Cytochrome c oxidase (COX) biogenesis in health and disease

Figure 1

Model of cytochrome c oxidase biogenesis

COX is the terminal oxidase of the respiratory chain. COX deficiency is the major cause of mitochondrial encephalomyopathies in humans. Our long-term goal is to attain a complete understanding of the pathways leading to COX assembly and their components as a prerequisite to the development of therapies for the management of disorders associated with COX deficiencies.

Figure 2

Translational regulation of cytochrome c oxidase biogenesis in S. cerevisiae

Translational regulation of COX assembly. COX is a hetero-oligomeric enzyme formed by subunits encoded in the nuclear and the mitochondrial DNA. Because COX contain highly reactive heme A and copper prosthetic groups, the biogenesis of this enzyme must be tightly regulated to prevent the accumulation of pro-oxidant assembly intermediates. Over the last few years we have used the yeast Saccharomyces cerevisiae to discover the existence of a negative feedback translational regulatory system. This system coordinates the synthesis of Cox1, a mtDNA-encoded catalytic subunit containing heme A and copper centers, with its assembly into the holoenzyme. With support from NIH and MDA, we identified a COX1 mRNA-specific translation activator, Mss51, as the key element of the system. Mss51 is a bi-functional protein that interacts with the 5’UTR of COX1 mRNA to promote translation and subsequently interacts with the newly synthesized Cox1 protein to facilitate its stability in pre-assembly complexes. Mss51 does not act alone. The general mitochondrial Hsp70 chaperone Ssc1 and the COX specific chaperones Cox14 and Cox25 dynamically interact with Mss51-containing complexes to coordinate Cox1 synthesis and assembly, and to facilitate Mss51 recycling between its two functions. More recently, we discovered that Mss51 binds heme. This finding, recently published in Cell Metabolism, has provided a key element for a regulatory mechanism that coordinates assembly of COX, the major oxygen-consuming mitochondrial enzyme, with heme and oxygen availability for respiration and aerobic energy production. Some ongoing projects include include: (i) Identification and characterization of the heme A insertase/s; (ii) Regulation of Mss51 function by Ssc1 (iii) Regulation of COX biogenesis by oxygen and reactive oxygen species.

Role of tween CX9C-motif proteins in redox balance and copper delivery to COX. Although several mitochondrial COX copper chaperones are already known, there are essential gaps in our knowledge regarding how copper reaches mitochondria, how copper is transported from a matrix pool to the intermembrane space, how copper is distributed to COX and mt-Sod1 (Cu-Zn superoxide dismutase) and how the metallation state of these two enzymes regulates cellular copper homeostasis. We have discovered two novel conserved mitochondrial copper chaperones, Cmc1 and Cmc2, from the twin-CX9C-motif family, required for the metallation of COX and also for the copper metallation of the mitochondrial intermembrane space portion of Sod1. We are currently working toward understanding how copper is partitioned between the two enzymes. Additionally, we continue the functional characterization of key CX9C-motif proteins in COX assembly in yeast and human cell lines.

Role of COX assembly factors in human cells. Most COX biogenetic factors were initially discovered in yeast but they have human homologues whose functions are in many instances poorly understood. We are taking advantage of the new advances in gene editing technology to create human cell lines knockout for specific COX assembly factors. The KO cell lines are subsequently characterized and used to express mutant or tagged versions of KO genes. Using this approach, we have already published in Human Molecular Genetics the characterization of human COX20 as a COX2 chaperone that assist the copper metallation of this subunit by the SCO1/2 proteins.