Translational control of mitochondrial gene expression

 描述（由申请人提供）：拟议研究的目标是确定线粒体内翻译激活的机制。线粒体基因表达的改变会损害细胞能量的产生，并产生活性氧，从而促进退行性疾病、衰老和癌症。因此，更清楚地了解这个系统是如何调节的，对于更好地了解线粒体疾病是必要的。为了进一步探索这一过程，第一个目标将阐明新发现的酵母翻译激活因子Mam 33激活细胞色素c氧化酶复合物的核心亚基COX 1翻译的机制。这些实验将确定1）足以进行Mam 33依赖性激活的COX1 mRNA区域，2）Mam 33是否直接结合COX1转录物，3）Mam 33是否与线粒体核糖体结合，以及4）Mam 33是否是限速的。这些信息将促进我们对Mam33和线粒体激活剂的理解，这些信息可以在以后用人类同源物进行测试。第二个目标将采用生物化学和遗传学策略来识别和表征与Mam33物理相互作用或功能相关的因子。这些结果将使我们能够将Mam33置于COX1表达途径中，并更好地定义其翻译激活机制。此外，Mam33的活动可能会被发现。第三个目的是了解Mam33活性的结构要求。将进行基于晶体结构特征和在来自其他生物体的同源物中发现的氨基酸保守性的突变分析。由于Cox1和Mam33在进化上是保守的，因此在酵母中获得的信息可能适用于人类线粒体疾病。此外，最近的数据也暗示了癌症中的人类同源物p32。因此，更好地了解酵母Mam33也可以提高我们对癌症的理解。

 DESCRIPTION (provided by applicant): The goal of the proposed research is to define the mechanisms of translational activation within mitochondria. Alterations in mitochondrial gene expression can compromise cellular energy production and generate reactive oxygen species that promote degenerative disease, aging, and cancer. Thus, a clearer understanding of how this system is regulated is necessary to better understand mitochondrial disease. To explore this process further, the first aim will elucidate the mechanism by which the newly discovered yeast translational activator Mam33 activates the translation of COX1, a core subunit of the cytochrome c oxidase complex. These experiments will determine 1) the region of COX1 mRNA sufficient for Mam33-dependent activation, 2) whether Mam33 directly binds the COX1 transcript, 3) if Mam33 associates with the mitochondrial ribosome, and 4) if Mam33 is rate-limiting. This information will advance our understanding of Mam33 and mitochondrial activators in general, which can be later tested with human homologues. The second aim will employ biochemical and genetic strategies to identify and characterize factors that physically interact with - or are functionally related to Mam33. These results will allow us to place Mam33 within the COX1 expression pathway and better define its translation activation mechanism. Additionally, unanticipated Mam33 activities may be revealed. The third aim is to understand the structural requirements for Mam33 activity. A mutational analysis based upon crystal structure features and amino acid conservation found in homologs from other organisms will be performed. Since both Cox1 and Mam33 are evolutionarily conserved, information gained in yeast will likely be applicable to human mitochondrial disorders. Furthermore, recent data also implicates the human homologue p32 in cancer. Thus, a better understanding of yeast Mam33 could also enhance our understanding of cancer.