| Following DNA damage, nuclear p53 induces the expression of PUMA (p53 upregulated modulator of apoptosis), a BH3-only protein that binds and inhibits the anti-apoptotic BCL-2 repertoire, including BCL-xL. Structural investigations of PUMA and the BCL-xL˙PUMA BH3 domain complex by X-ray crystallography and nuclear magnetic resonance (NMR) spectroscopy reveal a novel, PUMA-induced, domain-swapped dimerization of BCL-xL that requires a pi-stacking interaction between PUMA W71 and BCL-xL H113. PUMA is an intrinsically disordered protein, but upon interaction with BCL-xL, PUMA W71 and the PUMA BH3 domain residues fold into an alpha helix and subtly remodel BCL-xL to trigger its dimerization. Wild type PUMA or a PUMA mutant incapable of promoting BCL-xL dimerization (PUMA W71A) equivalently inhibit the anti-apoptotic BCL-2 repertoire to sensitize for death receptor-activated apoptosis, but only wild type PUMA promotes p53-dependent, DNA damage-induced apoptosis. Biochemical and cellular data demonstrate that PUMA-mediated structural remodeling and dimerization of BCL-xL modulates its affinity for cytosolic p53, providing a detailed mechanism of BCL-xL, cytosolic p53, and PUMA functional cooperation. Our data suggest that within the BCL-2 family, ligand binding-induced, domain-swapped dimerization is a critical control point to increase signal transduction complexity within the apoptotic pathways. |
