RNA SPLICING CONTROL IN ROUS SARCOMA VIRUS

劳斯肉瘤病毒中的 RNA 剪接控制

• 批准号: 6377180
• 负责人: MARK T MCNALLY
• 金额: $ 22.21万
• 依托单位: MEDICAL COLLEGE OF WISCONSIN
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-08-14 至 2003-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6377180
• 关键词: HeLa cells; RNA splicing; RNase protection assay; Rous sarcoma virus; crosslink; gel filtration chromatography; gene mutation; genetic regulatory element; protein purification; site directed mutagenesis; small nuclear ribonucleoproteins; transcription factor; ultraviolet radiation; virus genetics

DESCRIPTION (Adapted from Abstract): This application uses the Rous sarcoma virus (RSV) as a model to study regulation of RNA processing. RSV primary transcripts are spliced by the host RNA splicing machinery, but this process is controlled since the majority (75%) of the RNA remain unspliced to serve as the message for the gag-pol proteins and as genomic RNA for progeny virions. Viral proteins are not required for the normal processing indicating that the cis elements are in the RNA and host-derived trans-acting factors are responsible for the ratio of spliced to unspliced RNA. This study characterizes these cis and trans-acting factors. One control element has been identified, the Negative regulator of splicing (NRS), which is located in gag and is distinctive in that it is not located near any of the splice sites, yet suppresses both the env and src 3 splice sites (ss). Two subregions of the NRS have been identified; an upstream purine-rich region that binds essential splicing factors called SR proteins and an unknown factor (p55), and a downstream sequence that is required for binding of U11 snRNP, an RNP particle that recognizes the 5 ss in a new class of rare introns (AT-AC introns). U1 and U2, abundant snRNPs of the major splicing pathway also interact with the NRS RNA. Four specific aims are proposed. The first uses in vivo and in vitro approaches to further characterize the cis elements and trans-acting factors which govern NRS splicing inhibition. The second aim tests a hypothesis that the U11 snRNP, when bound to the NRS interacts nonproductively with the major pathway splicing factors present at the authentic 3 ss, excluding pairing with the authentic 5 ss. The third aim studies an element which suppresses src splicing, named the SSS. The final aim investigates the basis of the position dependence of the NRS and SSS in splicing inhibition. This may be effected by splice site competition or the position relative to the 5' cap structure.

描述（改编自摘要）：该应用程序使用劳斯肉瘤 病毒（RSV）作为研究 RNA 加工调控的模型。 RSV初级 转录本由宿主RNA剪接机器进行剪接，但这个过程 受到控制，因为大多数 (75%) RNA 保持未剪接状态以供使用 作为 gag-pol 蛋白的信息和后代的基因组 RNA 病毒体。 正常加工不需要病毒蛋白 表明顺式元件位于 RNA 中且源自宿主 反式作用因子决定剪接与未剪接的比例 核糖核酸。 这项研究表征了这些顺式和反式作用因子。 一 控制元件已被识别，剪接的负调节因子 (NRS)，位于 gag 中，其独特之处在于它不位于 靠近任何剪接位点，但同时抑制 env 和 src 3 剪接 站点（ss）。 NRS 的两个次区域已确定；上游 富含嘌呤的区域，结合称为 SR 蛋白的必需剪接因子 和一个未知因子（p55），以及所需的下游序列 U11 snRNP 的结合，一种 RNP 颗粒，可识别新的 5 ss 一类稀有内含子（AT-AC 内含子）。 U1 和 U2，丰富的 snRNP 主要剪接途径也与 NRS RNA 相互作用。 四个具体目标 被提议。 第一个使用体内和体外方法进一步 描述控制 NRS 的顺式元件和反式作用因子 剪接抑制。 第二个目标测试 U11 snRNP 的假设， 当与 NRS 结合时，与主要途径发生非生产性相互作用 真实 3 ss 中存在的剪接因子，不包括与 正品 5 SS。 第三个目标是研究抑制 src 的元素 拼接，命名为SSS。 最终目的是研究该理论的基础 NRS 和 SSS 在剪接抑制中的位置依赖性。 这可能是 受剪接位点竞争或相对于 5' 帽的位置的影响 结构。