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Structural investigation of Aquarius and associated splicing factors that couple pre-mRNA splicing to downstream RNA-processing events

Aquarius 的结构研究以及将前 mRNA 剪接与下游 RNA 加工事件耦合的相关剪接因子

基本信息

批准号：
241796087
负责人：
Dr. Vladimir Pena, Ph.D.
金额：
--
依托单位：
Institute of Cancer Research
依托单位国家：
德国
项目类别：
Research Grants
财政年份：
2013
资助国家：
德国
起止时间：
2012-12-31 至 2020-12-31
项目状态：
已结题

来源：
https://gepris.dfg.de/gepris/projekt/241796087?language=en
关键词：
Structural investigation Aquarius associated splicing

项目摘要

The spliceosome is a ribonucleoprotein machine that assembles on the pre mRNA and there catalyses the excision of introns and the ligation of exons. The entire splicing pathway is highly dynamic, with the spliceosome undergoing numerous remodelling steps driven by the motor activity of RNA helicases. Apart from participating in splicing per se, some of the spliceosomal components play pivotal roles in coupling splicing to other events of RNA metabolism.Aquarius is a massive spliceosomal helicase which binds the intron at a precise location and couples splicing to the deposition of the exon-junction complex on the mRNA as well as to the formation of snoRNPs from intronic segments. The structure of Aquarius is largely unknown and the mechanism that it employs for coupling is not understood.Converging evidence suggests that Aquarius is recruited to the spliceosome as part of a pentameric complex that we denote IBC (intron-binding complex). Subsequently, Aquarius and the other IBC components remain stably integrated into the spliceosome until the final stages of the splicing pathway. Elucidating the mechanistic role of Aquarius and, implicitly, that of the IBC in coupling spliceosomal remodelling to the downstream RNA-processing events would be greatly advanced by their 3D characterisation at atomic resolution. Therefore, our main goals are the crystallisation and X ray structure determination of Aquarius, of its complex with RNA and of the entire IBC. The crystallographic studies will be complemented with biochemical and biophysical analyses, in isolation and in the context of the whole spliceosome.

剪接体是一种核糖核蛋白机器，在前 mRNA 上组装并催化内含子的切除和外显子的连接。整个剪接途径是高度动态的，剪接体在 RNA 解旋酶的运动活动的驱动下经历许多重塑步骤。除了参与剪接本身之外，一些剪接体成分在将剪接与 RNA 代谢的其他事件偶联方面发挥着关键作用。Aquarius 是一种巨大的剪接体解旋酶，它在精确的位置结合内含子，并将剪接与外显子连接复合物在 mRNA 上的沉积以及从内含子片段形成 snoRNP 偶联。 Aquarius 的结构在很大程度上是未知的，并且它用于偶联的机制也不清楚。汇聚的证据表明，Aquarius 被招募到剪接体中，作为五聚体复合物的一部分，我们将其表示为 IBC（内含子结合复合物）。随后，Aquarius 和其他 IBC 组件保持稳定地整合到剪接体中，直到剪接途径的最后阶段。阐明 Aquarius 以及隐含的 IBC 在将剪接体重塑与下游 RNA 处理事件耦合中的机制作用将通过它们在原子分辨率下的 3D 表征得到极大的推进。因此，我们的主要目标是 Aquarius、其与 RNA 的复合物以及整个 IBC 的结晶和 X 射线结构测定。晶体学研究将辅以生化和生物物理分析，在整个剪接体的背景下进行单独的分析。