Evidence that BCL-2 represses apoptosis by regulating endoplasmic reticulum-associated Ca2+ fluxes.
BCL-2 is a 26-kDa integral membrane protein that represses apoptosis by an unknown mechanism. Recent findings indicate that Ca2+ release from the endoplasmic reticulum (ER) mediates apoptosis in mouse lymphoma cells. In view of growing evidence that BCL-2 localizes to the ER, as well as mitochondria and the perinuclear membrane, we investigated the possibility that BCL-2 represses apoptosis by regulating Ca2+ fluxes through the ER membrane. A cDNA encoding BCL-2 was introduced into WEHI7.2 cells and two subclones, W.Hb12 and W.Hb13, which express high and low levels of BCL-2 mRNA and protein, respectively, were isolated. WEHI7.2 cells underwent apoptosis in response to treatment with the glucocorticoid hormone dexamethasone, whereas W.Hb12 and W.Hb13 cells were protected from apoptosis, revealing a direct relationship between the level of BCL-2 expression and the degree of protection. Significantly, BCL-2 also blocked induction of apoptosis by thapsigargin (TG), a highly specific inhibitor of the ER-associated Ca2+ pump. TG completely inhibited ER Ca2+ pumping in both WEHI7.2 and W.Hb12 cells, but the release of Ca2+ into the cytosol after inhibition of ER Ca2+ pumping was significantly less in W.Hb12 cells than in WEHI7.2 cells, indicating that BCL-2 reduces Ca2+ efflux through the ER membrane. By reducing ER Ca2+ efflux, BCL-2 interfered with a signal for "capacitative" entry of extracellular Ca2+, preventing a sustained increase of cytosolic Ca2+ in TG-treated cells. These findings suggest that BCL-2 either directly or indirectly regulates the flux of Ca2+ across the ER membrane, thereby abrogating Ca2+ signaling of apoptosis.
Lam
M
M
Department of Medicine, Case Western Reserve University, Cleveland, OH 44106.
Dubyak
G
G
Chen
L
L
Nuñez
G
G
Miesfeld
R L
RL
Distelhorst
C W
CW
D017209:Apoptosis / Q000502:physiology*; D002118:Calcium / Q000378:metabolism*; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D002460:Cell Line; D002463:Cell Membrane Permeability; D002470:Cell Survival / Q000187:drug effects; D002999:Clone Cells; D003907:Dexamethasone / Q000494:pharmacology; D004072:Digitonin; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D011518:Proto-Oncogene Proteins / Q000096:biosynthesis / Q000378:metabolism*; D019253:Proto-Oncogene Proteins c-bcl-2; D011519:Proto-Oncogenes; D012333:RNA, Messenger / Q000096:biosynthesis*; D013729:Terpenes / Q000494:pharmacology; D019284:Thapsigargin; D013997:Time Factors; D014162:Transfection
D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger; D013729:Terpenes; D019284:Thapsigargin; D003907:Dexamethasone; D019204:GTP-Binding Proteins; D000252:Calcium-Transporting ATPases; D004072:Digitonin; D002118:Calcium