Thermal sensitivity of mitochondrial metabolism in two distinct mitotypes ofDrosophila simulans: evaluation of mitochondrial plasticity

Abstract

SUMMARYThe overall aim of this study was to (1) evaluate the adaptive value of mitochondrial DNA by comparing mitochondrial performance in populations possessing different haplotypes and distribution, and to (2) evaluate the sensitivity of different enzymes of the electron transport system (ETS) during temperature-induced changes. We measured the impact of temperature of mitochondrial respiration and several key enzymes of mitochondrial metabolism in two mitotypes (siII and siIII) of Drosophila simulans. The temperature dependencies of oxygen consumption for mitochondria isolated from flight muscle was assessed with complex I substrates (pyruvate + malate + proline) and with sn glycerol-3-phosphate (to reduce complex III via glycerophosphate dehydrogenase) in both coupled and uncoupled states. Activities of citrate synthase, cytochrome c oxidase (COX), catalase and aconitase, and the excess capacity of COX at high convergent pathway flux were also measured as a function of temperature. Overall, our results showed that functional differences between the two mitotypes are few. Results suggest that differences between the two mitotypes could hardly explain the temperature-specific differences measured in mitochondria performances. It suggests that some other factor(s) may be driving the maintenance of mitotypes. We also show that the different enzymes of the ETS have different thermal sensitivities. The catalytic capacities of these enzymes vary with temperature changes, and the corresponding involvement of the different steps on mitochondrial regulation probably varies with temperature. For example, the excess COX capacity is low, even non-existent, at high and intermediate temperatures (18°C, 24°C and 28°C) whereas it is quite high at a lower temperature (12°C), suggesting release of respiration control by COX at low temperature. Supplementary material available online at

Improved tolerance to salt and water stress in Drosophila melanogaster cells conferred by late embryogenesis abundant protein

“…Leak respiration rates in presence of high substrate concentrations were also identical for isolated mitochondria from both cell lines (Fig. 6) and matched well with respiration rates measured for mitochondria isolated from D. simulans flight muscle (Pichaud et al, 2010). Similar to mitochondria isolated from A. franciscana embryos (Menze et al, 2009) a high capacity to maintain functional integrity after freeze-thawing in presence of polyols (trehalose or sucrose) was observed for mitochondria isolated from Kc167-AfLEA1.3 cells.…”

“…Respiration measurements were performed first on fresh preparations using an Oxygraph-2K and about 10 -20 µl of resuspended mitochondria per assay in MiR05A at 25 °C. Respiration was induced by adding 5 mM pyruvate, 10 mM proline and 2 mM malate to provide electron flow through complex I (Pichaud et al, 2010). To engage the phosphorylation system 1 mM ADP was added followed by additions of 10 mM succinate and 10 mM sn-glycerol-3-phosphate to supply electrons to the ubiquinone pool via succinate dehydrogenase and glycerol-3-phosphate dehydrogenase and to maximally reduce complex III.…”

Experimental evidence that litter size imposes an oxidative challenge to offspring

“…Aconitase plays an important role in the tricarboxylic acid cycle and is principally located within mitochondria (Wiegand and Remington, 1986;Gardner et al, 1995); therefore, citrate synthase, a marker of mitochondrial density, was also measured (Pichaud et al, 2010). For both aconitase and citrate synthase activity assays, tissue samples were homogenised by macerating tissue into a fine powder in liquid nitrogen and suspended in 100 mmol l −1 potassium phosphate buffer.…”

“…For both aconitase and citrate synthase activity assays, tissue samples were homogenised by macerating tissue into a fine powder in liquid nitrogen and suspended in 100 mmol l −1 potassium phosphate buffer. For full method of both the aconitase and citrate synthase activity assays, see Garratt et al (2013) (see also Pichaud et al, 2010).…”

Physiological adaptations to reproduction II. Mitochondrial adjustments in livers of lactating mice

“…Homogenates were transferred in 100mmoll −1 imidazole-HCl (pH8), 0.1mmoll −1 5,5′-dithiobis(2-nitrobenzoic acid) (DTNB), 0.1mmoll −1 acetyl CoA and 0.15mmoll −1 oxaloacetate. Activities were determined by following the increase in absorbance due to the reduction of DTNB at 412nm (Pichaud et al, 2010). Protein concentration was determined using Coomassie Blue (Bradford, 1976) and results were expressed as Umg −1 proteins, where U is 1μmol of substrate transformed per minute.…”