CONTROL OF ARG-2 GENE EXPRESSION IN NEUROSPORA

神经孢子虫中 ARG-2 基因表达的控制

• 批准号: 2701566
• 负责人: MATTHEW Steven SACHS
• 金额: $ 24.3万
• 依托单位: OREGON GRADUATE INSTITUTE SCIENCE & TECH
• 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 2001-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2701566
• 关键词: Neurospora; arginine; cell free system; fungal genetics; gene expression; gene mutation; genetic mapping; genetic translation; messenger RNA; molecular cloning; nucleic acid sequence; open reading frames; site directed mutagenesis; synthetic peptide; transfer RNA; translation factor

DESCRIPTION: The long term goal of the research is to understand the molecular mechanisms by which different regulatory pathways interact to control expression of the Neurospora crassa arg-2 gene. The arg-2 gene encodes the mitochondrially localized small subunit of carbamoyl phosphate synthetase. The major goal of the application is to understand the mechanism of arginine-specific, negative translational control mediated by an upstream open reading frame (uORF) in arg-2 mRNA. The arg-2 uORF is essential for translational regulation in response to arginine availability. The primary amino acid sequence of the predicted uORF peptide, which is conserved in other uORFs, is critical for control. Arginine specific translational control through the arg-2 uORF will be assessed in vivo by analyzing levels of RNA and polypeptide synthesis and by examining the distribution of ribosomes on mRNA. Arginine specific translational control will be assessed in vitro using N. crassa cell free translation extracts programmed with synthetic RNAs. There are three specific aims. First, the trans-acting genes that are important for translational control will be identified. Mutants that show effects on arginine-specific regulation will be isolated and characterized. The genes will be placed into complementation groups, mapped, and isolated. The roles of the gene products in arginine-specific translational control will be determined. Second, site-specific mutagenesis will be used to determine the sequence requirements for translational control. The features of both the uORF sequence and the RNA important for translational control will be determined. Third, the mechanism of arginine-specific translational control will be determined. The ability of a synthetic uORF peptide to affect translational processes in trans will be analyzed. The form of arginine that elicits control-free arginine, an arginine metabolite, or charged tRNA-will be determined.

描述：研究的长期目标是了解