Conservation of an intact human immunodeficiency virus type 1 vif gene in vitro and in vivo

Abstract

Replication of vif-negative human immunodeficiency virus type 1 (HIV-1) is attenuated in certain cell lines and highly impaired in peripheral blood lymphocytes in vitro. To determine whether intact vif is positively selected during natural HIV-1 infection and to determine vif sequence variability, we employed PCR amplification, cloning, and sequencing to investigate the vif region of replicating virus in short-term-passage HIV-1 primary isolates from five asymptomatic individuals and from five persons with AIDS. A total of 46 vif clones were obtained and analyzed. Recombinant proviruses were constructed from selected vif clones from one patient and found to be fully infectious. We found that 38 of the 46 clones sequenced carried open vif reading frames and that there was a low degree of heterogeneity of vif genes within isolates from the same individual and among isolates from different donors. The cysteines previously found to be essential for vif protein function were conserved in all clones. A phylogenetic tree constructed from all available vif nucleotide sequences resulted in a virus grouping similar to those of gag and env. Direct sequencing of vif amplified by PCR from uncultured lymphocytes of 15 individuals at various stages of progression toward AIDS demonstrated vif open reading frames in 13 of 15 samples tested. There was no obvious correlation between disease status and the presence of an intact vif within this sample group at the time of sample procurement. The conservation of the vif open reading frame in vitro and in vivo and its limited variability following virus transmission in vitro are consistent with a role for vif in natural HIV-1 infection.

Characterization of Human Immunodeficiency Virus Type 1 vif Gene in Long-Term Asymptomatic Individuals

“…b Zhang LTNP correspond to nine LTNP described by Zhang et al (1997b). Progressors correspond to the eight late progressors of the present study and to seven AIDS progressors from Sova et al (1995) (see Fig. 1).…”

“…We report here our results on vif gene sequences from 42 LTNP untreated with anti-retroviral therapy (referred to here as the ALT cohort; (Candotti et al, 1999). This auxiliary gene is required for HIV-1 replication in primary lymphocytes and macrophages (Chowdhury et al, 1996;Fisher et al, 1987;Gabuzda et al, 1992Gabuzda et al, , 1994Sakai et al, 1988;Sova and Volsky, 1993;von Schwedler et al, 1993) and consequently is largely maintained intact in vivo in adults (Sova et al, 1995;Wieland et al, 1994;Zhang et al, 1997a) and in perinatally infected infants (Yedavalli et al, 1998). Our goal was to determine whether defined amino acid residues at particular positions of the predicted vif gene product were related to the viral load parameter among LTNP individuals.…”

“…Figure 1 shows the alignment of Vif amino acid sequences deduced from the nucleotide sequences. In addition to the Vif sequences from the eight control progressors of our cohort, seven Vif sequences, which were obtained by amplification of PBMC DNA from AIDS patients and were published by Sova et al (1995), were also aligned (Vif GMK1 to 4, VifGMK6 to 8). The major amino acid sequence of all subjects was found to be full-length, i.e., 192 amino acids long except for one subject (11007) who presents a Vif protein with 193 amino acids with an additional D at position 62.…”

“…An additional number 1 or 2 denotes the time of analysis after enrollment in the ALT cohort: 1, for 1 year; 2, for 2 years (Candotti et al, 1999). Sequences of AIDS progressors from the study of Sova et al (1995), which were obtained by PCR amplification from ex vivo PBMC, were added as reference sequences of progressors. Dots indicate amino acids identical to those of the YU-2 sequence (Li et al, 1992); dashes represent gaps.…”

vif-negative human immunodeficiency virus type 1 persistently replicates in primary macrophages, producing attenuated progeny virus

“…Recent studies (1,3,11,17,24,28,45) including our own (36), have refined these definitions by investigating the virus life cycle in primary lymphocytes and macrophages and have revealed a greater role for accessory genes than was first proposed. On the basis of the degree of HIV-1 attenuation by ablation of vif estimated here (3 to 4 orders of magnitude), the major role of vif in HIV-1 replication in both macrophages and lymphocytes in culture, and its maintenance in lentiviruses (30) and in HIV-1-infected persons (39), we propose that vif is a critical determinant of HIV-1 virulence during natural infection of humans. Consistent with this proposition, a long-term survivor of HIV-1 infection was shown recently to harbor virus carrying multiple mutations in accessory genes, including vif (27), and one of these mutations was the missense cysteine replacement described first by this laboratory (26).…”