Scientific Publications by FDA Staff

J Virol 2015 Feb;89(4):1975-85

Influenza M2 protein ion channel activity helps maintain the pandemic 2009 H1N1 hemagglutinin fusion competence during transport to the cell surface.

Alvarado-Facundo E, Gao Y, Ribas-Aparicio RM, Jimenez-Alberto A, Weiss CD, Wang W

Influenza virus hemagglutinin (HA) envelope protein mediates virus entry by first binding to cell surface receptors, then fusing viral and endosomal membranes during endocytosis. Cleavage of the HA precursor (HA0) into a surface receptor-binding subunit (HA1) and a fusion-inducing transmembrane subunit (HA2) by host cell enzymes primes HA for fusion competence by repositioning the fusion peptide to the newly created the N-terminus of HA2. We previously reported that influenza M2 protein enhances pandemic 2009 influenza A (H1N1) (pdm09) HA-pseudovirus infectivity, but the mechanism was unclear. Here, using cell-cell fusion and HA-pseudovirus infectivity assays, we found that the ion channel function of M2 was required for enhancing HA fusion and HA-pseudovirus infectivity. The M2 activity was only needed during HA biosynthesis, and proteolysis experiments indicate that M2 proton channel activity helps to protect (H1N1)pdm09 HA from premature conformational changes as it traversed low pH compartments during transport to the cell surface. While M2 has previously been shown to protect avian influenza from H5 and H7 subtypes that have HA with polybasic cleavage motifs, this study demonstrates that M2 can protect HA from human H1N1 strains that lack a polybasic cleavage motif. This finding suggests that M2 proton channel activity may play a wider role in preserving HA fusion competence among a variety of HA subtypes, including HA from emerging strains that may have reduced HA stability. IMPORTANCE: Influenza infects cells when the hemagglutinin surface protein (HA) undergoes irreversible pH-induced conformational changes after the virus is taken into the cell by endocytosis. HA fusion competence is primed when host cells enzymes cleave the HA precursor. The proton channel function of influenza M2 protein has previously been shown to protect avian influenza HA that contain a polybasic cleavage site from pH-induced conformation changes during biosynthesis, but this effect is less well understood for human influenza HA that lack polybasic cleavage sites. Using assays that focus on HA entry and fusion, we found that the M2 protein also protects the (H1N1)pdm09 influenza A virus HA from premature conformational changes as it transits low-pH compartments during biosynthesis. This work suggests that M2 may play a wider role in preserving HA function in a variety of influenza subtypes that infect humans and may be especially important for HA that are less stable.