Figure 1.

Introduction of tat/rev-targeting CRISPR-Cas9 protects T cells from new HIV-1 infection. (a) The target location of the designed gRNAs inside the HIV-1 genome. (b) gRNA-Cas9 constructs were transduced into T cell line MT-4 with a multiplicity of infection (MOI) of 10 and selected with puromycin for 14 days. Cells were lysed and immunoblotted with anti-Cas9 antibody and anti-β-actin as the loading control. Representative blot images and densitometry graph of Cas9 expression standardized to β-actin are shown. (c) As shown in the experimental timeline, gRNA-Cas9-transduced cells were challenged with HIV_NL4-3 for 2 days before washing with fresh medium. The supernatant p24 levels were measured at four days post-infection (dpi). Assays were performed four times, and average values (±SE) are shown. One-way ANOVA applied to all groups and Dunnett’s multiple comparison test for each group compared to wild type (WT) were used to calculate significance. *, p < 0.05; **, p < 0.01; ***, p < 0.001; ****, and p < 0.0001.