Anti-NDUFS4 monoclonal antibody, clone FQS23302

In mammalian cells the nuclear-encoded subunits of complex I are imported into mitochondria, where they are assembled with mt-DNA encoded subunits in the complex, or exchanged with pre-existing copies in the complex. The present work shows that in fibroblast cultures inhibition by KH7 of cAMP production in the mitochondrial matrix by soluble adenylyl cyclase (sAC) results in decreased amounts of free non-incorporated nuclear-encoded NDUFS4, NDUFV2 and NDUFA9 subunits of the catalytic moiety and inhibition of the activity of complex I. Addition of permeant 8-Br-cAMP prevents this effect of KH7. KH7 inhibits accumulation in isolated rat-liver mitochondria and incorporation in complex I of "in vitro" produced, radiolabeled NDUFS4 and NDUFV2 subunits. 8-Br-cAMP prevents also this effect of KH7. Use of protease inhibitors shows that intramitochondrial cAMP exerts this positive effect on complex I by preventing digestion of nuclear-encoded subunits by mitochondrial protease(s), whose activity is promoted by KH7 and KH89, an inhibitor of PICA. (C) 2014 Elsevier B.V. All rights reserved.

Mitochondrial diseases are a group of rare life-threatening diseases often caused by defects in the oxidative phosphorylation system. No effective treatment is available for these disorders. Therapeutic development is hampered by the high heterogeneity in genetic, biochemical, and clinical spectra of mitochondrial diseases and by limited preclinical resources to screen and identify effective treatment candidates. Alternative models of the pathology are essential to better understand mitochondrial diseases and to accelerate the development of new therapeutics. The fruit fly Drosophila melanogaster is a cost-and time-efficient model that can recapitulate a wide range of phenotypes observed in patients suffering from mitochondrial disorders. We targeted three important subunits of complex I of the mitochondrial oxidative phosphorylation system with the flexible UAS-Gal4 system and RNA interference (RNAi): NDUFS4 (ND-18), NDUFS7 (ND-20), and NDUFV1 (ND-51). Using two ubiquitous driver lines at two temperatures, we established a collection of phenotypes relevant to complex I deficiencies. Our data offer models and phenotypes with different levels of severity that can be used for future therapeutic screenings. These include qualitative phenotypes that are amenable to high-throughput drug screening and quantitative phenotypes that require more resources but are likely to have increased potential and sensitivity to show modulation by drug treatment.