Harvard Catalyst Profiles

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Functional Genomics of the Mitochondrial Proteome


Mitochondria are essential organelles for cellular metabolism and survival. As a result, mitochondrial dysfunction plays an important role in the pathophysiology of many metabolic disorders such as diabetes and obesity. The growing interest in mitochondria has stimulated large-scale proteomics efforts to define the mitochondrial proteome. Mammalian mitochondria harbor ~ 1000 proteins, but of these proteins, the function of ~ 300 remain largely unknown. The collaborative and interdisciplinary program in Metabolism at the University of Utah has established a platform that will enable efficient functional annotation of the mitochondrial proteome. Using bioinformatics tools, we have identified a subset of mitochondrial proteins of unknown function that are conserved from yeast to mammals. We hypothesize that evolutionary conservation provides powerful evidence for a critical roles for these poorly annotated proteins. We therefore began by screening yeast mutants that lack each of these proteins, for growth defects under conditions in which the yeast are dependent on aerobic metabolism, and evaluating further those mutations whose absence impaired mitochondrial respiratory function by determining mitochondrial localization and protein binding partners. This approach has already functionally characterized novel mitochondrial proteins and validated their roles in model organisms and in human disease. These include the succinate dehydrogenase assembly factor (Sdh5) which we proved to be a disease gene for familial paraganglioma, Vms1 a novel stress-responsive system for mitochondrial protein quality control and Coa4 which are novel assembly factors for the biogenesis of cytochrome c reductase and cytochrome oxidase, respectively. We now propose to expand our screen to functionally annotate the mitochondrial proteome and to study in depth a subset of mitochondrial proteins that are involved in electron transport chain assembly.

Aim 1 will expand our preliminary screening program by determining the impact of loss of function mutations in all other candidate proteins on growth, oxygen consumption, OXPHOS supercomplex formation, the metabolome, mitochondrial morphology and transition metal content. We will also determine the subcellular localization of each protein and identify potential binding partners by tandem affinity purification tagging. These data will be disseminated to the public via the SGD website. Aim 2 will focus on five recently identified outputs of our screen: Coa4, Sdhaf1, Acn9, Hig1, and Cfa1 (Ycl057) that are essential for electron transport chain assembly. Studies will define binding partners, identify genetic modifiers that suppress or exacerbate phenotypes in yeast, and use metabolomic profiling to identify enzymatic pathways that are regulated by these genes. Aim 3 will define the molecular mechanisms by which these OXPHOS assembly proteins regulate mitochondrial function in mammalian cells. Aim 4 will define the physiological role of these OXPHOS assembly proteins in the context of a multicellular model organism Drosophila. Aim 5 will generate mouse models of mitochondrial dysfunction, harboring tissue-restricted mutations these OXPHOS assembly proteins that will be utilized to define the in vivo physiology of these pathways. These mouse models will also be used to determine if oxidative stress and mitochondrial dysfunction will increase the risk for insulin resistance and lipotoxicity. These studies will significantly advance the functional annotation of the mitochondrial proteome and provide novel insights into the consequences of altered electron transport chain assembly on mitochondrial function and metabolism in vivo.

PUBLIC HEALTH RELEVANCE: Mitochondria play critical roles in cellular energy metabolism and in cell survival. A large number of diseases ranging from obesity, diabetes, heart failure and cancer are associated with mitochondrial dysfunction. The function of up to 30% of the mitochondrial proteome is unknown. This proposal will elucidate the function of these un-annotated mitochondrial proteins, providing new insights into the role of mitochondria in human disease.


Funded by the NIH National Center for Advancing Translational Sciences through its Clinical and Translational Science Awards Program, grant number UL1TR002541.