Comment[ArrayExpressAccession] E-GEOD-22572 Public Release Date 2010-08-11 Investigation Title TCF4 and CDX2, major transcription factors for intestinal function, converge on the same cis-regulatory regions Comment[Submitted Name] TCF4 and CDX2, major transcription factors for intestinal function, converge on the same cis-regulatory regions Experiment Description 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 over-representation 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. A series of ChIP-chip experiments identified the CDX2 cistrome and discovered and validated extensive co-binding with TCF4 in colon cancer cell lines Transcriptional profiling following shRNA-mediated CDX2 knockdown was employed to identify CDX2-dependent gene expression in the human colon cancer cell line Caco2 Date of Experiment Term Source Name EFO Term Source Version Term Source File http://efo.svn.sourceforge.net/viewvc/efo/trunk/src/efoinowl/efo.owl Person Last Name Verzi Michael Ramesh Person First Name Michael Verzi Shivdasani Person Mid Initials Person Email geo@ncbi.nlm.nih.gov Person Affiliation Dana Farber Cancer Institute Person Phone Person Fax Person Address Dana Farber Cancer Institute, 44 Binney st, Boston, MA, USA Person Roles submitter Person Roles Term Source REF Person Roles Term Accession Number Normalization Type Normalization Term Accession Number Normalization Term Source REF Replicate Type Replicate Term Accession Number Replicate Term Source REF Experimental Design Experimental Design Term Accession Number Experimental Design Term Source REF Quality Control Type Quality Control Term Accession Number Quality Control Term Source REF Protocol Name P-GSE22572-1 P-GSE22572-13 P-GSE22572-5 P-GSE22572-10 P-GSE22572-2 P-GSE22572-7 P-GSE22572-16 P-GSE22572-15 P-GSE22572-8 P-GSE22572-14 P-GSE22572-6 P-GSE22572-9 P-GSE22572-11 P-GSE22572-3 P-GSE22572-12 P-GSE22572-4 Protocol Description ID_REF =
VALUE = dChip normalized signal intensity Labeling, hybridization and scanning were done according to Affymetrix protocols labeling, hybridization, and scanning were done using manufacturer's instructions (Affymetrix) Caco-2 cells were grown in 10% FBS DMEM and grown until 1 day post confluent on 6 well plates Cells were grown in 10% FBS DMEM and grown until 1 day post confluent on 15 cm plates Model Based analysis of tiling arrays (MAT) was used to process the data as described previously: Johnson, et al, 2006, PMID: 16895995 The results file of comparison of shCDX2/shGFP contains dChip processed data with less stringent cutoffs to allow for larger selection of potentially regulated targets and is provided as a supplementary file on the Series record. Data were processed using dChip 2006 software using default Affymetrix normalization settings. .bar files were generated as the output of default MAT analysis and can be visualized using the publically available Affymetrix Ingtegrated Genome Browser Labeling, hybridization and scanning were done according to Affymetrix protocols labeling, hybridization, and scanning were done using manufacturer's instructions (Affymetrix) Cells were treated with lentivirus containing shCDX2 (MISSION TRCN13684 or shGFP (control)) when about 25% confluent and selected in 2ug/ml Puromycin until 1 day post confluent. Cells at this state are a heterogeneous population of dividing and differentiated CaCo2 cells. Trizol was used to extract RNA according to manufacturer's instructions Cells were fixed in 1% formaldehyde for 10 minutes at 37 degrees, then washed with cold PBS and scraped into PBS, and resuspended in ChIP lysis buffer and immunoprecipitated and processed as described by Carroll, et al, 2005, PMID: 16009131 and as described in our current work. Labeling, hybridization and scanning were done according to Affymetrix protocols labeling, hybridization, and scanning were done using manufacturer's instructions (Affymetrix) Protocol Software Protocol Hardware Protocol Contact Protocol Type bioassay_data_transformation hybridization hybridization grow grow feature_extraction feature_extraction feature_extraction feature_extraction image_aquisition image_aquisition specified_biomaterial_action nucleic_acid_extraction nucleic_acid_extraction labeling labeling Protocol Term Source REF Protocol Term Accession Number Experimental Factor Name CELL TYPE Experimental Factor Type cell type Experimental Factor Term Source REF Experimental Factor Term Accession Number Publication Title TCF4 and CDX2, major transcription factors for intestinal function, converge on the same cis-regulatory regions. Publication Author List Verzi MP, Hatzis P, Sulahian R, Philips J, Schuijers J, Shin H, Freed E, Lynch JP, Dang DT, Brown M, Clevers H, Liu XS, Shivdasani RA PubMed ID 20696899 Publication DOI 10.1073/pnas.1003822107 Publication Status Publication Status Term Source REF Publication Status Term Accession Number Comment[SecondaryAccession] GSE22572 Comment[GEOLastUpdateDate] 2010-08-26 Comment[AEExperimentType] ChIP-chip by tiling array transcription profiling by array Comment[GEOReleaseDate] 2010-08-10 Comment[ArrayExpressSubmissionDate] 2010-06-24 SDRF File E-GEOD-22572.sdrf.txt