Fig. 4

Distinct effects of KO of LEDGF/p75 or the components of DNA-PK on the early replicative stages of HIV. a Relative luciferase expression in 293T or different CRISPR/Cas9 modified cell lines transduced by 0.01 MOI of either HIV_wt or HIV_mut presented as: left plot—data normalized to the luciferase expression level in the parent 293T cell line transduced by HIV_wt; right plot—data normalized to the luciferase expression level in the 293T cells for HIV_wt and HIV_mut separately. b, c The level of total viral DNA (b) and integrated viral DNA (c) in 293T or different CRISPR/Cas9 modified cell lines transduced by either HIV_wt or HIV_mut measured by the standard qPCR analysis [45] and data was normalized to the data for CD3 gene to achieve a per cell normalization. d Gap repair efficiency in 293T or different CRISPR/Cas9 modified cell lines transduced by either HIV_wt or HIV_mut measured by the modified qPCR [39]. Data is shown as percentage of repaired proviral DNA relative to integrated proviral DNA (left plot) or as a repair efficiency of both pseudoviruses in CRISPR/Cas9 modified cell lines normalized to the repair efficiency in 293T cells (right plot) for HIV_wt and HIV_mut separately. e The siRNA mediated knockdown of Ku70, Ku80 or DNA-PKcs in 293T cells leads to a reduction in luciferase expression levels for HIV_wt and does not affect HIV_mut. Cells were transfected with each siRNA and 48 h later were transduced with 0.01 MOI of either HIV_wt or HIV_mut. 36 h later luciferase expression was measured and normalized to the luciferase levels in the control siRNA transfected cells for HIV_wt and HIV_mut separately. Means and SDs of (n = 3 for a–d and n = 4 for e) are plotted, significance was determined by two-tailed Student’s t-test, * = p < 0.05, ** = p < 0.01, *** = p < 0.001