Fig. 2
Top1 shRNA silencing or CRISPR/Cas9 depletion induces a reactivation of HIV-1 LTR promoter activity in J-Lat cells. A–C J-Lat A1 and J-Lat 10.6 cells were transduced with pLKO.1 vectors expressing a shRNA directed against Top1, Top2A or Top2B genes or a scrambled shRNA sequence (sh-) (transduction ratio of 300 µL vector / 2 × 106 cells). The % of GFP positive cells (n = 6 for J-Lat A1 and n = 3 for J-Lat 10.6) (A, B) or the level of mRNA coding for GFP (n = 3 for both cell lines) (C) were quantified 7 days post-transduction. D–F J-Lat A1 cells were transfected by a LentiCRISPRV2 plasmid expressing guide RNAs directed against Top1, Top2A or Top2B genes. The percentage of GFP-positive cells was measured after 10 days of puromycin selection (n = 9 for WT, n = 5 for Top1 bulk and n = 3 for TopA and Top2B bulk) (D). Three clones (K2, K18 and K30) were selected after transfection by a LentiCRISPRV2 targeting Top1 and analyzed for the % of GFP positive cells (n = 3) (E) or the level of mRNA coding for GFP (n = 3) (F)