Figure 4.

Respiration in complemented Arabidopsis grxs15 mutants. A, Root respiration rate of GRXS15 K83A line #3 (4.5-week old) and the respective WT grown to similar size (2-week old) after addition of the cytochrome c oxidase inhibitor KCN (4 mM) alone or together with the AOX inhibitor pGal (0.2 mM; n = 4; means ± sd). The statistical analysis (two-way ANOVA with post hoc Holm–Sidak comparisons for WT versus grxs15 mutant) indicated a significant difference in the respiration of mito from WT and GRXS15 K83A line #3; ***P ≤ 0.001. B, Respiratory complexes I, II, III, and V separated by Blue Native–PAGE and visualized with Coomassie staining in WT, GRXS15 K83A line #4, and GRXS15^amiR. Mito were purified from 4-week-old plants. C and D, O₂ consumption rates for purified mito from WT and GRXS15 K83A line #3 energized with succinate (left) or pyruvate/malate (right). O₂ consumption was measured before (blank) and after addition of mito. State II respiration was initiated by the addition of the respective substrate (State II; succinate (10 mM succinate, 0.25 mM ATP) or pyruvate/malate (10 mM pyruvate, 10 mM malate, 0.3 mM NAD, and 0.1 mM thiamine pyrophosphate). State III respiration was initiated by the addition of 50 μM ADP. State IV represents the respiration after ADP consumption and CCCP shows the respiration after addition of the protonophore carbonyl cyanide m-CCCP (10 μM), which uncouples electron transport from ATP synthesis. All results are based on three independent preparations of mito and are shown as means ± SEM.