GNPS - MITOMICS: defining mitochondrial protein functions through deep multi-omic profiling

Mitochondria are epicenters of eukaryotic metabolism and bioenergetics. Pioneering efforts in recent decades have established the core protein componentry of these organelles and have linked their dysfunction to over 150 distinct disorders. However, hundreds of mitochondrial proteins still lack clear functions, and ~40% of mitochondrial disorders remain unresolved. To establish a more complete functional compendium of human mitochondrial proteins, we profiled 203 CRISPR-mediated HAP1 cell knockout lines using mass spectrometry-based multi-omics analyses. This effort generated ~8.2 million distinct biomolecule measurements, providing a deep survey of the cellular responses to mitochondrial perturbations, and laying a foundation for mechanistic investigations into protein function. Guided by these data, we discovered that PYURF is a SAM-dependent methyltransferase chaperone that supports both complex I assembly and coenzyme Q biosynthesis, and that is disrupted in a previously unresolved multisystemic mitochondrial disorder. We further linked the putative zinc transporter SLC30A9 to mitoribosome and OxPhos integrity and established RAB5IF as the second gene harboring pathogenic variants causing cerebrofaciothoracic dysplasia. Our data - which can be explored through an interactive online resource - suggest biological roles for many other orphan mitochondrial proteins still lacking robust functional characterization, and define a rich cell signature of mitochondrial dysfunction that can support the genetic diagnosis of mitochondrial diseases.