Analysis of the role of conserved cysteine residues in the Bcl-2 oncoprotein

The Bcl-2 oncoprotein is an integral membrane protein localized primarily t o the outer membrane of the mitochondria. The precise molecular mechanism r esponsible for the antiapoptotic action of Bcl-2 remains unknown. Two cyste ine residues are found in Bcl-2 and these residues are well-conserved acros s species. The first cysteine (cys(155)) is located in the alpha5 domain, a region important for the ion channel properties of Bcl-2, while the second cysteine (cys(226)) is located in the carboxyl-terminal membrane anchor do main. In this study, we found that replacement of both cysteines with serin e residues generated a mutant protein that retained the ability to homodime rize and heterodimerize with proapoptotic Bar protein in vitro. In whole ce lls, the mutant protein efficiently heterodimerized with Bar, but exhibited impaired homodimerization relative to wild-type Bcl-2, The mutant protein was also less efficient than wild-type Bcl-2 at suppressing caspase activat ion, DNA fragmentation, and loss of viability during IL-3 withdrawal-induce d apoptosis, Together, the data indicate that the cysteine residues in Bcl- 2 contribute, but are not absolutely essential, to the ability of Bcl-2 to homodimerize, heterodimerize with Bar, and suppress apoptosis, (C) 2000 Aca demic Press.