Role of methyl-cytosine binding proteins in the gain and loss of pluripotency

甲基胞嘧啶结合蛋白(MBDS)在表观遗传效应的调节中起着核心作用。它们与甲基化的DNA结合，并招募组蛋白脱乙酰酶(HDAC)，这可能会导致染色质浓缩和转录沉默。我们之前已经证明，MeCP2水平在分化过程中增加，导致大规模的基因组重组。因此，我们正在研究Mbds在多能性的获得和丧失中的作用。我们已经产生并鉴定了一组特定的抗体，现在将完成我们对MBD在多潜能、分化和重编程(IPS)细胞中的表达和定位的系统分析。我们将进一步确定这些细胞中MBDS的动态，特别是对基因组甲基胞嘧啶羟化增加的反应，这是一种可能与表观遗传重新编程有关的新的DNA修饰。利用特定的突变体，我们将研究MBDS在控制染色质组织和多能性获得或丧失的动力学中的作用。在过去的几年里，我们已经将单分子显微镜应用于活细胞研究，现在将以高时间和空间分辨率研究染色质动力学。我们将在单分子水平上特别研究MBD染色质的相互作用，并比较多能细胞和分化细胞中的染色质动力学。这项研究应该阐明MBDS的作用，并将解决更高级别基因组组织和动态在多能性和重新编程中的功能相关性。

Methyl cytosine binding proteins (MBDs) play a central role in the mediation of epigenetic effects. They bind to methylated DNA and recruit histone deacetylases (HDACs), which may then cause chromatin condensation and transcriptional silencing. We have previously shown that MeCP2 levels increase during differentiation causing large scale genome reorganization. We are, therefore, investigating the role of MBDs in the gain and loss of pluripotency. We have generated and characterized a set of specific antibodies and will now complete our systematic analysis of MBD expression and localization in pluripotent, differentiated and reprogrammed (iPS) cells. We will further determine the dynamics of MBDs in these cells, especially in response to increased hydroxylation of genomic methyl cytosine, a novel DNA modification possibly linked to epigenetic reprogramming. Using specific mutants we will investigate the role of MBDs in controlling chromatin organization and dynamics in the gain or loss of pluripotency. Over the past years we have adapted single molecule microscopy to live cell studies and will now investigate chromatin dynamics with high temporal and spatial resolution. We will in particular study MBD chromatin interactions at the single molecule level and will compare chromatin dynamics in pluripotent versus differentiated cells. This study should clarify the role of MBDs and will address the functional relevance of higher order genome organization and dynamics in pluripotency and reprogramming.