Characterization of mitochondrial FOXRED1 in the assembly of respiratory chain complex I

Luke E Formosa, Masakazu Mimaki, Ann E Frazier, Matthew McKenzie, Tegan L Stait, David R Thorburn, David A Stroud, Michael T Ryan

Research output: Contribution to journalArticleResearchpeer-review

38 Citations (Scopus)


Human mitochondrial complex I is the largest enzyme of the respiratory chain and is composed of 44 different subunits. Complex I subunits are encoded by both nuclear and mitochondrial (mt) DNA and their assembly requires a number of additional proteins. FAD-dependent oxidoreductase domain-containing protein 1 (FOXRED1) was recently identified as a putative assembly factor and FOXRED1 mutations in patients cause complex I deficiency; however, its role in assembly is unknown. Here, we demonstrate that FOXRED1 is involved in mid-late stages of complex I assembly. In a patient with FOXRED1 mutations, the levels of mature complex I were markedly decreased, and a smaller approximately 475 kDa subcomplex was detected. In the absence of FOXRED1, mtDNA-encoded complex I subunits are still translated and transiently assembled into a late stage approximately 815 kDa intermediate; but instead of transitioning further to the mature complex I, the intermediate breaks down to an approximately 475 kDa complex. As the patient cells contained residual assembled complex I, we disrupted the FOXRED1 gene in HEK293T cells through TALEN-mediated gene editing. Cells lacking FOXRED1 had approximately 10 complex I levels, reduced complex I activity, and were unable to grow on galactose media. Interestingly, overexpression of FOXRED1 containing the patient mutations was able to rescue complex I assembly. In addition, FOXRED1 was found to co-immunoprecipitate with a number of complex I subunits. Our studies reveal that FOXRED1 is a crucial component in the productive assembly of complex I and that mutations in FOXRED1 leading to partial loss of function cause defects in complex I biogenesis.
Original languageEnglish
Pages (from-to)2952 - 2965
Number of pages14
JournalHuman Molecular Genetics
Issue number10
Publication statusPublished - 2015

Cite this