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

项目摘要 大多数真核生物的信使RNA前体（pre-mRNAs）必须经过广泛的共- 在它们被输出到细胞质之前， 并作为mRNA发挥作用。加工事件包括5 ′-末端加帽、剪接和3 ′-末端加帽。 末端裂解和多聚腺苷酸化。大多数前体mRNA的3 '末端加工需要 大量的蛋白质因子用于其执行，包括切割， 多聚腺苷酸化特异性因子（CPSF），裂解刺激因子（CstF），裂解 因子I和II，以及聚（A）聚合酶（PAP）。3 '端加工机械 酵母与哺乳动物中的酵母有相似之处，但也有显着差异。 复制依赖的组蛋白前体mRNA在其3 ′端附近含有一个保守的茎环， 结束并使用一种独特的机器进行加工，尽管它共享一些蛋白质， 与典型的前mRNA 3 ′末端加工机制的因素。 mRNA 5 ′-末端加帽发生在RNA聚合酶II转录的早期， 人们普遍认为，封顶总是进行到完成。我们最近 发现Rai 1/DXO蛋白质家族是mRNA加帽质量的一部分 监督机制。它们具有RNA 5 ′-末端焦磷酸水解酶（PPH） 和去帽活性，并帮助从细胞中去除不完全加帽的mRNA。 尽管对这些mRNA加工和质量控制因子进行了广泛的研究， 我们对其分子作用机制的了解仍存在重大差距。我们 将对蛋白质因子及其复合物进行结构研究，并评估 通过仔细的生化和功能实验进行结构观察。的 建议的项目将大大提高我们对这些重要事件的了解， mRNA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.