THE PROTEOLYTIC CLEAVAGE OF HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 NEF DOES NOT CORRELATE WITH ITS ABILITY TO STIMULATE VIRION INFECTIVITY

The Nef protein of human immunodeficiency virus type 1 (HIV-1) promote s virion infectivity through mechanisms that are yet ill defined. Some Nef is incorporated into particles, where it is cleaved by the viral protease between amino acids 57 and 58, The functional significance of this event, which liberates the C-terminal core domain of the protein from its membrane-associated N terminus, is unknown. To address this question, we examined the modalities of Nef virion association and pro cessing. We found that although significant levels of Nef were detecte d in HIV-1 virions partly in a cleaved form, cell-specific variations existed in the efficiency of Nef proteolytic processing, The virion as sociation of Nef was strongly enhanced by myristoylation but did not r equire other HIV-1-specific proteins, since Nef was efficiently incorp orated into and cleaved inside murine leukemia virus particles, Substi tuting alanine for tryptophan(57) decreased the efficiency of Nef proc essing, while mutating leucine(58) had little effect, In contrast, rep lacing both of these residue; simultaneously almost completely prevent ed this process, However, when the resulting mutants were compared wit h a wild-type control in viral infectivity assays, no correlation was found between the levels of cleavage and the ability to stimulate viri on infectivity. Furthermore, simian immunodeficiency virus Nef, which locks the sequence recognized by the protease and as a consequence is not cleaved despite its incorporation into virions, could stimulate th e infectivity of a nef-defective HIV-1 variant as efficiently as HIV-1 Nef. On these bases, we conclude that the proteolytic processing of N ef is not required for the ability of this protein to enhance, virion infectivity.