FIG 2.

Luciferase reporter constructs were generated to test the ability of the CTCF binding motif to block the LAT enhancer. (A) General schematic for construct design. Each CTCF/LTE construct contained the specific CTCF binding site positioned between the LAT enhancer element and the simian virus 40 (SV40) promoter in the pGL3 control vector. (B) Reporter assays in Vero cells. All transfections were done in triplicate wells and were repeated five times (n = 5). Data are presented as both a comparison of the LTE to the LTE/CTCF construct and of the CTCF compared to the LTE/CTCF construct (*, P < 0.001), comparing the reduction in luciferase expression levels between the LTE and LTE/CTCF constructs, determined by unpaired one-tailed Student's t tests in pairwise comparisons; #, 2-fold reduction in luciferase expression levels between the CTCF and LTE/CTCF constructs. All luciferase values were normalized to the value for the pGL3 control, which was set to 1. (C) Reporter assays done in Neuro 2a cells. All transfections were done in triplicate wells and were repeated five times (n = 5). Data are presented as both a comparison of the LTE to the LTE/CTCF construct and of the CTCF compared to the LTE/CTCF construct (*, P < 0.001), comparing the reduction in luciferase expression levels between the LTE and LTE/CTCF constructs, determined by unpaired one-tailed Student's t tests in pairwise comparisons; &, P < 0.01, comparing the reduction in luciferase expression levels between the CTCF and LTE/CTCF constructs.