BCL-2 INHIBITS HYDROGEN PEROXIDE-INDUCED ER CA2+ POOL DEPLETION

The mechanism by which Bcl-2 inhibits apoptosis is unknown, The Bcl-2 protein is localized to intracellular membranes, including the endopla smic reticulum (ER), The ER is the major intracellular reservoir of Ca 2+ in non-muscle cells, sequestering Ca2+ for use in intracellular sig naling, and is a prime target of oxidative damage, Because of the rece nt suggestion that Bcl-2 acts in an antioxidant pathway, we wondered w hether Bcl-2 might protect the ER Ca2+ pool in cells exposed to reacti ve oxygen species. To test this hypothesis, we assessed the effect of hydrogen peroxide (H2O2) treatment on the ER Ca2+ pool in WEH17.2 cell s, which do not express Bcl-2, and two stable transfectants, W.Hb13 an d W.Hb12, The Bcl-2 level by Western blotting is 4-fold higher in W.Hb 12 cells compared to W.Hb13 cells, The ER Ca2+ pool in H2O2-treated an d untreated cells was measured according to the amount of Ca2+ mobiliz ed from the ER lumen into the cytoplasm by thapsigargin (TG), a select ive inhibitor of the ER Ca2+-ATPase, H2O2 treatment produced a signifi cant reduction in the TG-mobilizable Ca2+ pool in WEH17.2 and W.Hb13 c ells, but not in W.Hb12 cells, indicating that overexpression of Bcl-2 preserves the integrity of the ER Ca2+ pool in cells exposed to react ive oxygen species.