Functions of Mammalian H1 Linker Histones in Gene Regulation and Chromatin Activity

这项建议的目的是阐明哺乳动物H1连接器组蛋白在基因调控中的功能 和染色质活性，并揭示11个变异体之间的功能差异 构成哺乳动物的H1基因家族。以及四个核心组蛋白(H_2A、H_2B、H_3和H_4) 将DNA组装成核小体，H1连接子组蛋白在组织DNA的结构中发挥关键作用 染色质纤维。与在理解核心组蛋白的作用方面的进展相比，我们对 连接子组蛋白在染色质生物学中的作用是相当有限的。我们对H1的S角色的大部分了解是 它来源于体外研究，而我们对其在体内的功能的了解仍然很不完整。我们 建议通过阐明基因的功能、作用机制和变种特有的活动来帮助填补这一空白 哺乳动物H1连接物组蛋白在体内的基因调控和染色质活性。 我们研究哺乳动物H1‘S体内功能的策略一直是产生和表征 小鼠胚胎干细胞和因几个H1基因失活而部分丧失H1的小鼠。通过 分析这些H1耗尽的模型，我们发现H1对于哺乳动物的发育是必不可少的，Play 在表观遗传调控中发挥意想不到的作用，对异染色质的完整性和沉默非常重要。 此外，我们对果蝇H1的研究表明，它是异染色质后期复制所必需的 在唾液腺多线染色体的内复制过程中。最近我们还发现，另外呢， 哺乳动物H1‘S除了在异染色质中发挥重要作用外，还在常染色质中发挥重要作用 它们抑制活性基因体染色质标记(H3K79、H3K36和H3K4的甲基化)的沉积，并影响 POL II转录延长和不同的mRNA剪接结果。 深入了解哺乳动物H1‘S参与基因调控的机制 和染色质活动，我们建议追求以下具体目标：(1)了解H1在 RNA POL II转录、mRNA剪接和基因体组蛋白H3赖氨酸甲基化；(2)确定其作用 S晚期H1的复制时机异染色质及其在有丝分裂至G1期的重组 转换；(3)确定H1变异体之间的功能差异及其N-末端和C-末端的贡献 域名和翻译后修饰。 这个项目的成功完成将为我们提供重要的见解，让我们了解这个专业 染色质的组成成分调节哺乳动物染色体的结构和活性。

The goals of this proposal are to elucidate the functions of mammalian H1 linker histones in gene regulation and chromatin activity during the cell cycle and to reveal functional differences amongst the 11 variants that constitute the mammalian H1 gene family. Along with the four core histones (H2A, H2B, H3 and H4) that assemble DNA into nucleosomes, the H1 linker histones play key roles in organizing the structure of the chromatin fiber. In comparison to the progress in understanding the roles of core histones, our knowledge about the functions of linker histones in chromatin biology is quite limited. Much of our knowledge about H1’s roles is derived from in vitro studies, whereas our understanding of its functions in vivo remains very incomplete. We propose to help fill this gap by elucidating the functions, mechanisms of action and variant-specific activities of mammalian H1 linker histones in gene regulation and chromatin activity in vivo. Our strategy for studying the in vivo functions of mammalian H1’s has been to generate and characterize mouse embryonic stem cells and mice that are partially depleted of H1 by inactivation of several H1 genes. By analyzing these H1-depleted models, we showed that H1 is essential for mammalian development, plays unexpected roles in epigenetic regulation, and is very important for the integrity and silencing of heterochromatin. Furthermore, our studies with Drosophila H1 showed that it is required for late replication of heterochromatin during endoreplication of salivary gland polytene chromosomes. Recently, we also discovered that, in addition to their important functions in heterochromatin, mammalian H1’s also play important roles in euchromatin where they inhibit deposition of active gene body chromatin marks (methylation of H3K79, H3K36 and H3K4) and affect Pol II transcription elongation and alternative mRNA splicing outcomes. To gain a deeper understanding of the mechanisms by which mammalian H1’s participate in gene regulation and chromatin activities, we propose to pursue the following specific aims: (1) Understand the functions of H1 in RNA Pol II transcription, mRNA splicing and gene body histone H3 lysine methylation; (2) Determine the roles of H1 in late S phase replication timing of heterochromatin and its reorganization during the mitosis to G1 phase transition; (3) Identify functional differences amongst H1 variants and the contributions of their N- and C-terminal domains and post-translational modifications. The successful completion of this project will provide important insights into the ways in which this major constituent of chromatin regulates the structure and activity of mammalian chromosomes.