Co-transcriptional Recruitment and Activity of the Spliceosomal snRNPs

该奖项是根据2009年美国复苏和再投资法案(公法111-5)资助的研究项目。在每个真核细胞的核心是核，这是一个高度组织成许多不同功能域的复杂结构，起着两个基本作用：维持基因组和分发遗传信息。在亚核结构域中，基因被转录成前信使RNA(Pre-mRNA)分子的位置。当它们被合成时，前mRNAs经历了一系列复杂的处理事件来产生信使RNAs，这些信使RNA随后用于蛋白质的合成。其中一项活动，剪接，需要五个主要的小核糖核蛋白颗粒(SNRNP)。尽管SnRNPs在剪接中起着关键的催化作用，但对调节它们招募到前mRNAs的细胞机制仍然知之甚少。这个项目的目的是将SNRNPs的分析转移到两栖类卵母细胞，这是一个独特的细胞系统，在这个系统中，可以在透射光学显微镜中直接显示单个转录位点和其他亚核结构域。使用这个体内系统的主要目的是确定SnRNPs与前mRNAs相互作用所需的特征元件及其共转录剪接活性。在蛙卵母细胞中，活跃转录的位置对应于灯刷染色体(LBC)的侧环。这项研究基于两种新开发的分析方法，这两种方法允许跟踪SnRNPs与LBC侧环的关联，并分别确定这些环上是否发生了前mRNA剪接。从长远来看，这些研究将有助于全面了解控制活跃转录单位的功能组织的细胞机制，以及不同的亚核结构域如何相互作用来调节前mRNA的加工。广泛的影响。两栖类卵母细胞的巨大尺寸和它的易操纵性使其特别适合积极的外展计划。事实上，向所有级别的学生(本科生和高中生)展示在荧光显微镜中活跃转录的染色体和基因总是能成功地抓住他们的兴趣，同时向他们介绍实验科学的原理。调查人员将利用这项研究继续培养与邻近高中和社区大学的教师的关系，为制定强有力的课程做出贡献，并为包括女性和少数族裔在内的学生提供研究实习机会。其目标是为教师提供科学支持，让学生接触学术环境中的研究，同时促进他们从小型高中和社区大学过渡到大型本科生校园。

This award is funded under the American Recovery and Reinvestment Act of 2009 (Public Law 111-5).Research project. At the heart of each eukaryotic cell is the nucleus, a complex structure highly organized into many distinct functional domains to perform two fundamental roles: the maintenance of the genome and the distribution of the genetic information. Among the sub-nuclear domains are the sites where genes are transcribed into pre-messenger RNA (pre-mRNA) molecules. As they are being synthesized, pre-mRNAs undergo a series of complex processing events to produce the messenger RNAs that are subsequently used for the synthesis of proteins. One of these events, splicing, requires the five major small ribonucleoprotein particles (snRNPs). Despite the critical catalytic role of snRNPs in splicing, little is still known about the cellular mechanisms that regulate their recruitment to pre-mRNAs. This program aims at moving the analysis of snRNPs to the amphibian oocyte, a unique cell system where direct visualization of individual transcription sites and other sub-nuclear domains is possible in the transmitted light microscope. The main objective, using this in vivo system, is to determine the characteristic elements of snRNPs required for their interactions with pre-mRNAs and their co-transcriptional splicing activity. In frog oocytes, the sites of active transcription correspond to the lateral loops of the lampbrush chromosomes (LBCs). The research is based on two newly developed assays, which permits one to follow the association of snRNPs with the lateral loops of LBCs and to determine whether pre-mRNA splicing occurs on these loops, respectively. In the long term, these studies will contribute to the general understanding of the cellular mechanisms that govern the functional organization of an active transcription unit, and how various subnuclear domains interact to regulate pre-mRNA processing.Broader Impacts.The giant size of an amphibian oocyte and its ease of manipulation make it particularly amenable to an active outreach program. Indeed, showing chromosomes and genes being actively transcribed in a fluorescence microscope to students of all levels (undergraduate and highschool) always succeeds at capturing their interest, while introducing them to the principles of experimental science. The investigator will use the research to continue fostering relations with teachers at neighboring high schools and community colleges, contributing to the development of strong curricula and providing their students, including women and minorities, with research internships. The goal is to provide teachers with scientific support and to expose students to research in an academic setting, while facilitating their transition from small high schools and community colleges to large undergraduate campuses.