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.
Mol Cell Biol. 2011 May;31(9):1833-47. doi: 10.1128/MCB.01331-10. Epub 2011 Feb 22.
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.
TRIM28（KAP1）在许多癌症中上调，并被牵连到转录激活和抑制中。通过染色质免疫沉淀和测序，我们表明 KAP1 结合位点分为几类，特别是锌指（ZNF）基因的 3'编码外显子和 ZNF 基因及其他基因的启动子区域。目前公认的模型是，KAP1 通过其 N 端 RBCC 结构域与 KRAB ZNF（含有 KRAB 结构域的 ZNF）的相互作用被招募到基因组。为了确定 KAP1 与 KRAB ZNF 的相互作用是否是 KAP1 被招募到基因组结合位点的机制，我们分析了表达标记野生型和突变型 KAP1 的稳定细胞系。令人惊讶的是，RBCC 结构域的缺失消除了 KAP1 与 ZNF 基因 3'外显子的结合，但 KAP1 与启动子区域的结合不受影响。使用 KAP1 敲低细胞，我们表明对 KAP1 反应最敏感的基因不是 ZNF 基因，而是间接靶基因或 KAP1 结合在转录起始位点 10 到 100kb 处。因此，我们的研究表明，KAP1 在其大多数最强结合位点处发挥作用不同于转录调节。
