Structural and functional analysis of the yeast heterochromatin complex SIR

真核生物将基因组包装成功能不同的染色质结构域，在控制DNA的复制、转录、DNA修复和染色体分离等所有过程中起着至关重要的作用。大的基因组区域具有一种被称为异染色质的抑制结构，这是由异染色质蛋白与核小体结合并通过未知的机制抑制转录而建立的。在高等真核生物中，这些区域通常存在于端粒和着丝粒上，在那里它们防止降解和重组，在那里它们对正确的染色体分离很重要。异染色质的原型存在于出芽酵母中，位于沉默交配型位点HML和HMR以及端粒处。这些位点上异染色质的建立和形成是由沉默信息调节器（SIR）复合物介导的，SIR复合物由NAD+依赖性组蛋白去乙酰化酶（HDAC） Sir2和组分Sir3和Sir4组成。要详细了解sir介导的抑制机制，就需要对复合物的分子知识。然而，虽然可以获得SIR复合物各个部分的信息，但却缺少完整的SIR复合物的结构。在这个项目中，Ehrenhofer-Murray和Spahn实验室的目标是解决SIR复合物的自由形式以及它与核小体结合时的结构。我们将把这些结构目标与基因实验结合起来，以评估酵母细胞中沉默结构的功能相关性。这项工作将是高度相关的，因为它将允许新的结构和机制的见解，抑制SIR复合体是如何实现的。

The packaging of eukaryotic genomes into functionally distinct chromatin domains plays a crucial role in controlling all processes on DNA like replication, transcription, DNA repair and chromosome segregation. Large genomic regions assume a repressive structure termed heterochromatin, which is established by heterochromatin proteins that bind to the nucleosomes and inhibit transcription by yet unknown mechanisms. In higher eukaryotes, such regions typically are found at the telomeres, where they prevent degradation and recombination, and centromeres, where they are important for proper chromosome segregation. An archetypal form of heterochromatin is found in the budding yeast Saccharomyces cerevisiae at the silent mating-type loci HML and HMR, and at the telomeres. The establishment and formation of heterochromatin at these loci is mediated by the silent information regulator (SIR) complex, which consists of the NAD+-dependent histone deacetylase (HDAC) Sir2 and the components Sir3 and Sir4. A detailed mechanistic understanding of SIR-mediated repression requires molecular knowledge the complex. However, while information is available for individual parts of the SIR complex, a structure of the complete SIR complex is missing. In this project, the Ehrenhofer-Murray and Spahn labs aim to solve the structure of the SIR complex in its free form as well as when it is bound to nucleosomes. We will combine these structural aims with genetic experiments to assess the functional relevance of the structure(s) for silencing in yeast cells. This work will be of high relevance, because it will allow novel structural and mechanistic insights into how repression by the SIR complex is achieved.