Characterization of an internal type-II NADH dehydrogenase from Chlamydomonas reinhardtii mitochondria

Abstract

Type-II NAD(P)H dehydrogenases form a multigene family that comprise six members in the green microalga Chlamydomonas. To date, only one enzyme (Nda2) located in the chloroplast has been characterized in this alga and demonstrated to participate in the reduction of the plastoquinone pool. We present here the functional characterization of Nda1. The enzyme is located on the inner face of the inner mitochondrial membrane. Its downregulation leads to a slight decrease of NADH:ferricyanide activity and of dark whole cell respiration. To determine whether the reduction of Nda1 combined with the lack of complex I would affect mitochondrial processes, double mutants affected in both Nda1 and complex I were isolated. Respiration and growth rates in heterotrophic conditions were significantly altered in the double mutants investigated, suggesting that Nda1 plays a role in the oxidation of matrix NADH in the absence of complex I.

Inactivation of genes coding for mitochondrial Nd7 and Nd9 complex I subunits in Chlamydomonas reinhardtii. Impact of complex I loss on respiration and energetic metabolism

“…Complex I-deficient mutants survive in photo-autotrophic conditions (light + mineral medium) due to the operation of rotenone-insensitive alternative NAD(P)H dehydrogenases (Lecler et al, 2012) and because their photosynthetic activity is barely affected (Cardol et al, 2003). Complex I-deficient mutants are also viable in heterotrophic conditions (darkness + acetate as a carbon source).…”

“…RNA interference also allowed us to gain insight into the role of other respiratory chain enzymes, such as F 1 F O ATP synthase (Lapaille et al, 2010), rotenone-insensitive alternative NAD(P)H dehydrogenase (Lecler et al, 2012) and cyanide-insensitive alternative oxidase (AOX) (Mathy et al, 2010).…”

Respiratory-deficient mutants of the unicellular green alga Chlamydomonas: A review

“…The NDA1-RNAi knock-down mutant presents a very mild phenotype, with only a slight decrease of respiration and no growth defect in heterotrophic conditions. In contrast, a double mutant affected in both Nda1 and complex I displayed strong alteration of respiration and growth rates in heterotrophic conditions, suggesting that Nda1 plays a role in the oxidation of matrix NADH in the absence of complex I [34].…”

“…The Chlamydomonas nuclear genome encodes six type II NAD(P)H dehydrogenases (Nda1, 2, 3, 5, 6, 7), and the localization of three of them has been determined: Nda2 and Nda3, are located in the chloroplast [32,33] while Nda1 is located at the inner side of the inner mitochondrial membrane [34].…”

Impaired Mitochondrial Transcription Termination Disrupts the Stromal Redox Poise in Chlamydomonas

“…In contrast to AOX1, expression of the NDA1 gene increased during cultivation of B13 in high light (3.11 6 1.05 at t 3h versus 1 at t 0 ; gray bars; P . 0.1) This gene encodes a matrix-facing rotenone-insensitive NAD(P)H dehydrogenase located in the inner mitochondrial membrane (Lecler et al, 2012). This strong difference between NDA1 transcript levels in low and high lightcultivated cells could not be seen in stm6 (2.49 6 0.36 at t 0 versus 2.96 6 0.49 at t 3h ; orange bars), which showed high levels already under low light conditions (2.49 6 0.36 versus 1; P , 0.05).…”