Uncovering the mechanisms of splice site choice

该项目旨在了解储存在DNA中的遗传信息是如何被准确处理以编码蛋白质组装的。为了将DNA中的信息用于细胞，DNA首先被复制到称为RNA的中间姊妹分子中，然后被解码成蛋白质序列。然而，为了有用，必须首先去除RNA的内部部分，然后通过一种称为RNA剪接的过程将周围的序列精确地粘贴在一起。剪接是由细胞中最大、最复杂的机器之一-剪接体执行的，剪接体由100多种蛋白质和5个RNA分子组成，它们在要剪接的区域组装到RNA上。由于将如此大的机器组装到RNA上的复杂性，关于剪接体准确选择RNA中将被剪接在一起的位置的机制仍有许多需要了解。寻找影响剪接位点选择的突变的基因实验是这项研究的重要和前沿组成部分，这些实验将由加州大学圣克鲁斯分校(UCSC)既定的夏季本科生培训项目的少数族裔本科生参与者进行。这一培训对这些学生进入加州大学洛杉矶分校教职员工研究实验室的职位以及未来进入博士课程将是重要的准备。该项目以微小的线虫秀丽线虫为模型，通过集成的遗传、基因组和生物化学方法来探索前mRNA剪接。在初步研究中，已经开发出独特的方法，揭示了在被移除的RNA区域的两端的剪接位点选择的新的调节现象，称为5‘和3’剪接位点。基因筛查已经被用来发现对调节5‘剪接位点选择很重要的因子的新作用。在体细胞和生殖细胞之间的相邻3‘剪接位点的选择性选择上已经发现了根本的差异。这项研究将通过解决以下两个问题来扩展这些观察：蛋白质因素如何调节5‘剪接位点的选择，以及在体细胞和生殖细胞之间3’剪接位点选择的机制差异是什么？这一结果有望为剪接机械如何准确选择5‘和3’剪接位点提供新的见解。

This project is aimed at understanding how the genetic information stored in DNA is accurately processed to code for protein assembly. In order for information in DNA to be used in the cell, the DNA is first copied into an intermediate sister molecule called RNA which is then decoded into protein sequence. However, in order to be useful, internal parts of the RNA must first be removed and the surrounding sequences precisely pasted together by a process called RNA splicing. Splicing is a carried out by one of the largest and most complicated machines in the cell, the spliceosome, which consists of over 100 proteins and 5 RNA molecules that assemble onto the RNA at the region to be spliced out. Because of the complexity of assembling such a large machine onto the RNA, there is still much to learn about the mechanisms by which the spliceosome accurately chooses the sites in the RNA that will be spliced together. Genetic experiments to search for mutations that affect splice site choice are an important and cutting-edge component of this research, and these experiments will be carried out by minority undergraduate participants in an established summer undergraduate training program at the University of California-Santa Cruz (UCSC). This training will be important in preparing these students for positions in UCSC faculty research labs, and for future admission into PhD programs.This project employs the microscopic worm Caenorhabditis elegans as a model to explore pre-mRNA splicing through integrated genetic, genomic and biochemical approaches. Unique approaches have been developed in the preliminary studies that uncovered new regulatory phenomena for splice site selection at both ends of the region of RNA to be removed, called the 5' and 3' splice sites. Genetic screens have been used to uncover new roles for factors important for the regulation of 5' splice site choice. Fundamental differences have been uncovered in alternative adjacent 3' splice site selection between somatic and germ cells. The research will expand on these observations by addressing the following two questions: how do protein factors regulate the choice of 5' splice sites, and what are the mechanistic differences in 3' splice site selection between soma and germ cells? The results are expected to provide novel insights into how the splicing machinery accurately chooses the 5' and 3' splice sites.