KRAB型锌指蛋白ZNF75D在基因转录调控及肿瘤发生发展中的作用

Krüppel-associated box （KRAB）-containing zinc-finger proteins（KRAB-ZFPs） contains the largest family of transcriptional regulators and over 400 KRAB-ZFPs are encoded in mammalian genomes.While the molecular mechanism of transcription regulation by KRAB-ZFPs has been clarified to some degree，little is known about their biological function or gene targets，which could be largely attributed to the difficulty in identifying DNA-binding sites recognized by long arrays of zinc fingers. In this proposal, our work focused on the identification and functional analysis of a KRAB-containing zinc-finger protein, ZNF75D, which contains SCAN, KRAB A and 5 C2H2 zinc finger domains. We demonstrated that ZNF75D is localized in nucleus and possesses intrinsic transcription repression activity. We also found that ZNF75D interacts with TIF1β (transcriptional intermediary factor 1β, also named KAP1 (KRAB-associated protein 1)), histone deacetylase HDAC1 and HDAC2, indicating ZNF75D maybe act as a transcription repressor by recruiting TIF1β/KAP1 and histone deacetylase complexes. Based on these interesting findings, to further elucidate its function, we endeavour to indentify putative DNA-binding sequences and potential downstream targets of ZNF75D by CASTing (Cyclic Amplification and Selection of Target) assays and ChIP-Seq (chromatin immunoprecipitation with massively parallel DNA sequencing). To further understand the molecular mechanisms underlying ZNF75D-mediated transcription repression, the ZNF75D-containing protein complexes will be identified by affinity purification and mass spectrometry. With the target genes of ZNF75D, its possible roles in tumorigenesis and tumor progression will be further studied. These findings may shed new light on transcriptional regulation by KRAB-ZFPs and might offer a potential new target for tumor therapy.

基因转录是由一系列转录因子和转录辅助因子参与的复杂过程。转录调控异常会导致多种疾病包括肿瘤的发生与发展。KRAB型锌指蛋白是哺乳动物最大的转录调控因子家族，虽然对其转录抑制的机制有所认知，但其多数成员的靶基因及生物学活性仍未知，主要原因之一是寻找锌指蛋白的DNA结合位点受限。我们在前期工作中克隆到一编码KRAB型锌指蛋白的基因ZNF75D。初步功能分析表明ZNF75D定位于细胞核，具有转录抑制活性且与转录中介因子TIF1β/KAP1及去乙酰化酶存在相互作用。本课题拟从利用ChIP-Seq技术高通量寻找ZNF75D全基因组结合位点和利用蛋白亲和层析联合质谱分析寻找ZNF75D相互作用蛋白入手，深入探讨ZNF75D参与转录调控的机理和生物学功能及其与肿瘤发生发展的关系，为进一步认识KRAB型锌指蛋白在基因转录调控中的作用及其与肿瘤发生发展的关系提供理论依据，为寻找新的肿瘤治疗靶点提供线索。