The Ca2+ concentration of the endoplasmic reticulum is a key determinant of ceramide-induced apoptosis:: significance for the molecular mechanism of Bcl-2 action

The mechanism of action of the anti-apoptotic oncogene Bcl-2 is still largely obscure. We have recently shown that the overexpression of Bcl-2 in HeLa cells reduces the Ca2+ concentration in the endoplasmic reticulum ([Ca2+](er)) by increasing the passive Ca2+ leak from the organelle, To investigate whether this Ca2+ depletion is part of the mechanism of action of Bcl-2, we mimicked the Bcl-2 effect on [Ca2+](er) by different pharmacological and molecular approaches. All conditions that lowered [Ca2+](er) protected HeLa cells from ceramide, a Bcl-2-sensitive apoptotic stimulus, while treatments that increased [Ca2+](er) had the opposite effect. Surprisingly, ceramide itself caused the release of Ca2+ from the endoplasmic reticulum and thus [Ca2+] increased both in the cytosol and in the mitochondrial matrix, paralleled by marked alterations in mitochondria morphology, The reduction of [Ca2+](er) levels, as well as the buffering of cytoplasmic [Ca2+] changes, prevented mitochondrial damage and protected cells from apoptosis, It is therefore concluded that the Bcl-2-dependent reduction of [Ca2+](er) is an important component of the anti-apoptotic program controlled by this oncogene.