J Biol Chem. 2019 Oct 07. pii: jbc.RA119.010317. [Epub ahead of print]
Hypoxia-inducible gene domain 1 (HIGD1) proteins are small integral membrane proteins, conserved from bacteria to humans, that associate with oxidative phosphorylation supercomplexes. Using yeast as a model organism, we have previously shown that its two HIGD1 proteins, Rcf1 and Rcf2, are required for the generation and maintenance of a normal membrane potential (Δψ) across the inner mitochondrial membrane (IMM). We have postulated that the lower Δψ observed in the absence of the HIGD1 proteins may be due to decreased proton pumping by complex IV (CIV) or to an enhanced leakage of protons across the IMM. Here, we measured the Δψ generated by complex III (CIII) to discriminate between these possibilities. First, we found that the decreased Δψ observed in the absence of the HIGD1 proteins cannot be due to decreased proton pumping by CIV, since CIII, operating alone, also exhibited a decreased Δψ when HIGD1 proteins were absent. Since CIII can neither lower its pumping stoichiometry nor transfer protons completely across the IMM, this result indicates that HIGD1 protein ablation enhances proton leakage across the IMM. Second, we demonstrate that this proton leakage occurs through CIV, since Δψ generation by CIII is restored when CIV is removed from the cell. Third, the proton leakage appeared to take place through an inactive population of CIV that accumulates when HIGD1 proteins are absent. We conclude that HIGD1 proteins in yeast prevent CIV inactivation, likely by preventing the loss of lipids bound within the Cox3 protein of CIV.
Keywords: Rcf; cytochrome c oxidase (Complex IV); hypoxia-inducible gene domain 1 (HIGD1); lipid-protein interaction; mitochondria; mitochondrial membrane potential; oxidative phosphorylation; proton leak; respiration; suicide inactivation