Role of histone tails in gene expression and cellular functions

在我的实验室里，我的研究项目的主要目的是研究染色质如何影响基因调控的过程。最近，我的实验室一直对理解H3和H4组蛋白尾部如何调节基因表达和其他细胞过程感兴趣。值得注意的是，非结构化的n端组蛋白尾部非常保守。组蛋白H3和H4尾部的序列最能说明这一点，从酵母到哺乳动物，这些序列几乎100%保守。组蛋白H3和H4尾部在其基本功能上是冗余的，因为任何一个都可以在另一个存在的情况下被删除，但它们不能一起被删除。此外，H3和H4尾部可以互换，分别位于H4和H3的核心。几个实验室的研究表明，H3和H4尾部对基因调控和DNA修复很重要，并提出这种机制需要对位于组蛋白尾部的几个关键残基（赖氨酸、精氨酸、丝氨酸和苏氨酸）进行翻译后修饰（甲基化、乙酰化、磷酸化等）。由于尾部内部和尾部之间的功能冗余，这些残基对H3和H4尾部特定功能的重要性尤其难以研究，因此阐明特定残基与组蛋白H3和H4尾部功能之间的联系将需要大量的点突变组合。为了解决这些复杂的问题，我的实验室利用H3和H4的n端尾部同时缺失是不可行的这一事实，建立了一个遗传筛选。以随机序列设计带n端尾的H3分子文库，筛选能恢复H3- h4突变株生存力的序列。我们成功地鉴定了人工合成的尾巴，不仅恢复了生存能力，而且在一定的应力条件下增加了原始菌株的适应度。因此，目前的研究计划将集中在这些合成尾巴的生物化学和遗传学特征上，以揭示它们的作用机制。这种筛选方法已被证明在识别蛋白质-蛋白质相互作用和理解复杂机制方面非常有用。***特别是，我们将重点筛选H4 n端尾部，正如我们对H3所做的那样，并使用蛋白质组学和遗传学方法寻找这些合成H3和H4组蛋白尾部的直接功能靶点。**

Significant effort from my research program in my laboratory has been aimed at studying how chromatin affects the process of gene regulation. More recently, my laboratory has been interested in understanding how H3 and H4 histone tails can regulate gene expression and other cellular processes. Remarkably, the non-structured N-terminal histone tails are extremely well conserved. This is best illustrated with the sequence of the histone H3 and H4 tails, which are practically 100% conserved from yeast to mammals. The histone H3 and H4 tails are redundant in their essential function, as either one can be deleted in the presence of the other, but they cannot be deleted together. Moreover, the H3 and H4 tails can be interchanged to reside on the core of H4 and H3 respectively. Work from several laboratories have shown that H3 and H4 tails are important for proper gene regulation and DNA repair and it is proposed that this mechanism requires the posttranslational modification (methylation, acetylation, phosphorylation, etc.) of several key residues (lysines, arginine, serines and threonines) located within the tail of histones. The importance of each of these residues for specific functionalities of H3 and H4 tails is particularly difficult to investigate because of the functional redundancy within and between tails, and thus the elucidation of the link between specific residues and histone H3 and H4 tail functions would require a tremendous number of combinations of point mutations. To address these complex issues, my laboratory has set up a genetic screen taking advantage of the fact that simultaneous deletion in the N-terminal tails of H3 and H4 is unviable. A library of H3 molecules bearing N-terminal tails was designed with random sequences and screened for sequences that could restore viability in H3-H4 mutant strains. We successfully identified synthetic tails that restored not only viability, but also increased the fitness of the original strain in certain stress conditions. The current research program will thus focus on the characterization of these synthetic tails both biochemically and genetically to uncover their mechanisms of action. Such screening methods have proven to be extremely useful in identifying protein-protein interactions and understanding complex mechanisms.***In particular, we will focus on screening for H4 N-terminal tails as we have done with H3, and use proteomics and genetics approaches to find direct functional targets of these synthetic H3 and H4 histone tails. **