UCLA

The cardiac mitochondrial proteome contains ~1,500 distinct proteins that carry out necessary metabolic and energetic processes in the heart. To sustain cardiac function, the mitochondrial proteome must be maintained in constant renewal, or turnover, especially under stress conditions. Disruptions of protein turnover can lead to protein damage and proteotoxicity, a hallmark of many heart disease etiologies. Current quantitative proteomics experiments largely focus on the measurement of the steady-state abundance, or changes therein, of proteins that are present in a system, and give little insights into the underlying regulations of protein synthesis, degradation, and homeostasis. Protein turnover rates provide this missing temporal dimension of information, and can inform on the potential mechanism through which protein abundance may permute during the development of disease (e.g., via increased synthesis or decreased degradation). Currently, such investigations are hampered by the fact that the technology to measure protein turnover in animals on a large scale has not been well developed. This dissertation outlines a new method to measure protein turnover half-life in the cardiac mitochondrion. Basic features of the regulation of protein turnover in the mitochondrion are discussed, and how protein dynamics permutes in early-stage heart failure after hypertrophic stimuli is described. In total, we measured the turnover rates of 2,986 proteins in the mouse heart under basal conditions, isoproterenol stimulus, and post-stimulus recovery, including 1,078 proteins from isolated mitochondria. The data revealed widespread, bidirectional changes in protein turnover in 35 functional categories, and further identified a number of novel candidate disease proteins with significantly up-regulated turnover rates in disease, including HK1, ALDH1B1, and PHB, which have been obscured from previous investigations due to their inconspicuous changes in steady-state abundance. Combinatorial analysis of protein expression and protein turnover data indicates that the remodeling heart is characterized by decreased turnover but increased expression of a cohort of mitochondrial proteins including FXN, LETM1, and CYC1, suggesting a potential class of candidate disease proteins whose impaired degradation is associated with remodeling. I further discuss the implications of the data to the cardiac remodeling process at large and how such investigations may be translated to human studies in the future. Taken together, the results suggest that comparisons of protein turnover rates can be a powerful new tool to understand the temporal dynamics of disease progression in the heart.