Alternative oxidase in durum wheat mitochondria. Activation by pyruvate, hydroxypyruvate and glyoxylate and physiological role

In order to gain a first insight into the alternative oxidase (AO) function in durum wheat mitochondria (DWM), we investigated some activation pathways of this enzyme in DWM purified from both etiolated shoots and green leaves. AO was activated when DWM were added with either pyruvate, known as an AO activator in other plant mitochondria, or alanine plus 2-oxoglutarate, which can generate intramitochondrial pyruvate and glutamate via transamination. In contrast, no AO activity was observed during oxidation of malate plus glutamate or succinate (which can generate malate). In this regard DWM differ from other plant mitochondria. Moreover, DWM were found: (i) to have a very low malic enzyme (ME) activity, (ii) to release oxaloacetate rather than pyruvate during malate oxidation and (iii) to poorly oxidise malate in the absence of glutamate, which removes oxaloacetate via transamination. Therefore, we show that, unlike other plant mitochondria, no pyruvate is generated inside DWM from malate via ME, allowing no AO activity. Other AO activators, alternative to pyruvate, were checked by evaluating the capability of several compounds to induce oxygen uptake and/or electrical membrane potential (ΔΨ) in cyanide-treated DWM. Hydroxypyruvate and glyoxylate, photorespiratory cycle intermediates, were found to be powerful AO activators, capable of inducing a maximal rate of cyanide-insensitive oxygen uptake 1.7 times and 2.3 times higher than pyruvate, respectively. These results suggest that in durum wheat a link may exist between AO activity and photorespiratory metabolism rather than malate metabolism. Moreover, we observed that AO activation resulted in both a partially coupled respiration and a reduction by half of the rate of superoxide anion generation; therefore, AO is expected to work as an antioxidative defence system when the photorespiratory cycle is highly active, as under environmental stress.