Mitochondrial complexome reveals quality-control pathways of protein import

Abstract

Mitochondria have crucial roles in cellular energetics, metabolism, signalling and quality control1-4. They contain around 1,000 different proteins that often assemble into complexes and supercomplexes such as respiratory complexes and preprotein translocases1,3-7. The composition ofthe mitochondrial proteome has been characterized1,3,5,6; however, the organization of mitochondrial proteins into stable and dynamic assemblies is poorly understood for major parts ofthe proteome1,4,7. Here we report quantitative mapping of mitochondrial protein assemblies using high-resolution complexome profiling of more than 90% ofthe yeast mitochondrial proteome, termed MitCOM. An analysis ofthe MitCOM dataset resolves >5,200 protein peaks with an average of six peaks per protein and demonstrates a notable complexity of mitochondrial protein assemblies with distinct appearance for respiration, metabolism, biogenesis, dynamics, regulation and redox processes. We detect interactors ofthe mitochondrial receptor for cytosolic ribosomes, of prohibitin scaffolds and ofrespiratory complexes. The identification of quality-control factors operating at the mitochondrial protein entry gate reveals pathways for preprotein ubiquitylation, deubiquitylation and degradation. Interactions between the peptidyl-tRNA hydrolase Pth2 and the entry gate led to the elucidation of a constitutive pathway for the removal of preproteins. The MitCOM dataset-which is accessible through an interactive profile viewer-is a comprehensive resource for the identification, organization and interaction of mitochondrial machineries and pathways.