CYTOCHROME C OXIDASE IN HEALTH AND DISEASE

描述(由申请人提供)：细胞色素c氧化酶(COX)缺乏是人类线粒体神经肌病最常见的原因。患有这些疾病的患者表现出不同的临床表型，包括Leigh综合征、肌肉无力和脑肌病。对COX生物发生的完整了解对于阐明这类疾病的分子基础是必不可少的。这项研究的主要目的是以酿酒酵母为模型，研究野生型细胞和进化保守组装因子突变的细胞中的COX组装。将实现几个具体目标。1)我们最近报道了人类SurMp的酵母同源物Shylp催化COX装配中间体的形成，Coxlp是COX的一个线粒体编码的催化亚单位。Shylp是大多数Leigh综合征的病因。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COX代谢因子的过程中调节Cox1 p的表达，如Mss 51p和Cox14p。这些蛋白调节COX表达的机制将被研究。适当标记的Mss51p和Cox14p将从高表达的酵母细胞中纯化。纯化蛋白质的可获得性将使有关其活性的假说能够被直接测试。3)参与COX组装调节Coxlp合成的蛋白质可能会在它们之间进行短暂或永久性的相互作用，以实现其功能。这些相互作用的性质将被描述。总而言之，酵母系统将被探索为一种破译复杂膜酶组装的一般原理的手段，该复合膜酶由来自两个空间分离的遗传源的亚单位多肽组成。酵母范例将被生物化学和遗传学手段所利用，以获得对Shylp和Surflp功能的全面了解，并阐明人类COX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.