Roles of S. pombe SR proteins in spliceosome function and assembly

描述（由申请人提供）：人类大多数基因表达的一个重要步骤是去除内含子，内含子通过大而动态的剪接体复合物从mrna前体剪接出来。研究表明，绝大多数人类遗传性疾病是由内含子的错误移除引起的。SR蛋白是人类重要的剪接因子，对剪接体的功能和调控至关重要。更具体地说，它们已被证明在剪接体组装以及剪接反应的后期步骤中发挥重要作用。SR蛋白水平的改变已被证明会导致许多人类疾病，如癌症和艾滋病毒。考虑到SR家族的9个蛋白、组织特异性表达水平以及在选择性剪接中的复杂作用，在人类中区分单个SR蛋白的作用已被证明是困难的。为了区分SR蛋白在遗传可处理系统中的全基因组作用，将利用单细胞模式生物裂糖酵母。在S. pombe中只有两种SR蛋白，其中许多内含子具有与人类相似的特征。鉴于S. pombe中许多内含子的不同内含子结构，只有少数内含子被测试依赖SR蛋白进行最佳剪接。此外，许多已被证明与人类SR蛋白相互作用的剪接因子也存在于S. pombe中，尽管这些相互作用仅被观察到少数，目前缺乏全面的分析。利用酵母特有的遗传技术以及适用于人类实验的生化分析，拟议的研究将确定SR蛋白如何与S. pombe （AIM1）中的剪接体相互作用，以及它们在有效去除该模式生物（AIM2）中存在的许多独特内含子的全基因组中起什么作用。此外，拟建的研究将通过分析特定内含子（AIM3）上的剪接体组装来确定SR蛋白如何影响S. pombe剪接的机制。对S. pombe中SR蛋白如何与多种内含子一起发挥作用的基本理解，将为SR蛋白的一般机制和剪接体对pre-mRNA加工的基本过程提供信息。

DESCRIPTION (provided by applicant): An essential step in the expression of the majority of genes in humans is the removal of introns, which are spliced out of pre-mRNA by the large and dynamic spliceosome complex. It has been indicated that a significant majority of hereditary disease in humans is caused by the incorrect removal of introns. SR proteins in humans are important splicing factors that have been shown to be essential for spliceosome function and regulation. More specifically, they have been shown to play essential roles in spliceosome assembly as well as later steps of the splicing reaction. Altered levels of SR proteins have been shown to result in many human diseases such as cancer and HIV. Distinguishing the roles of individual SR proteins has proven difficult in humans given the nine-protein SR family, tissue specific expression levels, and complex roles in alternative splicing. In order to distinguish the roles of SR proteins genome-wide in a genetically tractable system, the unicellular model organism Schizosaccharomyces pombe will be utilized. There are only two SR proteins in S. pombe with many introns containing similar characteristics with humans. Given the diverse intron structure of the many introns in S. pombe only a few introns have been tested for reliance on SR proteins for optimal splicing. In addition, many of the splicing factors that have been shown to interact with human SR proteins are also present in S. pombe although only a few of these interactions have been observed and a thorough analysis is currently lacking. Using genetic techniques that are uniquely available to yeast as well as biochemical assays adapted from experiments in humans, proposed studies will determine how SR proteins interact with the spliceosome in S. pombe (AIM1) and what role they play genome-wide in the efficient removal of the many unique introns that are present in this model organism (AIM2). In addition, proposed studies will determine the mechanism of how SR proteins influence splicing in S. pombe by analyzing spliceosome assembly on specific introns (AIM3). The fundamental understanding of how SR proteins function with the diverse set of introns in S. pombe will inform general mechanisms of SR proteins and the essential process of pre-mRNA processing by the spliceosome as a whole.