Molecular mechanisms of gene silencing by H4 methylation

描述（由申请人提供）：组蛋白负责细胞核中DMA的压实和功能组织。这些小的，进化保守的蛋白质是多种翻译后修饰的宿主，每种修饰通常与特定的dna模板化过程相关，例如基因调控。我们最近发现了一种由PR-Set7酶介导的新的哺乳动物基因沉默途径，该途径特异性地单甲基化组蛋白H4赖氨酸20 （H4K20）。我们的新发现表明，这种沉默途径是针对在生长和分化程序中起作用的某些基因的启动子。基于这些发现，我们的中心假设是H4K20单甲基化的一个主要作用是通过抑制促进生长和分化的特定基因集来维持细胞身份。本提案的目标是剖析这一基本基因沉默途径的分子机制，并确定这一途径在建立和维持细胞身份中的作用。我们最近发现PR-Set7与一种新的组蛋白H3赖氨酸9 （H3K9）甲基转移酶结合，在基因启动子上产生抑制单甲基- h4k20和H3K9“反尾组蛋白代码”。在目标1中，我们将使用已建立的分子和生化方法来鉴定和表征这种酶，并确定其在体内基因抑制中的作用。我们还发现了第一个已知的H4K20单甲基结合蛋白L(3)MBT，并确定这种相互作用对基因抑制至关重要。在目标2中，我们将定义L(3)MBT结合单甲基化H4K20和基因抑制所需的区域和氨基酸。在Aim 3中，我们将继续鉴定受该沉默途径调控的其他基因。这些基因作为体内模型来剖析该途径中每个组成部分对体内基因抑制的个体贡献。总的来说，这一建议将阐明参与关键生物过程的基本哺乳动物基因调控途径的分子机制，并可能对人类健康和疾病产生深远而广泛的影响。

DESCRIPTION (provided by applicant): The histone proteins are responsible for the compaction and functional organization of DMA in the nucleus. These small, evolutionary conserved proteins are host to a diverse array of post-translational modifications, each of which is typically associated with a specific DNA-templated process, such as gene regulation. We recently discovered a novel mammalian gene silencing pathway mediated by the PR-Set7 enzyme that specifically monomethylates histone H4 lysine 20 (H4K20). Our new findings indicate that this silencing pathway is targeted to the promoters of certain genes that function in growth and differentiation programs. Based on these findings, our central hypothesis is that one major role of H4K20 monomethylation is to maintain cellular identity by repressing specific sets of genes that promote growth and differentiation. The goal of this proposal is to dissect the molecular mechanisms of this fundamental gene silencing pathway with the long range goal of determining the role of this pathway in establishing and maintaining cellular identity. We recently discovered that PR-Set7 associates with a novel histone H3 lysine 9 (H3K9) methyltransferase to create a repressive monomethyl-H4K20 and H3K9 "trans-tail histone code" at gene promoters. In Aim 1 we will use established molecular and biochemical methods to identify and characterize this enzyme and determine its in vivo role in gene repression. We have also discovered the first known H4K20 monomethyl- binding protein, L(3)MBT, and determined that this interaction is essential for gene repression. In Aim 2 we will define the regions and amino acids of L(3)MBT required for binding monomethylated H4K20 and gene repression. In Aim 3 we will continue to identify additional genes regulated by this silencing pathway. These genes serve as the in vivo models to dissect apart the individual contributions of each of the components of this pathway on in vivo gene repression. Collectively, this proposal will illuminate the molecular mechanisms of a fundamental mammalian gene regulation pathway involved in critical biological processes and will likely have far-reaching and wide-spread impacts on human health and disease.