Direct Measurement of the Influenza A Virus M2 Protein Ion Channel Activity in Mammalian Cells

Abstract The influenza A virus M 2 integral membrane protein has an ion channel activity which is thought to play an essential role in the uncoating process of influenza virus in infected cells and, for some strains of influenza virus, in maintaining the hemagglutinin in its pH neutral form during transport through the trans Golgi network. To demonstrate directly that the M 2 protein forms an ion channel in mammalian cells, the M 2 protein was expressed in CV-1 cells by using an SV40-M 2 recombinant virus and the whole cell membrane currents were recorded. It was found that the whole cell current was activated by low pH and inhibited by the M 2 ion channel-specific blocker, amantadine hydrochloride. Expression of an altered M 2 protein that contains a deletion of four residues in the transmembrane domain (M 2 -del 28-31 ) and that when found in influenza virus confers amantadine resistance, resulted in a current that was activated by hyperpolarization of the membrane, was pH insensitive, and was resistant to block by amantadine. The data obtained in mammalian cells for the wild-type M 2 and M 2 -del 28-31 protein ion channel activities were very similar to those obtained when using the heterologous oocyte expression system.