HCOA3 Stabilizes Cytochrome c Oxidase 1 (COX1) and Promotes Cytochrome c Oxidase Assembly in Human Mitochondria

Paula Clemente,Susana Peralta,Alberto Cruz-Bermudez,Lucía Echevarría,Flavia Fontanesi,Antoni Barrientos,Miguel A Fernandez-Moreno,Rafael Garesse

doi:10.1074/jbc.m112.422220

Abstract

Cytochrome c oxidase (COX) or complex IV of the mitochondrial respiratory chain plays a fundamental role in energy production of aerobic cells. In humans, COX deficiency is the most frequent cause of mitochondrial encephalomyopathies. Human COX is composed of 13 subunits of dual genetic origin, whose assembly requires an increasing number of nuclear-encoded accessory proteins known as assembly factors. Here, we have identified and characterized human CCDC56, an 11.7-kDa mitochondrial transmembrane protein, as a new factor essential for COX biogenesis. CCDC56 shares sequence similarity with the yeast COX assembly factor Coa3 and was termed hCOA3. hCOA3-silenced cells display a severe COX functional alteration owing to a decreased stability of newly synthesized COX1 and an impairment in the holoenzyme assembly process. We show that hCOA3 physically interacts with both the mitochondrial translation machinery and COX structural subunits. We conclude that hCOA3 stabilizes COX1 co-translationally and promotes its assembly with COX partner subunits. Finally, our results identify hCOA3 as a new candidate when screening for genes responsible for mitochondrial diseases associated with COX deficiency.

Highlights

Of cytochrome c oxidase (COX), complex IV of the respiratory chain, requires a great number of accessory proteins known as assembly factors
Our results identify hCOA3 as a new candidate when screening for genes responsible for mitochondrial diseases associated with COX deficiency
To clarify whether hCOA3 plays a role on the stability or biogenesis of one or several of the mitochondrial DNA (mtDNA)-encoded COX subunits, we labeled mitochondrial translation products for 30 min with [35S]methionine and performed immunoprecipitation experiments with anti-FLAG M2 antibodies in HeLa cells either overexpressing hCOA3-FLAG or containing an empty pIRESpuro2 vector

Summary

Background

Of cytochrome c oxidase (COX), complex IV of the respiratory chain, requires a great number of accessory proteins known as assembly factors. More than 30 COX assembly factors have been identified in model organisms, mainly the yeast Saccharomyces cerevisiae, and in human patients with COX deficiencies They are required for every step of the formation of the complex, from translation of the individual subunits to incorporation of the prosthetic groups and assembly of the holoenzyme [6]. HCOA3 Participates in Cytochrome c Oxidase Biogenesis well as Cox and Coa, two COX1 stability/assembly factors Whether this regulatory mechanism is conserved in humans remains to be fully disclosed. We demonstrate that human CCDC56, hereafter termed hCOA3, is involved in the early steps of COX biogenesis by securing the stability and promoting the assembly of newly synthesized COX1 These results suggest that hCOA3 could play a role in coordinating COX1 synthesis and assembly. Our results identify hCOA3 as a new candidate when screening for genes responsible for mitochondrial diseases associated with COX deficiency

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION