The ESCRT pathway facilitates membrane fission events during enveloped virus budding, multivesicular body formation, and cytokinesis. To promote HIV budding and cytokinesis, the ALIX protein must bind and recruit CHMP4 subunits of the ESCRT-III complex, which in turn participate in essential membrane remodeling functions. This collaborative study from the Hill and Sundquist labs reports that the Bro1 domain of ALIX binds specifically to the C-termini of the human CHMP4 proteins (CHMP4A-C). Crystal structures of the three ALIXBro1-CHMP4 complexes revealed that the CHMP4 C-termini form amphipathic helices that bind across the conserved concave surface of ALIXBro1. ALIX-dependent HIV-1 budding is blocked by mutations in exposed ALIXBro1 residues that help form the binding sites for three essential hydrophobic residues displayed on one side of the CHMP4 recognition helix (M/L/IxxLxxW). The homologous CHMP1-3 classes of ESCRT-III proteins also have C-terminal amphipathic helices, but in those cases the three hydrophobic residues are arrayed with L/I/MxxxLxxxL spacing. Thus, the distinct patterns of hydrophobic residues provide a “code” that allows the different ESCRT-III subunits to bind different ESCRT pathway partners, with CHMP1-3 proteins binding MIT domain-containing proteins like VPS4 and Vta1/LIP5, and CHMP4 proteins binding Bro1 domain-containing proteins like ALIX.

Structures of the C-terminal helices from different human CHMP4 proteins in complex with the Bro1 domain of ALIX.