STABILITY AND TRANSLATION OF CYTOCHROME B MRNA IN YEAST

拟议研究的长期目标是更好地了解宿主 细胞-细胞器相互作用。具体地说，这项拨款针对的是 核编码蛋白在细胞色素b，COB， 酿酒酵母中线粒体基因的表达。Cbp1p与 COB mRNA未翻译前导末端促进稳定性的研究 而cbs1p和cbs2p与近端部分相互作用 促进翻译的领导者。这项建议的具体目的 1)鉴定与Cbp1p和COB mRNA相互作用的蛋白质以 通过反向分析促进mRNA的稳定性，这是一种双杂交筛选法 和生化分析。2)确定序列元素和/或 在COB mRNA区域很重要的二级结构， 通过定点突变和物理分析与cbplp相互作用 核糖核酸的体内结构。3)确定的方向性 体内COB mRNA的衰变开始阐明周转的机制。 抗核外溶解衰变的RNA结构将是 以间隔插入到信使核糖核酸中，以调查该消息是否 主要由5‘至3’或3‘至5’核酸外切酶或5‘依赖的核酸外切酶降解 核酸内切酶。4)确定序列元素和/或次要元素 COB信使核糖核酸所需区域的重要结构 通过连接子扫描突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 有助于加深对真核生物细胞器功能的认识 细胞。线粒体功能缺陷被认为与肌肉有关 衰弱的疾病、衰老和致命的遗传疾病。因此， 重要的是，线粒体核控制的细节 其功能有待进一步阐明。

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.