The heme molecule is an obligatory cofactor in the terminal enzyme complex of the electron transport chain, cytochrome oxidase. Heme is synthesized from heme by a multi-spanning inner membrane protein, heme synthase (Cox15 in the yeast). The insertion of heme is critical for cytochrome oxidase function and assembly, but this process has not been fully elucidated. To improve our understanding of heme insertion into cytochrome oxidase, here we investigated the protein-protein interactions that involve Cox15 in In addition to observing Cox15 in homooligomeric complexes, we found that a portion of Cox15 also associates with the mitochondrial respiratory supercomplexes. When supercomplex formation was abolished, as in the case of stalled cytochrome or cytochrome oxidase assembly, Cox15 maintained an interaction with select proteins from both respiratory complexes. In the case of stalled cytochrome assembly, Cox15 interacted with the late-assembling cytochrome oxidase subunit, Cox13. When cytochrome oxidase assembly was stalled, Cox15 unexpectedly maintained its interaction with the cytochrome protein, Cor1. Our results indicate that Cox15 and Cor1 continue to interact in the cytochrome dimer even in the absence of supercomplexes orwhen the supercomplexes are destabilized. These findings reveal that Cox15 not only associates with respiratory supercomplexes, but also interacts with the cytochrome dimer even in the absence of cytochrome oxidase.
Num1 is a multifunctional protein that both tethers mitochondria to the plasma membrane and anchors dynein to the cell cortex during nuclear inheritance. Previous work has examined the impact loss of Num1-based mitochondrial tethering has on dynein function in Saccharomyces cerevisiae; here, we elucidate its impact on mitochondrial function. We find that like mitochondria, Num1 is regulated by changes in metabolic state, with the protein levels and cortical distribution of Num1 differing between fermentative and respiratory growth conditions. In cells lacking Num1, we observe a reproducible respiratory growth defect, suggesting a role for Num1 in not only maintaining mitochondrial morphology, but also function. A structure-function approach revealed that, unexpectedly, Num1-mediated cortical dynein anchoring is important for normal growth under respiratory conditions. The severe respiratory growth defect in Δnum1 cells is not specifically due to dynein's canonical function in nuclear migration but is dependent on the presence of dynein, as deletion of DYN1 in Δnum1 cells partially rescues respiratory growth. We hypothesize that misregulated dynein present in cells that lack Num1 negatively impacts mitochondrial function resulting in defects in respiratory growth.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.