An ability to comprehensively track the assembly intermediates (AIs) of complex I (CI) biogenesis in Drosophila will enable the characterization of the precise mechanism(s) by which various CI regulators modulate CI assembly. Accordingly, we generated 21 novel antibodies to various mitochondrial proteins and used this resource to characterize the mechanism by which apoptosis-inducing factor (AIF) regulates CI biogenesis by tracking the AI profile observed when AIF expression is impaired. We find that when the AIF-Mia40 translocation complex is disrupted, the part of CI that transfers electrons to ubiquinone is synthesized but fails to progress in the CI biosynthetic pathway. This is associated with a reduction in intramitochondrial accumulation of the Mia40 substrate, MIC19. Importantly, knockdown of either MIC19 or MIC60, components of the mitochondrial contact site and cristae organizing system (MICOS), fully recapitulates the AI profile observed when AIF is inhibited. Thus, AIF's effect on CI assembly is principally due to compromised intramitochondrial transport of the MICOS complex.
© 2020 Murari et al.