GENETIC ANALYSIS OF TRNA SPLICING IN THE YEAST NUCLEUS

酵母细胞核中 TRNA 剪接的遗传分析

• 批准号: 2180250
• 负责人: MICHAEL R CULBERTSON
• 金额: $ 20.19万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 1988
• 资助国家: 美国
• 起止时间: 1988-09-01 至 1999-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2180250
• 关键词: RNA splicing; Saccharomyces cerevisiae; binding proteins; cell nucleus; confocal scanning microscopy; endonuclease; enzyme complex; enzyme structure; fungal genetics; gene expression; gene interaction; gene mutation; genetic manipulation; immunoaffinity chromatography; immunoelectron microscopy; immunofluorescence technique; laboratory rabbit; ligase; molecular cloning; nucleic acid sequence; polymerase chain reaction; posttranslational modifications; protein signal sequence; temperature sensitive mutant; transfer RNA

Intervening sequences are spliced from eukaryotic precursor-tRNAs through the action of three enzymes, a heterotrimeric endonuclease, a multi-functional but monomeric ligase, and an NAD-linked phosphatase. The endonuclease and ligase form a physical complex that is localized in the nuclear periphery in the vicinity of nuclear pores, suggesting the potential for functional coupling of splicing and nuclear export. Two genes, SEN1 and SEN2, were identified previously in the yeast Saccharomyces cerevisiae that are required for endonuclease activity. SEN1 codes for a nuclear-localized, positive effector of endonuclease activity that appears not to be a catalytic subunit of the enzyme. SEN2 codes for the 42 Kd catalytic subunit of the enzyme. We have isolated mutations in six new genes that affect SEN1 function. Mutations in two of the genes, SEN3 and SEN4, affect the expression of SEN1 at a post-translational level. One temperature-sensitive mutation, sen3-l, also causes excess accumulation of tRNA splicing intermediates in vivo. In addition, four genes have been identified that suppress the temperature-sensitive senl-l mutation when present in multiple copies and may encode products that interact with the SEN1 protein. By continuing with our analysis of SEN1 and SEN2 function and by initiating analyses of SEN3, SEN4, and the high-copy suppressor genes, we hope to provide critical information on the structure, organization, function, and intracellular location of the splicing complex. It is also possible that we may learn something about the import of splicing enzymes into the nucleus and the export of spliced tRNA products from the nucleus.

中间序列由真核前体trna剪接而成