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Alternative NADH dehydrogenase (NDH2): intermembrane-space-facing counterpart of mitochondrial complex I in the procyclic Trypanosoma brucei

Published online by Cambridge University Press:  30 October 2012

ZDENĚK VERNER ,
INGRID ŠKODOVÁ ,
SIMONA POLÁKOVÁ ,
VLADISLAVA ĎURIŠOVÁ-BENKOVIČOVÁ ,
ANTON HORVÁTH  and
JULIUS LUKEŠ
ZDENĚK VERNER
Affiliation:
Biology Centre, Institute of Parasitology, Czech Academy of Sciences, České Budějovice (Budweis), Czech Republic Faculty of Sciences, University of South Bohemia, České Budějovice (Budweis), Czech Republic Department of Biochemistry, Comenius University, Bratislava, Slovakia
INGRID ŠKODOVÁ
Affiliation:
Biology Centre, Institute of Parasitology, Czech Academy of Sciences, České Budějovice (Budweis), Czech Republic Department of Biochemistry, Comenius University, Bratislava, Slovakia
SIMONA POLÁKOVÁ
Affiliation:
Department of Zoology, DAPHNE ČR – Institute of Applied Ecology, České Budějovice (Budweis), Czech Republic
VLADISLAVA ĎURIŠOVÁ-BENKOVIČOVÁ
Affiliation:
Department of Biochemistry, Comenius University, Bratislava, Slovakia
ANTON HORVÁTH
Affiliation:
Department of Biochemistry, Comenius University, Bratislava, Slovakia
JULIUS LUKEŠ*
Affiliation:
Biology Centre, Institute of Parasitology, Czech Academy of Sciences, České Budějovice (Budweis), Czech Republic Faculty of Sciences, University of South Bohemia, České Budějovice (Budweis), Czech Republic
*
*Corresponding author: Biology Centre, Institute of Parasitology, Branišovská 31, 370 05 České Budějovice, Czech Republic. Tel: +420 387 775 416. Fax: +420 385 310 388. E-mail: [email protected]
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Summary

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.

Keywords

TrypanosomamitochondriondehydrogenaserespirationNDH2
Type
Research Article
Information
Parasitology , Volume 140 , Issue 3 , March 2013 , pp. 328 - 337
Copyright
Copyright © Cambridge University Press 2012

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