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可能会影响细胞的能量生产，并产生促进退行性疾病、衰老和癌症的活性氧物种。因此，更清楚地了解这一系统是如何被调控的，对于更好地理解线粒体疾病是必要的。为了进一步探索这一过程，第一个目的是阐明新发现的酵母翻译激活剂Mam33激活细胞色素C氧化酶复合体核心亚单位COX1翻译的机制。这些实验将决定1)COX1 mRNA的区域足以依赖于Mam33的激活，2)Mam33是否直接与COX1转录本结合，3)Mam33是否与线粒体核糖体结合，以及4)Mam33是否是限速的。这些信息将促进我们对Mam33和线粒体激活剂的总体理解，这些激活剂可以在以后用人类同源物进行测试。第二个目标将使用生化和遗传策略来识别和表征与Mam33物理上相互作用的因素-或者在功能上与Mam33相关的因素。这些结果将使我们能够将Mam33定位在COX1表达途径中，并更好地确定其翻译激活机制。此外，出乎意料的Mam33活动可能会被披露。第三个目标是了解Mam33活动的结构要求。将进行基于晶体结构特征和在其他生物同源物中发现的氨基酸保守性的突变分析。由于Cox1和Mam33在进化上都是保守的，在酵母中获得的信息很可能适用于人类线粒体疾病。此外，最近的数据还表明，人类同源基因p32与癌症有关。因此，更好地了解酵母Mam33也可以增强我们对癌症的理解。

 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.