Differential regulatory activities of viral protein X for anti-viral efficacy of nucleos(t)ide reverse transcriptase inhibitors in monocyte-derived macrophages and activated CD4(+) T cells

Abstract

Vpx encoded by HIV-2 and SIVsm enhances retroviral reverse transcription in macrophages in vitro by mediating the degradation of the host SAMHD1 protein that hydrolyzes dNTPs and by elevating cellular dNTP levels. Here we employed RT-SHIV constructs (SIV encoding HIV-1 RT) to investigate the contribution of Vpx to the potency of NRTIs, which compete against dNTPs, in monocyte-derived macrophages (MDMs) and activated CD4(+) T cells. Relative to HIV-1, both SIV and RT-SHIV exhibited reduced sensitivities to AZT, 3TC and TDF in MDMs but not in activated CD4(+) T cells. However, when SIV and RT-SHIV constructs not coding for Vpx were utilized, we observed greater sensitivities to all NRTIs tested using activated CD4(+) T cells relative to the Vpx-coding counterparts. This latter phenomenon was observed for AZT only when using MDMs. Our data suggest that Vpx in RT-SHIVs may underestimate the antiviral efficacy of NRTIs in a cell type dependent manner.

Keywords: Antiretrovirals; HIV-1; RT-SHIV; SAMHD1; SIV; Vpx.

Fig. 1

Dose–response of NRTIs in CD4⁺ Tcells challenged with VSV-G-pseudotyped HIV-1, SIV and RT-SHIV. Percent inhibition (y-axis) of CD4⁺ Tcells treated with increasing concentrations (x-axis) of AZT (A), 3TC (B), TDF (C) ddC (D), RAL (E), and ETR (F), and then challenged with VSV-G-pseudotyped HIV-1 (filled diamonds), SIV (filled squares) and RT-SHIV (filled circles). Dashed line transects curve to show where 50% inhibition occurs. Data points depict the means and standard error of the means from three to four independent experiments.

Fig. 2

Dose–response of NRTIs in activated CD4⁺ T cells challenged with molecular clones that code, or do not code, for Vpx. Percent inhibition (y-axis) of CD4⁺ T cells treated with increasing concentrations (x-axis) of AZT (A), 3TC (B), TDF (C), RAL (D) and ETR (E) before challenge with SIV (filled squares), SIVΔVpx (open squares), RT-SHIV (filled circles) and RT-SHIVΔVpx (open circles). Dashed line transects curve to show where 50% inhibition occurs. Significant differences in dose–response curves are indicated with an asterisk (*). Data points depict the means and standard error of the means from three to four independent experiments.

Fig. 3

Dose–response of ARVs in MDMs challenged with VSV-G-pseudotyped HIV-1, SIV and RT-SHIV expressing GFP in vitro. Percent inhibition (y-axis) of MDMs treated with increasing concentrations (x-axis) of AZT (A), 3TC (B), TDF (C) ddC (D), RAL (E), and ETR (F), and then challenged with VSV-G-pseudotyped HIV-1 (filled diamonds), SIV (filled squares) and RT-SHIV (filled circles). Dashed line transects curve to show where 50% inhibition occurs. Significant differences in dose–response curves are indicated with an asterisk (*). Data points depict the means and standard error of the means from four to five independent experiments.

Fig. 4

Dose–response of NRTIs in MDMs challenged with molecular clones that code, or do not code, for Vpx. Percent inhibition (y-axis) of MDMs treated with increasing concentrations (x-axis) of AZT (A), 3TC (B), TDF (C), RAL (D) and ETR (E) before challenge with SIV (filled squares), SIVΔVpx (open squares), RT-SHIV (filled circles) and RT-SHIVΔVpx (open circles). Dashed line transects curve to show where 50% inhibition occurs. Significant differences in dose–response curves are indicated with an asterisk (*). Data points depict the means and standard error of the means from four to five independent experiments.