Intergenic HA-NA interactions in influenza A virus: postreassortment substitutions of charged amino acid in the hemagglutinin of different subtypes

In our previous studies influenza A virus reassortants having neuraminidase (NA) gene of A/USSR/90/77 (H1N1) strain and hemagglutinin (HA) genes of H3 , H4 and H13 subtypes were shown to produce a low virus yield and to exhibi t a strong tendency to virion aggregation. More detailed studies with the u se of a H3N1 reassortant and its high-yield non-aggregating variants reveal ed that NA of A/USSR/90/77 strain is inefficient in the removal of the term inal sialic acid residues from the virion components, and that the ineffici ency of NA may be compensated by mutations in HA gene leading to a decrease of the receptor-binding affinity (Kaverin, N.V., Gambaryan, A.S., Bovin, N .V., Rudneva, I.A., Shilov, A.A., Khodova, O.M., Varich, N.L., Sinitsin, B. V., Makarova, N.L., Kaverin, N.V., 1998. Postreassortment changes in influe nza virus hemagglutinin restoring HA-NA functional match, Virology 244, 315 -321). The present report describes studies performed with the use of H2N1 and H4N1 reassortants having HA genes of A/Pintail/Primorie/695/76 (H2N3) a nd A/Duck/Czechoslovakia/56 (H4N6) strains respectively and NA gene of A/US SR/90/77 strain. The low-yield reassortants and their high-yield non-aggreg ating variants were studied in both direct and competitive binding assays w ith sialic acid-containing substrates. The non-aggregating variants were sh own to have a decreased affinity as compared to the initial reassortants to ward high-molecular-weight sialic acid-containing substrates. The sequencin g of HA genes revealed that all non-aggregating variants of H2N1 and H4N1 r eassortants had amino acid substitutions increasing the negative charge of the HA molecule in the vicinity of the receptor-binding pocket. The results suggest that the influenza virus reassortants containing low-functional NA undergo similar postreassortment changes irrespective of the HA subtype: t heir receptor-binding activity decreased due to negatively charged amino ac id substitutions in the vicinity of the receptor-binding pocket. (C) 2000 E lsevier Science B.V. All rights reserved.