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Jean-noël Freund - One of the best experts on this subject based on the ideXlab platform.
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Extending the functions of the homeotic Transcription Factor Cdx2 in the digestive system through nonTranscriptional activities.
World journal of gastroenterology, 2015Co-Authors: Jean-noël Freund, Isabelle Duluc, Isabelle Gross, Jean-marie Reimund, Claire Domon-dellAbstract:The homeoprotein encoded by the intestinal-specific Cdx2 gene is a major regulator of gut development and homeostasis, also involved in colon cancer as well as in intestinal-type metaplasias when it is abnormally expressed outside the gut. At the molecular level, structure/function studies have demonstrated that the Cdx2 protein is a Transcription Factor containing a conserved homeotic DNA-binding domain made of three alpha helixes arranged in a helix-turn-helix motif, preceded by a Transcriptional domain and followed by a regulatory domain. The protein interacts with several thousand sites on the chromatin and widely regulates intestinal functions in stem/progenitor cells as well as in mature differentiated cells. Yet, this Transcription Factor also acts trough original nonTranscriptional mechanisms. Indeed, the identification of novel protein partners of Cdx2 and also of a splicing variant revealed unexpected functions in the control of signaling pathways like the Wnt and NF-κB pathways, in double-strand break DNA repair and in premessenger RNA splicing. These novel functions of Cdx2 must be considered to fully understand the complexity of the role of Cdx2 in the healthy intestine and in diseases.
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Cdx2 regulation by the RNA-binding protein MEX3A: impact on intestinal differentiation and stemness
Nucleic Acids Research, 2013Co-Authors: Bruno Pereira, Sofia Sousa, Rita Barros, Laura Carreto, Patrícia Oliveira, Carla Oliveira, Nicolas T. Chartier, Michelina Plateroti, Jean-pierre Rouault, Jean-noël FreundAbstract:The homeobox Transcription Factor Cdx2 plays a crucial role in intestinal cell fate specification, both during normal development and in tumorigenic processes involving intestinal reprogramming. The Cdx2 regulatory network is intricate, but it has not yet been fully uncovered. Through genome-wide screening of a 3D culture system, the RNA-binding protein MEX3A was identified as putatively involved in Cdx2 regulation; therefore, its biological relevance was addressed by setting up cell-based assays together with expression studies in murine intestine. We demonstrate here that MEX3A has a repressive function by controlling Cdx2 levels in gastric and colorectal cellular models. This is dependent on the interaction with a specific binding determinant present in Cdx2 mRNA 3'untranslated region. We have further determined that MEX3A impairs intestinal differentiation and cellular polarization, affects cell cycle progression and promotes increased expression of intestinal stem cell markers, namely LGR5, BMI1 and MSI1. Finally, we show that MEX3A is expressed in mouse intestine, supporting an in vivo context for interaction with Cdx2 and modulation of stem cell properties. Therefore, we describe a novel Cdx2 post-Transcriptional regulatory mechanism, through the RNA-binding protein MEX3A, with a major impact in intestinal differentiation, polarity and stemness, likely contributing to intestinal homeostasis and carcinogenesis.
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Cdx2 Controls Expression of the Protocadherin Mucdhl, an Inhibitor of Growth and β-Catenin Activity in Colon Cancer Cells
Gastroenterology, 2011Co-Authors: Isabelle Hinkel, Jean-noël Freund, Isabelle Duluc, Elisabeth Martin, Dominique Guenot, Isabelle GrossAbstract:Background & Aims The intestine-specific homeobox Transcription Factor Cdx2 is an important determinant of intestinal identity in the embryonic endoderm and regulates the balance between proliferation and differentiation in the adult intestinal epithelium. Human colon tumors often lose Cdx2 expression, and heterozygous inactivation of Cdx2 in mice increases colon tumorigenesis. We sought to identify Cdx2 target genes to determine how it contributes to intestinal homeostasis. Methods We used expression profiling analysis to identify genes that are regulated by Cdx2 in colon cancer cells lines. Regulation and function of a potential target gene were further investigated using various cell assays. Results In colon cancer cell lines, Cdx2 directly regulated the Transcription of the gene that encodes the protocadherin Mucdhl. Mucdhl localized to the apex of differentiated cells in the intestinal epithelium, and its expression was reduced in most human colon tumors. Overexpression of Mucdhl inhibited low-density proliferation of colon cancer cells and reduced tumor formation in nude mice. One isoform of Mucdhl interacted with β-catenin and inhibited its Transcriptional activity. Conclusions The Transcription Factor Cdx2 activates expression of the protocadherin Mucdhl, which interacts with β-catenin and regulates activities of intestinal cells. Loss of Cdx2 expression in colon cancer cells might reduce expression of Mucdhl and thereby lead to tumor formation.
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Expression and localisation of insulin receptor substrate 2 in normal intestine and colorectal tumours. Regulation by intestine-specific Transcription Factor Cdx2
Gut, 2009Co-Authors: Salvatore Modica, Isabelle Duluc, Annalisa Morgano, Lorena Salvatore, Michele Petruzzelli, Marie-therese Vanier, Rosa Valanzano, Diana L. Esposito, Giuseppe Palasciano, Jean-noël FreundAbstract:Background and aims: Self-renewal and differentiation of intestinal epithelium is a tightly regulated process, whose perturbations are implicated in human colorectal tumourigenesis. The insulin/insulin-like growth Factor (IGF) signalling pathway may play an important role in intestinal epithelium homeostasis. Insulin receptor substrate 2 (IRS2) is a poorly characterised component in this pathway. Methods: Using complementary in vitro and in vivo human and murine models, expression (mRNA and protein levels), localisation (immunohistochemistry) and regulation of IRS2 were investigated in the normal intestine and colorectal tumours. In silico analysis of the human IRS2 promoter was performed together with reporter and chromatin immunoprecipitation assays. Results: Significant IRS2 expression was detected in the intestine, with specific protein localisation in the villus region of the ileum and in the surface epithelium of the colon. In human HT29 and Caco2 cells, IRS2 mRNA levels increased with spontaneous and induced differentiation, together with Cdx2 (caudal-related homeobox protein 2), P21 and KLF4 (Kruppel-like Factor 4). Adenoviral infection with human Cdx2 induced IRS2 expression in APC- (adenomatous polyposis coli) and β-catenin-mutated cells. On the other hand, IRS2 downregulation was observed in differentiated enterocytes after adenoviral infection with short hairpin Cdx2 (shCdx2), in the intestine of Cdx2 heterozygous mice and in colorectal tumours of Apc Min/+ and patients with familial adenomatous polyposis (FAP). The human IRS2 promoter region presents several Cdx2-binding sites where Cdx2 immunoprecipitated in vivo. IRS2 reporters were functionally activated via Cdx2 and blocked via a dominant-negative Cdx2 protein. Conclusions: Combining gain- and loss-of-function approaches, an intriguing scenario is presented whereby IRS2 is significantly expressed in the apical intestinal compartment and is directly controlled by Cdx2 in normal intestine and tumours.
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Phosphorylation of the homeotic tumor suppressor Cdx2 mediates its ubiquitin-dependent proteasome degradation.
Oncogene, 2005Co-Authors: Isabelle Gross, Isabelle Duluc, Elisabeth Martin, Claire Domon-dell, Benoit Lhermitte, Christian Gaiddon, Michele Kedinger, Jean-noël FreundAbstract:The Caudal-related homeodomain Transcription Factor Cdx2 plays a key role in intestinal cell fate determination. Reduction of Cdx2 expression is a feature of many human colon carcinomas and inactivation of one Cdx2 allele facilitates the development of invasive adenocarcinoma in the murine colon. Here, we investigated the post-translational regulation of Cdx2. We showed that various forms of Cdx2 coexist in the intestine and colon cancer cell lines, some of them being phosphorylated forms. We found that cyclin-dependent kinase 2 phosphorylated Cdx2 in vitro and in vivo. Using site-specific mutagenesis, we identified serine 281 as a new key residue for Cdx2 phosphorylation. Intriguingly, serine 281 belongs to a conserved motif of four evenly spaced serines (the 4S motif) similar to the one controlling beta-catenin degradation by the proteasome pathway. A nonphosphorylated mutant Cdx2 lacking the 4S motif (4S>A) exhibited reduced polyubiquitination upon proteasome inhibition and increased stability compared to wild-type Cdx2. In addition, we found that this mutant was less efficient to suppress colony formation than wild-type Cdx2. Thus, our data highlight a novel post-translational mechanism controlling Cdx2 degradation via phosphorylation and polyubiquitination, which may be of importance for intestinal development and cancer.
John P Lynch - One of the best experts on this subject based on the ideXlab platform.
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intestine specific Transcription Factor Cdx2 induces e cadherin function by enhancing the trafficking of e cadherin to the cell membrane
American Journal of Physiology-gastrointestinal and Liver Physiology, 2010Co-Authors: Shinsuke Funakoshi, Duyen T Dang, Jianping Kong, Mary Ann S Crissey, Long H Dang, John P LynchAbstract:Cdx2 is an intestine-specific Transcription Factor required for normal intestinal epithelium development. Cdx2 regulates the expression of intestine-specific genes and induces cell adhesion and columnar morphogenesis. Cdx2 also has tumor-suppressor properties, including the reduction of colon cancer cell proliferation and cell invasion, the latter due to its effects on cell adhesion. E-cadherin is a cell adhesion protein required for adherens junction formation and the establishment of intestinal cell polarity. The objective of this study was to elucidate the mechanism by which Cdx2 regulates E-cadherin function. Two colon cancer cell lines were identified in which Cdx2 expression was associated with increased cell-cell adhesion and diminished cell migration. In both cell lines, Cdx2 did not directly alter E-cadherin levels but increased its trafficking to the cell membrane compartment. Cdx2 enhanced this trafficking by altering receptor tyrosine kinase (RTK) activity. Cdx2 expression diminished phosphorylated Abl and phosphorylated Rac levels, which are downstream effectors of RTKs. Specific chemical inhibition or short interfering RNA (shRNA) knockdown of c-Abl kinase phenocopied Cdx2's cell-cell adhesion effects. In Colo 205 cells, Cdx2 reduced PDGF receptor and IGF-I receptor activation. This was mediated by caveolin-1, which was induced by Cdx2. Targeted shRNA knockdown of caveolin-1 restored PDGF receptor and reversed E-cadherin membrane trafficking, despite Cdx2 expression. We conclude that Cdx2 regulates E-cadherin function indirectly by disrupting RTK activity and enhancing E-cadherin trafficking to the cell membrane compartment. This novel mechanism advances Cdx2's prodifferentiation and antitumor properties and suggests that Cdx2 may broadly regulate RTK activity in normal intestinal epithelium by modulating membrane trafficking of proteins.
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tcf4 and Cdx2 major Transcription Factors for intestinal function converge on the same cis regulatory regions
Proceedings of the National Academy of Sciences of the United States of America, 2010Co-Authors: Michael P. Verzi, Hyunjin Shin, Rita Sulahian, Pantelis Hatzis, Juliet Philips, Jurian Schuijers, Ellen Freed, John P Lynch, Duyen T Dang, Myles BrownAbstract:Surprisingly few pathways signal between cells, raising questions about mechanisms for tissue-specific responses. In particular, Wnt ligands signal in many mammalian tissues, including the intestinal epithelium, where constitutive signaling causes cancer. Genome-wide analysis of DNA cis-regulatory regions bound by the intestine-restricted Transcription Factor Cdx2 in colonic cells uncovered highly significant overrepresentation of sequences that bind TCF4, a Transcriptional effector of intestinal Wnt signaling. Chromatin immunoprecipitation confirmed TCF4 occupancy at most such sites and co-occupancy of Cdx2 and TCF4 across short distances. A region spanning the single nucleotide polymorphism rs6983267, which lies within a MYC enhancer and confers colorectal cancer risk in humans, represented one of many co-occupied sites. Co-occupancy correlated with intestine-specific gene expression and Cdx2 loss reduced TCF4 binding. These results implicate Cdx2 in directing TCF4 binding in intestinal cells. Co-occupancy of regulatory regions by signal-effector and tissue-restricted Transcription Factors may represent a general mechanism for ubiquitous signaling pathways to achieve tissue-specific outcomes.
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the intestine specific Transcription Factor Cdx2 inhibits β catenin tcf Transcriptional activity by disrupting the β catenin tcf protein complex
Carcinogenesis, 2010Co-Authors: Rong-jun Guo, Jianping Kong, Shinsuke Funakoshi, Hannah H. Lee, John P LynchAbstract:Cdx2 is an intestine-specific Transcription Factor known to regulate proliferation and differentiation. We have reported previously that Cdx2 limits the proliferation of human colon cancer cells by inhibiting the Transcriptional activity of the beta-catenin-T-cell Factor (TCF) bipartite complex. Herein we further elucidate this mechanism. Studies with a classic Cdx2 target gene and a canonical Wnt/beta-catenin/TCF reporter suggest that Cdx2 regulates these promoters by distinctly different processes. Specifically, inhibition of beta-catenin/TCF activity by Cdx2 does not require Cdx2 Transcriptional activity. Instead, Cdx2 binds beta-catenin and disrupts its interaction with the DNA-binding TCF Factors, thereby silencing beta-catenin/TCF target gene expression. Using Cdx2 mutants, we map the Cdx2 domains required for the inhibition of beta-catenin/TCF activity. We identify a subdomain in the N-terminus that is highly conserved and when mutated significantly reduces Cdx2 inhibition of beta-catenin/TCF Transcriptional activity. Mutation of this subdomain also abrogates Cdx2's anti-proliferative effects in colon cancer cells. In summary, we conclude that Cdx2 binds beta-catenin and disrupts the beta-catenin-TCF complex. Considering the pivotal role of beta-catenin/TCF activity in driving proliferation of normal intestinal epithelial and colon cancer cells, our findings suggest a novel mechanism for Cdx2-mediated regulation of Wnt/beta-catenin signaling and cell proliferation.
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The intestine-specific Transcription Factor Cdx2 inhibits β-catenin/TCF Transcriptional activity by disrupting the β-catenin–TCF protein complex
Carcinogenesis, 2009Co-Authors: Rong-jun Guo, Jianping Kong, Shinsuke Funakoshi, Hannah H. Lee, John P LynchAbstract:Cdx2 is an intestine-specific Transcription Factor known to regulate proliferation and differentiation. We have reported previously that Cdx2 limits the proliferation of human colon cancer cells by inhibiting the Transcriptional activity of the beta-catenin-T-cell Factor (TCF) bipartite complex. Herein we further elucidate this mechanism. Studies with a classic Cdx2 target gene and a canonical Wnt/beta-catenin/TCF reporter suggest that Cdx2 regulates these promoters by distinctly different processes. Specifically, inhibition of beta-catenin/TCF activity by Cdx2 does not require Cdx2 Transcriptional activity. Instead, Cdx2 binds beta-catenin and disrupts its interaction with the DNA-binding TCF Factors, thereby silencing beta-catenin/TCF target gene expression. Using Cdx2 mutants, we map the Cdx2 domains required for the inhibition of beta-catenin/TCF activity. We identify a subdomain in the N-terminus that is highly conserved and when mutated significantly reduces Cdx2 inhibition of beta-catenin/TCF Transcriptional activity. Mutation of this subdomain also abrogates Cdx2's anti-proliferative effects in colon cancer cells. In summary, we conclude that Cdx2 binds beta-catenin and disrupts the beta-catenin-TCF complex. Considering the pivotal role of beta-catenin/TCF activity in driving proliferation of normal intestinal epithelial and colon cancer cells, our findings suggest a novel mechanism for Cdx2-mediated regulation of Wnt/beta-catenin signaling and cell proliferation.
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S1760 The Homeodomain Transcription Factor Cdx2 Regulates E Cadherin Activity By Enhancing Its Localization from the Cytoplasm to the Cell Membrane
Gastroenterology, 2009Co-Authors: Shinsuke Funakoshi, Duyen T Dang, Jianping Kong, Long H Dang, Sanjay R. Hegde, John P LynchAbstract:trafficking. Methods: In the present study, we employed fluorescence resonance energy transfer (FRET) and Zeiss 510 confocal microscopy to study direct binding of NHE3 and NHERF2 in living or fixed polarized kidney epithelial cells (OK cells) that were transiently transfected with CFP-NHERF2 and YFP-NHE3. Results: Under basal conditions, NHERF2 and NHE3 exhibited 10-20% FRET signaling solely at the microvilli (an average of 10 regions of interest (ROIs) in the microvilli of each cells) but not at the juxtanuclear region. As a negative control, there was no FRET signaling with expression of YFP-GPI and CFP-NHERF2. Further, treatment with 10 μMCa2+ ionophore, which increases intracellular Ca2+, abolished the NHERF2/NHE3 FRET signaling within a minute. To further elucidate the role of Ca2+ signaling in regulation of NHE3 trafficking, we treated Caco-2/HA-NHE3 cells with carbachol (10 uM) that activates M3 muscarinic receptors and increases intracellular Ca2+ and monitored its effect on the trafficking of NHE3 via total internal reflection fluorescence (TIRF) microscopy. Carbachol reduced the surface level of NHE3 with an onset within one minute. Conclusions: 1) Under basal conditions, NHE3 binds NHERF2 in the OK cell microvilli and this binding can be quantified by acceptor photobleaching FRET. 2) Within one minute of elevation of intracellular Ca2+, the FRET signal between NHE3 and NHERF2 is abolished in OK cells. 3) Carbachol causes a similarly rapid reduction of surface NHE3 observed via TIRF. We conclude that elevation of intracellular Ca2+ leads to dissociation of NHERF2 from NHE3 at the microvillus which allows NHE3 transport activity to be inhibited by facilitating endocytosis.
Hubert Schorle - One of the best experts on this subject based on the ideXlab platform.
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the Transcription Factor tcfap2c ap 2γ cooperates with Cdx2 to maintain trophectoderm formation
Molecular and Cellular Biology, 2010Co-Authors: Peter Kuckenberg, Sandra Buhl, Tatiana Woynecki, Betina Van Fürden, E. N. Tolkunova, Friederike Seiffe, Markus Moser, Alexey Tomilin, Elke Winterhager, Hubert SchorleAbstract:In mammals, cell lineage specification is established at the blastocyst stage. At this stage, Transcription Factor Cdx2 represses pluripotency genes, thus promoting extraembryonic trophoblast fate. Recently, Transcription Factor Gata3 was shown to act in a parallel pathway in promoting trophoblast cell fate, suggesting that there are more Factors working in the trophoblast lineage. Here, we report that the Transcription Factor Tcfap2c is expressed at a high level in the trophectoderm and is able to induce trophoblast fate in embryonic stem cells. Trophoblast fate induced by Tcfap2c does not require Cdx2 and vice versa, suggesting that the molecules act in alternative pathways. However, both Tcfap2c and Cdx2 are required for the upregulation of Elf5, a marker of trophoblast stem cell maintenance, suggesting that both Factors are required for stable trophoblast induction. Tcfap2c-induced trophoblast-like cells are stable in long-term culture, indicating that they are capable of self-renewal. Tcfap2c-controlled trophoblast maintenance involves the induction of Cdx2 and the repression of the pluripotency Factor Nanog. Tcfap2c-induced trophoblast-like cells differentiate to trophoblast derivatives in vitro and contribute to the trophectoderm in blastocysts in vivo. Taken together, these observations suggest that Tcfap2c and Cdx2 cooperate to override the pluripotency program and establish the extraembryonic trophoblast maintenance program in murine embryos.
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The Transcription Factor TCFAP2C/AP-2γ Cooperates with Cdx2 To Maintain Trophectoderm Formation
Molecular and cellular biology, 2010Co-Authors: Peter Kuckenberg, Sandra Buhl, Tatiana Woynecki, Betina Van Fürden, E. N. Tolkunova, Friederike Seiffe, Markus Moser, Alexey Tomilin, Elke Winterhager, Hubert SchorleAbstract:In mammals, cell lineage specification is established at the blastocyst stage. At this stage, Transcription Factor Cdx2 represses pluripotency genes, thus promoting extraembryonic trophoblast fate. Recently, Transcription Factor Gata3 was shown to act in a parallel pathway in promoting trophoblast cell fate, suggesting that there are more Factors working in the trophoblast lineage. Here, we report that the Transcription Factor Tcfap2c is expressed at a high level in the trophectoderm and is able to induce trophoblast fate in embryonic stem cells. Trophoblast fate induced by Tcfap2c does not require Cdx2 and vice versa, suggesting that the molecules act in alternative pathways. However, both Tcfap2c and Cdx2 are required for the upregulation of Elf5, a marker of trophoblast stem cell maintenance, suggesting that both Factors are required for stable trophoblast induction. Tcfap2c-induced trophoblast-like cells are stable in long-term culture, indicating that they are capable of self-renewal. Tcfap2c-controlled trophoblast maintenance involves the induction of Cdx2 and the repression of the pluripotency Factor Nanog. Tcfap2c-induced trophoblast-like cells differentiate to trophoblast derivatives in vitro and contribute to the trophectoderm in blastocysts in vivo. Taken together, these observations suggest that Tcfap2c and Cdx2 cooperate to override the pluripotency program and establish the extraembryonic trophoblast maintenance program in murine embryos.
Michael P. Verzi - One of the best experts on this subject based on the ideXlab platform.
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Transcription Factors gata4 and hnf4a control distinct aspects of intestinal homeostasis in conjunction with Transcription Factor Cdx2
Journal of Biological Chemistry, 2015Co-Authors: Adrianna San K Roman, Michael P. Verzi, Ramesh A. Shivdasani, Boaz E Aronson, Stephen D KrasinskiAbstract:Distinct groups of Transcription Factors (TFs) assemble at tissue-specific cis-regulatory sites, implying that different TF combinations may control different genes and cellular functions. Within such combinations, TFs that specify or maintain a lineage and are therefore considered master regulators may play a key role. Gene enhancers often attract these tissue-restricted TFs, as well as TFs that are expressed more broadly. However, the contributions of the individual TFs to combinatorial regulatory activity have not been examined critically in many cases in vivo. We address this question using a genetic approach in mice to inactivate the intestine-specifying and intestine-restricted Factor Cdx2 alone or in combination with its more broadly expressed partner Factors, GATA4 and HNF4A. Compared with single mutants, each combination produced significantly greater defects and rapid lethality through distinct anomalies. Intestines lacking Gata4 and Cdx2 were deficient in crypt cell replication, whereas combined loss of Hnf4a and Cdx2 specifically impaired viability and maturation of villus enterocytes. Integrated analysis of TF binding and of transcripts affected in Hnf4a;Cdx2 compound-mutant intestines indicated that this TF pair controls genes required to construct the apical brush border and absorb nutrients, including dietary lipids. This study thus defines combinatorial TF activities, their specific requirements during tissue homeostasis, and modules of Transcriptional targets in intestinal epithelial cells in vivo.
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Intestinal Master Transcription Factor Cdx2 Controls Chromatin Access for Partner Transcription Factor Binding
Molecular and cellular biology, 2012Co-Authors: Michael P. Verzi, Hyunjin Shin, Adrianna K. San Roman, X. Shirley Liu, Ramesh A. ShivdasaniAbstract:Tissue-specific gene expression requires modulation of nucleosomes, allowing Transcription Factors to occupy cis elements that are accessible only in selected tissues. Master Transcription Factors control cell-specific genes and define cellular identities, but it is unclear if they possess special abilities to regulate cell-specific chromatin and if such abilities might underlie lineage determination and maintenance. One prevailing view is that several Transcription Factors enable chromatin access in combination. The homeodomain protein Cdx2 specifies the embryonic intestinal epithelium, through unknown mechanisms, and partners with Transcription Factors such as HNF4A in the adult intestine. We examined enhancer chromatin and gene expression following Cdx2 or Hnf4a excision in mouse intestines. HNF4A loss did not affect Cdx2 binding or chromatin, whereas Cdx2 depletion modified chromatin significantly at Cdx2-bound enhancers, disrupted HNF4A occupancy, and abrogated expression of neighboring genes. Thus, Cdx2 maintains Transcription-permissive chromatin, illustrating a powerful and dominant effect on enhancer configuration in an adult tissue. Similar, hierarchical control of cell-specific chromatin states is probably a general property of master Transcription Factors.
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Differentiation-Specific Histone Modifications Reveal Dynamic Chromatin Interactions and Partners for the Intestinal Transcription Factor Cdx2
Developmental cell, 2010Co-Authors: Michael P. Verzi, Hyunjin Shin, X. Shirley Liu, Rita Sulahian, Clifford A. Meyer, Robert K. Montgomery, James C. Fleet, Myles Brown, Ramesh A. ShivdasaniAbstract:Cell differentiation requires remodeling of tissue-specific gene loci and activities of key Transcriptional regulators, which are recognized for their dominant control over cellular programs. Using epigenomic methods, we characterized enhancer elements specifically modified in differentiating intestinal epithelial cells and found enrichment of Transcription Factor-binding motifs corresponding to Cdx2, a critical regulator of the intestine. Directed investigation revealed surprising lability in Cdx2 occupancy of the genome, with redistribution from hundreds of sites occupied only in proliferating cells to thousands of new sites in differentiated cells. Knockout mice confirmed distinct Cdx2 requirements in dividing and mature adult intestinal cells, including responsibility for the active enhancer configuration associated with maturity. Dynamic Cdx2 occupancy corresponds with condition-specific gene expression and, importantly, to differential co-occupancy with other tissue-restricted Transcription Factors, such as GATA6 and HNF4A. These results reveal dynamic, context-specific functions and mechanisms of a prominent Transcriptional regulator within a cell lineage.
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tcf4 and Cdx2 major Transcription Factors for intestinal function converge on the same cis regulatory regions
Proceedings of the National Academy of Sciences of the United States of America, 2010Co-Authors: Michael P. Verzi, Hyunjin Shin, Rita Sulahian, Pantelis Hatzis, Juliet Philips, Jurian Schuijers, Ellen Freed, John P Lynch, Duyen T Dang, Myles BrownAbstract:Surprisingly few pathways signal between cells, raising questions about mechanisms for tissue-specific responses. In particular, Wnt ligands signal in many mammalian tissues, including the intestinal epithelium, where constitutive signaling causes cancer. Genome-wide analysis of DNA cis-regulatory regions bound by the intestine-restricted Transcription Factor Cdx2 in colonic cells uncovered highly significant overrepresentation of sequences that bind TCF4, a Transcriptional effector of intestinal Wnt signaling. Chromatin immunoprecipitation confirmed TCF4 occupancy at most such sites and co-occupancy of Cdx2 and TCF4 across short distances. A region spanning the single nucleotide polymorphism rs6983267, which lies within a MYC enhancer and confers colorectal cancer risk in humans, represented one of many co-occupied sites. Co-occupancy correlated with intestine-specific gene expression and Cdx2 loss reduced TCF4 binding. These results implicate Cdx2 in directing TCF4 binding in intestinal cells. Co-occupancy of regulatory regions by signal-effector and tissue-restricted Transcription Factors may represent a general mechanism for ubiquitous signaling pathways to achieve tissue-specific outcomes.
Ramesh A. Shivdasani - One of the best experts on this subject based on the ideXlab platform.
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The lineage-specific Transcription Factor Cdx2 navigates dynamic chromatin to control distinct stages of intestine development.
Development (Cambridge England), 2019Co-Authors: Namit Kumar, Yu Hwai Tsai, Lei Chen, Anbo Zhou, Kushal K. Banerjee, Madhurima Saxena, Sha Huang, Jinchuan Xing, Natalie H. Toke, Ramesh A. ShivdasaniAbstract:Lineage-restricted Transcription Factors, such as the intestine-specifying Factor Cdx2, often have dual requirements across developmental time. Embryonic loss of Cdx2 triggers homeotic transformation of intestinal fate, whereas adult-onset loss compromises crucial physiological functions but preserves intestinal identity. It is unclear how such diverse requirements are executed across the developmental continuum. Using primary and engineered human tissues, mouse genetics, and a multi-omics approach, we demonstrate that divergent Cdx2 loss-of-function phenotypes in embryonic versus adult intestines correspond to divergent Cdx2 chromatin-binding profiles in embryonic versus adult stages. Cdx2 binds and activates distinct target genes in developing versus adult mouse and human intestinal cells. We find that temporal shifts in chromatin accessibility correspond to these context-specific Cdx2 activities. Thus, Cdx2 is not sufficient to activate a mature intestinal program; rather, Cdx2 responds to its environment, targeting stage-specific genes to contribute to either intestinal patterning or mature intestinal function. This study provides insights into the mechanisms through which lineage-specific regulatory Factors achieve divergent functions over developmental time.
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The Lineage-Specific Transcription Factor Cdx2 Navigates Dynamic Chromatin to Control Distinct Stages of Intestine Development
2018Co-Authors: Namit Kumar, Ramesh A. Shivdasani, Yu Hwai Tsai, Lei Chen, Anbo Zhou, Kushal K. Banerjee, Madhurima Saxena, Sha Huang, Jinchuan Xing, Jason R. SpenceAbstract:Lineage-restricted Transcription Factors, such as the intestine-specifying Factor Cdx2, often have dual requirements across developmental time. Embryonic-loss of Cdx2 triggers homeotic transformation of intestinal fate, while adult-onset Cdx2-loss compromises critical physiological functions but preserves intestinal identity. It is unclear how such diverse requirements are executed across the developmental continuum. Using primary and engineered human tissues, mouse genetics, and a multi-omics approach, we demonstrate that divergent Cdx2 loss-of-function phenotypes in embryonic versus adult intestines correspond to divergent Cdx2 chromatin-binding profiles in embryonic versus adult stages. Cdx2 binds and activates distinct target genes in developing versus adult mouse and human intestinal cells. We find that temporal shifts in chromatin accessibility correspond to these context-specific Cdx2 activities. Thus, Cdx2 is not sufficient to activate a mature intestinal program, but rather, Cdx2 responds to its environment, targeting stage-specific genes to contribute to either intestinal patterning or maturity. This study provides insights into the mechanisms through which lineage-specific regulatory Factors achieve divergent functions over developmental time.
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Transcription Factors gata4 and hnf4a control distinct aspects of intestinal homeostasis in conjunction with Transcription Factor Cdx2
Journal of Biological Chemistry, 2015Co-Authors: Adrianna San K Roman, Michael P. Verzi, Ramesh A. Shivdasani, Boaz E Aronson, Stephen D KrasinskiAbstract:Distinct groups of Transcription Factors (TFs) assemble at tissue-specific cis-regulatory sites, implying that different TF combinations may control different genes and cellular functions. Within such combinations, TFs that specify or maintain a lineage and are therefore considered master regulators may play a key role. Gene enhancers often attract these tissue-restricted TFs, as well as TFs that are expressed more broadly. However, the contributions of the individual TFs to combinatorial regulatory activity have not been examined critically in many cases in vivo. We address this question using a genetic approach in mice to inactivate the intestine-specifying and intestine-restricted Factor Cdx2 alone or in combination with its more broadly expressed partner Factors, GATA4 and HNF4A. Compared with single mutants, each combination produced significantly greater defects and rapid lethality through distinct anomalies. Intestines lacking Gata4 and Cdx2 were deficient in crypt cell replication, whereas combined loss of Hnf4a and Cdx2 specifically impaired viability and maturation of villus enterocytes. Integrated analysis of TF binding and of transcripts affected in Hnf4a;Cdx2 compound-mutant intestines indicated that this TF pair controls genes required to construct the apical brush border and absorb nutrients, including dietary lipids. This study thus defines combinatorial TF activities, their specific requirements during tissue homeostasis, and modules of Transcriptional targets in intestinal epithelial cells in vivo.
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Intestinal Master Transcription Factor Cdx2 Controls Chromatin Access for Partner Transcription Factor Binding
Molecular and cellular biology, 2012Co-Authors: Michael P. Verzi, Hyunjin Shin, Adrianna K. San Roman, X. Shirley Liu, Ramesh A. ShivdasaniAbstract:Tissue-specific gene expression requires modulation of nucleosomes, allowing Transcription Factors to occupy cis elements that are accessible only in selected tissues. Master Transcription Factors control cell-specific genes and define cellular identities, but it is unclear if they possess special abilities to regulate cell-specific chromatin and if such abilities might underlie lineage determination and maintenance. One prevailing view is that several Transcription Factors enable chromatin access in combination. The homeodomain protein Cdx2 specifies the embryonic intestinal epithelium, through unknown mechanisms, and partners with Transcription Factors such as HNF4A in the adult intestine. We examined enhancer chromatin and gene expression following Cdx2 or Hnf4a excision in mouse intestines. HNF4A loss did not affect Cdx2 binding or chromatin, whereas Cdx2 depletion modified chromatin significantly at Cdx2-bound enhancers, disrupted HNF4A occupancy, and abrogated expression of neighboring genes. Thus, Cdx2 maintains Transcription-permissive chromatin, illustrating a powerful and dominant effect on enhancer configuration in an adult tissue. Similar, hierarchical control of cell-specific chromatin states is probably a general property of master Transcription Factors.
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Differentiation-Specific Histone Modifications Reveal Dynamic Chromatin Interactions and Partners for the Intestinal Transcription Factor Cdx2
Developmental cell, 2010Co-Authors: Michael P. Verzi, Hyunjin Shin, X. Shirley Liu, Rita Sulahian, Clifford A. Meyer, Robert K. Montgomery, James C. Fleet, Myles Brown, Ramesh A. ShivdasaniAbstract:Cell differentiation requires remodeling of tissue-specific gene loci and activities of key Transcriptional regulators, which are recognized for their dominant control over cellular programs. Using epigenomic methods, we characterized enhancer elements specifically modified in differentiating intestinal epithelial cells and found enrichment of Transcription Factor-binding motifs corresponding to Cdx2, a critical regulator of the intestine. Directed investigation revealed surprising lability in Cdx2 occupancy of the genome, with redistribution from hundreds of sites occupied only in proliferating cells to thousands of new sites in differentiated cells. Knockout mice confirmed distinct Cdx2 requirements in dividing and mature adult intestinal cells, including responsibility for the active enhancer configuration associated with maturity. Dynamic Cdx2 occupancy corresponds with condition-specific gene expression and, importantly, to differential co-occupancy with other tissue-restricted Transcription Factors, such as GATA6 and HNF4A. These results reveal dynamic, context-specific functions and mechanisms of a prominent Transcriptional regulator within a cell lineage.