ABSTRACT
The nucleocapsid (NC) domain of the retrovirus Gag protein plays several important roles in the viral life cycle, including virus assembly, viral genomic RNA encapsidation, primer tRNA placement, and enhancement of viral reverse transcription. In this study, deletion of NC domain of human immunodeficiency virus type 1 (HIV-1) Gag was found to drastically reduce virus particle production in CD4+ T cells. Cellular fractionation experiments showed that although most of the uncleaved wild-type HIV-1 Gag, unmyristylated Gag, and p6Gag domain-truncated Gag molecules copurified with the host cell cytoskeleton, most of the mutant Gag molecules lacking both the NC and p6Gag domains failed to cofractionate with cytoskeleton. In wild-type virus-infected cells, in which the viral protease was active, the cleaved NCp7 copurified with the cytoskeleton, whereas most of the MAp17 and CAp24 did not. Monoclonal antibody against actin coimmunoprecipitated full-length Gag and p6Gag domain-truncated Gag molecules from cell lysates but failed to precipitate the truncated mutant Gag molecules lacking NC plus p6Gag. Purified recombinant NCp7, but not CAp24, was able to bind F-actin in cosedimentation experiments. Furthermore, wild-type NCp7 and a zinc finger mutant NCp7(F16A), like a cellular actin-binding protein (the villin headpiece), bound F-actin in a dose-dependent fashion in vitro. Taken together, these results suggest that HIV-1 NCp7 can bind F-actin directly and that interaction between HIV-1 Gag and the actin cytoskeleton through the NC domain may play an important role in HIV-1 assembly and/or other steps of the viral life cycle.
Cyclophilin A Plays Distinct Roles in Human Immunodeficiency Virus Type 1 Entry and Postentry Events, as Revealed by Spinoculation
