RNA HELICASES INVOLVED IN YEAST MRNA SPLICING

参与酵母 mRNA 剪接的 RNA 解旋酶

基本信息

批准号：
2701583
负责人：
Tien-Hsien CHANG
金额：
$ 11.42万
依托单位：
OHIO STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
1993
资助国家：
美国
起止时间：
1993-09-01 至 2000-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/2701583
关键词：
RNA splicing fungal genetics gene complementation genetic strain helicase messenger RNA molecular cloning mutant phenotype precursor mRNA suppressor mutations temperature sensitive mutant yeasts

项目摘要

RNA helicases are enzymes which catalyze the separation of strands in RNA duplexes or the removal of secondary structure in single-stranded RNA. The reaction requires hydrolysis of nucleoside triphosphates as an energy source. A growing number of putative RNA helicase genes involved in a variety of physiological functions such as translation initiation, ribosome assembly, mRNA splicing, and germinal line cell differentiation have been found in organisms ranging from E. coli to human. Many of these putative RNA helicases share a series of sequence elements, most noticeably the DEAD-Box motif. In yeast Saccharomyces cerevisiae, there are at least 16 putative RNA helicase genes belong to this DEAD-Box Protein (DBP) gene family. Multiple DBP genes have also been discovered in E. coli, Drosophila, and HeLa cells. Thus, this gene family appears to have evolved very early and is highly conserved in evolution. The unexpected diversity of the putative RNA helicase genes in yeast as well as in other organisms immediately raises a number of intriguing questions regarding their cellular functions and biochemical properties. A crucial question, in particular, is how and what RNAs are functionally regulated by these enzymes. This proposal will address these questions by focusing on studying the biochemistry and cellular functions of two yeast RNA helicases, PRP28 and SPP81/DED1, which are implicated in nuclear pre-mRNA splicing. Other yeast DBP genes with unknown functions will also be investigated. It is anticipated that these studies will provide novel information about the RNA helicases, of which little is known at the moment. Specific aims are: [1] To construct conditional mutants for SPP81/DED1, as well as for other essential DBP genes with unknown functions. [2] To characterize the splicing phenotypes of these conditional mutants. [3] To identify interacting components of the RNA helicases by isolating extragenic suppressors for conditional mutants. [4] To identify the cellular substrates for PRP28 and SPP81/DED1. [5] To study the biochemical properties of PRP28.

RNA解旋酶是酶，可催化RNA中的链分离双链体或单链RNA中二级结构的去除。这反应需要将核苷三磷酸水解为能量来源。越来越多的推定RNA解旋酶基因参与A 多种生理功能，例如翻译启动，核糖体组装，mRNA剪接和生发线细胞分化已经在大肠杆菌到人类的生物中发现。其中许多推定的RNA解旋酶共享一系列序列元素，大多数明显的死盒主题。在酿酒酵母的酵母菌中至少16个假定的RNA解旋酶基因属于此死盒蛋白质（DBP）基因家族。在大肠杆菌，果蝇和HeLa细胞。因此，这个基因家族似乎进化很早，并且在进化中是高度保守的。这假定的RNA解旋酶基因的意外多样性也多样性就像其他生物一样，立即提出了许多有趣的问题关于它们的细胞功能和生化特性。关键特别是问题是如何和哪些RNA在功能上调节通过这些酶。该建议将通过关注来解决这些问题研究两个酵母RNA的生物化学和细胞功能解旋酶，PRP28和SPP81/DED1，与核前MRNA有关剪接。其他功能未知的酵母DBP基因也将是调查。预计这些研究将提供新颖有关RNA解旋酶的信息，其中知之甚少片刻。具体目的是：[1]为SPP81/DED1构造条件突变体以及其他具有未知功能的必需DBP基因。 [2]到表征这些条件突变体的剪接表型。 [3]到通过隔离来识别RNA解旋酶的相互作用成分有条件突变体的基因抑制剂。 [4]确定 PRP28和SPP81/DED1的细胞底物。 [5]研究生化 PRP28的属性。