ROLE OF ACTIN MICROFILAMENTS IN BLACK CREEK CANAL VIRUS MORPHOGENESIS

We have investigated the involvement of cytoskeletal proteins in the m orphogenesis of Black Creek Canal virus (BCCV), a New World hantavirus . Immunofluorescent staining of BCCV-infected cells revealed a filamen tous pattern of virus antigen, the appearance of which was sensitive t o treatment with cytochalasin D, an actin microfilament-depolymerizing drug. Double immunofluorescence staining of BCCV-infected Vero cells with anti-BCCV nucleocapsid (N) monoclonal antibody and phalloidin rev ealed a colocalization of the BCCV N protein with actin microfilaments . A similar, though less prominent, filamentous pattern was observed i n BHK21 cells transiently expressing the BCCV N protein alone but not in cells expressing the BCCV G1 and G2 glycoproteins. Moreover, the as sociation of the N protein with actin microfilaments,vas confirmed by coimmunoprecipitation with beta-actin-specific antibody. Treatment of the BCCV-infected Vero cells at 3 days postinfection with cytochalasin D decreased the yield of released BCCV by 94% relative to the yield f rom untreated cells. Pretreatment of Vero cells with cytochalasin D pr ior to and during BCCV adsorption and entry had no effect on the outco me of virus production. These results indicate that actin filaments ma y play an important role in hantavirus assembly and/or release.