Role of multi zinc finger proteins in the dynamic change of the nuclear architecture during cell cycle and differentiation

多锌指蛋白BORIS和CTCF是独特的保守因子，在转录调控、染色质进入不同结构域和印迹中发挥作用。BORIS在睾丸中不表达CTCF的细胞中表达。另一方面，BORIS的异常上调可能与肿瘤发生有关。因此，虽然这些蛋白质与基因组中的相似位点结合，但它们可能具有不同的作用。我们已经产生了可诱导的BORIS和CTCF敲除小鼠，并正在小鼠中产生GFP-（或生物素）标记的BORIS和CTCF敲除。从可诱导敲除的小鼠细胞系中分离出可转染（突变）多种锌指蛋白和/或具有特定结合位点的DNA结构的细胞系。使用这些工具，我们将对不同类型的小鼠、组织和细胞进行显微（实时）成像分析（Galjart组）、生物素标记蛋白的亲和纯化（Galjart组和Renakwitz组）和结构分析，如DNA环形成（Renkawitz组）。本研究旨在了解多锌指蛋白在细胞周期中的动态行为，以及这种行为与这些蛋白维持染色质结构的相关性。

The multi-zinc finger proteins BORIS and CTCF are unique and conserved factors with a role in transcriptional regulation, the organization of chromatin into distinct domains and imprinting. BORIS is expressed in the testis in cells that do not express CTCF. Abnormal upregulation of BORIS, on the other hand, may be linked to tumorigenesis. Thus, while binding to similar sites in the genome, these proteins could have distinct roles. We have generated inducible BORIS and CTCF knock out mice and are generating GFP- (or biotin)-tagged BORIS and CTCF knock in mice. From the inducible knock out mice cell lines have been isolated, which can be transfected with (mutant) multi-zinc finger proteins and/or DNA contructs with particular binding sites. Using these tools we will perform microscopic (live) imaging analysis (group Galjart), affinity purifications of biotin-tagged proteins (groups Galjart and Renakwitz) and structural analysis, like DNA loop formation (group Renkawitz) on the different types of mice, tissues and cells. This proposal aims at understanding the dynamic behaviour of both multi-zinc finger proteins during the cell cycle and the relevance of this behaviour and of these proteins for the maintenance of chromatin structure.