Aging Mitochondrial Interactome

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Changes in mitochondrial function play a central role in age-related pathologies, loss of resilience, and the decline in quality of life in older adults. As we age there is a shift in our mitochondria toward a reduced ability to generate ATP and increased oxidant production. These changes lead to disruption of redox and energy homeostasis, altered metabolite levels, and increased sensitivity to permeability transition, all of which contribute to tissue dysfunction. Mitochondria are dynamic organelles that continuously adapt to changing cellular demands by altering protein assembly and interactions to modify their function. Despite the obvious importance, little is known about how age-related changes in mitochondrial protein interactions (interactome) underlie changes in function with age. To address this fundamental question, we propose to apply a state of the art quantitative chemical cross-linking with mass spectrometry (qXL-MS) strategy to quantify changes in the mitochondrial interactome with age. By combining this innovative qXL-MS approach with detailed assays of mitochondrial metabolism and interventions we have developed over the last several years to manipulate mitochondria in vivo and in vitro, we are uniquely positioned to identify the molecular level changes in mitochondrial interactome that underlie age-related mitochondrial dysfunction. Our preliminary data indicate disruption of multiple protein interaction networks involved in ADP transport, ATP synthesis, and substrate supply to the electron transport system in aged heart and skeletal muscle. These changes are associated with previously demonstrated decreases in ATP production and lower sensitivity to ADP. Furthermore, we have shown that a mitochondrial targeted intervention (SS-31) that reverses mitochondrial dysfunction in heart and skeletal muscle, specifically interacts with many of the same protein complexes that our interactome studies reveal are disrupted in aging, including the ANT and complexes IV and V of the electron transport system. Our overall hypothesis tested in this proposal is that changes in the mitochondrial interactome with age underlie decreased ATP production and increased oxidant production in mitochondria from aged heart and skeletal muscle. Aim 1 applies XL-MS and protein and site-specific mitochondrial assays to quantify changes in the mitochondrial interactome with age and those induced by pro-oxidant treatment in vivo and in vitro in mouse heart and skeletal muscle. Aim 2 quantifies the effect on the mitochondrial interactome of two well established mitochondrial targeted interventions, SS-31 and mitochondrial targeted catalase, to identify the most important age-related changes in the protein interaction networks. Aim 3 tests whether changes in the mitochondrial interactome identified in aims 1 and 2 translate into aged human skeletal muscle. The mitochondrial interactome, function, and effects of SS-31 from older adults separated into low and high performing groups are compared with young adults. Results from these experiments will have a significant impact and have the potential to transform the field by providing the first test of how disruption of mitochondrial protein interactions contribute to age-related mitochondrial dysfunction. 1