title: Functional analysis of KAP1 genomic recruitment.
abstract: TRIM28 (KAP1) is upregulated in many cancers and has been implicated in both transcriptional activation and repression. Using chromatin immunoprecipitation and sequencing, we show that KAP1 binding sites fall into several categories, specifically, the 3' coding exons of zinc finger (ZNF) genes and promoter regions of ZNFs and other genes. The currently accepted model is that KAP1 is recruited to the genome via interaction of its N-terminal RBCC domain with KRAB ZNFs (KRAB domain containing ZNFs). To determine whether the interaction of KAP1 with KRAB ZNFs is the mechanism by which KAP1 is recruited to genomic binding sites, we analyzed stable cell lines that express tagged wild-type and mutant KAP1. Surprisingly, deletion of the RBCC domain abolished KAP1 binding to the 3' exons of ZNF genes but KAP1 binding to promoter regions was unaffected. Using KAP1 knockdown cells, we showed that the genes most responsive to KAP1 were not ZNF genes but instead were either indirect targets or had KAP1 bound 10 to 100 kb from the transcription start site. Therefore, our studies suggest that KAP1 plays a role distinct from transcriptional regulation at the majority of its strongest binding sites.
authors.affiliation: Department of Biochemistry & Molecular Biology, Norris Comprehensive Cancer Center, 1450 Biggy Street, NRT 6503, Mail Code 9601, University of Southern California, Los Angeles, CA 90089, USA.
authors.forename: Sushma
authors.initials: S
authors.lastname: Iyengar
authors.forename: Alexey V
authors.initials: AV
authors.lastname: Ivanov
authors.forename: Victor X
authors.initials: VX
authors.lastname: Jin
authors.forename: Frank J
authors.initials: FJ
authors.lastname: Rauscher
authors.forename: Peggy J
authors.initials: PJ
authors.lastname: Farnham
mesh_terms: D001665:Binding Sites; D002460:Cell Line; D045744:Cell Line, Tumor; D000090266:Chromobox Protein Homolog 5; D002868:Chromosomal Proteins, Non-Histone / Q000378:metabolism; D005786:Gene Expression Regulation; D011495:Histone-Lysine N-Methyltransferase; D006657:Histones / Q000378:metabolism; D006801:Humans; D009154:Mutation; D011401:Promoter Regions, Genetic; D011496:Protein Methyltransferases / Q000378:metabolism; D012097:Repressor Proteins / Q000032:analysis / Q000235:genetics* / Q000378:metabolism*; D000076183:Tripartite Motif-Containing Protein 28; D016335:Zinc Fingers
chemical_list: D002868:Chromosomal Proteins, Non-Histone; D006657:Histones; D012097:Repressor Proteins; C422407:ZNF350 protein, human; D000090266:Chromobox Protein Homolog 5; D011496:Protein Methyltransferases; D011495:Histone-Lysine N-Methyltransferase; C454062:SETDB1 protein, human; C101116:TRIM28 protein, human; D000076183:Tripartite Motif-Containing Protein 28