Epigenetic Mechanisms of Retrotransposon Silencing

项目摘要/摘要 反转录转座子是人类遗传密码的古老组成部分，经过数百万年的自我复制 现在占我们基因组的45%。反转录转座子激活对细胞活力有严重影响- 反转录转座子本质上是诱变的，即使表达“死亡的”反转录转座子也能触发先天的 引起炎症的免疫反应。因此，细胞进化出错综复杂的表观遗传机制。 通过将反转录转座子隔离在构成的异染色质中来抑制它们的表达，这是 标记了H3K9甲基化和DNA CpG甲基化的表观遗传修饰。H3K9的调节失调 甲基化和CpG甲基化导致反转录转座子的广泛重新激活，并在 人类疾病。然而，人们对引发这种疾病的确切分子机制知之甚少。 反转录转座子沉默或保持反转录转座子处于沉默状态。这项研究的总体目标是 程序是定义细胞用来抑制反转录转座子的精确分子机制 表情。细胞反转录转座子沉默机制的一个关键成分是组蛋白赖氨酸 甲基转移酶Setdb-1，它在反转录转座子上沉积H3K9甲基标记。因为它的核心作用， Setdb-1的活性受到一组复杂的分子玩家的严格调控，这些分子玩家控制着它的催化活性， 逆转座子序列的募集及其将组蛋白甲基化传播组蛋白甲基化的能力 反转录转座子。适当控制Setdb-1活性是逆转座子沉默的关键，但许多 Setdb-1调控的某些方面仍然是个谜。我们将使用体外生化重组，Cryo-EM 以及基因组学来破译Setbd-1调控不足的精确机制。概述的工作 在这项提案中将解释Setdb-1如何被催化激活以进行逆转座子沉默，Setdb-1如何 阅读反转录转座子的表观遗传密码以及Setdb-1如何被招募到反转录转座子序列以 建立H3K9甲基化模式。

Project Summary/Abstract Retrotransposons are ancient components of our genetic code that have self-replicated over millions of years and now make up 45% of our genome. Retrotransposon activation has serious implications for cell viability – retrotransposition is mutagenic by its nature and even expression of “dead” retrotransposons can trigger innate immune responses that cause inflammation. Therefore, the cell has evolved intricate epigenetic mechanisms to silence expression of retrotransposons by sequestering them in constitutive heterochromatin, which is marked the epigenetic modifications of H3K9 methylation and DNA CpG methylation. Dysregulation of H3K9 methylation and CpG methylation causes widespread re-activation of retrotransposons and occurs in multiple human diseases. However, very little is known about the precise molecular mechanisms that initiate retrotransposon silencing or maintain retrotransposons in a silent state. The broad goal of this research program is to define the precise molecular mechanisms the cell uses to suppress retrotransposon expression. A key component of the cells retrotransposon silencing machinery is the histone lysine methyltransferase Setdb-1, which deposits H3K9 methyl marks at retrotransposons. Because of its central role, Setdb-1 activity is tightly regulated by a complex cast of molecular players that control its catalytic activation, recruitment to retrotransposon sequences and its ability to spread histone methylation across the length of retrotransposons. Appropriate control of Setdb-1 activity is critical for retrotransposon silencing, but many aspects of Setdb-1 regulation remain enigmatic. We will use in vitro biochemical reconstitutions, Cryo-EM and genomics to decipher the precise mechanisms that underly regulation of Setbd-1. The work outlined in this proposal will explain how Setdb-1 is catalytically activated for retrotransposon silencing, how Setdb-1 reads the epigenetic code on retrotransposons and how Setdb-1 is recruited to retrotransposon sequences to establish H3K9 methylation patterns.