EXPRESSION AND FUNCTION OF U1 RNA ISOFORMS DURING DEVELOPMENT

这项建议代表着我们继续努力推动 有前途的少数族裔学生将在生物医学科学领域寻求职业。 为了实现这一目标，学生们已经并将接受全面的培训 分子生物学的各个方面。为此所采用的科学项目 目的涉及U1-的结构和功能表征 家蚕蛾的相关序列及其转录本。U1 RNA是六种重要的小核RNA之一，U1-U6，它 已知与六到十种蛋白质结合形成小分子 参与转化的核糖核蛋白颗粒(SnRNP) 通过剪接将hnRNA转化为mRNA。我们的实验室有部分 描述了该生物体中与U1相关的三类基因。 一类是转录不活跃的假基因； 另一种是以高度重复的U1相关序列为代表的 发育和组织特异性转录，表明在 基因表达的调节。这篇文章中概述的实验 提案将允许我们的学生阐明U1亚型所起的作用 在RNA剪接水平上调节基因表达。他们的 对hnRNA剪接的参与将在发育过程中确定 分期的丝腺、毛囊和几个细胞系。U1亚型将 被提纯并在体外用于重建实验 它们差异组装成SnRNP/hnRNP颗粒，与 含有内含子的丝素和绒毛膜RNA，剪接能力将 被调查。将进行体内实验来研究 其中U1异构体的差异组装和高效剪接 分期细胞类型和细胞系。就像β-地中海贫血的情况一样 人类的RNA剪接突变，差异剪接是由 U1异构体的效率和/或特异性可能被证明是 一些人类疾病。蚕蛾，有一个研究得很好的发育 分子生物学，加上高度专业化的细胞类型，提供了一种 我们的学生有独特的机会来研究这些U1的功能- 相关序列。具体来说，少数族裔学生将被激励去 通过让他们接触生物医学科学来追求他们的事业 相关文献，并培训他们设计实验，制定 协议，进行实验并公布结果。

This proposal represents a continuation of our efforts to motivate promising minority students to seek careers in the biomedical sciences. To achieve this goal, students have been and will be trained in all aspects of molecular biology. The scientific project employed for this purpose involves the structural and functional characterization of U1- related sequences and their transcripts in the silkmoth Bombyx mori. U1 RNA is one of six prominent small nuclear RNAs (snRNAs), U1-U6, which are known to associate with six to ten proteins to form small ribonucleoprotein particles (snRNPs) which participate in the conversion of hnRNA to mRNA by splicing. Our laboratory has partially characterized three categories of U1-related genes in this organism. One category is composed of transcriptionally inactive pseudogenes; another, is represented by a highly reiterated U1-related sequences are developmentally and tissue-specific transcribed, suggesting a role in the regulation of gene expression. The experiments outlined in this proposal will allow our students to elucidate the role U1 isoforms play in the regulation of gene expression at the RNA splicing level. Their involvement in hnRNA splicing will be determined using developmentally staged silk glands, follicles, and several cell lines. U1 isoforms will be purified and used in vitro in reconstitution experiments in which their differential assembly into snRNP/hnRNP particles, association with intron-containing fibroin and chorion RNAs, the splicing capacities will be investigated. In vivo experiments will be performed to study the differential assembly and splicing efficiently of U1 isoforms in these staged cell types and cell lines. As with the case of beta-thalassemia RNA splicing mutations in humans, differential splicing as a result of U1 isoforms efficiency and/or specificity may prove to be the basis of some human maladies. The silkmoth , with a well-studies developmental and molecular biology, plus highly specialized cell types, provides a unique opportunity for our students to examine the function of these U1- related sequences. Specifically, minority students will be motivated to pursue a career in the biomedical sciences by exposing them to the pertinent literature, and training them to design experiments, formulate protocols, conduct experiments and publish the results.