1. Fig. 1 HIV transcription profiles in freshly isolated CD4+ T cells and PBMCs from HIV-infected patients on ART suggest blocks to HIV transcriptional elongation, completion, and multiple splicing.
HIV transcription profiles in freshly isolated CD4+ T cells and PBMCs from HIV-infected patients on ART suggest blocks to HIV transcriptional elongation, completion, and multiple splicing. (A to C) Total RNA from unstimulated [day 0 (d0)] CD4+ T cells (A and C) and PBMCs (B) was used for a polyadenylation-RT-ddPCR assay for the TAR loop (found in all HIV transcripts) and RT-ddPCR assays for HIV sequence regions suggesting transcriptional interference (read-through transcripts), transcriptional elongation (long LTR), completion of transcription (polyA), and multiple splicing (Tat-Rev). Each transcript was normalized to 1 μg of cellular RNA (~106 cells) and plotted on a log scale. (C) To determine whether there are blocks to more distal elongation beyond the long LTR region, additional assays were developed for HIV Pol and Nef, and all seven transcripts were measured in six HIV-infected patients on ART. (D and E) To evaluate how deletions or hypermutations in the provirus could affect the measured HIV RNA quantities, HIV DNA was measured using the same ddPCR assays (except Tat-Rev and polyA, which are RNA-specific), and each HIV RNA was normalized to the corresponding HIV DNA (polyA was normalized to the read-through assay, which uses the same forward primer/probe). Bars indicate the median. Comparisons between transcripts were performed using the Wilcoxon signed-rank test.
Steven A. Yukl et al., Sci Transl Med 2018;10:eaap9927
Published by AAAS