Sarco(endo)plasmic reticulum Ca2+ pump isoform SERCA3 is more resistant than SERCA2b to peroxide
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abstract
Sarco(endo)plasmic reticulum Ca2+ pumps are encoded by genes SERCA1, SERCA2, and SERCA3. Most tissues express SERCA2 Ca2+ pumps (splice SERCA2b) which are inactivated by reactive oxygen. In contrast, SERCA3 is expressed in tissues such as tracheal epithelium, mast cells, lymphoid cells, and aortic endothelium, which are frequently exposed to oxidative stress. Therefore, we compared SERCA3 and SERCA2b proteins for their sensitivity to oxidation. We isolated microsomes from HEK-293 cells overexpressing SERCA3 or SERCA2b. We incubated the microsomes with different concentrations of hydrogen peroxide and then determined Ca2+ pump activities in them in the following three assay systems: ATP-dependent oxalate-stimulated azide-insensitive 45Ca2+ uptake by the microsomal vesicles, Ca(2+)-Mg(2+)-ATPase, and Ca(2+)-dependent acylphosphate formation. Peroxide inhibited the pump activities in microsomes with half-maximal inhibitory concentration (IC50) values of 69 +/- 14, 66 +/- 13, and 84 +/- 15 microM for the 45Ca2+ uptake, Ca(2+)-Mg(2+)-ATPase, and the acylphosphate formation reactions, respectively. However, for microsomes from SERCA3-expressing cells, the corresponding values of IC50 for peroxide were 274 +/- 47, 857 +/- 110, and 746 +/- 40 microM. Thus, in each assay system, the resistance to inactivation by peroxide was significantly (P < 0.05) higher for the SERCA3 protein than for SERCA2b. The SERCA3 resistance to oxidants may aid the cells expressing it to function during exposure to oxidative stress.
