Histone mRNA Regulation in Development

摘要 在细胞周期中，组蛋白生物合成的适当调节对于协调 染色质组装和DNA复制。组蛋白的过量生产或生产不足 蛋白质导致复制应激和基因组不稳定， 癌症的发展。在本提案中，我们对以下方面进行了全面分析： 复制依赖性（RD）组蛋白mRNA的生物合成，以及这一过程是如何 在动物发育的背景下，在细胞周期中调节。RD-组蛋白mRNA 是动物细胞中唯一缺乏polyA尾的真核mRNA，而是以 保守的茎环结构。相比之下，组蛋白变体如H3.3 和H2A.Z由不受细胞周期调节的多聚腺苷酸化mRNA编码。 独特的RD-组蛋白mRNA 3'端的产生是由一种蛋白酶的活性引起的。 进化保守的前RNA加工因子。编码这五种基因 RD-组蛋白在后生动物基因组中聚集，并且转录和前体蛋白在后生动物基因组中聚集。 组蛋白mRNA生物合成所需的mRNA加工因子被组织成 在这些基因簇上组装的核体（组蛋白基因座体或HLB）。我们 将决定RD-组蛋白mRNA的协调合成的要求 在果蝇中使用生物化学和遗传学方法，特别关注 HLB在组蛋白转录和前mRNA加工中的作用。组蛋白 基因簇提供了一个系统，在该系统中，人们可以很容易地研究一个紧密相关的基因的表达。 在mRNA生物合成的所有水平上的一组受调控的基因，从组织中 通过激活转录、前mRNA加工和 转录终止我们的果蝇实验范式允许在体内 基因表达中这些基本过程的研究， 在其他实验系统中不可用。

ABSTRACT Proper regulation of histone biosynthesis during the cell cycle is critical for coordinating chromatin assembly and DNA replication. Overproduction or underproduction of histone protein results in replication stress and genome instability that contributes to the development of cancer. In this proposal, we undertake a comprehensive analysis of replication-dependent (RD) histone mRNA biosynthesis and how this process is regulated during the cell cycle in the context of animal development. RD-histone mRNAs are the only eukaryotic mRNAs in animal cells that lack a polyA tail, ending instead in a conserved stem loop structure. In contrast, mRNAs for histone variants such as H3.3 and H2A.Z are encoded by polyadenylated mRNAs that are not cell cycle regulated. Generation of the unique RD-histone mRNA 3’ end results from the activity of an evolutionary conserved set of pre-RNA processing factors. The genes encoding all five RD-histone proteins are clustered in metazoan genomes, and transcription and pre- mRNA processing factors required for histone mRNA biosynthesis are organized into a nuclear body (the histone locus body or HLB) that assembles at these gene clusters. We will determine the requirements for the coordinate synthesis of the RD-histone mRNAs using both biochemical and genetic approaches in Drosophila, with a particular focus on the role that the HLB plays in histone transcription and pre-mRNA processing. Histone gene clusters provide a system in which one can readily study the expression of a tightly regulated set of genes at all levels of mRNA biosynthesis, from the organization of genes within the nucleus through activation of transcription, pre-mRNA processing and transcription termination. Our Drosophila experimental paradigm permits the in vivo interrogation of these fundamental processes in gene expression in ways that are unavailable in other experimental systems.