Regulation of succinate-ubiquinone reductase and fumarate reductase activities in human complex II by phosphorylation of its flavoprotein subunit - PubMed (original) (raw)

Regulation of succinate-ubiquinone reductase and fumarate reductase activities in human complex II by phosphorylation of its flavoprotein subunit

Eriko Tomitsuka et al. Proc Jpn Acad Ser B Phys Biol Sci. 2009.

Abstract

Complex II (succinate-ubiquinone reductase; SQR) is a mitochondrial respiratory chain enzyme that is directly involved in the TCA cycle. Complex II exerts a reverse reaction, fumarate reductase (FRD) activity, in various species such as bacteria, parasitic helminths and shellfish, but the existence of FRD activity in humans has not been previously reported. Here, we describe the detection of FRD activity in human cancer cells. The activity level was low, but distinct, and it increased significantly when the cells were cultured under hypoxic and glucose-deprived conditions. Treatment with phosphatase caused the dephosphorylation of flavoprotein subunit (Fp) with a concomitant increase in SQR activity, whereas FRD activity decreased. On the other hand, treatment with protein kinase caused an increase in FRD activity and a decrease in SQR activity. These data suggest that modification of the Fp subunit regulates both the SQR and FRD activities of complex II and that the phosphorylation of Fp might be important for maintaining mitochondrial energy metabolism within the tumor microenvironment.

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Figures

Fig. 1.

Fig. 1.

Effects of phosphatase and protein kinase on complex II. A) Effects of phosphatase and protein kinase on SQR and FRD activities in DLD-1 mitochondria. a) SQR and FRD activities after treatment with Antarctic Phosphatase in solubilized DLD-1 mitochondria. b) SQR and FRD activities after treatment with the protein kinase A catalytic subunit in solubilized DLD-1 mitochondria. The open squares show the ratio of SQR activity versus the control value, while the closed circle show the ratio of FRD activity versus the control in the absence of phosphatase/kinase. All data were expressed as the means ± S.D. of three experiments. Statistically significant differences with respect to the control value (p < 0.05) are shown by an asterisk (Student’s _t_-test). B) Separation of Fp proteins. The detection of Fp was performed using an anti-bovine heart Fp monoclonal antibody after two-dimensional gel electrophoresis in solubilized DLD-1 mitochondria. a) Untreated, b) treated with 0.5 Units Antarctic Phosphatase, and c) treated with 0.2 Units protein kinase A.

Fig. 2.

Fig. 2.

Complex II activities in mitochondria from DLD-1 under tumor-mimicking microenvironmental conditions. A) SQR activities, B) FRD activities, and C) ratios of FRD/SQR. The closed circles show the enzyme activities in solubilized mitochondria from DLD-1 cultured under hypoxic and glucose-depleted conditions. The open circles show the enzyme activities in solubilized mitochondria from DLD-1 cultured under hypoxic conditions. The open triangles show the enzyme activities in solubilized mitochondria from DLD-1 cultured under glucose-depleted conditions. All the data were expressed as the means ± S.D. of three experiments. Statistically significant differences with respect to the baseline values (p < 0.05) are shown by an asterisk (Student’s _t_-test).

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