TRANS-SPLICING IN C ELEGANS

描述(改编自申请者摘要)：核mRNA加工 是由剪接体催化的，剪接体由小核组成 核糖核蛋白颗粒(SnRNPs)和其他蛋白质。在C。 这个过程本质上类似于更高级的剪接 真核生物，除了内含子往往要小得多。线虫也 执行另一种剪接体催化的mRNA剪接事件，反式- 剪接，其中两个独立基因的RNA产物是剪接 共同形成单个成熟的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，即前mRNA 其他的产物通过反式剪接得到一个22个核苷酸的外显子1。这 外显子是由100个核苷酸的RNA分子SL RNA捐赠的，它本身就是 包装成单链核糖核酸。线虫有两个不同的SL单链RNP，每个都是 将其5‘22个核苷酸传递给一组不同的前-mRNAs。这个 研究人员正在探索正常(顺式)剪接之间的关系 和反式剪接。为什么某些基因的前信使核糖核酸产物是受体 对于SL RNA的剪接，其他的不是；特异性在哪里 SL1和SL2剪接的决定因素位于SL SnRNP和On上 接受者是前病毒吗？调查人员已经证明，当搬进 在适当的上下文中，内含子3‘剪接位点足以转化为 将传统基因转化为反式剪接基因。相反，调查员 已经表明有可能将反式剪接的基因转化为 通过在其反式基因的上游插入一个5‘剪接位点来获得常规基因. 拼接位置。在当前的申请中，调查者提出了一种 介绍了一系列基因突变的转化实验 变成蠕虫，并对它们所经历的剪接事件进行测试。以下是 具体问题将会获得解答；(I)。分支点是什么？3‘ 反式剪接的剪接位点序列要求？(Ii)。你还好吗？ 选择了反式剪接位点吗？(三)。是否存在最小或最大超子 长度？(研究人员定义了反式剪接转录本的区域 在它的5‘端和反式剪接位点之间是一个“外显子”。(四)。是一种 反式剪接所需的Outtron中的整体高A+U百分比？(五)。 什么因素会影响顺式和反式之间的竞争？ 在Outtron中存在5‘剪接点时进行剪接吗？在这种情况下 5‘剪接部位与反式剪接部位的距离有多重要？ (Vi)。在哪里是关于接收者RNA的信息，它导致 SL1或SL2的特定剪接？(七)。关于SL1的信息在哪里 RNA和SL2 RNA用于特异的反式剪接？

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?