STABILITY AND TRANSLATION OF CYTOCHROME B MRNA IN YEAST

这项研究的长期目标是更好地了解宿主 细胞-细胞器相互作用 具体来说，这笔赠款涉及以下角色 细胞色素B，CO B， 酿酒酵母线粒体mRNA。cbp1p与 COB mRNA的非翻译前导序列的远端部分，以促进稳定性 而Cbs1p和Cbs2p与近端部分交互 的领导人，以促进翻译。本提案的具体目标 1）鉴定与Cbp1p和COB mRNA相互作用的蛋白质， 通过逆转录分析、双杂交筛选、 和生化分析。 2)为了确定序列元素和/或 在COB mRNA区域中重要的二级结构， 通过定点诱变和物理分析与CBPLP相互作用 RNA的体内结构。3)确定…的方向 COB mRNA在体内的衰减到开始，以阐明周转的机制。 一种能抵抗核酸外切衰变的RNA结构将被 在mRNA中间隔插入，以调查信息是否 主要由5 '至3'或3 '至5'外切核酸酶或5 '依赖性核酸酶降解。 核酸内切酶。4)为了确定序列元素和/或次级 在COB mRNA区域中重要的结构， 通过接头扫描诱变和物理分析进行翻译， RNA的体内结构。这些研究将扩大 对RNA-蛋白质相互作用的具体和一般的理解将 有助于进一步了解细胞器的功能，在真核生物 细胞 线粒体功能的缺陷与肌肉 消耗性疾病、衰老和致命的遗传性疾病。因此，在本发明中， 重要的是，核控制线粒体的细节 功能被阐明。

The long-term goal of the proposed research is to better understand host cell-organelle interactions. Specifically this grant addresses the role of nuclear-coded proteins in the expression of the cytochrome b, COB, mitochondrial mRNA in Saccharomyces cerevisiae. cbp1p interacts with the distal portion of the untranslated leader of COB mRNA to promote stability of the message, while cbs1p and Cbs2p interact with the proximal portion of the leader to promote translation. The Specific Aims of this proposal are: 1) To identify proteins that interact with Cbp1p and COB mRNA to promote stability of the mRNA via reversion analyses, a dihybrid screen and biochemical analyses. 2) To determine the sequence element and/or secondary structures that are important in the region of COB mRNA that interacts with cbplp via site-directed mutagenesis and a physical analysis of the in vivo structure of the RNA. 3) To determine the directionality of decay of COB mRNA in vivo to begin to elucidate the mechanism of turnover. An RNA structure that is resistant to exonucleolytic decay will be inserted at intervals in the mRNA to investigate whether the message is primarily degraded by 5' to 3' or 3' to 5' exonucleases or 5'-dependent endonucleases. 4) To determine the sequence element and/or secondary structures that are important in the regions of COB mRNA that are required for translation via linker-scanning mutagenesis and physical analysis of the in vivo structure of the RNA. These studies will expand the understanding of RNA-protein interactions specifically and in general will contribute to furthering the knowledge of organelle function in eukaryotic cells. Defects in mitochondrial function have been implicated in muscle wasting diseases, aging and lethal genetically inherited disorders. Thus, it is important that the details of nuclear control of mitochondrial function be elucidated.