Cytochrome c Oxidase Assembly in Health and Disease

描述(申请人提供)：细胞色素c氧化酶(COX)缺乏是人类线粒体脑肌病最常见的原因，也与神经退化和衰老有关。更好地了解COX的生物发生对于阐明这些疾病的分子基础是至关重要的。本研究的主要目的是以酿酒酵母和培养的人类细胞为研究模型，研究COX生物发生的相关因素和机制。我们的长期目标是全面了解导致COX组装的途径及其成分，作为开发治疗COX缺乏症相关疾病的治疗的先决条件。在我们的前一期拨款中，我们在理解如何调控COX生物发生方面取得了重大进展。真核细胞环氧合酶是由具有核和线粒体双重遗传来源的多肽组成的多聚酶，它的组装涉及大量核编码的辅助因子。COX表现出其组成亚基的协同积累。未组装的亚基具有产生活性氧物种的高风险，其积累受到翻译后降解的限制。我们揭示了在COX生物发生过程中亚基化学计量积累的另一个贡献。我们的数据支持存在一种调控机制，通过该机制，线粒体DNA编码的COX亚单位1(Cox1)的合成受到其在酿酒酵母中的组装伙伴的可用性的调节。因此，这一建议的中心假设是存在依赖于COX组装的对Cox1合成的调节，其独特的性质反过来提供了催化多亚单位组装的手段。在酿酒酵母中，调控系统涉及特定的COX1 mRNA翻译激活子、特定的Cox1伴侣和组装因子以及一般的伴侣，它们在复杂的相互作用中共同作用，我们刚刚开始描述这种相互作用。调节系统的中心元件是双功能的COX1mRNA翻译激活子和Cox1伴侣Mss51，它从酵母到人类都是保守的。提出了三个特定的目标来表征最近发现的新的Mss-51相互作用的成员(即Ssc1和Cox25)以及与酿酒酵母中的翻译调控系统及其从酵母到人类的保护有关的复杂性水平(即Mss 51是一个血红素结合蛋白)。目的#1-揭示Mss-51与血红素结合的机制和作用目的#2-研究Mss-51与线粒体伴侣蛋白相互作用调节Cox1合成的机制/S目标#3-确定酵母和人类Mss-51的功能等价性

DESCRIPTION (provided by applicant): Cytochrome c oxidase (COX) deficiency is the most frequent cause of mitochondrial encephalomyopathies in humans and has also been associated to neurodegeneration and aging. A better understanding of COX biogenesis is essential for elucidating the molecular basis underlying these groups of diseases. The main objective of the proposed research is to investigate the players and mechanisms involved in COX biogenesis using the yeast Saccharomyces cerevisiae and cultured human cells as research models. Our long-term goal is to attain a complete understanding of the pathways leading to COX assembly and their components as a prerequisite to the development of therapies for the management of disorders associated with COX deficiencies. In the previous term of our grant we made significant progress in the understanding of how COX biogenesis is regulated. Eukaryotic COX is a multimeric enzyme formed by polypeptides of dual genetic origin (nuclear and mitochondrial) which assembly involves a large number of nuclear-encoded auxiliary factors. COX display a concerted accumulation of its constitutive subunits. Unassembled subunits bear a high risk to produce reactive oxygen species and their accumulation is limited by posttranslational degradation. We have revealed another contribution to the stoichiometric accumulation of subunits during COX biogenesis. Our data support the existence of a regulatory mechanism by which the synthesis of mtDNA-encoded COX subunit 1 (Cox1) is regulated by the availability of its assembly partners in the yeast Saccharomyces cerevisiae. Thus, the central hypothesis of this proposal is the existence of a COX assembly dependent regulation of Cox1 synthesis, which unique properties, in turn, offer a means to catalyze multiple- subunit assembly. In S. cerevisiae, the regulatory system involves specific COX1 mRNA translational activators, specific Cox1 chaperones and assembly factors as well as general chaperones acting together in a sophisticated interplay that we are just beginning to characterize. The central element of the regulatory system is the bi-functional COX1 mRNA translational activator and Cox1 chaperone Mss51 which is conserved from yeast to humans. Three specific aims are proposed to characterize recently identified new Mss51-interacting players (i.e. Ssc1 and Cox25) and levels of complexity (i.e. Mss51 is a heme binding protein) concerning the translational regulatory system in S. cerevisiae and its conservation from yeast to humans. Aim # 1 - Disclose the mechanism and role of heme binding by Mss51 Aim #2 - Investigate the mechanism/s by which the interactions of Mss51 with mitochondrial chaperones regulate Cox1 synthesis Aim # 3 - Determine the functional equivalence of yeast and human Mss51