Interconnections with and within the trypanosomal respiratory chain revealed by complexome profiling

Abstract

Proteins are frequently integrated into multicomponent complexes that execute the elaborate processes supporting life. Thus, a protein’s function can only be defined by the company it keeps within a complex. Proteomes provide informative protein inventories but lack information about protein quaternary structures. Complexome profiling (CP) has been transformative in capturing the comprehensive population structure of complexes at a given moment within a cell. We have employed CP to chart the multiprotein complex landscape of two life cycle stages of Trypanosoma brucei. These data have allowed the observation of previously hidden interactions with and within the mitochondrial respiratory chain. We have found (1) an exceptional case of a SLC25 solute transporter that interacts with NADH dehydrogenase, (2) two ATP synthase subunit g paralogs that are intriguingly excluded from the enzyme’s dimers, and (3) refined the known composition of ubiquinol:cytochrome c oxidoreductase by addition of missing subunits and removing an incorrectly assigned subunit, which more likely acts to insert the iron-sulfur co-factor into the complex. Further investigation into ubiquinol:cytochrome c oxidoreductase assembly revealed crosstalk between incorporation of its nuclear subunits with mitochondrial translation, possibly facilitating a hitherto unknown quality control mechanism. These discoveries demonstrate the power of our CP data for generation and testing of hypotheses about the mitochondrial and other organellar multiprotein complexes of T. brucei, a protist of medical and evolutionary importance. Statement for broader audience Mitochondria are powerhouses thanks to enclosing the respiratory chain (RC), a collection of multiprotein complexes. Their complete make-up and interactions beyond the RC remain mysterious. We have cataloged the proteins of RC and other mitochondrial complexes in unicellular Trypanosoma, which remodels its RC during its life cycle. We reveal and investigate interesting interconnections uncovered by our ‘complexome’ to expand the known interactions that the RC engages in to keep the power running in mitochondria.