title: Identification of β-catenin-interacting proteins in nuclear fractions of native rat collecting duct cells.
abstract: The gene encoding the aquaporin-2 water channel is regulated transcriptionally in response to vasopressin. In the renal collecting duct, vasopressin stimulates the nuclear translocation and phosphorylation (at Ser552) of β-catenin, a multifunctional protein that acts as a transcriptional coregulator in the nucleus. The purpose of this study was to identify β-catenin-interacting proteins that might be involved in transcriptional regulation in rat inner medullary collecting duct (IMCD) cells, using experimental and computational approaches. We used a standard chromatin immunoprecipitation procedure coupled to mass spectrometry (ChIP-MS) in a nuclear fraction isolated from rat IMCD suspensions. Over four biological replicates, we reproducibly identified 43 β-catenin-binding proteins, including several known β-catenin-binding partners as well as novel interacting proteins. Multiple proteins involved in transcriptional regulation were identified (Taf1, Jup, Tdrd3, Cdh1, Cenpj, and several histones). Many of the identified β-catenin-binding partners were found in prior studies to translocate to the nucleus in response to vasopressin. There was only one DNA-binding transcription factor (TF), specifically Taf1, part of the RNA-polymerase II preinitiation complex. To identify sequence-specific TFs that might interact with β-catenin, Bayes' theorem was used to integrate data from several information sources. The analysis identified several TFs with potential binding sites in theAqp2gene promoter that could interact with β-catenin in the regulation ofAqp2gene transcription, specifically Jun, Junb, Jund, Atf1, Atf2, Mef2d, Usf1, Max, Pou2f1, and Rxra. The findings provide information necessary for modeling the transcriptional response to vasopressin.
authors.affiliation: Epithelial Systems Biology Laboratory, Systems Biology Center, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland.
authors.forename: Jacqueline R
authors.initials: JR
authors.lastname: Hwang
authors.affiliation: Epithelial Systems Biology Laboratory, Systems Biology Center, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland.
authors.forename: Chung-Lin
authors.initials: CL
authors.lastname: Chou
authors.affiliation: Epithelial Systems Biology Laboratory, Systems Biology Center, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland.
authors.forename: Barbara
authors.initials: B
authors.lastname: Medvar
authors.affiliation: Epithelial Systems Biology Laboratory, Systems Biology Center, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland knepperm@nhlbi.nih.gov.
authors.forename: Mark A
authors.initials: MA
authors.lastname: Knepper
authors.affiliation: Epithelial Systems Biology Laboratory, Systems Biology Center, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland.
authors.forename: Hyun Jun
authors.initials: HJ
authors.lastname: Jung
mesh_terms: D000818:Animals; D051399:Aquaporin 2 / Q000235:genetics / Q000378:metabolism; D001499:Bayes Theorem; D001665:Binding Sites; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism*; D047369:Chromatin Immunoprecipitation; D002853:Chromatography, Liquid; D019295:Computational Biology; D030541:Databases, Genetic; D005786:Gene Expression Regulation; D007685:Kidney Tubules, Collecting / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D060066:Protein Interaction Maps; D040901:Proteomics / Q000379:methods; D017207:Rats, Sprague-Dawley; D018511:Systems Integration; D053719:Tandem Mass Spectrometry; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic; D014667:Vasopressins / Q000494:pharmacology; D051176:beta Catenin / Q000235:genetics / Q000378:metabolism*
chemical_list: C496025:Aqp2 protein, rat; D051399:Aquaporin 2; C495268:Ctnnb1 protein, rat; D014157:Transcription Factors; D051176:beta Catenin; D014667:Vasopressins
keywords: aquaporin-2; kidney; transcription; transcriptional coregulator