title: The influenza virus ion channel and maturation cofactor M2 is a cholesterol-binding protein.
abstract: The influenza-virus M2 protein has proton channel activity required for virus uncoating and maturation of hemagglutinin (HA) through low-pH compartments. The proton channel is cytotoxic in heterologous expression systems and can be blocked with rimantadine. In an independent, rimantadine-resistant function, M2, interacting with the M1 protein, controls the shape of virus particles. These bud from cholesterol-rich membrane rafts where viral glycoproteins and matrix (M1)/RNP complexes assemble. We demonstrate that M2 preparations from influenza virus-infected cells and from a baculovirus expression system contain 0.5-0.9 molecules of cholesterol per monomer. Sequence analyses of the membrane-proximal M2 endodomain reveal interfacial hydrophobicity, a cholesterol-binding motif first identified in peripheral benzodiazepine receptor and human immunodeficiency virus gp41, and an overlapping phosphatidylinositol 4,5-bisphosphate-binding motif. M2 induced rimantadine-reversible cytotoxicity in intrinsically cholesterol-free E. coli, and purified E. coli-expressed M2 functionally reconstituted into cholesterol-free liposomes supported rimantadine-sensitive proton translocation. Therefore, cholesterol was nonessential for M2 ion-channel function and cytotoxicity and for the effect of rimantadine. Only about 5-8% of both M2 preparations, regardless of cholesterol content, associated with detergent-resistant membranes. Cholesterol affinity and palmitoylation, in combination with a short transmembrane segment suggest M2 is a peripheral raft protein. Preference for the raft/non-raft interface may determine colocalization with HA during apical transport, the low level of M2 incorporated into the viral envelope and its undisclosed role in virus budding for which a model is presented. M2 may promote clustering and merger of rafts and the pinching-off (fission) of virus particles.
authors.affiliation: Abteilung Virologie, Institut für Mikrobiologie und Hygiene, Universität des Saarlandes, Homburg/Saar, 66421 Homburg, Germany.
authors.forename: Cornelia
authors.initials: C
authors.lastname: Schroeder
authors.forename: Harald
authors.initials: H
authors.lastname: Heider
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Möncke-Buchner
authors.forename: Tse-I
authors.initials: TI
authors.lastname: Lin
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D002784:Cholesterol / Q000737:chemistry* / Q000378:metabolism*; D009980:Influenza A virus / Q000378:metabolism*; D007313:Insecta / Q000821:virology; D007473:Ion Channels / Q000502:physiology*; D008954:Models, Biological; D008956:Models, Chemical; D011485:Protein Binding; D014763:Viral Matrix Proteins / Q000737:chemistry* / Q000378:metabolism*; D000086082:Viroporin Proteins
chemical_list: D002784:Cholesterol; D007473:Ion Channels; D014763:Viral Matrix Proteins; C488937:M2 protein, Influenza A virus; D000086082:Viroporin Proteins