Understanding the structure and function of U1 snRNP

了解 U1 snRNP 的结构和功能

基本信息

批准号：
10200085
负责人：
RUI ZHAO
金额：
$ 45.17万
依托单位：
UNIVERSITY OF COLORADO DENVER
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-08-01 至 2023-06-30
项目状态：
已结题

项目摘要

Abstract Pre-mRNA splicing is essential for gene expression in all eukaryotes and errors in splicing cause genetic disorders and many other diseases. A thorough understanding of the molecular mechanisms of pre-mRNA splicing has the potential to provide useful approaches for human disease therapy. The splicing of introns is carried out through two transesterification reactions catalyzed by the spliceosome, a large RNA/protein complex composed of five snRNPs (U1, U2, U4, U5, U6) and many non-snRNP related protein factors. Despite the recent determination of multiple high resolution cryo EM structures of spliceosome complexes at later stages of the splicing pathway, there is a lack of structural and mechanistic understanding of the initial intron recognition and early spliceosome assembly events. U1 snRNP is critical for the initial recognition of 5' splice site (ss). Due to its important role in 5' ss recognition, U1 snRNP is a frequent target of the action of alternative splicing factors that either facilitate or prevent U1 snRNP from binding to 5' ss. Much of what we know today about the molecular mechanism and regulation of 5' ss recognition comes from genetic, biochemical, and structural studies of two commonly used model systems, S. cerevisiae (yeast) and human U1 snRNP. Intriguingly, the yeast U1 snRNP is much more complex than the human U1 snRNP. Yeast U1 snRNA contains a 3.5-fold larger RNA and seven additional proteins, most of which have human homologs that are weakly associated with U1 snRNP and are involved in alternative splicing. We have recently determined the cryoEM structure of yeast U1 snRNP to 3.6 Å resolution, generating many interesting hypotheses on how human alternative splicing factors recruit U1 snRNP and the unexpected role of human PrpF39 in alternative splicing. We will test these hypotheses using a combination of biochemistry and molecular biology approaches. We will also decipher the molecular mechanism of concerted recognition of all intron elements through a combination of structural biology and biochemical approaches. Results from this project will significantly advance our understanding of the molecular mechanism of intron definition and alternative splicing.

抽象的 mRNA 前体剪接对于所有真核生物的基因表达至关重要，剪接错误会导致遗传全面了解前 mRNA 的分子机制。剪接有可能为人类疾病治疗提供有用的方法。通过剪接体（一种大的 RNA/蛋白质）催化的两个酯交换反应进行由五个 snRNP（U1、U2、U4、U5、U6）和许多非 snRNP 相关蛋白因子组成的复合物。最近确定了剪接体复合物的多个高分辨率冷冻电镜结构在剪接途径的各个阶段，缺乏对初始内含子的结构和机械理解识别和早期剪接体组装事件。 U1 snRNP 对于 5' 剪接位点 (ss) 的初始识别至关重要由于其在 5' ss 识别中的重要作用， U1 snRNP 是促进或阻止 U1 的选择性剪接因子作用的常见目标 snRNP 与 5' ss 的结合是我们今天所了解的有关 5' 分子机制和调控的大部分内容。 ss识别来自于两个常用模型系统的遗传、生化和结构研究，酿酒酵母（酵母）和人类 U1 snRNP 有趣的是，酵母 U1 snRNP 比酵母复杂得多。酵母 U1 snRNP 含有 3.5 倍大的 RNA 和 7 种附加蛋白质，其中大部分是其具有与 U1 snRNP 弱相关的人类同源物并参与替代我们最近以 3.6 Å 分辨率确定了酵母 U1 snRNP 的冷冻电镜结构，生成了关于人类选择性剪接因子如何招募 U1 snRNP 的许多有趣的假设以及意想不到的情况人类 PrpF39 在选择性剪接中的作用我们将使用以下组合来测试这些假设。我们还将破译生物化学和分子生物学方法的分子机制。通过结构生物学和生化方法的结合识别所有内含子元件。该项目的结果将显着促进我们对内含子分子机制的理解定义和选择性剪接。