Abstract
Recent studies have demonstrated that the anti-apoptotic proteins, Bcl-2 and Bcl-xl, with the carboxyl-terminal hydrophobic domain removed, form cation-selective channels in the lipid bilayer reconstitution system. However, the regulatory properties of these channels are unknown. In this study, we investigated the ion-conducting properties of full-length Bcl-xl in the lipid bilayer reconstitution system. Our findings indicate that Bcl-xl forms a cation-selective channel that conducts sodium but not calcium and that Bcl-xl channel activity is reversibly inhibited by luminal calcium with a half-dissociation constant of approximately 60 microM. This calcium-dependent regulation of the Bcl-xl channel provides new insights into the roles of calcium and Bcl-2-related proteins in the programmed cell death pathway.
Highlights
Recent studies have demonstrated that the anti-apoptotic proteins, Bcl-2 and Bcl-xl, with the carboxyl-terminal hydrophobic domain removed, form cation-selective channels in the lipid bilayer reconstitution system
In light of our recent evidence that depletion of cellular calcium abrogates the anti-apoptotic effect of Bcl-2 [16], we set out to examine whether calcium is able to regulate the channel activity of anti-apoptotic Bcl-2 family members in a lipid bilayer reconstitution system
To characterize the single channel function of full-length Bcl-xl, purified protein was reconstituted in the lipid bilayer, and the selected current traces shown in Fig. 2 were recorded with 200 mM NaCl present in the cis-solution and 50 mM NaCl present in the trans-solution
Summary
Recent studies have demonstrated that the anti-apoptotic proteins, Bcl-2 and Bcl-xl, with the carboxyl-terminal hydrophobic domain removed, form cation-selective channels in the lipid bilayer reconstitution system. Bcl-xl, Bcl-2 and Bax, have been shown to form ion channels using the lipid bilayer reconstitution system (18 –21) These studies all utilized proteins from which the carboxyl-terminal hydrophobic domain had been deleted. It seems paradoxical that Bcl-2 and Bcl-xl, which function to inhibit apoptosis, should structurally and functionally resemble bacterial toxins whose main purpose is to form pores in cell membranes and destroy cells by disrupting ion homeostasis This suggests that the ability of Bcl-2 and Bcl-xl to promote cell survival may not derive solely from their ability to form a channel. We observed that Bcl-xl channel activity is inhibited by calcium, indicating for the first time that Bcl-xl forms a calcium-regulated cation channel
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