FIG. 5.

Effects of single and multiple NFI mutations on TGF-β modulation of the HPV16 URR. The entire HPV16 URR (Fig. 1) was cloned into the luciferase reporter vector pGL3 (Promega) (pGL3/URR) where various NFI sites were mutated from GCCAA to GCAGA, which is unable to bind NFI. These constructs were transfected into TGF-β-sensitive HKc/HPV16 and treated with and without 40 pM TGF-β for 42 h. Luciferase activity was determined 68 h posttransfection. Corrected percent TGF-β inhibition for each construct was determined by subtracting the percent TGF-β inhibition obtained by transfection of a promoterless pGL3 plasmid from the total TGF-β inhibition obtained for each reporter construct. The specific NFI site(s) that was mutated and the percent reduction of basal URR activity are shown for each mutant construct at the bottom of the figure.