PERSISTENT INFECTION OF MT-4 CELLS BY HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 BECOMES INCREASINGLY LIKELY WITH IN-VITRO SERIAL PASSAGE OF WILD-TYPE BUT NOT NEF MUTANT VIRUS

Our previous studies have shown that human immunodeficiency virus type 1 (HIV-1), with mutations in accessory genes such as vif; vpr or vpu, can generate persistent infection of MT-4 cells, whereas infection by wild-type or nef mutant HIV-1 causes extensive cell death. The possib ility of generating a naturally attenuated form of HIV-1 with reduced cytopathogenicity in MT-4 cells was examined by in vitro serial passag e of the wild-type and a nef mutant form of HIV-1, each derived from t he infectious molecular clone pNL432. The ability to cause persistent infection was observed after four passages of wild-type HIV-1 with the frequency of persistence becoming progressively higher with serial pa ssage. In contrast, persistent infection was not observed even after 5 0 passages of the nef mutant virus. Sequence analysis of the accessory gene loci in genomes recovered from the persistent infections caused by passaged virus revealed mutations in vif and vpr, but not in vpu. T he processing of the Env precursor to mature forms was not modified in any of the passages of either wild-type or nef mutant HIV-1. However, when compared with acute infections caused by similarly passaged viru s of both wild-type and nef mutant HIV-1, persistent infections by pas saged wild-type HIV-1 showed a significant decrease in the cell surfac e expression and function of Env. Cell surface CD4 was only partially down-regulated on cells acutely infected with the passaged viruses, wh ereas on cells persistently infected with passaged wild-type HIV-1 it was completely downregulated. These results suggest that, during seria l passage of HIV-I, mutations accumulate at least in the accessory gen es vif and vpr in parallel with a lesser interaction between cell surf ace Env and CD4 molecules, and lead to the generation of less cytopath ogenic viruses capable of persistent infection. Our results also sugge st an important role for the nef gene product in the generation of HIV -1 strains that are less cytopathogenic.