Mitochondrial respiratory chain (MRC) deficiencies frequently cause encephalomyopathy. The MRC is responsible for the aerobic generation of energy within the cell and includes four multimeric enzymatic complexes (CI to CIV). These respiratory complexes can be organized as individual entities or together assembled into supercomplexes (in human, CI+CIII2, CIII2+CIV and CI+CIII2+CIV1-n). The major goal of our research is to identify the SC assembly factors involved and the physiological relevance of this dynamic organization, which remain largely undefined.

Here, we focus on two members of the Hypoxia Inducible Gene Domain family, HIGD1A and HIGD2A, which are the mammalian homologs of yeast Rcf1, required for CIII-CIV association. We hypothesize that HIGD proteins could regulate MRC complex and supercomplex biogenesis, to adapt to changing environmental conditions, especially during hypoxia and oxidative stress.

We found that HIGD1A and HIGD2A are mitochondrial inner membrane proteins, upregulated upon hypoxia and oxidative stress. Both proteins associate with CIV, and HIGD1A additionally assembles into supercomplexes. We generated human HEK293T cell lines knock-out (KO) for HIGD1A or HIGD2A, or both. In every KO line, the steady-state levels of CIII and CIV were reduced, correlating with attenuated respiratory capacity, enzymatic activities, and ROS production. However, the accumulation of a CIV subassembly exclusively in HIGD2A-KO cells suggests a primary role of HIGD2A in CIV biogenesis, whereas HIGD1A-KO cells have major decreases in CIII2 and CIII-containing supercomplexes. Overexpression of HIGD1A in HIGD2A-KO partially restored CIV assembly and respiration rate, but not vice versa, suggesting a more general role of HIGD1A on MRC biogenesis. Our results also propose a model in which both HIGD proteins participate in CIV assembly in a sequential manner, in which HIGD2A participates in early-assembly stages before HIGD1A.

Current investigations will determine whether these roles are modified and/or extended under hypoxic and oxidative stress conditions, relevant to mitochondrial encephalomyopathies, neurodegenerative disorders, stroke and cancer.

Authors: Alba Timon-Gomez and Antoni Barrientos