Cholesterol binding to the transmembrane region of a group 2 HA of Influenza virus is essential for virus replication affecting both virus assembly and HA’s fusion activity

Abstract

Hemagglutinin (HA) of Influenza virus is incorporated into cholesterol enriched, 1 nanodomains of the plasma membrane. Phylogenetic group 2 HAs contain the 2 conserved cholesterol consensus motif (CCM) YKLW in the transmembrane region. 3 We previously reported that mutations in the CCM retarded intracellular transport of 4 HA and decreased its nanodomain association. Here we analyzed whether cholesterol 5 interacts with the CCM. Incorporation of photocholesterol into HA was significantly 6 reduced if the whole CCM is replaced by alanine, both using immunoprecipitated HA 7 and when HA is embedded in the membrane. Next, we used reverse genetics to 8 investigate the significance of the CCM for virus replication. No virus was rescued if 9 the whole motif is exchanged (YKLW4A); single (LA) or double (YK2A and LW2A)