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Matthew J. Sikora - One of the best experts on this subject based on the ideXlab platform.
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WNT4 balances development vs disease in gynecologic tissues and women s health
Endocrinology, 2021Co-Authors: Lauren M Pitzer, Marisa R Moroney, Natalie J Nokoff, Matthew J. SikoraAbstract:The WNT family of proteins is crucial in numerous developmental pathways and tissue homeostasis. WNT4, in particular, is uniquely implicated in the development of the female phenotype in the fetus, and in the maintenance of mullerian and reproductive tissues. WNT4 dysfunction or dysregulation can drive sex-reversal syndromes, highlighting the key role of WNT4 in sex determination. WNT4 is also critical in gynecologic pathologies later in life, including several cancers, uterine fibroids, endometriosis, and infertility. The role of WNT4 in normal decidualization, implantation, and gestation is being increasingly appreciated, while aberrant activation of WNT4 signaling is being linked both to gynecologic and breast cancers. Notably, single-nucleotide polymorphisms (SNPs) at the WNT4 gene locus are strongly associated with these pathologies and may functionally link estrogen and estrogen receptor signaling to upregulation and activation of WNT4 signaling. Importantly, in each of these developmental and disease states, WNT4 gene expression and downstream WNT4 signaling are regulated and executed by myriad tissue-specific pathways. Here, we review the roles of WNT4 in women's health with a focus on sex development, and gynecologic and breast pathologies, and our understanding of how WNT4 signaling is controlled in these contexts. Defining WNT4 functions provides a unique opportunity to link sex-specific signaling pathways to women's health and disease.
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estrogen regulation of mtor signaling and mitochondrial function in invasive lobular carcinoma cell lines requires WNT4
Cancers, 2020Co-Authors: Madeleine T Shackleford, Deviyani M. Rao, Steffi Oesterreich, Evelyn K Bordeaux, Hannah M Hicks, Christina G Towers, Joseph L Sottnik, Matthew J. SikoraAbstract:Invasive lobular carcinoma of the breast (ILC) is strongly estrogen-driven and represents a unique context for estrogen receptor (ER) signaling. In ILC, ER controls the expression of the Wnt ligand WNT4, which is critical for endocrine response and anti-estrogen resistance. However, signaling mediated by WNT4 is cell type- and tissue-specific, and has not been explored in ILC. We utilized reverse phase protein array (RPPA) to characterize ER and WNT4-driven signaling in ILC cells and identified that WNT4 mediates downstream mTOR signaling via phosphorylation of S6 Kinase. Additionally, ER and WNT4 control levels of MCL-1, which is associated with regulation of mitochondrial function. In this context, WNT4 knockdown led to decreased ATP production and increased mitochondrial fragmentation. WNT4 regulation of both mTOR signaling and MCL-1 were also observed in anti-estrogen resistant models of ILC. We identified that high WNT4 expression is associated with similar mTOR pathway activation in ILC and serous ovarian cancer tumors, suggesting that WNT4 signaling is active in multiple tumor types. The identified downstream pathways offer insight into WNT4 signaling and represent potential targets to overcome anti-estrogen resistance for patients with ILC.
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wnt family member 4 WNT4 and wnt3a activate cell autonomous wnt signaling independent of porcupine o acyltransferase or wnt secretion
Journal of Biological Chemistry, 2019Co-Authors: Deviyani M. Rao, Rebecca Ferguson, Madeleine T Shackleford, Evelyn K Bordeaux, Joseph L Sottnik, Tomomi M Yamamoto, Elizabeth A Wellberg, Benjamin G Bitler, Matthew J. SikoraAbstract:Porcupine O-acyltransferase (PORCN) is considered essential for Wnt secretion and signaling. However, we observed that PORCN inhibition does not phenocopy the effects of WNT4 knockdown in WNT4-dependent breast cancer cells. This suggests a unique relationship between PORCN and WNT4 signaling. To examine the role of PORCN in WNT4 signaling, here we overexpressed WNT4 or WNT3A in breast cancer, ovarian cancer, and fibrosarcoma cell lines. Conditioned media from these lines and co-culture systems were used to assess the dependence of Wnt secretion and activity on the critical Wnt secretion proteins PORCN and Wnt ligand secretion (WLS) mediator. We observed that WLS is universally required for Wnt secretion and paracrine signaling. In contrast, the dependence of WNT3A secretion and activity on PORCN varied across the cell lines, and WNT4 secretion was PORCN-independent in all models. Surprisingly, WNT4 did not exhibit paracrine activity in any tested context. Absent the expected paracrine activity of secreted WNT4, we identified cell-autonomous Wnt signaling activation by WNT4 and WNT3A, independent of PORCN or Wnt secretion. The PORCN-independent, cell-autonomous Wnt signaling demonstrated here may be critical in WNT4-driven cellular contexts or in those that are considered to have dysfunctional Wnt signaling.
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estrogen controls mtor signaling and mitochondrial function via WNT4 in lobular carcinoma cells
bioRxiv, 2019Co-Authors: Madeleine T Shackleford, Deviyani M. Rao, Steffi Oesterreich, Evelyn K Bordeaux, Hannah M Hicks, Matthew J. SikoraAbstract:Abstract Invasive lobular carcinoma of the breast (ILC) is strongly estrogen-driven, and represents a unique context for estrogen receptor (ER) signaling. In ILC, ER controls the expression of the Wnt ligand WNT4, which is critical for endocrine response and anti-estrogen resistance, yet signaling mediated by WNT4 is poorly understood. We utilized reverse phase protein array (RPPA) to characterize ER and WNT4-driven signaling in ILC cells, and identified WNT4 as a mediator of downstream mTOR signaling via p70-S6K. Independent of mTOR/p70-S6K, ER and WNT4 control levels of MCL-1, which is associated with mitochondrial function. In this context, knockdown of WNT4 caused accumulation of reactive oxygen species and decreased ATP production that precede cell death. WNT4 regulation of both mTOR signaling and MCL-1 levels was also observed in anti-estrogen resistant models of ILC. Further, we identified that high WNT4 expression is associated with similar mTOR pathway activation in serous ovarian cancer tumors, suggesting that this WNT4 pathway is important in multiple tumor types. The identified downstream pathways represent potential targets to inhibit WNT4 signaling in ovarian cancer and overcome anti-estrogen resistance for patients with ILC.
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WNT4 and WNT3A activate cell autonomous Wnt signaling independent of secretion
2018Co-Authors: Deviyani M. Rao, Rebecca Ferguson, Matthew J. SikoraAbstract:Background: Wnt protein secretion and signaling canonically require the enzyme PORCN, suggesting PORCN inhibitors can effectively treat Wnt-dependent forms of cancer. We tested the ability of PORCN inhibitors to inhibit the growth of WNT4-dependent breast cancer cell lines, but found PORCN inhibitors did not phenocopy WNT4 knockdown. This led us to investigate the role of PORCN in WNT4 signaling. Methods: Breast cancer cell line MDA MB 134VI, fibrosarcoma cell line HT1080, and PORCN-knockout HT1080 were used as model systems for WNT4 signaling (i.e. derived from WNT4-responsive tissues; breast and bone, respectively), along with additional breast and ovarian cancer cell lines. Using these models, WNT3A or WNT4 were constitutively over-expressed. Conditioned medium was used to assess Wnt secretion and paracrine activity, and dependence on the Wnt secretion proteins PORCN and Wntless (WLS). Secreted Wnt protein activity was compared to recombinant Wnt protein. Results: Whereas WLS was universally required for Wnt secretion and paracrine signaling, the role of PORCN varied across contexts. PORCN was not required for WNT3A secretion in all cell lines, but PORCN was universally required for paracrine activity of WNT3A. In contrast, WNT4 secretion was PORCN-independent in all models tested. However, while recombinant WNT4 activated Wnt signaling in treated cells, secreted WNT4 did not present paracrine activity in any tested context. Absent the expected paracrine activity of secreted WNT4, we examined Wnt signaling directly in cells over-expressing Wnt proteins. This revealed cell autonomous activation of Wnt signaling, independent of secretion, by both WNT4 and WNT3A. Direct transfection of Wnt protein activated non-canonical Wnt signaling independent of membrane receptor activation. Conclusions: PORCN is differentially required for secretion and paracrine activity of WNT4 and WNT3A, specific to the cell type expressing Wnt proteins and the paracrine recipient of Wnt signals. Wnt proteins can activate cell autonomous signaling, independent of PORCN and of secretion. This PORCN-independent mode of Wnt signaling may be critical in cell contexts that are otherwise considered to have dysfunctional Wnt signaling.
Steffi Oesterreich - One of the best experts on this subject based on the ideXlab platform.
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estrogen regulation of mtor signaling and mitochondrial function in invasive lobular carcinoma cell lines requires WNT4
Cancers, 2020Co-Authors: Madeleine T Shackleford, Deviyani M. Rao, Steffi Oesterreich, Evelyn K Bordeaux, Hannah M Hicks, Christina G Towers, Joseph L Sottnik, Matthew J. SikoraAbstract:Invasive lobular carcinoma of the breast (ILC) is strongly estrogen-driven and represents a unique context for estrogen receptor (ER) signaling. In ILC, ER controls the expression of the Wnt ligand WNT4, which is critical for endocrine response and anti-estrogen resistance. However, signaling mediated by WNT4 is cell type- and tissue-specific, and has not been explored in ILC. We utilized reverse phase protein array (RPPA) to characterize ER and WNT4-driven signaling in ILC cells and identified that WNT4 mediates downstream mTOR signaling via phosphorylation of S6 Kinase. Additionally, ER and WNT4 control levels of MCL-1, which is associated with regulation of mitochondrial function. In this context, WNT4 knockdown led to decreased ATP production and increased mitochondrial fragmentation. WNT4 regulation of both mTOR signaling and MCL-1 were also observed in anti-estrogen resistant models of ILC. We identified that high WNT4 expression is associated with similar mTOR pathway activation in ILC and serous ovarian cancer tumors, suggesting that WNT4 signaling is active in multiple tumor types. The identified downstream pathways offer insight into WNT4 signaling and represent potential targets to overcome anti-estrogen resistance for patients with ILC.
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estrogen controls mtor signaling and mitochondrial function via WNT4 in lobular carcinoma cells
bioRxiv, 2019Co-Authors: Madeleine T Shackleford, Deviyani M. Rao, Steffi Oesterreich, Evelyn K Bordeaux, Hannah M Hicks, Matthew J. SikoraAbstract:Abstract Invasive lobular carcinoma of the breast (ILC) is strongly estrogen-driven, and represents a unique context for estrogen receptor (ER) signaling. In ILC, ER controls the expression of the Wnt ligand WNT4, which is critical for endocrine response and anti-estrogen resistance, yet signaling mediated by WNT4 is poorly understood. We utilized reverse phase protein array (RPPA) to characterize ER and WNT4-driven signaling in ILC cells, and identified WNT4 as a mediator of downstream mTOR signaling via p70-S6K. Independent of mTOR/p70-S6K, ER and WNT4 control levels of MCL-1, which is associated with mitochondrial function. In this context, knockdown of WNT4 caused accumulation of reactive oxygen species and decreased ATP production that precede cell death. WNT4 regulation of both mTOR signaling and MCL-1 levels was also observed in anti-estrogen resistant models of ILC. Further, we identified that high WNT4 expression is associated with similar mTOR pathway activation in serous ovarian cancer tumors, suggesting that this WNT4 pathway is important in multiple tumor types. The identified downstream pathways represent potential targets to inhibit WNT4 signaling in ovarian cancer and overcome anti-estrogen resistance for patients with ILC.
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WNT4 mediates estrogen receptor signaling and endocrine resistance in invasive lobular carcinoma cell lines
Breast Cancer Research, 2016Co-Authors: Matthew J. Sikora, Caroline M Alexander, Britta M Jacobsen, Kevin M Levine, Jian Chen, Nancy E Davidson, Adrian V Lee, Steffi OesterreichAbstract:Invasive lobular carcinoma (ILC) of the breast typically presents with clinical biomarkers consistent with a favorable response to endocrine therapies, and over 90 % of ILC cases express the estrogen receptor (ER). However, a subset of ILC cases may be resistant to endocrine therapies, suggesting that ER biology is unique in ILC. Using ILC cell lines, we previously demonstrated that ER regulates a distinct gene expression program in ILC cells, and we hypothesized that these ER-driven pathways modulate the endocrine response in ILC. One potential novel pathway is via the Wnt ligand WNT4, a critical signaling molecule in mammary gland development regulated by the progesterone receptor. The ILC cell lines MDA-MB-134-VI, SUM44PE, and BCK4 were used to assess WNT4 gene expression and regulation, as well as the role of WNT4 in estrogen-regulated proliferation. To assess these mechanisms in the context of endocrine resistance, we developed novel ILC endocrine-resistant long-term estrogen-deprived (ILC-LTED) models. ILC and ILC-LTED cell lines were used to identify upstream regulators and downstream signaling effectors of WNT4 signaling. ILC cells co-opted WNT4 signaling by placing it under direct ER control. We observed that ER regulation of WNT4 correlated with use of an ER binding site at the WNT4 locus, specifically in ILC cells. Further, WNT4 was required for endocrine response in ILC cells, as WNT4 knockdown blocked estrogen-induced proliferation. ILC-LTED cells remained dependent on WNT4 for proliferation, by either maintaining ER function and WNT4 regulation or uncoupling WNT4 from ER and upregulating WNT4 expression. In the latter case, WNT4 expression was driven by activated nuclear factor kappa-B signaling in ILC-LTED cells. In ILC and ILC-LTED cells, WNT4 led to suppression of CDKN1A/p21, which is critical for ILC cell proliferation. CDKN1A knockdown partially reversed the effects of WNT4 knockdown. WNT4 drives a novel signaling pathway in ILC cells, with a critical role in estrogen-induced growth that may also mediate endocrine resistance. WNT4 signaling may represent a novel target to modulate endocrine response specifically for patients with ILC.
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abstract 862 WNT4 mediates endocrine response and resistance in invasive lobular carcinoma cell lines and patient tumor explants
Cancer Research, 2016Co-Authors: Matthew J. Sikora, Caroline M Alexander, Courtney L Andersen, Priscilla M Mcauliffe, Steffi OesterreichAbstract:Invasive lobular carcinoma (ILC) is a breast cancer subtype affecting ∼30,000 U.S. women annually. Over 90% of ILC are estrogen receptor (ER)-positive; however, endocrine therapy may have poorer efficacy in a subset of ILC patients versus invasive ductal carcinoma (IDC) patients. This prompted us to assess global ER activity in ILC cell lines MDA MB 134VI (MM134) and SUM44PE (44PE) to identify novel mediators of ER signaling. These analyses identified the Wnt ligand WNT4 as an ILC-specific ER target gene, with an ILC-specific ER binding site (ERBS) at the WNT4 locus. Considering the critical role of WNT4 in normal mammary gland expansion, we hypothesize that ILC cells utilize WNT4 signaling to drive endocrine response and resistance. We assessed whether WNT4 is necessary for ILC cell growth using siRNA. WNT4 knockdown completely blocked estrogen-induced growth in ILC cells but not IDC cells. In parallel, the WNT4 ERBS was only occupied in ILC cells in response to estrogen, but progesterone-induced WNT4 in IDC was not associated with this ERBS. This suggests that, via the ILC-specific WNT4 ERBS, ILC cells drive estrogen-regulated proliferation by hijacking a developmental Wnt pathway. Wnt pathways typically activate â-catenin; however, we observed â-catenin dysfunction in ILC cells and that WNT4 cannot activate â-catenin. Thus, WNT4 signals in ILC cells via a novel non-canonical pathway. Using long-term estrogen-deprived (LTED) variants of MM134 and 44PE (4 and 2 lines, respectively), we assessed WNT4 in ILC endocrine resistance. WNT4 is over-expressed, but uncoupled from ER, in all MM134-LTED. Conversely, WNT4 is reduced in 44PE-LTED but remains ER-regulated; ER occupies the WNT4 ERBS only in 44PE-LTED cells and not MM134-LTED. Using siRNA, MM134-LTED (high WNT4) are growth-inhibited by WNT4 knockdown, while 44PE-LTED (low WNT4) are insensitive. However, WNT4 knockdown sensitizes 44PE-LTED to endocrine therapy. Taken together, uncoupling and upregulating WNT4 or WNT4/ER cross-talk may represent convergent endocrine resistance mechanisms in ILC. To assess the role of WNT4 in patient ILC, we examined WNT4 protein in archival breast tumors and observed that WNT4 is frequently expressed in ILC and IDC tumors. We also examined WNT4 regulation and endocrine response in patient tumor explants. We observed ER regulation of WNT4 directly in ILC tissues that correlated with sensitivity to fulvestrant but resistance to tamoxifen; this may mimic clinical endocrine resistance. Ongoing studies are assessing WNT4 as a biomarker and mediator of endocrine resistance in ILC. Clinical observations suggest that ER regulates unique pathways in ILC. We identified WNT4 as a downstream effector of endocrine signaling in ILC, with critical roles in both estrogen-induced growth and endocrine resistance. WNT4 signaling may represent a novel target to modulate endocrine response specifically for ILC patients. Citation Format: Matthew J. Sikora, Courtney L. Andersen, Caroline M. Alexander, Priscilla M. McAuliffe, Steffi Oesterreich. WNT4 mediates endocrine response and resistance in invasive lobular carcinoma cell lines and patient tumor explants. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 862.
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abstract p3 04 02 invasive lobular carcinoma cell lines utilize WNT4 signaling to mediate estrogen induced growth
Cancer Research, 2016Co-Authors: Matthew J. Sikora, Steffi OesterreichAbstract:Invasive lobular carcinoma (ILC) is a histological subtype of breast cancer representing 10-15% of newly diagnosed breast tumors. Over 90% of ILC are estrogen receptor (ER)-positive, however, endocrine response and estrogen signaling are not well understood in ILC. Retrospective analyses suggest that ILC patients treated with endocrine therapy have poorer outcomes than invasive ductal carcinoma (IDC) patients, and that ILC patients may not benefit from adjuvant tamoxifen. Based on these observations, we hypothesize that ER regulates unique signaling pathways in ILC cells that control growth and endocrine response. To identify putative targets that regulate endocrine response in ILC, we assessed genome-wide ER-mediated gene expression and ER genomic binding in the ILC cell lines MDA MB 134VI (MM134) and SUM44PE (SUM44). Among ILC-specific estrogen-regulated genes, the most strongly induced was the secreted ligand WNT4. Additionally, we identified an ILC-specific ER binding site (ERBS) at WNT4, suggesting that WNT4 is directly controlled by ER in ILC cells. Direct ER-regulation of WNT4 is in contrast to control of WNT4 by progesterone receptor (PR) during mammary gland development; however, further ER ChIP experiments suggest that ILC cells place WNT4 under ER control via unique use of the WNT4 ERBS. Thus, ILC cells may hijack a tightly regulated developmental program to drive estrogen-regulated cell phenotypes. Based on the role of WNT4 in mammary gland growth and expansion, we hypothesized that WNT4 is required for estrogen-induced growth in ILC cells. To test this, we used siRNA to knock down WNT4 in ILC and IDC cell lines. Using either of two siRNAs, WNT4 knockdown completely blocked estrogen-induced growth in ILC cells, but not IDC cells. Consistent with this, WNT4 knockdown abrogated estrogen-regulation of a subset of ER-target genes in MM134 cells, suggesting that these genes are downstream of WNT4 signaling. Wnt signaling typically acts via the canonical, β-catenin-dependent pathway; however, we observed that β-catenin is dysfunctional in ILC cells, and WNT4 cannot activate β-catenin signaling in cancer cell lines. This suggests that WNT4 regulates estrogen-induced growth in ILC cells via a novel non-canonical signaling pathway. Clinical observations suggest that ER regulates unique downstream pathways in ILC. We identified WNT4 as a putative downstream effector of ER signaling in ILC, as WNT4 is strongly-induced and directly regulated by ER specifically in ILC cells. Further, WNT4 is necessary for estrogen-induced growth in ILC cells, and likely signals via a non-canonical signaling pathway. Targeting WNT4 signaling represents a novel approach to modulate endocrine response specifically for ILC patients. Future studies will focus on characterizing the signaling pathway controlled by WNT4 in order to identify putative therapeutic targets. Citation Format: Sikora MJ, Oesterreich S. Invasive lobular carcinoma cell lines utilize WNT4 signaling to mediate estrogen-induced growth. [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P3-04-02.
Blanche Capel - One of the best experts on this subject based on the ideXlab platform.
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testis development requires the repression of WNT4 by fgf signaling
Developmental Biology, 2012Co-Authors: Samantha Jameson, Yitzu Lin, Blanche CapelAbstract:The bipotential gonad expresses genes associated with both the male and female pathways. Adoption of the male testicular fate is associated with the repression of many female genes including WNT4. However, the importance of repression of WNT4 to the establishment of male development was not previously determined. Deletion of either Fgf9 or Fgfr2 in an XY gonad resulted in up-regulation of WNT4 and male-to-female sex reversal. We investigated whether the deletion if WNT4 could rescue sex reversal in Fgf9 and Fgfr2 mutants. XY Fgf9/WNT4 and Fgfr2/WNT4 double mutants developed testes with male somatic and germ cells present, suggesting that the primary role of Fgf signaling is the repression of female-promoting genes. Thus, the decision to adopt the male fate is based not only on whether male genes, such as Sox9, are expressed, but also on the active repression of female genes, such as WNT4. Because loss of WNT4 results in the up-regulation of Fgf9, we also tested the possibility that derepression of Fgf9 was responsible for the aspects of male development observed in XX WNT4 mutants. However, we found that the relationship between these two signaling factors is not symmetric: loss of Fgf9 in XX WNT4−/− gonads does not rescue their partial female-to-male sex-reversal.
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a microarray analysis of the xx WNT4 mutant gonad targeted at the identification of genes involved in testis vascular differentiation
Gene Expression Patterns, 2007Co-Authors: Douglas Coveney, Andrea J Ross, Jesse Slone, Blanche CapelAbstract:One of the earliest morphological changes during testicular differentiation is the establishment of an XY specific vasculature. The testis vascular system is derived from mesonephric endothelial cells that migrate into the gonad. In the XX gonad, mesonephric cell migration and testis vascular development are inhibited by WNT4 signaling. In WNT4 mutant XX gonads, endothelial cells migrate from the mesonephros and form a male-like coelomic vessel. Interestingly, this process occurs in the absence of other obvious features of testis differentiation, suggesting that WNT4 specifically inhibits XY vascular development. Consequently, the XX WNT4 mutant mice presented an opportunity to focus a gene expression screen on the processes of mesonephric cell migration and testicular vascular development. We compared differences in gene expression between XY WNT4+/+ and XX WNT4+/+ gonads and between XX WNT4+/+ and XX WNT4+/+ gonads to identify sets of genes similarly upregulated in wildtype XY gonads and XX mutant gonads or upregulated in XX gonads as compared to XY gonads and XX mutant gonads. We show that several genes identified in the first set are expressed in vascular domains, and have predicted functions related to cell migration or vascular development. However, the expression patterns and known functions of other genes are not consistent with roles in these processes. This screen has identified candidates for regulation of sex specific vascular development, and has implicated a role for WNT4 signaling in the development of Sertoli and germ cell lineages not immediately obvious from previous phenotypic analyses.
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follistatin operates downstream of WNT4 in mammalian ovary organogenesis
Developmental Dynamics, 2004Co-Authors: Humphrey H C Yao, Amanda Swain, Martin M Matzuk, Carolina J Jorgez, Douglas B Menke, David C Page, Blanche CapelAbstract: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 TGFbeta 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.
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follistatin operates downstream of WNT4 in mammalian ovary organogenesis
Developmental Dynamics, 2004Co-Authors: Martin M Matzuk, Amanda Swain, Carolina J Jorgez, Douglas B Menke, David L Page, Blanche CapelAbstract:Follistatin operates downstream of WNT4 in mammalian ovary organogenesis 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 TGFbeta superfamily binding protein, is a downstream component of Wnt 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. (C) 2004 Wiley-Liss, Inc.
Caroline M Alexander - One of the best experts on this subject based on the ideXlab platform.
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WNT4 mediates estrogen receptor signaling and endocrine resistance in invasive lobular carcinoma cell lines
Breast Cancer Research, 2016Co-Authors: Matthew J. Sikora, Caroline M Alexander, Britta M Jacobsen, Kevin M Levine, Jian Chen, Nancy E Davidson, Adrian V Lee, Steffi OesterreichAbstract:Invasive lobular carcinoma (ILC) of the breast typically presents with clinical biomarkers consistent with a favorable response to endocrine therapies, and over 90 % of ILC cases express the estrogen receptor (ER). However, a subset of ILC cases may be resistant to endocrine therapies, suggesting that ER biology is unique in ILC. Using ILC cell lines, we previously demonstrated that ER regulates a distinct gene expression program in ILC cells, and we hypothesized that these ER-driven pathways modulate the endocrine response in ILC. One potential novel pathway is via the Wnt ligand WNT4, a critical signaling molecule in mammary gland development regulated by the progesterone receptor. The ILC cell lines MDA-MB-134-VI, SUM44PE, and BCK4 were used to assess WNT4 gene expression and regulation, as well as the role of WNT4 in estrogen-regulated proliferation. To assess these mechanisms in the context of endocrine resistance, we developed novel ILC endocrine-resistant long-term estrogen-deprived (ILC-LTED) models. ILC and ILC-LTED cell lines were used to identify upstream regulators and downstream signaling effectors of WNT4 signaling. ILC cells co-opted WNT4 signaling by placing it under direct ER control. We observed that ER regulation of WNT4 correlated with use of an ER binding site at the WNT4 locus, specifically in ILC cells. Further, WNT4 was required for endocrine response in ILC cells, as WNT4 knockdown blocked estrogen-induced proliferation. ILC-LTED cells remained dependent on WNT4 for proliferation, by either maintaining ER function and WNT4 regulation or uncoupling WNT4 from ER and upregulating WNT4 expression. In the latter case, WNT4 expression was driven by activated nuclear factor kappa-B signaling in ILC-LTED cells. In ILC and ILC-LTED cells, WNT4 led to suppression of CDKN1A/p21, which is critical for ILC cell proliferation. CDKN1A knockdown partially reversed the effects of WNT4 knockdown. WNT4 drives a novel signaling pathway in ILC cells, with a critical role in estrogen-induced growth that may also mediate endocrine resistance. WNT4 signaling may represent a novel target to modulate endocrine response specifically for patients with ILC.
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abstract 862 WNT4 mediates endocrine response and resistance in invasive lobular carcinoma cell lines and patient tumor explants
Cancer Research, 2016Co-Authors: Matthew J. Sikora, Caroline M Alexander, Courtney L Andersen, Priscilla M Mcauliffe, Steffi OesterreichAbstract:Invasive lobular carcinoma (ILC) is a breast cancer subtype affecting ∼30,000 U.S. women annually. Over 90% of ILC are estrogen receptor (ER)-positive; however, endocrine therapy may have poorer efficacy in a subset of ILC patients versus invasive ductal carcinoma (IDC) patients. This prompted us to assess global ER activity in ILC cell lines MDA MB 134VI (MM134) and SUM44PE (44PE) to identify novel mediators of ER signaling. These analyses identified the Wnt ligand WNT4 as an ILC-specific ER target gene, with an ILC-specific ER binding site (ERBS) at the WNT4 locus. Considering the critical role of WNT4 in normal mammary gland expansion, we hypothesize that ILC cells utilize WNT4 signaling to drive endocrine response and resistance. We assessed whether WNT4 is necessary for ILC cell growth using siRNA. WNT4 knockdown completely blocked estrogen-induced growth in ILC cells but not IDC cells. In parallel, the WNT4 ERBS was only occupied in ILC cells in response to estrogen, but progesterone-induced WNT4 in IDC was not associated with this ERBS. This suggests that, via the ILC-specific WNT4 ERBS, ILC cells drive estrogen-regulated proliferation by hijacking a developmental Wnt pathway. Wnt pathways typically activate â-catenin; however, we observed â-catenin dysfunction in ILC cells and that WNT4 cannot activate â-catenin. Thus, WNT4 signals in ILC cells via a novel non-canonical pathway. Using long-term estrogen-deprived (LTED) variants of MM134 and 44PE (4 and 2 lines, respectively), we assessed WNT4 in ILC endocrine resistance. WNT4 is over-expressed, but uncoupled from ER, in all MM134-LTED. Conversely, WNT4 is reduced in 44PE-LTED but remains ER-regulated; ER occupies the WNT4 ERBS only in 44PE-LTED cells and not MM134-LTED. Using siRNA, MM134-LTED (high WNT4) are growth-inhibited by WNT4 knockdown, while 44PE-LTED (low WNT4) are insensitive. However, WNT4 knockdown sensitizes 44PE-LTED to endocrine therapy. Taken together, uncoupling and upregulating WNT4 or WNT4/ER cross-talk may represent convergent endocrine resistance mechanisms in ILC. To assess the role of WNT4 in patient ILC, we examined WNT4 protein in archival breast tumors and observed that WNT4 is frequently expressed in ILC and IDC tumors. We also examined WNT4 regulation and endocrine response in patient tumor explants. We observed ER regulation of WNT4 directly in ILC tissues that correlated with sensitivity to fulvestrant but resistance to tamoxifen; this may mimic clinical endocrine resistance. Ongoing studies are assessing WNT4 as a biomarker and mediator of endocrine resistance in ILC. Clinical observations suggest that ER regulates unique pathways in ILC. We identified WNT4 as a downstream effector of endocrine signaling in ILC, with critical roles in both estrogen-induced growth and endocrine resistance. WNT4 signaling may represent a novel target to modulate endocrine response specifically for ILC patients. Citation Format: Matthew J. Sikora, Courtney L. Andersen, Caroline M. Alexander, Priscilla M. McAuliffe, Steffi Oesterreich. WNT4 mediates endocrine response and resistance in invasive lobular carcinoma cell lines and patient tumor explants. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 862.
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abstract a35 WNT4 signaling mediates endocrine response and resistance in invasive lobular carcinoma cells
Molecular Cancer Research, 2016Co-Authors: Matthew J. Sikora, Caroline M Alexander, Amir Bahreini, Steffi OesterreichAbstract:Invasive lobular carcinoma (ILC) is a histological subtype of breast cancer, affecting ~30,000 U.S. women annually. Over 90% of ILC are estrogen receptor (ER)-positive, however, endocrine therapy may have poorer efficacy in a subset of ILC patients compared to invasive ductal carcinoma (IDC) patients. Based on these observations, we assessed genome-wide ER-mediated gene expression and ER genomic binding in ILC cell lines MDA MB 134VI (MM134) and SUM44PE (44PE), to identify novel mediators of ER signaling and putative therapeutic targets specifically in ILC. Among ILC-specific estrogen-regulated genes, the most strongly induced was the Wnt ligand WNT4. In parallel, we identified an ILC-specific ER binding site (ERBS) at WNT4, suggesting that WNT4 is directly ER-controlled in ILC cells. We hypothesized that this would be an analog to progesterone-controlled WNT4 in mammary gland expansion, and assessed whether WNT4 is necessary for estrogen-induced growth in ILC cells. Using siRNAs, knockdown of WNT4 completely blocked estrogen-induced growth in ILC cells, but not IDC cells. Consistent with this, we found that the WNT4 ERBS is only occupied in ILC cells that strongly upregulate WNT4 in response to estrogen, whereas progesterone-regulated WNT4 expression in T47D cells was not associated with ER binding at the WNT4 ERBS. These data suggest that, via an ILC-specific ERBS at WNT4, ILC cells can drive estrogen-regulated proliferation by hijacking a developmental Wnt pathway. Canonical Wnt pathways activate β-catenin; however, we observed β-catenin dysfunction in ILC cells, and that WNT4 cannot activate β-catenin in cell lines. Thus, WNT4 regulates estrogen-induced growth in ILC cells via a novel non-canonical pathway. Using long-term estrogen-deprived (LTED) variants of MM134 and 44PE (4 and 2 lines, respectively), we assessed WNT4 in ILC endocrine resistance. WNT4 is over-expressed but uncoupled from ER in all MM134-LTED; expression is greatly reduced in 44PE-LTED, but weakly estrogen-regulated. Consistent with regulation, ER occupies the WNT4 ERBS only in 44PE-LTED cells. However, 44PE-LTED (low WNT4) are resistant to growth inhibition by WNT4 siRNA, while MM134-LTED (high WNT4) are growth-inhibited. Taken together, uncoupling and upregulating WNT4 may be necessary for LTED growth in MM134. Clinical observations suggest that ER regulates unique downstream pathways in ILC. We identified WNT4 as a putative downstream effector of endocrine signaling in ILC, having critical roles in both estrogen-induced growth and endocrine resistance. Targeting WNT4 signaling represents a novel approach to modulate endocrine response specifically for ILC patients. Citation Format: Matthew J. Sikora, Amir Bahreini, Caroline M. Alexander, Steffi Oesterreich. WNT4 signaling mediates endocrine response and resistance in invasive lobular carcinoma cells. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Breast Cancer Research; Oct 17-20, 2015; Bellevue, WA. Philadelphia (PA): AACR; Mol Cancer Res 2016;14(2_Suppl):Abstract nr A35.
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abstract p3 04 05 invasive lobular carcinoma cell lines utilize WNT4 signaling to mediate estrogen induced growth
Cancer Research, 2015Co-Authors: Matthew J. Sikora, Caroline M Alexander, Amir Bahreini, Steffi OesterreichAbstract:Invasive lobular carcinoma (ILC) is a histological subtype of breast cancer representing 10-15% of newly diagnosed breast tumors. Over 90% of ILC are ER-positive, however, endocrine response and estrogen signaling are not well described in ILC. Retrospective analyses suggest that ILC patients treated with endocrine therapy have poorer outcomes than similar invasive ductal carcinoma (IDC) patients, and that ILC patients may not benefit from adjuvant tamoxifen. Additionally, we recently identified ILC-specific ER-target genes and de novo tamoxifen resistance driven by ER in ILC model systems. Based on these observations, we hypothesize that ILC-specific signaling pathways driven by ER mediate growth and endocrine resistance in ILC cells. Among ILC-specific estrogen-regulated genes in the ILC cell lines MDA MB 134VI (MM134) and SUM44PE (SUM44), Wnt signaling genes were highly differentially expressed. The secreted ligand WNT4 was the most strongly estrogen-induced gene in ILC cells. The frizzled receptor FZD7 is also strongly induced in ILC cells, but only transiently induced in the ER-positive IDC cell line MCF-7. Among IDC cell lines, either WNT4 or FZD7 is over-expressed in ER-positive or ER-negative cells, respectively. Conversely, MM134 and SUM44 over-express both WNT4 and FZD7. Also, we identified an ILC-specific ER binding site at WNT4; located in intron 1, this site contains a predicted estrogen response element. Direct WNT4 regulation and parallel regulation of pathway genes suggests that ER controls a WNT4 signaling pathway in ILC cells. In samples from the Cancer Genome Atlas, WNT4 and FZD7 are each over-expressed in ER-positive ILC versus IDC; co-expression is also enriched only in ILC. These observations suggest that a WNT4 signaling pathway may be specifically active in ILC tumors. To assess whether WNT4 is necessary for estrogen-induced growth, we used siRNA to knock down WNT4. Using either of two siRNAs, WNT4 knockdown completely blocks estrogen-induced growth in ILC cells, but not IDC cells. Consistent with this, WNT4 knockdown abrogated estrogen-regulation of a subset of ER-target genes in MM134 cells; induction or repression was inhibited by WNT4 knockdown prior to estrogen treatment. Thus, a subset of estrogen-induced gene expression changes is mediated by WNT4 signaling. Though Wnt signaling typically acts via the canonical, β-catenin-dependent pathway, we observed that β-catenin signaling is dysfunctional in ILC cells. Additionally, WNT4 over-expression or recombinant protein cannot activate canonical Wnt signaling in breast cancer cell lines. This suggests that WNT4 signaling mediates estrogen-induced growth in ILC cells via a novel non-canonical signaling pathway. Wnt signaling pathway genes including WNT4 are uniquely regulated in ILC cell lines, and are over-expressed in ILC tumors, suggesting that a WNT4-driven pathway may be active specifically in ILC. WNT4 is necessary for estrogen-mediated growth in ILC cells, and likely signaling via a novel non-canonical signaling pathway. Targeting WNT4 signaling represents a novel approach to modulate endocrine response specifically for ILC patients. Future studies will focus on identifying the signaling pathway controlled by WNT4 in order to identify novel therapeutic targets. Citation Format: Matthew J Sikora, Amir Bahreini, Caroline M Alexander, Steffi Oesterreich. Invasive lobular carcinoma cell lines utilize WNT4 signaling to mediate estrogen-induced growth [abstract]. In: Proceedings of the Thirty-Seventh Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2014 Dec 9-13; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2015;75(9 Suppl):Abstract nr P3-04-05.
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WNT4 is not sufficient to induce lobuloalveolar mammary development
BMC Developmental Biology, 2009Co-Authors: Young Chul Kim, Rod J Clark, Francisco Pelegri, Caroline M AlexanderAbstract:Background Brisken et al (2000) showed that WNT4 null mammary glands were deficient in early lobuloalveolar mammary outgrowth during pregnancy, and implicated WNT4 as an effector for the progesterone-induced mammary growth program. Though ectopic Wnt1 signaling is known to be mitogenic and oncogenic, no endogenously expressed Wnt ligands have ever been directly implicated in mammary growth and morphogenesis. Therefore, we generated conditional transgenic mice to test whether WNT4 can stimulate mammary epithelial cell growth.
Milan K Bagchi - One of the best experts on this subject based on the ideXlab platform.
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WNT4 is a key regulator of normal postnatal uterine development and progesterone signaling during embryo implantation and decidualization in the mouse
The FASEB Journal, 2011Co-Authors: Heather L. Franco, John P Lydon, Daisy Dai, Kevin Y Lee, Cory A Rubel, Dennis R Roop, Derek Boerboom, Jae Wook Jeong, Indrani C Bagchi, Milan K BagchiAbstract:WNT4, a member of the Wnt family of ligands, is critical for the development of the female reproductive tract. Analysis of WNT4 expression in the adult uterus during pregnancy indicates that it may play a role in the regulation of endometrial stromal cell proliferation, survival, and differentiation, which is required to support the developing embryo. To investigate the role of WNT4 in adult uterine physiology, conditional ablation of WNT4 using the PRcre mouse model was accomplished. Ablation of WNT4 rendered female mice subfertile due to a defect in embryo implantation and subsequent defects in endometrial stromal cell survival, differentiation, and responsiveness to progesterone signaling. In addition to altered stromal cell function, the uteri of PRcre/+WNT4f/f (WNT4d/d) mice displayed altered epithelial differentiation characterized by a reduction in the number of uterine glands and the emergence of a p63-positive basal cell layer beneath the columnar luminal epithelial cells. The altered epithelial cell phenotype was further escalated by chronic estrogen treatment, which caused squamous cell metaplasia of the uterine epithelium in the WNT4d/d mice. Thus, WNT4 is a critical regulator not only of proper postnatal uterine development, but also embryo implantation and decidualization.—Franco, H. L., Dai, D., Lee, K. Y., Rubel, C. S., Roop, D., Boerboom, D., Jeong, J.-W., Lydon, J.-P., Bagchi, I. C., Bagchi, M. K., DeMayo, F. J. WNT4 is a key regulator of normal postnatal uterine development and progesterone signaling during embryo implantation and decidualization in the mouse.
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WNT4 is a key regulator of normal postnatal uterine development and progesterone signaling during embryo implantation and decidualization in the mouse
The FASEB Journal, 2011Co-Authors: Heather L. Franco, John P Lydon, Daisy Dai, Kevin Y Lee, Cory A Rubel, Dennis R Roop, Derek Boerboom, Jae Wook Jeong, Indrani C Bagchi, Milan K BagchiAbstract:WNT4, a member of the Wnt family of ligands, is critical for the development of the female reproductive tract. Analysis of WNT4 expression in the adult uterus during pregnancy indicates that it may play a role in the regulation of endometrial stromal cell proliferation, survival, and differentiation, which is required to support the developing embryo. To investigate the role of WNT4 in adult uterine physiology, conditional ablation of WNT4 using the PR(cre) mouse model was accomplished. Ablation of WNT4 rendered female mice subfertile due to a defect in embryo implantation and subsequent defects in endometrial stromal cell survival, differentiation, and responsiveness to progesterone signaling. In addition to altered stromal cell function, the uteri of PR(cre/+)WNT4(f/f) (WNT4(d/d)) mice displayed altered epithelial differentiation characterized by a reduction in the number of uterine glands and the emergence of a p63-positive basal cell layer beneath the columnar luminal epithelial cells. The altered epithelial cell phenotype was further escalated by chronic estrogen treatment, which caused squamous cell metaplasia of the uterine epithelium in the WNT4(d/d) mice. Thus, WNT4 is a critical regulator not only of proper postnatal uterine development, but also embryo implantation and decidualization.
