Complex Involved In mRNA Decay in Yeast

参与酵母 mRNA 衰变的复合物

• 批准号: 6682684
• 负责人: STUART W PELTZ
• 金额: $ 30.39万
• 依托单位: UNIV OF MED/DENT NJ-R W JOHNSON MED SCH
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-05-01 至 2007-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6682684
• 关键词: SDS polyacrylamide gel electrophoresis; Saccharomyces cerevisiae; adenosine diphosphate; binding sites; fungal genetics; genetic transcription; immunoprecipitation; messenger RNA; microarray technology; nucleic acid metabolism; nucleic acid sequence; phosphorylation; posttranscriptional RNA processing; posttranslational modifications; protein protein interaction; stimulus /response; stress proteins; translation factor

DESCRIPTION (provided by applicant): The decay of mRNA is central to the post-transcriptional regulation of gene expression. The expression of many clinically relevant genes, including cytokines, proto-oncogenes and growth factors, is regulated at the level of mRNA turnover through AU-rich elements (AREs) in their 3' untranslated regions. Therefore, studies of this process provide valuable insight into the deregulation of cellular mechanisms in some cancers and immune disorders. This proposal aims to use the yeast, Saccharomyces cerevisiae, as a model system to study the phenomenon of ARE-mediated mRNA decay. The enzymes and pathways of mRNA turnover are well characterized in this organism. Regulation of AREmediated mRNA decay in yeast occurs in response to at least two cellular stimuli and involves three identified factors; Publp, Cthlp and Cth2p. The goals of this proposal are to determine how (i) the ARE-binding complex regulates mRNA decay rates (ii) the ARE-binding complex is modulated in response to cellular stimuli and (iii) the complex of factors that assembles on the ARE is specified its sequence context. The first part of the proposal concentrates on characterization of the three ARE-binding proteins and the interactions among them and between them and the ARE. One important aspect focuses on dissecting the interaction between the Cth proteins and Pablp. In addition, experiments in this Aim will determine how these factors are post-translationally modified in response to cellular conditions. Aim II of the proposal is to isolate ARE-binding complexes formed in vivo using both RNA tags and protein tags. Together the results of this part of the study will identify novel components of the ARE-binding complex and give us insight into how these complexes are altered in response to cellular conditions. The final Aim of the proposal is to identify novel AU-rich elements in yeast through microarray analysis of mRNA decay rates in strains lacking the known ARE-binding factors and by analysis of candidate ARE-containing transcripts discovered by searching the Transterm database. The goal is to amass a set of AU-rich elements large enough to facilitate computer analysis of the sequences to identify commonalities between them that might specify the proteins they interact with and their regulation.

描述（由申请人提供）：mRNA的降解是基因表达的转录后调节的核心。许多临床相关基因（包括细胞因子、原癌基因和生长因子）的表达在mRNA周转水平上通过其3'非翻译区中的富含AU的元件（战神）来调节。因此，对这一过程的研究为某些癌症和免疫疾病中细胞机制的失调提供了有价值的见解。本研究拟以酿酒酵母为模型系统，研究ARE介导的mRNA降解现象。mRNA周转的酶和途径在这种生物体中得到了很好的表征。在酵母中ARE介导的mRNA衰变的调节响应于至少两种细胞刺激而发生，并且涉及三种鉴定的因子; Publp、Cthlp和Cth 2 p。该提议的目标是确定（i）ARE结合复合物如何调节mRNA衰减速率（ii）ARE结合复合物响应于细胞刺激而被调节，以及（iii）在ARE上组装的因子的复合物被指定其序列背景。该提案的第一部分集中在三个ARE结合蛋白的特性和它们之间的相互作用，它们和ARE之间。一个重要的方面集中在解剖Cth蛋白和Pablp之间的相互作用。此外，本目标中的实验将确定这些因子如何响应于细胞条件而进行后修饰。目的II的建议是分离ARE结合复合物在体内形成的RNA标签和蛋白质标签。这部分研究的结果将共同确定ARE结合复合物的新组分，并使我们深入了解这些复合物如何响应细胞条件而改变。该提案的最终目的是通过微阵列分析缺乏已知ARE结合因子的菌株中的mRNA衰减率，并通过分析通过搜索Transterm数据库发现的候选含ARE转录本，来确定酵母中新的富含AU的元素。目标是积累一组足够大的富含AU的元素，以便于计算机分析序列，以确定它们之间的共性，这些共性可能指定它们与之相互作用的蛋白质及其调控。