Structural and functional studies of mRNA processing, stability and quality control

项目摘要 大多数真核信使RNA前体(Pre-mRNAs)必须经历广泛的协同作用 在它们被输出到细胞质之前，在核中进行转录处理 并作为mRNAs发挥作用。加工事件包括5美分末端封口、拼接和3美分- 末端切割和多聚腺苷酸化。大多数前mRNAs的3‘端处理需要 它的执行需要大量的蛋白质因子，包括卵裂和 多聚腺苷酸特异性因子(CPSF)、切割刺激因子(CstF)、切割 因子I和因子II，以及多聚(A)聚合酶(PAP)。中国的3‘端加工机械 酵母与哺乳动物有相似之处，但也有显著的不同。 复制依赖的组蛋白前mRNAs在其3？ 结束并使用一种独特的机械来处理它，尽管它共享一些蛋白质 与规范的Pre-mRNA3‘末端处理机制有关的因素。 在RNA聚合酶II转录的早期，mRNA5‘端封顶，并且它 人们普遍认为，封顶总是会进行到完成。我们最近做了 发现Rai1/DXO蛋白家族是mRNA封顶质量的一部分 监督机制。它们可以拥有RNA5‘端焦磷水解酶(PPH) 以及去包裹活动，并帮助从细胞中移除未完全封顶的mRNA。 尽管对这些mRNA的加工和质量控制因素进行了广泛的研究， 我们对它们的分子作用机制的认识仍然存在很大差距。我们 将对蛋白质因子及其复合体进行结构研究，并评估 通过仔细的生化和机能实验进行结构观察。这个 拟议的项目将大大加深我们对这些重要事件的了解 信使核糖核酸生命周期。

Project Summary Most eukaryotic messenger RNA precursors (pre-mRNAs) must undergo extensive co- transcriptional processing in the nucleus before they can be exported to the cytoplasm and function as mRNAs. The processing events include 5¢-end capping, splicing, and 3¢- end cleavage and polyadenylation. The 3’-end processing of most pre-mRNAs requires a large number of protein factors for its execution, including cleavage and polyadenylation specificity factor (CPSF), cleavage stimulation factor (CstF), cleavage factors I and II, and poly(A) polymerase (PAP). The 3’-end processing machinery in yeast has similarity to that in mammals, although there are also significant differences. Replication-dependent histone pre-mRNAs contain a conserved stem-loop near their 3¢- end and employ a distinct machinery for its processing, although it shares some protein factors with the canonical pre-mRNA 3¢-end processing machinery. mRNA 5¢-end capping occurs early during transcription by RNA polymerase II, and it was generally believed that capping always proceeds to completion. We have recently discovered that the Rai1/DXO family of proteins are part of a mRNA capping quality surveillance mechanism. They can possess RNA 5¢-end pyrophosphohydrolase (PPH) and decapping activities, and help remove incompletely capped mRNAs from cells. Despite the extensive studies on these mRNA processing and quality control factors, significant gaps remain in our knowledge on their molecular mechanisms of action. We will carry out structural studies on the protein factors and their complexes, and assess the structural observations by careful biochemical and functional experiments. The proposed project will greatly enhance our understanding of these important events in mRNA lifecycle.