THE M1 AND M2 PROTEINS OF INFLUENZA-A VIRUS ARE IMPORTANT DETERMINANTS IN FILAMENTOUS PARTICLE FORMATION

Influenza A virus is highly pleomorphic with particles exhibiting eith er spherical or filamentous morphology. The mechanisms behind this ple omorphism and its importance in viral pathogenesis are not clearly und erstood. We have observed that budding of filamentous influenza A/Udor n virus particles can be readily visualized by immunofluorescence micr oscopy. Filamentous particle formation was inhibited by treatment of c ells with the anti-M2 14C2 antibody, but was not inhibited with the is otype identical 5C4 anti-M2 antibody or by anti-neuraminidase antibody . To further explore the viral determinants of filamentous particle fo rmation, we investigated the morphology and growth characteristics of three variants of A/Udorn/72 virus, which had previously been selected for their resistance to growth inhibition by the 14C2 anti-M2 monoclo nal antibody. Two of the variant viruses, 5A and 10A, contain single a mino acid substitutions in the cytoplasmic domain of the M2 protein, w hereas the 1A variant contains a single amino acid substitution in the viral matrix protein, M1. Variants 5A and 10A both were found to reta in the filamentous particle phenotype found in the parental strain A/U dorn/72, and the production of filamentous virions by both variants wa s resistant to inhibition by the 14C2 antibody. However, immunofluores cence and electron microscopy revealed that the variant 1A was compose d almost exclusively of spherical particles. The 1A variant displayed higher viral yields and a larger plaque size than the filamentous viru ses. In addition, we separated distinct populations highly enriched in spherical or filamentous particles by velocity gradient centrifugatio n. Analysis of the protein compositions of these particles revealed th at the NP:M1 or NP:HA ratios in filamentous particles were significant ly lower than in spherical particles, but the filaments have higher le vels of NP per particle. The spherical and filamentous particles were found to have similar specific infectivity. These results indicate tha t the filamentous morphology of the A/Udorn virus depends upon the mat rix (M1) and/or M2 proteins. (C) 1998 Academic Press.