, Nirmala; Quijada, Pearl; Mohsin, Sadia; Joyo, Anya; Samse, Kaitlen; Monsanto, Megan; De La Torre, Andrea; Avitabile, Daniele; Ormachea, Lucia; McGregor, Michael J.; Tsai, Emily J; Sussman, Mark A.

    2015-01-01

    BACKGROUND Functional decline in stem cell-mediated regeneration contributes to aging associated with cellular senescence in c-kit+ cardiac progenitor cells (CPCs). Clinical implementation of CPC-based therapy with elderly patients would benefit tremendously from understanding molecular characteristics of senescence to antagonize aging. Nucleostemin (NS) is a nucleolar protein regulating stem cell proliferation and pluripotency. OBJECTIVES The goal is to demonstrate that NS preserves characteristics associated with “stemness” in CPCs and antagonizes myocardial senescence and aging. METHODS CPCs isolated from human fetal (FhCPC) and adult failing (AhCPC) hearts, as well as young (YCPC) and old mice (OCPC), were studied for senescence characteristics and NS expression. Heterozygous knockout mice with one functional allele of NS (NS+/−) were used to demonstrate that NS preserves myocardial structure and function and slows characteristics of aging. RESULTS NS expression is decreased in AhCPCs relative to FhCPC, correlating with lowered proliferation potential and shortened telomere length. AhCPC characteristics resemble OCPCs, which have a phenotype induced by NS silencing, resulting in cell flattening, senescence, multinucleated cells, decreased S phase progression, diminished expression of stemness markers and up-regulation of p53 and p16. CPC senescence resulting from NS loss is partially p53 dependent and is rescued by concurrent silencing of p53. Mechanistically, NS induction correlates with Pim-1 kinase-mediated stabilization of c-Myc. Engineering OCPCs and AhCPCs to overexpress NS decreases senescent and multinucleated cells, restores morphology, and antagonizes senescence, thereby preserving phenotypic properties of “stemness.” Early cardiac aging with decline in cardiac function, increase in senescence markers p53 and p16, telomere attrition, and accompanied CPC exhaustion is evident in NS+/− mice. CONCLUSIONS Youthful properties and antagonism of

  10. Convergent evolution of escape from hepaciviral antagonism in primates.

    PubMed

    Patel, Maulik R; Loo, Yueh-Ming; Horner, Stacy M; Gale, Michael; Malik, Harmit S

    2012-01-01

    The ability to mount an interferon response on sensing viral infection is a critical component of mammalian innate immunity. Several viruses directly antagonize viral sensing pathways to block activation of the host immune response. Here, we show that recurrent viral antagonism has shaped the evolution of the host protein MAVS--a crucial component of the viral-sensing pathway in primates. From sequencing and phylogenetic analyses of MAVS from 21 simian primates, we found that MAVS has evolved under strong positive selection. We focused on how this positive selection has shaped MAVS' susceptibility to Hepatitis C virus (HCV). We functionally tested MAVS proteins from diverse primate species for their ability to resist antagonism by HCV, which uses its protease NS3/4A to cleave human MAVS. We found that MAVS from multiple primates are resistant to inhibition by the HCV protease. This resistance maps to single changes within the protease cleavage site in MAVS, which protect MAVS from getting cleaved by the HCV protease. Remarkably, most of these changes have been independently acquired at a single residue 506 that evolved under positive selection. We show that "escape" mutations lower affinity of the NS3 protease for MAVS and allow it to better restrict HCV replication. We further show that NS3 proteases from all other primate hepaciviruses, including the highly divergent GBV-A and GBV-C viruses, are functionally similar to HCV. We conclude that convergent evolution at residue 506 in multiple primates has resulted in escape from antagonism by hepaciviruses. Our study provides a model whereby insights into the ancient history of viral infections in primates can be gained using extant host and virus genes. Our analyses also provide a means by which primates might clear infections by extant hepaciviruses like HCV.

  11. CXCR4 Antagonism Attenuates the Development of Diabetic Cardiac Fibrosis.

    PubMed

    Chu, Po-Yin; Walder, Ken; Horlock, Duncan; Williams, David; Nelson, Erin; Byrne, Melissa; Jandeleit-Dahm, Karin; Zimmet, Paul; Kaye, David M

    2015-01-01

    Heart failure (HF) is an increasingly recognized complication of diabetes. Cardiac fibrosis is an important causative mechanism of HF associated with diabetes. Recent data indicate that inflammation may be particularly important in the pathogenesis of cardiovascular fibrosis. We sought to determine the mechanism by which cardiac fibrosis develops and to specifically investigate the role of the CXCR4 axis in this process. Animals with type I diabetes (streptozotocin treated mice) or type II diabetes (Israeli Sand-rats) and controls were randomized to treatment with a CXCR4 antagonist, candesartan or vehicle control. Additional groups of mice also underwent bone marrow transplantation (GFP+ donor marrow) to investigate the potential role of bone marrow derived cell mobilization in the pathogenesis of cardiac fibrosis. Both type I and II models of diabetes were accompanied by the development of significant cardiac fibrosis. CXCR4 antagonism markedly reduced cardiac fibrosis in both models of diabetes, similar in magnitude to that seen with candesartan. In contrast to candesartan, the anti-fibrotic actions of CXCR4 antagonism occurred in a blood pressure independent manner. Whilst the induction of diabetes did not increase the overall myocardial burden of GFP+ cells, it was accompanied by an increase in GFP+ cells expressing the fibroblast marker alpha-smooth muscle actin and this was attenuated by CXCR4 antagonism. CXCR4 antagonism was also accompanied by increased levels of circulating regulatory T cells. Taken together the current data indicate that pharmacological inhibition of CXCR4 significantly reduces diabetes induced cardiac fibrosis, providing a potentially important therapeutic approach.

  12. STUDIES OF PYRIDOXAL-PENICILLAMINE ANTAGONISM IN THE HUMAN,

    DTIC Science & Technology

    The pyridoxal-L- penicillamine antagonism previously studied by others in lower animals has now been studied in man by means of urinary xanthurenic...studies involved urinary XA determinations, with and without tryptophan-loading tests, and with varying dosage combinations of penicillamines and...pyridoxine hydrochloride. It was found that urinary XA excretions remained normal after DL- penicillamine or DL-tryptophan alone but increased when the two

  13. Activation of Alternative Wnt Signaling Pathways in Human Mammary Gland and Breast Cancer Cells

    DTIC Science & Technology

    2006-06-01

    signalling, the antagonistic role of Wnt5a on canonical Wnt signalling, and the fact that the genes regulated by either of these pathways differ in...differentiation, apoptosis, and migration. Wnt/Frizzled signaling is now linked to human hereditary disorders with retinal vascular defects, implicating...www.physiologyonline.org fact that the role of FrzA in vascular biology is not well understood. Wnt Signaling Comes into Play in Human Vascular

  14. Multiple Wnt genes are required for posterior patterning in the short germ embryo of Tribolium castaneum

    PubMed Central

    Bolognesi, Renata; Farzana, Laila; Fischer, Tamara D.; Brown, Susan J.

    2008-01-01

    Summary wingless (wg)/Wnt family genes encode secreted glycoproteins essential for the development of virtually all metazoans. In short germ insects, including the red flour beetle, Tribolium castaneum, the segment-polarity function of wg is conserved [1]. Wnt signalling is also implicated in posterior patterning and germband elongation [2–4], but despite its expression in the posterior growth zone, Wnt1/wg alone is not responsible for these functions; [1–3]. Tribolium contains additional Wnt family genes of unknown function that are also expressed in the growth zone [5]. After depleting one of these, Tc-WntD/8, we found a small percentage of embryos lacking abdominal segments. Additional removal of Tc-Wnt1 significantly enhanced this phenotype, suggesting functional redundancy. Seeking alternative methods to deplete Wnt signal, we performed RNAi with other components of the Wnt pathway including wntless (wls) and porcupine (porc), which process Wnt ligands, and pangolin (pan), which transduces the signal to the nucleus. Tc-wls RNAi caused segmentation defects similar to Tc-Wnt1, but not Tc-WntD/8 RNAi, indicating that the effects of Tc-WntD/8 depletion are Tc-wls-independent. In contrast, depletion of Tc-porc and Tc-pan resulted in embryos resembling those of double Tc-Wnt1,Tc-WntD/8 RNAi, suggesting Tc-porc is essential for the function of both ligands and that they signal through the canonical pathway. Our results provide the first evidence of functional redundancy between Wnt ligands in posterior patterning in short germ insects. This Wnt function appears to be conserved in other arthropods [6] and vertebrates [7–9]. PMID:18926702

  15. RING finger protein PLR-1 blocks Wnt signaling by altering trafficking of Wnt Receptors

    NASA Astrophysics Data System (ADS)

    Robinson, Ryan E.

    Secreted Wnt proteins control a wide range of essential developmental processes, including axon guidance and establishment of anteroposterior neuronal polarity. We identified a transmembrane RING finger protein, PLR-1, that governs the response to Wnts by reducing the cell surface levels of Wnt receptors Frizzled, CAM-1 and LIN-18 in Caenorhabditis elegans. Frizzled, CAM-1 and LIN-18 are normally enriched at the plasma membrane where they are capable of detecting and responding to extracellular Wnts. However, when PLR-1 is expressed Frizzled, CAM-1 and LIN-18 are no longer detected at the cell surface and instead colocalize with PLR-1 in endosomes and Golgi. PLR-1 is related to a broad family of transmembrane proteins that contain a lumenal protease associated domain and a cytosolic RING finger. The RING finger is a hallmark of one type of E3 ubiquitin ligase and monoubiquitination is commonly used to regulate protein trafficking. Protease associated domains are largely thought to mediate interactions between proteins. To identify the domains responsible for PLR-1 regulation of Frizzled from the cell surface we utilized a series of fluorescently tagged fusion proteins and protein truncations containing various domains from PLR-1 and Frizzled. Our data suggests that PLR-1 and Frizzled interact and form a complex via their respective extracellular/lumenal domains, and that ubiqiuitination of Frizzled by PLR-1 targets the Frizzled/PLR-1 complex to the endosome.

  16. Activation of the Wnt Pathway by Mycobacterium tuberculosis: A Wnt–Wnt Situation

    PubMed Central

    Villaseñor, Tomás; Madrid-Paulino, Edgardo; Maldonado-Bravo, Rafael; Urbán-Aragón, Antonio; Pérez-Martínez, Leonor; Pedraza-Alva, Gustavo

    2017-01-01

    Mycobacterium tuberculosis (M. tuberculosis), an intracellular pathogenic Gram-positive bacterium, is the cause of tuberculosis (TB), a major worldwide human infectious disease. The innate immune system is the first host defense against M. tuberculosis. The recognition of this pathogen is mediated by several classes of pattern recognition receptors expressed on the host innate immune cells, including Toll-like receptors, Nod-like receptors, and C-type lectin receptors like Dectin-1, the Mannose receptor, and DC-SIGN. M. tuberculosis interaction with any of these receptors activates multiple signaling pathways among which the protein kinase C, the MAPK, and the NFκB pathways have been widely studied. These pathways have been implicated in macrophage invasion, M. tuberculosis survival, and impaired immune response, thus promoting a successful infection and disease. Interestingly, the Wnt signaling pathway, classically regarded as a pathway involved in the control of cell proliferation, migration, and differentiation in embryonic development, has recently been involved in immunoregulatory mechanisms in infectious and inflammatory diseases, such as TB, sepsis, psoriasis, rheumatoid arthritis, and atherosclerosis. In this review, we present the current knowledge supporting a role for the Wnt signaling pathway during macrophage infection by M. tuberculosis and the regulation of the immune response against M. tuberculosis. Understanding the cross talk between different signaling pathways activated by M. tuberculosis will impact on the search for new therapeutic targets to fuel the rational design of drugs aimed to restore the immunological response against M. tuberculosis. PMID:28203237

  17. Wnt/beta-Catenin Signaling and Small Molecule Inhibitors

    PubMed Central

    Voronkov, Andrey; Krauss, Stefan

    2012-01-01

    Wnt/β-catenin signaling is a branch of a functional network that dates back to the first metazoans and it is involved in a broad range of biological systems including stem cells, embryonic development and adult organs. Deregulation of components involved in Wnt/β-catenin signaling has been implicated in a wide spectrum of diseases including a number of cancers and degenerative diseases. The key mediator of Wnt signaling, β-catenin, serves several cellular functions. It functions in a dynamic mode at multiple cellular locations, including the plasma membrane, where β-catenin contributes to the stabilization of intercellular adhesive complexes, the cytoplasm where β-catenin levels are regulated and the nucleus where β-catenin is involved in transcriptional regulation and chromatin interactions. Central effectors of β-catenin levels are a family of cysteine-rich secreted glycoproteins, known as Wnt morphogens. Through the LRP5/6-Frizzled receptor complex, Wnts regulate the location and activity of the destruction complex and consequently intracellular β- catenin levels. However, β-catenin levels and their effects on transcriptional programs are also influenced by multiple other factors including hypoxia, inflammation, hepatocyte growth factor-mediated signaling, and the cell adhesion molecule E-cadherin. The broad implications of Wnt/β-catenin signaling in development, in the adult body and in disease render the pathway a prime target for pharmacological research and development. The intricate regulation of β-catenin at its various locations provides alternative points for therapeutic interventions. PMID:23016862

  18. Differential niche and Wnt requirements during acute myeloid leukemia progression.

    PubMed

    Lane, Steven W; Wang, Yingzi J; Lo Celso, Cristina; Ragu, Christine; Bullinger, Lars; Sykes, Stephen M; Ferraro, Francesca; Shterental, Sebastian; Lin, Charles P; Gilliland, D Gary; Scadden, David T; Armstrong, Scott A; Williams, David A

    2011-09-08

    Hematopoietic stem cells (HSCs) engage in complex bidirectional signals with the hematopoietic microenvironment (HM), and there is emerging evidence that leukemia stem cells (LSCs) may use similar interactions. Using a syngeneic retroviral model of MLL-AF9 induced acute myeloid leukemia (AML), we have identified 2 different stages of leukemia progression, propagated by "pre-LSCs" and established leukemia (LSCs) and compared the homing properties of these distinctive entities to that of normal HSCs. The homing and microlocalization of pre-LSCs was most similar to long-term HSCs and was dependent on cell-intrinsic Wnt signaling. In contrast, the homing of established LSCs was most similar to that of committed myeloid progenitors and distinct from HSCs. Although osteoblast-derived Dickkopf-1, a potent Wnt inhibitor known to impair HSC function, dramatically impaired normal HSC localization within the bone marrow, it did not affect pre-LSCs, LSC homing, or AML development. Mechanistically, cell-intrinsic Wnt activation was observed in human and murine AML samples, explaining the independence of MLL-AF9 LSCs from niche-derived Wnt signals. These data identify differential engagement of HM associated with leukemic progression and identify an LSC niche that is physically distinct and independent of the constraints of Wnt signaling that apply to normal HSCs.

  19. Differential niche and Wnt requirements during acute myeloid leukemia progression

    PubMed Central

    Lane, Steven W.; Wang, Yingzi J.; Lo Celso, Cristina; Ragu, Christine; Bullinger, Lars; Sykes, Stephen M.; Ferraro, Francesca; Shterental, Sebastian; Lin, Charles P.; Gilliland, D. Gary

    2011-01-01

    Hematopoietic stem cells (HSCs) engage in complex bidirectional signals with the hematopoietic microenvironment (HM), and there is emerging evidence that leukemia stem cells (LSCs) may use similar interactions. Using a syngeneic retroviral model of MLL-AF9 induced acute myeloid leukemia (AML), we have identified 2 different stages of leukemia progression, propagated by “pre-LSCs” and established leukemia (LSCs) and compared the homing properties of these distinctive entities to that of normal HSCs. The homing and microlocalization of pre-LSCs was most similar to long-term HSCs and was dependent on cell-intrinsic Wnt signaling. In contrast, the homing of established LSCs was most similar to that of committed myeloid progenitors and distinct from HSCs. Although osteoblast-derived Dickkopf-1, a potent Wnt inhibitor known to impair HSC function, dramatically impaired normal HSC localization within the bone marrow, it did not affect pre-LSCs, LSC homing, or AML development. Mechanistically, cell-intrinsic Wnt activation was observed in human and murine AML samples, explaining the independence of MLL-AF9 LSCs from niche-derived Wnt signals. These data identify differential engagement of HM associated with leukemic progression and identify an LSC niche that is physically distinct and independent of the constraints of Wnt signaling that apply to normal HSCs. PMID:21765021

  20. Neurodevelopment in Schizophrenia: The Role of the Wnt Pathways

    PubMed Central

    Panaccione, Isabella; Napoletano, Flavia; Forte, Alberto Maria; Kotzalidis, Giorgio D.; Del Casale, Antonio; Rapinesi, Chiara; Brugnoli, Chiara; Serata, Daniele; Caccia, Federica; Cuomo, Ilaria; Ambrosi, Elisa; Simonetti, Alessio; Savoja, Valeria; De Chiara, Lavinia; Danese, Emanuela; Manfredi, Giovanni; Janiri, Delfina; Motolese, Marta; Nicoletti, Ferdinando; Girardi, Paolo; Sani, Gabriele

    2013-01-01

    Objectives. To review the role of Wnt pathways in the neurodevelopment of schizophrenia. Methods: Systematic PubMed search, using as keywords all the terms related to the Wnt pathways and crossing them with each of the following areas: normal neurodevelopment and physiology, neurodevelopmental theory of schizophrenia, schizophrenia, and antipsychotic drug action. Results: Neurodevelopmental, behavioural, genetic, and psychopharmacological data point to the possible involvement of Wnt systems, especially the canonical pathway, in the pathophysiology of schizophrenia and in the mechanism of antipsychotic drug action. The molecules most consistently found to be associated with abnormalities or in antipsychotic drug action are Akt1, glycogen synthase kinase3beta, and beta-catenin. However, the extent to which they contribute to the pathophysiology of schizophrenia or to antipsychotic action remains to be established. Conclusions: The study of the involvement of Wnt pathway abnormalities in schizophrenia may help in understanding this multifaceted clinical entity; the development of Wnt-related pharmacological targets must await the collection of more data. PMID:24403877

  1. Glucocorticoid suppresses the canonical Wnt signal in cultured human osteoblasts

    SciTech Connect

    Ohnaka, Keizo . E-mail: oonaka@geriat.med.kyushu-u.ac.jp; Tanabe, Mizuho; Kawate, Hisaya; Nawata, Hajime; Takayanagi, Ryoichi

    2005-04-01

    To explore the mechanism of glucocorticoid-induced osteoporosis, we investigated the effect of glucocorticoid on canonical Wnt signaling that emerged as a novel key pathway for promoting bone formation. Wnt3a increased the T-cell factor (Tcf)/lymphoid enhancer factor (Lef)-dependent transcriptional activity in primary cultured human osteoblasts. Dexamethasone suppressed this transcriptional activity in a dose-dependent manner, while 1,25-dihydroxyvitamin D3 increased this transcriptional activity. LiCl, an inhibitor of glycogen synthase kinase-3{beta}, also enhanced the Tcf/Lef-dependent transcriptional activity, which was, however, not inhibited by dexamethasone. The addition of anti-dickkopf-1 antibody partially restored the transcriptional activity suppressed by dexamethasone. Dexamethasone decreased the cytosolic amount of {beta}-catenin accumulated by Wnt3a and also inhibited the nuclear translocation of {beta}-catenin induced by Wnt3a. These data suggest that glucocorticoid suppresses the canonical Wnt signal in cultured human osteoblasts, partially through the enhancement of the dickkopf-1 production.

  2. Wnt signaling is required for long-term memory formation

    PubMed Central

    Tan, Ying; Yu, Dinghui; Busto, Germain U.; Wilson, Curtis; Davis, Ronald L.

    2013-01-01

    SUMMARY Wnt signaling regulates synaptic plasticity and neurogenesis in the adult nervous system, suggesting a potential role in behavioral processes. Here, we probed the requirement for Wnt signaling during olfactory memory formation in Drosophila using an inducible RNA interference approach. Interfering with β-catenin expression in the adult mushroom body neurons specifically impaired long-term memory without altering short-term memory. The impairment was reversible, rescued with expression of a wild-type β-catenin transgene, and correlated with a disruption of a cellular long-term memory trace. Inhibition of wingless, a Wnt ligand, and arrow, a Wnt co-receptor, also impaired long-term memory. Wingless expression in wild type flies was transiently elevated in the brain after long-term memory conditioning. Thus, inhibiting three key components of the Wnt signaling pathway in the adult mushroom bodies impairs long-term memory, collectively indicating that this pathway mechanistically underlies this specific form of memory. PMID:24035392

  3. The Polycystin complex mediates WNT/Ca2+ signaling

    PubMed Central

    Nesin, Vasyl; Tran, Uyen; Outeda, Patricia; Bai, Chang-Xi; Keeling, Jacob; Maskey, Dipak; Watnick, Terry; Wessely, Oliver; Tsiokas, Leonidas

    2016-01-01

    WNT ligands induce Ca2+ signaling on target cells. PKD1 (Polycystin 1) is considered an orphan, atypical G protein coupled receptor complexed with TRPP2 (Polycystin 2 or PKD2), a Ca2+-permeable ion channel. Inactivating mutations in their genes cause autosomal dominant polycystic kidney disease (ADPKD), one of the most common genetic diseases. Here, we show that WNTs bind to the extracellular domain of PKD1 and induce whole cell currents and Ca2+ influx dependent on TRPP2. Pathogenic PKD1 or PKD2 mutations that abrogate complex formation, compromise cell surface expression of PKD1, or reduce TRPP2 channel activity suppress activation by WNTs. Pkd2−/− fibroblasts lack WNT-induced Ca2+ currents and are unable to polarize during directed cell migration. In Xenopus embryos, PKD1, Dishevelled 2 (DVL2), and WNT9A act within the same pathway to preserve normal tubulogenesis. These data define PKD1 as a WNT (co)receptor and implicate defective WNT/Ca2+ signaling as one of the causes of ADPKD. PMID:27214281

  4. Wnt and planar cell polarity signaling in cystic renal disease.

    PubMed

    Goggolidou, Paraskevi

    2014-01-01

    Cystic kidney diseases can cause end stage renal disease, affecting millions of individuals worldwide. They may arise early or later in life, are characterized by a spectrum of symptoms and can be caused by diverse genetic defects. The primary cilium, a microtubule-based organelle that can serve as a signaling antenna, has been demonstrated to have a significant role in ensuring correct kidney development and function. In the kidney, one of the signaling pathways that requires the cilium for normal development is Wnt signaling. In this review, the roles of primary cilia in relation to canonical and non-canonical Wnt/PCP signaling in cystic renal disease are described. The evidence of the associations between cilia, Wnt signaling and cystic renal disease is discussed and the significance of planar cell polarity-related mechanisms in cystic kidney disease is presented. Although defective Wnt signaling is not the only cause of renal disease, research is increasingly highlighting its importance, encouraging the development of Wnt-associated diagnostic and prognostic tools for cystic renal disease.

  5. Pantothenate and pantetheine antagonize the antitubercular activity of pyrazinamide.

    PubMed

    Dillon, Nicholas A; Peterson, Nicholas D; Rosen, Brandon C; Baughn, Anthony D

    2014-12-01

    Pyrazinamide (PZA) is a first-line tuberculosis drug that inhibits the growth of Mycobacterium tuberculosis via an as yet undefined mechanism. An M. tuberculosis laboratory strain that was auxotrophic for pantothenate was found to be insensitive to PZA and to the active form, pyrazinoic acid (POA). To determine whether this phenotype was strain or condition specific, the effect of pantothenate supplementation on PZA activity was assessed using prototrophic strains of M. tuberculosis. It was found that pantothenate and other β-alanine-containing metabolites abolished PZA and POA susceptibility, suggesting that POA might selectively target pantothenate synthesis. However, when the pantothenate-auxotrophic strain was cultivated using a subantagonistic concentration of pantetheine in lieu of pantothenate, susceptibility to PZA and POA was restored. In addition, we found that β-alanine could not antagonize PZA and POA activity against the pantothenate-auxotrophic strain, indicating that the antagonism is specific to pantothenate. Moreover, pantothenate-mediated antagonism was observed for structurally related compounds, including n-propyl pyrazinoate, 5-chloropyrazinamide, and nicotinamide, but not for nicotinic acid or isoniazid. Taken together, these data demonstrate that while pantothenate can interfere with the action of PZA, pantothenate synthesis is not directly targeted by PZA. Our findings suggest that targeting of pantothenate synthesis has the potential to enhance PZA efficacy and possibly to restore PZA susceptibility in isolates with panD-linked resistance.

  6. Examining Docking Interactions on ERK2 with Modular Peptide Substrates

    PubMed Central

    Lee, Sunbae; Warthaka, Mangalika; Yan, Chunli; Kaoud, Tamer S; Ren, Pengyu; Dalby, Kevin N

    2012-01-01

    ERK2 primarily recognizes substrates through two recruitment sites, which lie outside the active site cleft of the kinase. These recruitment sites bind modular-docking sequences called docking sites and are potentially attractive sites for the development of non-ATP competitive inhibitors. The D-recruitment site (DRS) and the F-recruitment site (FRS) bind D-sites and F-sites, respectively. For example, peptides that target the FRS have been proposed to inhibit all ERK2 activity (J. Biol. Chem. 2001, 276, 965-973), however it has not been established whether this inhibition is steric or allosteric in origin. To facilitate inhibitor design and to examine potential coupling of recruitment sites to other ligand recognition sites within ERK2 Energetic coupling within ERK2 was investigated using two new modular peptide substrates for ERK2. Modeling shows that one peptide (Sub-D) recognizes the DRS, while the other peptide (Sub-F) binds the FRS. A steady-state kinetic analysis reveals little evidence of thermodynamic linkage between peptide substrate and ATP. Both peptides are phosphorylated through a random-order sequential mechanism with a kcat/Km comparable to Ets-1, a bona fide ERK2 substrate. Occupancy of the FRS with a peptide containing a modular docking sequence has no effect on the intrinsic ability of ERK2 to phosphorylate Sub-D. Occupancy of the DRS with a peptide containing a modular docking sequence has a slight effect (1.3 ± 0.1-fold increase in kcat) on the intrinsic ability of ERK2 to phosphorylate Sub-F. These data suggest that while docking interactions at the DRS and the FRS are energetically uncoupled, the DRS can exhibit weak communication to the active site. In addition, they suggest that peptides bound to the FRS inhibit the phosphorylation of protein substrates through a steric mechanism. The modeling and kinetic data suggest that the recruitment of ERK2 to cellular locations via its DRS may facilitate the formation of F-site selective ERK2

  7. SNP-SNP interactions between WNT4 and WNT5A were associated with obesity related traits in Han Chinese Population

    PubMed Central

    Dong, Shan-Shan; Hu, Wei-Xin; Yang, Tie-Lin; Chen, Xiao-Feng; Yan, Han; Chen, Xiang-Ding; Tan, Li-Jun; Tian, Qing; Deng, Hong-Wen; Guo, Yan

    2017-01-01

    Considering the biological roles of WNT4 and WNT5A involved in adipogenesis, we aimed to investigate whether SNPs in WNT4 and WNT5A contribute to obesity related traits in Han Chinese population. Targeted genomic sequence for WNT4 and WNT5A was determined in 100 Han Chinese subjects and tag SNPs were selected. Both single SNP and SNP × SNP interaction association analyses with body mass index (BMI) were evaluated in the 100 subjects and another independent sample of 1,627 Han Chinese subjects. Meta-analyses were performed and multiple testing corrections were carried out using the Bonferroni method. Consistent with the Genetic Investigation of ANthropometric Traits (GIANT) dataset results, we didn’t detect significant association signals in single SNP association analyses. However, the interaction between rs2072920 and rs11918967, was associated with BMI after multiple testing corrections (combined P = 2.20 × 10−4). The signal was also significant in each contributing data set. SNP rs2072920 is located in the 3′-UTR of WNT4 and SNP rs11918967 is located in the intron of WNT5A. Functional annotation results revealed that both SNPs might be involved in transcriptional regulation of gene expression. Our results suggest that a combined effect of SNPs via WNT4-WNT5A interaction may affect the variation of BMI in Han Chinese population. PMID:28272483

  8. Loss of Wnt8b has no overt effect on hippocampus development but leads to altered Wnt gene expression levels in dorsomedial telencephalon

    PubMed Central

    McLaughlin, David; Nichols, Jennifer; Price, David J.; Theil, Thomas; Mason, John O.

    2015-01-01

    Wnt signalling proteins regulate many aspects of animal development. We have investigated the function of mouse Wnt8b during forebrain development. Wnt8b is expressed in a highly restricted pattern including the prospective hippocampus and hypothalamus. Mutant mice lacking Wnt8b are viable and healthy. The size and morphology of the hippocampus appeared normal in mutant embryos and adults and we found no evidence of hypothalamic defects in mutants. Wnt8b is also expressed in the neurogenic region of the adult dentate gyrus, however cell proliferation was unchanged in Wnt8b−/− mutants. Mutant embryos did, however, display altered levels of expression of other Wnt genes normally expressed in forebrain. The spatial expression patterns of other Wnt genes and the overall level of canonical Wnt activity were indistinguishable from wild types. Thus, loss of Wnt8b does not give rise to an overt morphological phenotype, but does affect expression levels of other Wnts in developing forebrain. PMID:19890917

  9. Abnormal epigenetic regulation of the gene expression levels of Wnt2b and Wnt7b: Implications for neural tube defects.

    PubMed

    Bai, Baoling; Chen, Shuyuan; Zhang, Qin; Jiang, Qian; Li, Huili

    2016-01-01

    The association between Wnt genes and neural tube defects (NTDs) is recognized, however, it remains to be fully elucidated. Our previous study demonstrated that epigenetic mechanisms are affected in human NTDs. Therefore, the present study aimed to evaluate whether Wnt2b and Wnt7b are susceptible to abnormal epigenetic modification in NTDs, using chromatin immunoprecipitation assays to evaluate histone enrichments and the MassARRAY platform to detect the methylation levels of target regions within Wnt genes. The results demonstrated that the transcriptional activities of Wnt2b and Wnt7b were abnormally upregulated in mouse fetuses with NTDs and, in the GC‑rich promoters of these genes, histone 3 lysine 4 (H3K4) acetylation was enriched, whereas H3K27 trimethylation was reduced. Furthermore, several CpG sites in the altered histone modification of target regions were significantly hypomethylated. The present study also detected abnormal epigenetic modifications of these Wnt genes in human NTDs. In conclusion, the present study detected abnormal upregulation in the levels of Wnt2b and Wnt7b, and hypothesized that the alterations may be due to the ectopic opening of chromatin structure. These results improve understanding of the dysregulation of epigenetic modification of Wnt genes in NTDs.

  10. Distinct requirements of wls, wnt9a, wnt5b and gpc4 in regulating chondrocyte maturation and timing of endochondral ossification

    PubMed Central

    Ling, Irving TC; Rochard, Lucie; Liao, Eric C.

    2017-01-01

    Formation of the mandible requires progressive morphologic change, proliferation, differentiation and organization of chondrocytes preceding osteogenesis. The Wnt signaling pathway is involved in regulating bone development and maintenance. Chondrocytes that are fated to become bone require Wnt to polarize and orientate appropriately to initiate the endochondral ossification program. Although the canonical Wnt signaling has been well studied in the context of bone development, the effects of non-canonical Wnt signaling in regulating the timing of cartilage maturation and subsequent bone formation in shaping ventral craniofacial structure is not fully understood.. Here we examined the role of the non-canonical Wnt signaling pathway (wls, gpc4, wnt5b and wnt9a) in regulating zebrafish Meckel’s cartilage maturation to the onset of osteogenic differentiation. We found that disruption of wls resulted in a significant loss of craniofacial bone, whereas lack of gpc4, wnt5b and wnt9a resulted in severely delayed endochondral ossification. This study demonstrates the importance of the non-canonical Wnt pathway in regulating coordinated ventral cartilage morphogenesis and ossification. PMID:27908786

  11. Loss of Wnt8b has no overt effect on hippocampus development but leads to altered Wnt gene expression levels in dorsomedial telencephalon.

    PubMed

    Fotaki, Vassiliki; Larralde, Osmany; Zeng, Shaoju; McLaughlin, David; Nichols, Jennifer; Price, David J; Theil, Thomas; Mason, John O

    2010-01-01

    Wnt signalling proteins regulate many aspects of animal development. We have investigated the function of mouse Wnt8b during forebrain development. Wnt8b is expressed in a highly restricted pattern including the prospective hippocampus and hypothalamus. Mutant mice lacking Wnt8b are viable and healthy. The size and morphology of the hippocampus appeared normal in mutant embryos and adults, and we found no evidence of hypothalamic defects in mutants. Wnt8b is also expressed in the neurogenic region of the adult dentate gyrus, however, cell proliferation was unchanged in Wnt8b(-/-) mutants. Mutant embryos did, however, display altered levels of expression of other Wnt genes normally expressed in forebrain. The spatial expression patterns of other Wnt genes and the overall level of canonical Wnt activity were indistinguishable from wild-types. Thus, loss of Wnt8b does not give rise to an overt morphological phenotype, but does affect expression levels of other Wnts in developing forebrain. (c) 2009 Wiley-Liss, Inc.

  12. Activation of Wnt11 by Transforming Growth Factor-β Drives Mesenchymal Gene Expression through Non-canonical Wnt Protein Signaling in Renal Epithelial Cells*

    PubMed Central

    Zhang, Peng; Cai, Yi; Soofi, Abdul; Dressler, Gregory R.

    2012-01-01

    Transforming growth factor β1 (TGF-β) promotes renal interstitial fibrosis in vivo and the expression of mesenchymal genes in vitro; however, most of its direct targets in epithelial cells are still elusive. In a screen for genes directly activated by TGF-β, we found that components of the Wnt signaling pathway, especially Wnt11, were targets of activation by TGF-β and Smad3 in primary renal epithelial cells. In gain and loss of function experiments, Wnt11 mediated the actions of TGF-β through enhanced activation of mesenchymal marker genes, such as Zeb1, Snail1, Pai1, and αSMA, without affecting Smad3 phosphorylation. Inhibition of Wnt11 by receptor knockdown or treatment with Wnt inhibitors limited the effects of TGF-β on gene expression. We found no evidence that Wnt11 activated the canonical Wnt signaling pathway in renal epithelial cells; rather, the function of Wnt11 was mediated by the c-Jun N-terminal kinase (JNK) pathway. Consistent with the in vitro results, all the TGF-β, Wnt11, and JNK targets were activated in a unilateral ureteral obstruction (UUO) model of renal fibrosis in vivo. Our findings demonstrated cooperativity among the TGF-β, Wnt11, and JNK signaling pathways and suggest new targets for anti-fibrotic therapy in renal tissue. PMID:22556418

  13. Activation of Wnt11 by transforming growth factor-β drives mesenchymal gene expression through non-canonical Wnt protein signaling in renal epithelial cells.

    PubMed

    Zhang, Peng; Cai, Yi; Soofi, Abdul; Dressler, Gregory R

    2012-06-15

    Transforming growth factor β1 (TGF-β) promotes renal interstitial fibrosis in vivo and the expression of mesenchymal genes in vitro; however, most of its direct targets in epithelial cells are still elusive. In a screen for genes directly activated by TGF-β, we found that components of the Wnt signaling pathway, especially Wnt11, were targets of activation by TGF-β and Smad3 in primary renal epithelial cells. In gain and loss of function experiments, Wnt11 mediated the actions of TGF-β through enhanced activation of mesenchymal marker genes, such as Zeb1, Snail1, Pai1, and αSMA, without affecting Smad3 phosphorylation. Inhibition of Wnt11 by receptor knockdown or treatment with Wnt inhibitors limited the effects of TGF-β on gene expression. We found no evidence that Wnt11 activated the canonical Wnt signaling pathway in renal epithelial cells; rather, the function of Wnt11 was mediated by the c-Jun N-terminal kinase (JNK) pathway. Consistent with the in vitro results, all the TGF-β, Wnt11, and JNK targets were activated in a unilateral ureteral obstruction (UUO) model of renal fibrosis in vivo. Our findings demonstrated cooperativity among the TGF-β, Wnt11, and JNK signaling pathways and suggest new targets for anti-fibrotic therapy in renal tissue.

  14. WNT5A promotes stemness characteristics in nasopharyngeal carcinoma cells leading to metastasis and tumorigenesis.

    PubMed

    Qin, Li; Yin, Yan-Tao; Zheng, Fang-Jing; Peng, Li-Xia; Yang, Chang-Fu; Bao, Ying-Na; Liang, Ying-Ying; Li, Xin-Jian; Xiang, Yan-Qun; Sun, Rui; Li, An-Hua; Zou, Ru-Hai; Pei, Xiao-Qing; Huang, Bi-Jun; Kang, Tie-Bang; Liao, Duan-Fang; Zeng, Yi-Xin; Williams, Bart O; Qian, Chao-Nan

    2015-04-30

    Nasopharyngeal carcinoma (NPC) has the highest metastasis rate among head and neck cancers with unclear mechanism. WNT5A belongs to the WNT family of cysteine-rich secreted glycoproteins. Our previous high-throughput gene expression profiling revealed that WNT5A was up-regulated in highly metastatic cells. In the present study, we first confirmed the elevated expression of WNT5A in metastatic NPC tissues at both the mRNA and protein levels. We then found that WNT5A promoted epithelial-mesenchymal transition (EMT) in NPC cells, induced the accumulation of CD24-CD44+ cells and side population, which are believed to be cancer stem cell characteristics. Moreover, WNT5A promoted the migration and invasion of NPC cells in vitro, while in vivo treatment with recombinant WNT5A promoted lung metastasis. Knocking down WNT5A diminished NPC tumorigenesis in vivo. When elevated expression of WNT5A coincided with the elevated expression of vimentin in the primary NPC, the patients had a poorer prognosis. Among major signaling pathways, protein kinase C (PKC) signaling was activated by WNT5A in NPC cells. A positive feedback loop between WNT5A and phospho-PKC to promote EMT was also revealed. Taken together, these data suggest that WNT5A is an important molecule in promoting stem cell characteristics in NPC, leading to tumorigenesis and metastasis.

  15. Immunoexpression of Wnt/β-catenin signaling pathway proteins in ameloblastoma and calcifying cystic odontogenic tumor.

    PubMed

    Dutra, Sabrina-Nogueira; Pires, Fábio-Ramôa; Armada, Luciana; Azevedo, Rebeca-Souza

    2017-01-01

    Wnt/β-catenin signaling pathway is essential for the beginning of odontogenesis and may be involved in the development and progression of some odontogenic tumors. Thus, the aim of this study was to comparatively evaluate the immunohistochemical expression of Wnt/β-catenin signaling pathway proteins in a series of AME and CCOT. Immunohistochemical reactions were performed using antibodies against Wnt1, Wnt5a and β-catenin in 17 cases of solid AME and 6 cases of CCOT. In the AME group, Wnt1 and Wnt5a were identified in the epithelium in most of the cases, and β-catenin was mainly identified in the cytoplasm of the tumoral cells. In the CCOT group, Wnt1 and Wnt5a were identified in the epithelium and in the ghost cells in almost all the cases, and β-catenin was mainly identified in the cytoplasm and in the nuclei of the tumoral cells. These results contribute to support the importance of Wnt/β-catenin signaling pathway proteins in AME and CCOT tumorigenesis. The abnormal expression of cytoplasmic and/or nuclear β-catenin appears to contribute to the development of both AME and CCOT. In addition, it is possible that Wnt1 and Wnt5a expression in ghost cells can contribute to its histogenesis in CCOT. Key words:Ameloblastoma, β-catenin, calcifying cystic odontogenic tumor, immunohistochemistry, Wnt.

  16. Genome-wide analysis of a Wnt1-regulated transcriptional network implicates neurodegenerative pathways.

    PubMed

    Wexler, Eric M; Rosen, Ezra; Lu, Daning; Osborn, Gregory E; Martin, Elizabeth; Raybould, Helen; Geschwind, Daniel H

    2011-10-04

    Wnt proteins are critical to mammalian brain development and function. The canonical Wnt signaling pathway involves the stabilization and nuclear translocation of β-catenin; however, Wnt also signals through alternative, noncanonical pathways. To gain a systems-level, genome-wide view of Wnt signaling, we analyzed Wnt1-stimulated changes in gene expression by transcriptional microarray analysis in cultured human neural progenitor (hNP) cells at multiple time points over a 72-hour time course. We observed a widespread oscillatory-like pattern of changes in gene expression, involving components of both the canonical and the noncanonical Wnt signaling pathways. A higher-order, systems-level analysis that combined independent component analysis, waveform analysis, and mutual information-based network construction revealed effects on pathways related to cell death and neurodegenerative disease. Wnt effectors were tightly clustered with presenilin1 (PSEN1) and granulin (GRN), which cause dominantly inherited forms of Alzheimer's disease and frontotemporal dementia (FTD), respectively. We further explored a potential link between Wnt1 and GRN and found that Wnt1 decreased GRN expression by hNPs. Conversely, GRN knockdown increased WNT1 expression, demonstrating that Wnt and GRN reciprocally regulate each other. Finally, we provided in vivo validation of the in vitro findings by analyzing gene expression data from individuals with FTD. These unbiased and genome-wide analyses provide evidence for a connection between Wnt signaling and the transcriptional regulation of neurodegenerative disease genes.

  17. The role of Ppt/Wnt5 in regulating cell shape and movement during zebrafish gastrulation.

    PubMed

    Kilian, Beate; Mansukoski, Hannu; Barbosa, Filipa Carreira; Ulrich, Florian; Tada, Masazumi; Heisenberg, Carl Philipp

    2003-04-01

    Wnt genes play important roles in regulating patterning and morphogenesis during vertebrate gastrulation. In zebrafish, slb/wnt11 is required for convergence and extension movements, but not cell fate specification during gastrulation. To determine if other Wnt genes functionally interact with slb/wnt11, we analysed the role of ppt/wnt5 during zebrafish gastrulation. ppt/wnt5 is maternally provided and zygotically expressed at all stages during gastrulation. The analysis of ppt mutant embryos reveals that Ppt/Wnt5 regulates cell elongation and convergent extension movements in posterior regions of the gastrula, while its function in more anterior regions is largely redundant to that of Slb/Wnt11. Frizzled-2 functions downstream of ppt/wnt5, indicating that it might act as a receptor for Ppt/Wnt5 in this process. The characterisation of the role of Ppt/Wnt5 provides insight into the functional diversity of Wnt genes in regulating vertebrate gastrulation movements.

  18. Gpr177/mouse Wntless is essential for Wnt-mediated craniofacial and brain development.

    PubMed

    Fu, Jiang; Ivy Yu, Hsiao-Man; Maruyama, Takamitsu; Mirando, Anthony J; Hsu, Wei

    2011-02-01

    We have previously demonstrated that Gpr177, the mouse orthologue of Drosophila Wls/Evi/Srt, is required for establishment of the anterior-posterior axis. The Gpr177 null phenotype is highly reminiscent to the loss of Wnt3, the earliest abnormality among all Wnt knockouts in mice. The expression of Gpr177 in various cell types and tissues lead us to hypothesize that reciprocal regulation of Wnt and Gpr177 is essential for the Wnt-dependent developmental and pathogenic processes. Here, we create a new mouse strain permitting conditional inactivation of Gpr177. The loss of Gpr177 in the Wnt1-expressing cells causes mid/hindbrain and craniofacial defects which are far more severe than the Wnt1 knockout, but resemble the double knockout of Wnt1 and Wnt3a as well as β-catenin deletion in the Wnt1-expressing cells. Our findings demonstrate the importance of Gpr177 in Wnt1-mediated development of the mouse embryo, suggesting an overlapping function of Wnt family members in the Wnt1-expressing cells.

  19. Reciprocal regulation of Wnt and Gpr177/mouse Wntless is required for embryonic axis formation.

    PubMed

    Fu, Jiang; Jiang, Ming; Mirando, Anthony J; Yu, Hsiao-Man Ivy; Hsu, Wei

    2009-11-03

    Members of the Wnt family are secreted glycoproteins that trigger cellular signals essential for proper development of organisms. Cellular signaling induced by Wnt proteins is involved in diverse developmental processes and human diseases. Previous studies have generated an enormous wealth of knowledge on the events in signal-receiving cells. However, relatively little is known about the making of Wnt in signal-producing cells. Here, we describe that Gpr177, the mouse orthologue of Drosophila Wls, is expressed during formation of embryonic axes. Embryos with deficient Gpr177 exhibit defects in establishment of the body axis, a phenotype highly reminiscent to the loss of Wnt3. Although many different mammalian Wnt proteins are required for a wide range of developmental processes, the Wnt3 ablation exhibits the earliest developmental abnormality. This suggests that the Gpr177-mediated Wnt production cannot be substituted. As a direct target of Wnt, Gpr177 is activated by beta-catenin and LEF/TCF-dependent transcription. This activation alters the cellular distributions of Gpr177 which binds to Wnt proteins and assists their sorting and secretion in a feedback regulatory mechanism. Our findings demonstrate that the loss of Gpr177 affects Wnt production in the signal-producing cells, leading to alterations of Wnt signaling in the signal-receiving cells. A reciprocal regulation of Wnt and Gpr177 is essential for the patterning of the anterior-posterior axis during mammalian development.

  20. Immunoexpression of Wnt/β-catenin signaling pathway proteins in ameloblastoma and calcifying cystic odontogenic tumor

    PubMed Central

    Dutra, Sabrina-Nogueira; Pires, Fábio-Ramôa; Armada, Luciana

    2017-01-01

    Background Wnt/β-catenin signaling pathway is essential for the beginning of odontogenesis and may be involved in the development and progression of some odontogenic tumors. Thus, the aim of this study was to comparatively evaluate the immunohistochemical expression of Wnt/β-catenin signaling pathway proteins in a series of AME and CCOT. Material and Methods Immunohistochemical reactions were performed using antibodies against Wnt1, Wnt5a and β-catenin in 17 cases of solid AME and 6 cases of CCOT. Results In the AME group, Wnt1 and Wnt5a were identified in the epithelium in most of the cases, and β-catenin was mainly identified in the cytoplasm of the tumoral cells. In the CCOT group, Wnt1 and Wnt5a were identified in the epithelium and in the ghost cells in almost all the cases, and β-catenin was mainly identified in the cytoplasm and in the nuclei of the tumoral cells. Conclusions These results contribute to support the importance of Wnt/β-catenin signaling pathway proteins in AME and CCOT tumorigenesis. The abnormal expression of cytoplasmic and/or nuclear β-catenin appears to contribute to the development of both AME and CCOT. In addition, it is possible that Wnt1 and Wnt5a expression in ghost cells can contribute to its histogenesis in CCOT. Key words:Ameloblastoma, β-catenin, calcifying cystic odontogenic tumor, immunohistochemistry, Wnt. PMID:28149478

  1. ERK5 signalling rescues intestinal epithelial turnover and tumour cell proliferation upon ERK1/2 abrogation

    PubMed Central

    de Jong, Petrus R.; Taniguchi, Koji; Harris, Alexandra R.; Bertin, Samuel; Takahashi, Naoki; Duong, Jen; Campos, Alejandro D.; Powis, Garth; Corr, Maripat; Karin, Michael; Raz, Eyal

    2016-01-01

    The ERK1/2 MAPK signalling module integrates extracellular cues that induce proliferation and differentiation of epithelial lineages, and is an established oncogenic driver, particularly in the intestine. However, the interrelation of the ERK1/2 module relative to other signalling pathways in intestinal epithelial cells and colorectal cancer (CRC) is unclear. Here we show that loss of Erk1/2 in intestinal epithelial cells results in defects in nutrient absorption, epithelial cell migration and secretory cell differentiation. However, intestinal epithelial cell proliferation is not impeded, implying compensatory mechanisms. Genetic deletion of Erk1/2 or pharmacological targeting of MEK1/2 results in supraphysiological activity of the ERK5 pathway. Furthermore, targeting both pathways causes a more effective suppression of cell proliferation in murine intestinal organoids and human CRC lines. These results suggest that ERK5 provides a common bypass route in intestinal epithelial cells, which rescues cell proliferation upon abrogation of ERK1/2 signalling, with therapeutic implications in CRC. PMID:27187615

  2. ERK inhibition sensitizes cancer cells to oleanolic acid-induced apoptosis through ERK/Nrf2/ROS pathway.

    PubMed

    Liu, Jia; Ma, Leina; Chen, Xiao; Wang, Jianxun; Yu, Tao; Gong, Ying; Ma, Aiguo; Zheng, Lanhong; Liang, Hui

    2016-06-01

    Oleanolic acid (OA) is a natural triterpenoid that is widely distributed in edible and medicinal plants. OA exerts anti-tumor activity on a wide range of cancer cells primarily through inducing apoptosis. Dysregulated ERK signaling is closely complicated in the biology of cancer, such as metastasis, proliferation, and survival, and it can be activated by various stimuli. In this study, we found that OA induced the activation of ERK in cancer cells. ERK activation compromised the apoptosis induced by OA. Blocking ERK activation by U0126 or siRNAs was able to potentiate the pro-apoptotic activity of OA on cancer cells. OA was shown to promote ERK-dependent Nrf2 expression in cancer cells, and in turn, Nrf2 expression was able to suppress OA-induced ROS generation. Blockade of Nrf2 expression was able to increase ROS levels and apoptotic death in cancer cells. In conclusion, we provided evidences that ERK activation is a mechanism underlying the resistance of cancer cells to OA-induced apoptosis and targeting ERK is a promising strategy to enhance the anti-tumor efficacy of OA.

  3. Role of Wnt/β-catenin, Wnt/c-Jun N-terminal kinase and Wnt/Ca2+ pathways in cisplatin-induced chemoresistance in ovarian cancer

    PubMed Central

    Huang, Lu; Jin, Ye; Feng, Shujun; Zou, Yuqing; Xu, Sainan; Qiu, Shuang; Li, Ling; Zheng, Jianhua

    2016-01-01

    The aim of the present study was to explore the expression of Wnt signaling proteins β-catenin, c-Jun N-terminal kinase (JNK) and Ca2+/calmodulin-dependent protein kinase II (CaMKII) in ovarian cancer cells, and assess the correlation between this expression and cisplatin-induced chemoresistance. SKOV3 ovarian carcinoma cells and SKOV3/DDP (cisplatin resistant) cells were treated with cisplatin in the absence or presence of a Wnt signaling activator (CHIR-99021, glycogen synthase kinase 3β inhibitor) or inhibitor (XAV-939, tankyrase inhibitor). Following incubation for 48 h, cell viability, proliferation and cytotoxicity were measured using a sensitive colorimetric cell counting kit. Expression levels of β-catenin, JNK and CaMKII were detected by western blot and immunofluorescence staining. The results of the current study identified that β-catenin and JNK expression levels were significantly higher (P<0.01 and P<0.05 respectively), while CaMKII expression was lower (P>0.05), in SKOV3/DDP cells compared with SKOV3 cells. Moreover, following treatment with 20 µM cisplatin, reduced expression of β-catenin and JNK (P<0.05 and P<0.01 respectively), and increased expression of CaMKII (P<0.01), was observed in SKOV3 and SKOV3/DPP cell lines. Furthermore, inhibition of β-catenin signaling by XAV-939 effectively reversed cisplatin chemoresistance in SKOV3/DDP cells. Similarly, XAV-939 downregulated JNK expression (P<0.001), but upregulated CaMKII expression (P<0.001), in SKOV3/DDP cells. In conclusion, abnormal activation of Wnt/β-catenin and Wnt/JNK signaling pathways in ovarian cancer cells promotes cisplatin resistance, while the Wnt/Ca2+ signaling pathway reduces cisplatin resistance. This indicates that β-catenin, JNK and CaMKII are potential therapeutic targets in chemoresistant ovarian cancers. PMID:28101169

  4. Regulation of ERK5 by insulin and angiotensin-II in vascular smooth muscle cells

    SciTech Connect

    Sharma, Girish; Goalstone, Marc Lee; E-mail: Marc.Goalstone@uchsc.edu

    2007-03-23

    ERK5 is involved in proliferation of vascular smooth muscle cells (VSMC). The proliferative actions of insulin and angiotensin-II (A-II) in VSMC are mediated in part by ERK1/2. We hypothesized that insulin and A-II also regulate ERK5 activity in VSMC. Acute treatment (<60 min) with insulin or A-II increased phosphorylation of ERK1/2 at 15 min and ERK5 at 5 min. Chronic treatment ({<=}8 h) with insulin increased ERK1/2 phosphorylation by 4 h and ERK5 by 8 h. A-II-stimulated phosphorylation of ERK1/2 by 8 h and ERK5 by 4 h. The EC{sub 50} for insulin treatment effecting ERK1/2 and ERK5 phosphorylation was 1.5 and 0.1 nM, whereas the EC{sub 50} for A-II was 2 nM, each. Insulin plus A-II induced an additive effect only on ERK5 phosphorylation. Inhibition of insulin- and A-II-stimulated phosphorylation of ERK5 and ERK1/2 by PD98059 and Wortmannin exhibited differential and time-dependent effects. Taken together, these data indicate that insulin and A-II regulate the activity of ERK5, but different from that seen for ERK1/2.

  5. Genomic insights into WNT/β-catenin signaling

    PubMed Central

    Rosenbluh, Joseph; Wang, Xiaoxing; Hahn, William C.

    2014-01-01

    The canonical WNT pathway regulates the stability of the proto-oncogene β-catenin and is aberrantly activated in many cancer types. Studies in a wide range of experimental models confirm that β-catenin activity is required for tumor initiation in cancers where this pathway is deregulated. However, to date this pathway has proven to be challenging to target therapeutically. Moreover, several lines of evidence suggest that other components and regulators of β-catenin exist. Here we will describe recent structural and functional studies describing genomic alterations and new regulators of β-catenin that lead to aberrant activation of the WNT/β-catenin pathway. These findings provide new insights into the biology of WNT/β-catenin signaling and suggest potential therapeutic opportunities. PMID:24365576

  6. Crossroads of Wnt and Hippo in epithelial tissues.

    PubMed

    Bernascone, Ilenia; Martin-Belmonte, Fernando

    2013-08-01

    Epithelial tissues undergo constant growth and differentiation during embryonic development and to replace damaged tissue in adult organs. These processes are governed by different signaling pathways that ultimately control the expression of genes associated with cell proliferation, patterning, and death. One essential pathway is Wnt, which controls tubulogenesis in several epithelial organs. Recently, Wnt has been closely linked to other signaling pathways, such as Hippo, that orchestrate proliferation and apoptosis to control organ size. There is evidence that epithelial cell junctions may sequester the transcription factors that act downstream of these signaling pathways, which would represent an important aspect of their functional regulation and their influence on cell behavior. Here, we review the transcriptional control exerted by the Wnt and Hippo signaling pathways during epithelial growth, patterning, and differentiation and recent advances in understanding of the regulation and crosstalk of these pathways in epithelial tissues.

  7. IGFBP‐3 inhibits Wnt signaling in metastatic melanoma cells