Figure 4

Treatment of cells with latency reversing drugs induces Cas9 expression and cleavage of integrated viral DNA in Jurkat 2D10 cells.

2D10 cells expressing LTR_−80/+66-Cas9 were treated with control (empty) or lentivirus plasmid expressing gRNAs A/B and 24 hours later cells were treated with PMA (P), TSA (T) or both (P/T) for 16 hours, as indicated. Protein studies for the expression of Cas9-Flag, α-tubulin and GFP (indicative of the integrated HIV-1 genome) was determined by Western blot (a). Genomic DNA for the detection of the level of excision within the integrated LTR DNA by Cas9 and gRNA A/B was assessed by PCR and the excision efficiency was determined as described in Fig. 3 legend (b). GFP reporter assay, by flow cytometry and representative scatter plot is shown (c).