Permeation and activation of the M-2 ion channel of influenza A virus

The M-2 ion channel protein of influenza A virus is essential for mediating protein-protein dissociation during the virus uncoating process that occur s when the virus is in the acidic environment of the lumen of the secondary endosome. The difficulty of determining the ion selectivity of this minima listic ion channel is due in part to the fact that the channel activity is so great that it causes local acidification in the expressing cells and a c onsequent alteration of reversal voltage, V-rev. We have confirmed the high proton selectivity of the channel (1.5-2.0 x 10(6)) in both oocytes and ma mmalian cells by using four methods as follows: 1) comparison of V-rev with proton equilibrium potential; 2) measurement of pH(in) and V-rev while Na- out(+) was replaced; 3) measurements with limiting external buffer concentr ation to limit proton currents specifically; and 4) comparison of measureme nts of M-2-expressing cells with cells exposed to a protonophore. Increased currents at low pH(out) are due to true activation and not merely increase d [H+](out) because increased pH(out) stops the outward current of acidifie d cells. Although the proton conductance is the biologically relevant condu ctance in an influenza virus-infected cell, experiments employing methods 1 -3 show that the channel is also capable of conducting NH4+, probably by a different mechanism from H+.