PERIPHERAL-BLOOD MONONUCLEAR-CELLS PRODUCE NORMAL AMOUNTS OF DEFECTIVE VIF(-) HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 PARTICLES WHICH ARE RESTRICTED FOR THE PRERETROTRANSCRIPTION STEPS

Previous studies have demonstrated the absence of viral replication of Vif(-) mutants in stimulated primary blood mononuclear cells (PBMC). Human immunodeficiency virus type 1 strain NDK Vif(-) mutants were pro pagated on the semipermissive CEM cell line, and the viral stock obtai ned was compared with the wild-type virus during a single cycle in PBM C. The Vif(-) virus was able to enter PBMC with the same efficiency as the wild type, as demonstrated by quantification of the strong-stop c DNA, and retrotranscription was observed for both viruses within 4 h p ostinfection. Using a PCR assay with an Alu-long terminal repeat pair of primers, we detected integration for both the wild-type and Vif(-) viruses. We then used qualitative and quantitative reverse transcripti on-mediated PCR techniques to study the steady-state level of intracel lular and extracellular viral RNAs. All mRNA species were detected in PBMC infected with the wild-type virus or with the Vif(-) virus 36 h p ostinfection. Furthermore, quantification of viral RNA released from i nfected cells demonstrated similar levels of virus produced after a un ique cycle of replication. However, the Vif(-) virus obtained after on e replication cycle in PBMC was unable to initiate retrotranscription in permissive target tells. These data strongly suggest that the failu re to infect target cells is due to a defect in the formation of the v iral particle in PBMC.