STRUCTURE AND EXPRESSION OF EUKARYOTIC GENES

反式剪接涉及将两个独立的RNA分子连接到 一份连续的成绩单。锥虫、线虫和吸虫 进行反式剪接。线虫中的剪接前导RNA(SL RNA) 将其5‘22个核苷酸从一致的供体剪接位点转移到 共识受体剪接位点位于5-非翻译区 一种前信使核糖核酸。因此，一些mRNAs携带22个核苷酸长。 5‘拼接的前导序列。 已鉴定出两种蛋白质，它们与mRNAs特异性结合， 包含SL，但无法与缺少SL的mRNA结合。建议的 研究将定义这些分子之间的详细相互作用 SL结合蛋白和SL核苷酸序列。这些方法包括 RNA的化学修饰和复合体的足迹。 还将选择与蛋白质结合的变异rna分子。 亲和力很强。这些变体将有助于定义一致的RNA 序列和结构将提供对体系结构的洞察 分子间的相互作用。这些研究将有助于建立 RNA蛋白质相互作用的基本规则。 结合蛋白的生物学功能及其相互作用 带有SL的mRNA将在线虫中进行遗传学研究。基因 编码SL蛋白的基因将被鉴定出来。DNA序列同源性 将寻找可能与SL结合相关的相关基因 蛋白质与其他已知功能的RNA结合蛋白的结合。基因 转座子会破坏结合蛋白的编码 诱变。这些突变线虫的表型缺乏 将对结合蛋白进行研究以确定无效表型 并确定这些蛋白质是否是必需的。突变将是 以阐明它们的生物学功能 线虫的生长发育及其对 已知功能的编码蛋白结构域，例如显性负值 与核苷酸结合位点相关的表型。

Trans-splicing involves the ligation of two separate RNA molecules into a single contiguous transcript. Trypanosomes, nematodes and trematodes carry out trans-splicing. In nematodes, the spliced leader RNA (SL RNA) transfers its 5'22 nucleotides from a consensus donor splice site to a consensus acceptor splice site located in the 5-untranslated region of a pre-mRNA. Consequently, some mRNAs carry a 22 nucleotide long 5'spliced leader sequence. Two proteins have been identified that bind specifically to mRNAs that contain an SL but fail to bind to mRNAs that lack an SL. The proposed research will define the detailed molecular interactions between these SL binding proteins and the SL nucleotide sequence. The methods include chemical modifications of the RNA and footprinting of the complex. Variant RNA molecules will also be selected that bind to the proteins with high affinity. These variants will help define a consensus RNA sequence and structure that will provide insight into the architecture of the molecular interactions. These studies will help establish the basic rules for RNA protein interactions. The biological function of the binding proteins and their interactions with SL on the mRNA will be studied genetically in C. elegans. The genes encoding the SL proteins will be identified. DNA sequence homologies with related genes will be sought that might relate the SL binding proteins to other RNA binding proteins of known function. The genes encoding the binding proteins will be disrupted by transposon mutagenesis. The phenotypes of those mutant nematodes lacking the binding proteins will be studied in order to define the null phenotype and to determine whether the proteins are essential. Mutations will be derived in the genes in order to elucidate their biological functions in the growth and development of C. elegans with particular attention to encoded protein domains of known functions, e.g. dominant negative phenotypes associated with nucleotide binding sites.