CYTOCHROME C OXIDASE IN HEALTH AND DISEASE

描述（由申请方提供）：细胞色素c氧化酶（考克斯）缺乏是人类线粒体神经肌病的最常见原因。患有这些疾病的患者呈现异质性临床表型，包括Leigh综合征、肌无力和脑肌病。对考克斯生物发生的完整理解对于阐明这组疾病的分子基础是必不可少的。拟议的研究的主要目的是使用酵母酿酒酵母作为一个模型，调查考克斯组装在野生型细胞和进化保守的组装因子突变的细胞。将努力实现若干具体目标。1)我们最近报道，Shylp，人SurMp的酵母同源物，负责大多数情况下的利氏综合征，催化形成一个考克斯组装中间体涉及Coxlp，一个神经编码的催化亚基的考克斯。Shylp在Coxlp表达中的作用将被研究。2)最近的证据表明，该中间体在涉及其他考克斯代谢因子如Mss 51 p和Cox14 p的过程中调节Cox1 p的表达。这些蛋白调节考克斯表达的机制将被研究。适当标记的Mss51p和Cox14p将从过表达的酵母细胞中纯化。纯化蛋白质的可用性将允许直接测试关于其活性的假设。3)参与通过考克斯组装调节Coxlp合成的蛋白质可能在它们之间短暂或永久地相互作用以执行它们的功能。这些相互作用的性质将被描述。总之，酵母系统将被探索作为一种手段，破译的一般原则，在组装一个复杂的膜酶组成的亚基多肽来源于两个空间上分离的遗传来源。酵母范例将利用生物化学和遗传学手段，以获得一个完整的理解的功能Shylp，因此Surflp以及，并澄清人类考克斯缺陷的分子基础。

DESCRIPTION (provided by applicant): Cytochrome c oxidase (COX) deficiency is the most frequent cause of mitochondrial neuromyopathies in humans. Patients afflicted with these diseases present heterogeneous clinical phenotypes, including Leigh syndrome, muscle weakness and encephalomyopathy. A complete understanding of COX biogenesis is essential for elucidating the molecular basis underlying this group of diseases. The main objective of the proposed research is to use the yeast Saccharomyces cerevisiae as a model to investigate COX assembly in wild type cells and in cells with mutations in evolutionary conserved assembly factors. Several specific aims will be pursued. 1) We have recently reported that Shylp, the yeast homologue of human SurMp, responsible for most cases of Leigh's syndrome, catalyzes the formation of a COX assembly intermediate involving Coxlp, a mitochondrially encoded catalytic subunit of COX. The role of Shylp in expression of Coxlp will be studied. 2) More recent evidence indicates that this intermediate regulates Cox1 p expression in a process involving other COX metabolism factors, such as Mss51p and Cox14p. The mechanisms by which these proteins regulate COX expression will be studied. Appropriately tagged Mss51p and Cox14p will be purified from over-expressing yeast cells. The availability of purified proteins will permit hypotheses concerning their activities to be tested directly. 3) The proteins involved in regulation of Coxlp synthesis by COX assembly are likely to interact transiently or permanently among them to perform their functions. The nature of these interactions will be characterized. In summary, the yeast system will be explored as a means of deciphering the general principles operating in the assembly of a complex membrane enzyme composed of subunit polypeptides derived from two spatially separated genetic sources. The yeast paradigm will be exploited by biochemical and genetic means to gain a complete understanding of the function of Shylp and therefore of Surflp as well, and to clarify the molecular basis of human COX deficiencies.