Mitochondrial Parts, Pathways, and Pathogenesis ABSTRACT Mitochondria are ancient organelles central to eukaryotic life. Defects in these organelles underlie a spectrum of human diseases, ranging from rare inborn errors of metabolism to common age- associated degenerative diseases. The long-term goal of our laboratory is to achieve a systems- level understanding of this organelle that can serve as a foundation for studying mitochondria in health and in disease. NIGMS has provided the foundational funding for our laboratory which has enabled our ongoing efforts to systematically define the mitochondria's protein parts, organize them into functional pathways, and identify the proteins and pathways that underlie human disease. We have made tremendous progress, including producing MitoCarta, the most widely used catalog of mitochondrial protein parts. We have linked dozens of previously unstudied proteins to biological pathways and human diseases. However the protein inventory is not yet comprehensive and moreover contains hundreds of poorly studied proteins unlinked to any biological process in health or disease. In the next 5 years we aim to leverage emerging technologies in proteomics, Bayesian statistics, CRISPR, and high-resolution metabolomics, and human genetics to unveil the function of the dark corners of the mitochondrial proteome, while also pioneering new methods that can be generalized to other pathways within the cell.