ADCK2 Haploinsufficiency Reduces Mitochondrial Lipid Oxidation and Causes Myopathy Associated with CoQ Deficiency.

Fatty acids and glucose are the main bioenergetic substrates in mammals. Impairment of mitochondrial fatty acid oxidation causes mitochondrial myopathy leading to decreased physical performance. Here, we report that haploinsufficiency of ADCK2, a member of the aarF domain-containing mitochondrial protein kinase family, in human is associated with liver dysfunction and severe mitochondrial myopathy with lipid droplets in skeletal muscle. In order to better understand the etiology of this rare disorder, we generated a heterozygous Adck2 knockout mouse model to perform in vivo and cellular studies using integrated analysis of physiological and omics data (transcriptomics-metabolomics). The data showed that Adck2+/- mice exhibited impaired fatty acid oxidation, liver dysfunction, and mitochondrial myopathy in skeletal muscle resulting in lower physical performance. Significant decrease in Coenzyme Q (CoQ) biosynthesis was observed and supplementation with CoQ partially rescued the phenotype both in the human subject and mouse model. These results indicate that ADCK2 is involved in organismal fatty acid metabolism and in CoQ biosynthesis in skeletal muscle. We propose that patients with isolated myopathies and myopathies involving lipid accumulation be tested for possible ADCK2 defect as they are likely to be responsive to CoQ supplementation.

Coenzyme Q is irreplaceable by demethoxy-coenzyme Q in plasma membrane of Caenorhabditis elegans.

A procedure was developed to isolate fractions enriched in plasma membrane from Caenorhabditis elegans. Coenzyme Q9 (Q9) was found in plasma membrane isolated from either wild-type or long-lived qm30 and qm51 clk-1 mutant strains of Caenorhabditis elegans, along with dietary coenzyme Q8 (Q8) and the biosynthetic intermediate demethoxy-Q9 (DMQ9). NADH was able to reduce both Q8 and Q9, but not DMQ9. Our results indicate that DMQ9 cannot achieve the same redox role of Q9 in plasma membrane, suggesting that proportion of all these Q isoforms in plasma membrane must be an important factor in establishing the clk-1 mutant phenotype.

Silencing of ubiquinone biosynthesis genes extends life span in Caenorhabditis elegans.

Ubiquinone (coenzyme Q; Q) is a key factor in the mitochondria electron transport chain, but it also functions as an antioxidant and as a cofactor of mitochondrial uncoupling proteins. Furthermore, Q isoforms balance in Caenorhabditis elegans is determined by both dietary intake and endogenous biosynthesis. In the absence of synthesis, withdrawal of dietary Q8 in adulthood extends life span. Thus, Q plays an important role in the aging process and understanding its synthesis acquires a new impetus. We have identified by RNA interference (RNAi) eight genes, including clk-1, involved in ubiquinone biosynthesis in C. elegans feeding animals with dsRNA-containing Escherichia coli HT115 strains. Silenced C. elegans showed lower levels of both endogenous Q9 and Q8 provided by diet, produced less superoxide without a significant modification of mitochondrial electron chain, and extended life span compared with non-interfered animals. E. coli strains harboring dsRNA also interfered with their own Q8 biosynthesis. These findings suggest that more efficient electron transport between a lower amount of Q and electron transport capacity of the mitochondrial complexes leads to less production of reactive oxygen species that contributes to extension of life span in the nematode C. elegans.

C. elegans knockouts in ubiquinone biosynthesis genes result in different phenotypes during larval development.

Ubiquinone is an essential molecule in aerobic organisms to achieve both, ATP synthesis and antioxidant defence. Mutants in genes responsible of ubiquinone biosynthesis lead to non-respiring petite yeast. In C. elegans, coq-7/clk-1 but not coq-3 mutants live longer than wild type showing a 'slowed' phenotype. In this paper we demonstrate that absence in ubiquinone in coq-1, coq-2 or coq-8 mutants lead to larval development arrest, slowed pharyngeal pumping, eventual paralysis and cell death. All these features emerge during larval development, whereas embryo development appeared similar to that of wild type individuals. Dietary coenzyme Q did not restore any of the alterations found in these coq mutants. These phenomena suggest that coenzyme Q mutants unable to synthesize this molecule develop a deleterious phenotype leading to lethality. On the contrary, phenotype of C. elegans coq-7/clk-1 mutants may be a unique phenotype than can not generalize to mutants in ubiquinone biosynthesis. This particular phenotype may not be based on the absence of endogenous coenzyme Q, but to the simultaneous presence of dietary coenzyme Q and the its biosynthesis intermediate demethoxy-coenzyme Q.

Caenorhabditis elegans ubiquinone biosynthesis genes.

Ubiquinone (coenzyme Q, Q) is an essential lipid electron carrier in the mitochondria respiratory chain, and also functions as antioxidant and participates as a cofactor of mitochondrial uncoupling proteins. Caernorhabditis elegans synthesize Q9, but both dietary Q8 intake and endogenous Q9 biosynthesis determine Q balance. Thus, it is of current interest to know the regulatory mechanisms of Q9 biosynthesis in this nematode. Here we review results that leaded to identification of genes involved in Q9 biosynthesis in this nematode using the RNA interference technology. C. elegans coq genes were silenced and depletion of Q content was observed, indicating that the genes related here participate in Q9 biosynthesis. Silenced populations showed an extension of adult life span, probably by the decrease of endogenous oxidative stress produced in mitochondria. We also report the heterologous complementation of C. elegans coq-5 and coq-7 genes in their homologue yeast coq null mutants, leading to restore its ability to growth in non-fermentable sugars. These complemented yeast strains accumulated Q6 but also the intermediate demethoxy-Q6. These findings support the conservative functional homology of these genes.

A novel plasma membrane quinone reductase and NAD(P)H:quinone oxidoreductase 1 are upregulated by serum withdrawal in human promyelocytic HL-60 cells.

We have studied changes in plasma membrane NAD(P)H:quinone oxidoreductases of HL-60 cells under serum withdrawal conditions, as a model to analyze cell responses to oxidative stress. Highly enriched plasma membrane fractions were obtained from cell homogenates. A major part of NADH-quinone oxidoreductase in the plasma membrane was insensitive to micromolar concentrations of dicumarol, a specific inhibitor of the NAD(P)H:quinone oxidoreductase 1 (NQOI, DT-diaphorase), and only a minor portion was characterized as DT-diaphorase. An enzyme with properties of a cytochrome b5 reductase accounted for most dicumarol-resistant quinone reductase activity in HL-60 plasma membranes. The enzyme used mainly NADH as donor, it reduced coenzyme Q0 through a one-electron mechanism with generation of superoxide, and its inhibition profile by p-hydroxymercuribenzoate was similar to that of authentic cytochrome b5 reductase. Both NQO1 and a novel dicumarol-insensitive quinone reductase that was not accounted by a cytochrome b5 reductase were significantly increased in plasma membranes after serum deprivation, showing a peak at 32 h of treatment. The reductase was specific for NADH, did not generate superoxide during quinone reduction, and was significantly resistant to p-hydroxymercuribenzoate. The function of this novel quinone reductase remains to be elucidated whereas dicumarol inhibition of NQO1 strongly potentiated growth arrest and decreased viability of HL-60 cells in the absence of serum. Our results demonstrate that upregulation of two-electron quinone reductases at the plasma membrane is a mechanism evoked by cells for defense against oxidative stress caused by serum withdrawal.