Overlapping Role of Respiratory Supercomplex Factor Rcf2 and Its N-terminal Homolog Rcf3 in Saccharomyces cerevisiae

Abstract

Edited by Linda SpremulliThe mitochondrial electron transport chain consists of individual protein complexes arranged into large macromolecular structures, termed respiratory chain supercomplexes or respirasomes. In the yeast Saccharomyces cerevisiae, respiratory chain supercomplexes form by association of the bc 1 complex with the cytochrome c oxidase. Formation and maintenance of these assemblies are promoted by specific respiratory supercomplex factors, the Rcf proteins. For these proteins a regulatory function in bridging the electron transfer within supercomplexes has been proposed. Here we report on the maturation of Rcf2 into an N-and C-terminal peptide. We show that the previously uncharacterized Rcf3 (YBR255c-A) is a homolog of the N-terminal Rcf2 peptide, whereas Rcf1 is homologous to the C-terminal portion. Both Rcf3 and the C-terminal fragment of Rcf2 associate with monomeric cytochrome c oxidase and respiratory chain supercomplexes. A lack of Rcf2 and Rcf3 increases oxygen flux through the respiratory chain by up-regulation of the cytochrome c oxidase activity. A double gene deletion of RCF2 and RCF3 affects cellular survival under non-fermentable growth conditions, suggesting an overlapping role for both proteins in the regulation of the OXPHOS activity. Furthermore, our data suggest an association of all three Rcf proteins with the bc 1 complex in the absence of a functional cytochrome c oxidase and identify a supercomplex independent interaction network of the Rcf proteins.Eukaryotic cells cover their energy demands mainly with ATP generated by oxidative phosphorylation in mitochondria.

Definition of a High-Confidence Mitochondrial Proteome at Quantitative Scale

“…Rcf3 may thus function as mediator between respiratory chain III–IV supercomplexes and MICOS. (6) Yil077c assembled into large blue native gel complexes that are characteristic for III–IV supercomplexes (Figures 7A and 7D, right panel), in agreement with the co-purification of Yil077c with Rcf3 (Figure 7C) (Römpler et al., 2016). Tagged Yil077c, named Rci37 (respiratory chain interacting protein of 37 kDa), co-purified subunits of complexes III and IV and additionally the subunits Afg3 and Yta12 of the inner membrane m-AAA protease (Rugarli and Langer, 2012), followed by the prohibitin complex (Figure 7D).…”

“…Tmh11 interacts with oligomerization-promoting subunits of MICOS and the F 1 F o -ATP synthase (Arnold et al., 1998, Barbot et al., 2015, Bohnert et al., 2015, Rabl et al., 2009), pointing to an interaction of Tmh11 with a special pool of the MICOS core component Mic10. We found that Rcf3 associates not only with respiratory chain supercomplexes (Römpler et al., 2016) but also with the entire MICOS complex, indicating a role of Rcf3 in linking respiratory chain and the cristae organizing system. Mco10 was found as partner protein of the F 1 F o -ATP synthase, and Coq21 was identified as component of the coenzyme Q biosynthesis cluster.…”

NMR Study of Rcf2 Reveals an Unusual Dimeric Topology in Detergent Micelles

“…Formation, maintenance, and regulation of respiratory supercomplexes in S. cerevisiae are promoted by specific factors, the so‐called respiratory supercomplex factor 1 and 2 (Rcf1 and Rcf2) proteins . Rcf2 is a 224‐residue integral membrane protein, located in the mitochondrial inner membrane .…”

“…Formation, maintenance, and regulation of respiratory supercomplexes in S. cerevisiae are promoted by specific factors, the so‐called respiratory supercomplex factor 1 and 2 (Rcf1 and Rcf2) proteins . Rcf2 is a 224‐residue integral membrane protein, located in the mitochondrial inner membrane . Both Rcf1 and Rcf2 have a conserved domain at the N terminus that belongs to the hypoxia‐induced gene 1 family, the function of which might be related to the regulation of supercomplexes in higher eukaryotes as well .…”

“…Both Rcf1 and Rcf2 have a conserved domain at the N terminus that belongs to the hypoxia‐induced gene 1 family, the function of which might be related to the regulation of supercomplexes in higher eukaryotes as well . Biochemical studies have revealed that the C‐terminal fragment of Rcf2 associates with supercomplexes through complex IV . The Rcf2 protein, however, has not been observed in the low‐resolution cryo‐EM structure of the III 2 IV 2 supercomplex in yeast .…”