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Molecular and Functional Effects of Loss of Cytochrome c Oxidase Subunit 8A

Daria Rotko1,2,a, Alexei P. Kudin1,b, Gábor Zsurka1,3,c, Bogusz Kulawiak2,d, Adam Szewczyk2,e, and Wolfram S. Kunz1,3,f*

1Institute of Experimental Epileptology and Cognition Research, Life & Brain Center, University of Bonn, Venusberg-Campus 1, 53127 Bonn, Germany

2Laboratory of Intracellular Ion Channels, Nencki Institute of Experimental Biology, Polish Academy of Sciences, 02-093 Warsaw, Poland

3Department of Epileptology, University Bonn Medical Center, Venusberg-Campus 1, 53127 Bonn, Germany

* To whom correspondence should be addressed.

Received November 2, 2020; Revised November 23, 2020; Accepted November 23, 2020
In this work we studied molecular and functional effects of the loss of the smallest nuclear encoded subunit of cytochrome c oxidase COX8A in fibroblasts from a patient with a homozygous splice site mutation and in CRISPR/Cas9 genome-edited HEK293T cells. In both cellular model systems, between 20 to 30% of the residual enzymatic activity of cytochrome c oxidase (COX) was detectable. In immunoblots of BN-PAGE separated mitochondria from both cellular models almost no monomers and dimers of the fully assembled COX could be visualized. Interestingly, supercomplexes of COX formed with complex III and also with complexes I and III retained considerable immunoreactivity, while nearly no immunoreactivity attributable to subassemblies was found. That indicates that COX lacking subunit 8A is stabilized in supercomplexes, while monomers and dimers are rapidly degraded. With transcriptome analysis by 3′-RNA sequencing we failed to detect in our cellular models of COX8A deficiency transcriptional changes of genes involved in the mitochondrial unfolded protein response (mtUPR) and the integrated stress response (ISR). Thus, our data strongly suggest that the smallest subunit of cytochrome c oxidase COX8A is required for maintenance of the structural stability of COX monomers and dimers.
KEY WORDS: mitochondria, cytochrome c oxidase, subunit 8A, respiratory chain super complexes

DOI: 10.1134/S0006297921010041