Abstract
Mitochondria are essential organelles whose functions depend on coordinated multiprotein complexes, yet their composition and organization remain incomplete. Here, we present a large-scale map of mitochondrial protein complexes by integrating affinity purification of 740 endogenously GFP-tagged mitochondrial proteins with biochemical co-fractionation of mitochondrial extracts from yeast (Saccharomyces cerevisiae) grown under respiratory conditions. Mass spectrometry identifies 13,716 high-confidence protein associations and defines 556 heteromeric complexes, many previously unknown. These assemblies reveal factors involved in coenzyme Q6 biosynthesis, membrane contact sites, phospholipid transport, and coordination with the MICOS complex during respiration. We further link 538 assemblies to 294 candidate human disease genes and construct a conservation map of 852,146 predicted mitochondrial interactions across 271 genomes, and validate key predictions in human cell lines and mouse brain tissue. Together, this work provides a comprehensive mitochondrial interactome, assigning functions to poorly characterized proteins, and offering insights into mitochondrial biology and disease-associated assemblies.
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Acknowledgements
We are grateful to Dr. Jodi Nunnari’s laboratory at the University of California Davis, USA, for providing GFP-tagged yeast library strains and for guidance on the affinity purification procedure. We also thank members from the Schuldiner and Babu groups for their assistance. M.T.M. was supported by a Canadian Institutes of Health Research (CIHR) postdoctoral fellowship. I.G.C. is the recipient of an EMBO long-term fellowship (ALTF-580-2017). M.B. acknowledges funding from the Chancellors Research Chair in Network Biology. M.S. holds the Dr. Gilbert Omenn and Martha Darling Professorial Chair in Molecular Genetics. This work was funded by the National Science Foundation Grant (MCB-2343997) to C.F.C., Natural Sciences and Engineering Research Council of Canada (DG-123456) to A.G., the Ontario Genomics Institute to I.S., the University of Puerto Rico Medical Sciences Campus to J.R.R, the Deutsche Forschungsgemeinschaft (DFG, DFGRA 1028/11-1) to M.S., as well as the CIHR (MOP-125952; RSN-124512, 132191; FDN-154318) and the Canada Foundation for Innovation to M.B.
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Jessulat, M., Phanse, S., Aoki, H. et al. Global mitochondrial connectivity map reveals the landscape of yeast functional assemblies and conserved protein communities. Nat Commun (2026). https://doi.org/10.1038/s41467-026-72525-2
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DOI: https://doi.org/10.1038/s41467-026-72525-2
