Structural biology of the Bcl-2 family and its mimicry by viral proteins

Abstract

Intrinsic apoptosis in mammals is regulated by protein-protein interactions among the B-cell lymphoma-2 (Bcl-2) family. The sequences, structures and binding specificity between pro-survival Bcl-2 proteins and their pro-apoptotic Bcl-2 homology 3 motif only (BH3-only) protein antagonists are now well understood. In contrast, our understanding of the mode of action of Bax and Bak, the two necessary proteins for apoptosis is incomplete. Bax and Bak are isostructural with pro-survival Bcl-2 proteins and also interact with BH3-only proteins, albeit weakly. Two sites have been identified; the in-groove interaction analogous to the pro-survival BH3-only interaction and a site on the opposite molecular face. Interaction of Bax or Bak with activator BH3-only proteins and mitochondrial membranes triggers a series of ill-defined conformational changes initiating their oligomerization and mitochondrial outer membrane permeabilization. Many actions of the mammalian pro-survival Bcl-2 family are mimicked by viruses. By expressing proteins mimicking mammalian pro-survival Bcl-2 family proteins, viruses neutralize death-inducing members of the Bcl-2 family and evade host cell apoptosis during replication. Remarkably, structural elements are preserved in viral Bcl-2 proteins even though there is in many cases little discernible sequence conservation with their mammalian counterparts. Some viral Bcl-2 proteins are dimeric, but they have distinct structures to those observed for mammalian Bcl-2 proteins. Furthermore, viral Bcl-2 proteins modulate innate immune responses regulated by NF-κB through an interface separate from the canonical BH3-binding groove. Our increasing structural understanding of the viral Bcl-2 proteins is leading to new insights in the cellular Bcl-2 network by exploring potential alternate functional modes in the cellular context. We compare the cellular and viral Bcl-2 proteins and discuss how alterations in their structure, sequence and binding specificity lead to differences in behavior, and together with the intrinsic structural plasticity in the Bcl-2 fold enable exquisite control over critical cellular signaling pathways.

Figure 1

Points of intervention of viral Bcl-2 proteins in mammalian Bcl-2-regulated apoptosis. Shown is a simplified schematic view of apoptosis induction in mammalian cells and its inhibition by pro-survival viral Bcl-2 (vBcl-2) proteins. Apoptosis may be initiated by activation of cell surface receptors (extrinsic apoptosis or death receptor-initiated apoptosis) or by intracellular mechanisms (‘intrinsic' or mitochondrial pathway). In either case, a proteolytic cascade of caspases is initiated that destroys the cell. A network of interactions between Bcl-2 proteins controls mitochondrial membrane integrity. BH3-only proteins may either inhibit pro-survival proteins or activate the two key Bcl-2 family members, Bax and Bak, to disrupt the mitochondrial outer membrane and release caspase-activating factors. Viral Bcl-2 proteins bind Bax or Bak or BH3-only proteins to prevent apoptosis induction. cBcl-2, cellular Bcl-2

Figure 2

Structural comparison of human and viral Bcl-2 proteins and their complexes with Bak BH3 peptides. (a) Human Bcl-x_L (pdb accession code: 1MAZ) with the BH-motifs colored BH1 purple; BH2, red; BH3, green. (b) Epstein-Barr virus BHRF1 (1Q59), (c) Myxoma virus M11L (2BJX), (d) Vaccinia virus F1L (2VTY), (e) Bcl-x_L:Bak (1BXL), (f) BHRF1:Bak (2XPX), (g) M11L:Bak (2JBY), (h) Vaccinia virus N1L (2UXE). The Bak BH3 peptide is shown in orange. Structures were aligned on Bcl-x_L in a. Protomers of the F1L and N1L dimers are depicted in different colors

Figure 3

Structures of Bcl-2 dimers. Dimers may be induced by treatment with heat, pH, surfactant or BH3-ligand. (a and b) Bcl-x_L dimers; (c and d) Bax dimers. (a) Bcl-x_L dimer (2B48). The BH motifs are indicated in color as Figure 1 and the helices indicated with the alternate protomer helices denoted with a prime (′) (b) Beclin-1: Bcl-x_L heterotetramer (2P1L). (c) Octylmaltoside-induced Bax dimer (4BD7) (c) CHAPS-induced alternate dimeric Bax core (4BDU) that contains only the central helices (α2–α5) of Bax and lacks both N-terminal residues and the C-terminal TM region. For clarity, the GFP-fusion tag on each chain is not shown. BH motifs have been colored as in Figure 1. The structures depicted in a and c have been structurally aligned with Bcl-x_L in the same orientation as Figure 1 and the Bcl-x_L dimer in b was aligned over helices α5–α8 on the structure in a