title: Alternative NADH dehydrogenase (NDH2): intermembrane-space-facing counterpart of mitochondrial complex I in the procyclic Trypanosoma brucei.
abstract: The respiratory chain of the procyclic stage of Trypanosoma brucei contains the standard complexes I through IV, as well as several alternative enzymes contributing to electron flow. In this work, we studied the function of an alternative NADH : ubiquinone oxidoreductase (NDH2). Depletion of target mRNA was achieved using RNA interference (RNAi). In the non-induced and RNAi-induced cell growth, membrane potential change, alteration in production of reactive oxygen species, overall respiration, enzymatic activities of complexes I, III and/or IV and distribution of NADH : ubiquinone oxidoreductase activities in glycerol gradient fractions were measured. Finally, respiration using different substrates was tested on digitonin-permeabilized cells. The induced RNAi cell line exhibited slower growth, decreased mitochondrial membrane potential and lower sensitivity of respiration to inhibitors. Mitochondrial glycerol-3-phosphate dehydrogenase was the only enzymatic activity that has significantly changed in the interfered cells. This elevation as well as a decrease of respiration using NADH was confirmed on digitonin-permeabilized cells. The data presented here together with previously published findings on complex I led us to propose that NDH2 is the major NADH : ubiquinone oxidoreductase responsible for cytosolic and not for mitochondrial NAD+ regeneration in the mitochondrion of procyclic T. brucei.
authors.affiliation: Biology Centre, Institute of Parasitology, Czech Academy of Sciences, České Budějovice (Budweis), Czech Republic.
authors.forename: Zdeněk
authors.initials: Z
authors.lastname: Verner
authors.forename: Ingrid
authors.initials: I
authors.lastname: Skodová
authors.forename: Simona
authors.initials: S
authors.lastname: Poláková
authors.forename: Vladislava
authors.initials: V
authors.lastname: Durišová-Benkovičová
authors.forename: Anton
authors.initials: A
authors.lastname: Horváth
authors.forename: Julius
authors.initials: J
authors.lastname: Lukeš
mesh_terms: D000818:Animals; D003600:Cytosol / Q000201:enzymology; D004579:Electron Transport; D042967:Electron Transport Complex I; D007425:Intracellular Membranes / Q000378:metabolism; D008564:Membrane Potentials; D008928:Mitochondria / Q000201:enzymology*; D009243:NAD / Q000378:metabolism; D009245:NADH Dehydrogenase / Q000235:genetics / Q000378:metabolism*; D010088:Oxidoreductases / Q000235:genetics / Q000378:metabolism; D010101:Oxygen Consumption; D015800:Protozoan Proteins / Q000378:metabolism; D014346:Trypanosoma brucei brucei / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development / Q000502:physiology
chemical_list: D015800:Protozoan Proteins; D009243:NAD; D010088:Oxidoreductases; D009245:NADH Dehydrogenase; D042967:Electron Transport Complex I