Abstract
The COX7A2L (Supercomplex Assembly Factor I, SCAFI) protein has been proposed to be a mitochondrial supercomplex assembly factor required for respirasome (supercomplex containing complexes I, III, and IV) formation. In the C57BL/6 mouse strain a homozygous in-frame 6-base-pair deletion in the COX7a2l/SCAF1 gene resulting in unstable protein and suggesting loss of function was previously identified. The loss of SCAFI was shown to impede respirasome formation, a major concern for the use of C57BL mouse strains in mitochondrial research. In contradiction, another recent study suggested that supercomplex formation is independent of SCAFI isoforms. We investigated whether SCAFI isoform status affected the disease severity and supercomplex formation in the liver of Bcs1lc.232A>G knock-in mice with incomplete complex III assembly. In homozygotes (Bcs1lG/G) of mixed (C57BL/6:129/Sv) genetic background, the lifespan was similar in mice with wild-type SCAFI allele and in those homozygous (SCAFIshort/short) for the deleted SCAF1 variant (34±3 days; n = 6 vs. 32±2 days; n = 7, respectively). SCAFI heterozygosity (SCAFIlong/short) resulted in decreased SCAFI protein but respirasome assembly was unaffected. Congenic (C57BL/6) mice were of the genotype SCAFIshort/short and had no detectable SCAFI protein. In their liver mitochondria, respirasome composition was altered as compared to mixed background mice. Complex IV was mainly present as monomers and dimers, and only low amounts were found in combination with complex I and complex III or with precomplex III. The main supercomplex in the liver mitochondria of C57BL/6 mice comprised only complexes I and III. In conclusion, in liver mitochondria of C57BL/6 mice, supercomplexes had markedly reduced amount of, but were not completely depleted of, complex IV, supporting a role for COX7A2L/SCAFI in supercomplex assembly. However, the disease progression of the Bcs1l mutant mice was unrelated to SCAFI isoforms and supercomplex composition, suggesting that other genetic factors contribute to the different survival in the different genetic backgrounds.
Highlights
Mitochondrial respiratory chain (RC) consists of four complexes (CI-CIV), which use the energy of electron transport to generate a proton gradient that drives ATP synthesis by the ATP synthase
supercomplex assembly factor I (SCAFI) genotyping showed that 129/Sv wild-type mice were homozygous for the long allele encoding a 113-amino acid protein, whereas mice of the C57BL/6 backcrossed strain were homozygous for the short allele predicting a 111-amino acid protein that was undetectable on Western blots (Fig 2A and 2B)
Our hypothesis that the longer survival of homozygous Bcs1l mutant mice in mixed background would be related to presence of long SCAFI isoform was not supported
Summary
Mitochondrial respiratory chain (RC) consists of four complexes (CI-CIV), which use the energy of electron transport to generate a proton gradient that drives ATP synthesis by the ATP synthase (complex V, CV). The presence of larger conglomerates (supercomplexes) of the RC complexes was first suggested in 2000 [1] and has been supported by several other studies. The study suggested that supercomplex formation might have important effects on the phenotype of mouse disease models [9]. A contradictory study found that CIV was present in supercomplexes and RC function was normal in mitochondria of wildtype C57BL/6 mice with short SCAFI alleles [10]. It is not known whether the short/short genotype has an effect on the phenotype and disease progress in mouse models of RC complex deficiencies
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