Abstact

Respiratory complex I (NADH:ubiquinone oxidoreductase) is essential for cellular energy production and NAD+ homeostasis. Complex I mutations cause neuromuscular, mitochondrial diseases, such as Leigh Syndrome, but their molecular-level consequences remain poorly understood. Here, we use a popular complex I-linked mitochondrial disease model, the ndufs4-/- mouse, to define the structural, biochemical, and functional consequences of the absence of subunit NDUFS4. Cryo-EM analyses of the complex I from ndufs4-/- mouse hearts revealed a loose association of the NADH-dehydrogenase module, and discrete classes containing either assembly factor NDUFAF2 or subunit NDUFS6. Subunit NDUFA12, which replaces its paralogue NDUFAF2 in mature complex I, is absent from all classes, compounding the deletion of NDUFS4 and preventing maturation of an NDUFS4-free enzyme. We propose that NDUFAF2 recruits the NADH-dehydrogenase module during assembly of the complex. Taken together, the findings provide new molecular-level understanding of the ndufs4-/- mouse model and complex I-linked mitochondrial disease.