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Abstract
Isolated complex I deficiencies are one of the most commonly observed biochemical features in patients suffering from mitochondrial disorders. In the majority of these clinical cases the molecular bases of the diseases remain unknown suggesting the involvement of unidentified factors that are critical for complex I function. The Saccharomyces cerevisiae NDI1 gene, encoding the mitochondrial internal NADH dehydrogenase was previously shown to complement a complex I deficient strain in Caenorhabditis elegans with notable improvements in reproduction and whole organism respiration. These features indicate that Ndi1p can functionally integrate the respiratory chain, allowing complex I deficiency complementation. Taking into account the Ndi1p ability to bypass complex I, we evaluate the possibility to extend the range of defects/mutations causing complex I deficiencies that can be alleviated by NDI1 expression. We report here that NDI1 expressing animals unexpectedly exhibit a slightly shortened lifespan, a reduction in the progeny, and a depletion of the mitochondrial genome. However, Ndi1p is expressed and targeted to the mitochondria as a functional protein that confers rotenone resistance to those animals without affecting their respiration rate and ATP content. We show that the severe embryonic lethality level caused by the RNAi knockdowns of complex I structural subunit encoding genes (e.g., NDUFV1, NDUFS1, NDUFS6, NDUFS8, or GRIM-19 human orthologs) in wild type animals is significantly reduced in the Ndi1p expressing worm. All together these results open up the perspective to identify new genes involved in complex I function, assembly, or regulation by screening an RNAi library of genes leading to embryonic lethality that should be rescued by NDI1 expression.
Highlights
In humans, NADH: ubiquinone oxidoreductase is composed of at least 45 different subunits encoded by both mitochondrial and nuclear genomes making it the largest of the five respiratory chain complexes
Mitochondrial dysfunctions caused by respiratory chain complex I deficiency have been shown to possibly originate from deleterious mutations in either the nuclear or the mitochondrial genomes, resulting in a wide spectrum of human diseases ranging from leber hereditary optic neuropathy (LHON), that targets one organ, to diseases affecting several organs as the mitochondrial encephalomyopathy associated with cardiomyopathy (Fassone and Rahman, 2012)
As Ndi1p is a mitochondrial NADH dehydrogenase that does not exist in C. elegans, its expression was investigated by Western blot analysis using a specific antibody directed against S. cerevisiae Ndi1p
Summary
NADH: ubiquinone oxidoreductase (complex I) is composed of at least 45 different subunits encoded by both mitochondrial and nuclear genomes making it the largest of the five respiratory chain complexes. Identification of new complex I related genes chain through ubiquinone as electron acceptor This huge complex is composed of structural subunits organized in three well-defined functional modules: the N module involved in oxidizing NADH, the Q module involved in reducing ubiquinone and the P module dedicated to the proton translocation (Brandt, 2006). A so challenging task that only around 20% of the patients harboring a complex I deficiency carry a mutation in one of the known complex I subunit encoding genes (Rotig, 2010). A recent exome sequencing screening, for mutations in 75 complex I associated genes in about 150 patients with a biochemically defined complex I deficiency identified mutations in 50% of the cases and causative mutations in less than 20% of the patients (Haack et al, 2012)
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