Mitochondria are centers of metabolism and signaling whose function is essential to all but a few eukaryotic cell types. General dysfunction of these organelles is implicated in a wide range of inborn errors of metabolism, and in an increasing number of common human diseases. Elucidation of the biochemical functions of these disease-related proteins has become a bottleneck in understanding mitochondrial pathophysiology. This project aims to widen this biomedical bottleneck by accelerating the functional annotation of key disease- related orphan mitochondrial proteins (OMPs), thereby taking an important first step toward rational therapeutic design for multiple diseases. We will do so by establishing a robust computation- and mass spectrometry-based platform for systematically annotating ~100 of these disease-related OMPs, and then to employ rigorous molecular and structural biology methods to define the functions of select proteins at biochemical depth. Our annotation pipeline involves: 1) establishing an extensive network of condition-specific protein-protein interactions using affinity purification mass spectrometry in mammalian cell lines, 2) connecting the yeast orthologs of these OMPs with proteins and pathways of known function through whole-proteome and metabolome correlation profiling, and 3) interpreting these data in the context of other large-scale resources using a Bayesian framework. We will first leverage these tools to further elucidate the functions of OMPs associated with the biosynthesis of coenzyme Q (CoQ) and the assembly of complex I (CI)-core components of the mitochondrial oxidative phosphorylation machinery whose deficiency is associated with multiple human diseases. Prior studies have identified nine proteins that are associated with CoQ biosynthesis (COQ1-9), and seven associated with CI maturation/assembly (NDUFAF1-7). However, many of these proteins are OMPs with no clear biochemical roles in these pathways, and at least three steps in the CoQ pathway are likely fulfilled by yet-to-be-identified mitochondrial proteins. Our initial approaches above already have shed light on the functions of two disease-related CoQ-related OMPs (COQ8 and COQ9) and on three other novel CoQ- and CI- related proteins identified by our systematic analyses. Here, we will employ biochemical and structural biology approaches to investigate the functions of these new proteins, and to test the hypothesis that COQ9 uses its lipid-binding capacity to present CoQ intermediates to other enzymes in the pathway. In future years, we will employ similar approaches to investigate the functions of additional OMPs related to the respiratory chain in our own laboratories; will continue working with our network of collaborators that have expertise in areas distinct to our own; and will actively provide our data to the larger mitochondrial community via an interactive website to further accelerate investigations of OMPs connected to a broad range of mitochondrial processes and human diseases. In doing so, our work promises to shed new insight into basic mitochondrial function and to widen a key bottleneck in understanding mitochondrial dysfunction and its associated pathophysiology.
Mitochondrial dysfunction is implicated in a wide range of rare and common human diseases, yet the biological functions of the mutated proteins underlying these diseases are predominantly unknown. This proposal aims to blend large-scale experimental and computational approaches with focused biochemical and structural biology techniques to systematically elucidate the functions of many such disease-related, orphan mitochondrial proteins. Completion of these goals will be important steps toward establishing a more complete understanding of basic mitochondrial biology and the molecular etiology of various mitochondrial diseases.
| Jha, Pooja; McDevitt, Molly T; Gupta, Rahul et al. (2018) Systems Analyses Reveal Physiological Roles and Genetic Regulators of Liver Lipid Species. Cell Syst 6:722-733.e6 |
| Lapointe, Christopher P; Stefely, Jonathan A; Jochem, Adam et al. (2018) Multi-omics Reveal Specific Targets of the RNA-Binding Protein Puf3p and Its Orchestration of Mitochondrial Biogenesis. Cell Syst 6:125-135.e6 |
| Schaffer, Leah V; Rensvold, Jarred W; Shortreed, Michael R et al. (2018) Identification and Quantification of Murine Mitochondrial Proteoforms Using an Integrated Top-Down and Intact-Mass Strategy. J Proteome Res 17:3526-3536 |
| Guo, Xiao; Niemi, Natalie M; Hutchins, Paul D et al. (2017) Ptc7p Dephosphorylates Select Mitochondrial Proteins to Enhance Metabolic Function. Cell Rep 18:307-313 |
| Veling, Mike T; Reidenbach, Andrew G; Freiberger, Elyse C et al. (2017) Multi-omic Mitoprotease Profiling Defines a Role for Oct1p in Coenzyme Q Production. Mol Cell 68:970-977.e11 |
| Stefely, Jonathan A; Pagliarini, David J (2017) Biochemistry of Mitochondrial Coenzyme Q Biosynthesis. Trends Biochem Sci 42:824-843 |
| Chung, Jacky; Wittig, Johannes G; Ghamari, Alireza et al. (2017) Erythropoietin signaling regulates heme biosynthesis. Elife 6: |
| Stefely, Jonathan A; Kwiecien, Nicholas W; Freiberger, Elyse C et al. (2016) Mitochondrial protein functions elucidated by multi-omic mass spectrometry profiling. Nat Biotechnol 34:1191-1197 |
| Floyd, Brendan J; Wilkerson, Emily M; Veling, Mike T et al. (2016) Mitochondrial Protein Interaction Mapping Identifies Regulators of Respiratory Chain Function. Mol Cell 63:621-632 |
| Rensvold, Jarred W; Krautkramer, Kimberly A; Dowell, James A et al. (2016) Iron Deprivation Induces Transcriptional Regulation of Mitochondrial Biogenesis. J Biol Chem 291:20827-20837 |
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