TRANS-SPLICING IN C ELEGANS

线虫中的反式剪接

• 批准号: 3300963
• 负责人: Thomas Blumenthal
• 金额: $ 18.68万
• 依托单位: TRUSTEES OF INDIANA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-05-01 至 1995-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3300963
• 关键词: Caenorhabditis elegans; RNA splicing; genetic manipulation; molecular cloning; nucleic acid sequence; precursor mRNA; small nuclear ribonucleoproteins; spliceosomes

DESCRIPTION (Adapted from Applicant's Abstract): Nuclear mRNA processing is catalyzed by spliceosomes which are composed of small nuclear ribonucleoprotein particles (snRNPs) and additional proteins. In C. elegans this process is essentially similar to splicing in higher eucaryotes, except the introns tend to be much smaller. Nematodes also perform an alternative spliceosome-catalyzed mRNA splicing event, trans- splicing, in which the RNA products of two separate gene are splices together to form a single mature mRNA. While some genes are conventional in that they initiate transcription at the 5' end of exon 1, the pre mRNA products of others receive a 22 nucleotide exon 1 by trans-splicing. This exon is donated by a 100 nucleotide RNA molecule, SL RNA, which is itself packaged as a snRNA. C. elegans has two distinct SL snrnps, each of which delivers its 5' 22 nucleotides to a distinct group of pre-mRNAS. The investigator is exploring the relationship between normal (cis-) splicing and trans-splicing. Why are the pre-mRNA products of some genes recipients for SL RNA splicing which others are not; Where do the specificity determinants for SL1 vs. SL2 splicing reside, both on the SL snRNP and on the recipient pre-mRNAs? The investigator has shown that, when moved into an appropriate context, an intron 3' splice site is sufficient to convert a conventional gene into a trans-spliced gene. Conversely the investigator has shown that it is possible to convert a trans-spliced gene into a conventional gene by inserting a 5' splice site upstream of its trans- splicing site. In the current application the investigator proposes a series of transformation experiments in which genes are mutated, introduced into worms, and tested for the splicing events they undergo. The following specific questions will be answered; (I). What are the branch point and 3' splice site sequence requirements for trans-splicing? (II). How is the trans-splice site chosen? (III). Is there a minimal or maximal outron length? (The investigator defines the region of a trans-spliced transcript between its 5' end and the trans-splice site as an "outron." (IV). Is an overall high percent A+U in the outron required for trans-splicing? (V). What factors influence the competition that occurs between cis-and trans- splicing when a 5' splice site is present in an outron? In this situation how critical is distance of the 5' splice site from the trans-splice site? (VI). Where is the information on the recipient RNA that results in specific splicing of SL1 or SL2? (VII). Where is the information on SL1 RNA and SL2 RNA for specific trans-splicing?

描述（改编自申请人的摘要）：核mRNA加工 由剪接体催化，剪接体由小的核 核糖核蛋白颗粒（snRNP）和另外的蛋白质。 in C. 这一过程基本上类似于高等生物中的剪接。 真核生物，除了内含子往往要小得多。 线虫也 进行选择性剪接体催化的mRNA剪接事件，反式 剪接，其中两个独立基因的RNA产物是剪接 形成一个成熟的mRNA。 虽然有些基因是传统的 因为它们在外显子1的5 ′端启动转录，前mRNA 其它的产物通过反式剪接接收22个核苷酸的外显子1。 这 外显子由100个核苷酸的RNA分子SL RNA提供，SL RNA本身是 包装成snRNA C. elegans有两个不同的SL snrnps，每一个都 将其5 ′ 22个核苷酸递送至不同的pre-mRNAS组。 的 研究人员正在探索正常（顺式）剪接 和反式剪接。 为什么某些基因的前mRNA产物是受体 对于SL RNA剪接，其他人没有;特异性在哪里 SL1与SL2剪接的决定因素存在于SL snRNP和 受体前体mRNA 研究人员表示，当进入 在适当的情况下，内含子3 '剪接位点足以将 一个常规的基因变成一个反式剪接的基因。 相反，研究人员 已经表明，有可能将反式剪接的基因转化为 通过将5 '剪接位点插入其反式上游， 剪接位点 在本申请中，研究者提出了一个 一系列基因突变的转化实验，介绍了 并测试它们所经历的剪接事件。 以下 将回答具体问题;（一）。 什么是分支点和3 ' 反式剪接的剪接位点序列要求？ （二）. 怎么样 选择反式剪接位点（三）、 有最小或最大的outron吗 长度？ (The研究者定义了反式剪接转录本的区域 在其5 ′端和反式剪接位点之间作为"外区"。"（四）。 是一 反式剪接所需的外区中的总体高百分比A + U？ （五）. 哪些因素影响顺式和反式之间的竞争？ 当一个5 '剪接位点存在于一个外区时， 在这种情况下 5 '剪接位点与反式剪接位点的距离有多重要？ （六）。 导致以下结果的受体RNA信息在哪里 特异性剪接的SL1或SL2？ （七）。 SL1的信息在哪里 RNA和SL2 RNA的特异性反式剪接？