Selective regulation of IP3-receptor-mediated Ca2+ signaling and apoptosis by the BH4 domain of Bcl-2 versus Bcl-Xl

Abstract

Antiapoptotic B-cell lymphoma 2 (Bcl-2) targets the inositol 1,4,5-trisphosphate receptor (IP₃R) via its BH4 domain, thereby suppressing IP₃R Ca²⁺-flux properties and protecting against Ca²⁺-dependent apoptosis. Here, we directly compared IP₃R inhibition by BH4-Bcl-2 and BH4-Bcl-Xl. In contrast to BH4-Bcl-2, BH4-Bcl-Xl neither bound the modulatory domain of IP₃R nor inhibited IP₃-induced Ca²⁺ release (IICR) in permeabilized and intact cells. We identified a critical residue in BH4-Bcl-2 (Lys17) not conserved in BH4-Bcl-Xl (Asp11). Changing Lys17 into Asp in BH4-Bcl-2 completely abolished its IP₃R-binding and -inhibitory properties, whereas changing Asp11 into Lys in BH4-Bcl-Xl induced IP₃R binding and inhibition. This difference in IP₃R regulation between BH4-Bcl-2 and BH4-Bcl-Xl controls their antiapoptotic action. Although both BH4-Bcl-2 and BH4-Bcl-Xl had antiapoptotic activity, BH4-Bcl-2 was more potent than BH4-Bcl-Xl. The effect of BH4-Bcl-2, but not of BH4-Bcl-Xl, depended on its binding to IP₃Rs. In agreement with the IP₃R-binding properties, the antiapoptotic activity of BH4-Bcl-2 and BH4-Bcl-Xl was modulated by the Lys/Asp substitutions. Changing Lys17 into Asp in full-length Bcl-2 significantly decreased its binding to the IP₃R, its ability to inhibit IICR and its protection against apoptotic stimuli. A single amino-acid difference between BH4-Bcl-2 and BH4-Bcl-Xl therefore underlies differential regulation of IP₃Rs and Ca²⁺-driven apoptosis by these functional domains. Mutating this residue affects the function of Bcl-2 in Ca²⁺ signaling and apoptosis.