FIG 5.

(A) HeLa cells were transfected with FLAG-WT and E2 Y138 mutants. After 48 h, ChIP assays were performed using the ChIP-IT express chromatin immunoprecipitation kit (Active Motif) with FLAG antibodies. HPV-18 LCR DNA was amplified using RT-PCR, and values were normalized to those of input DNA. Control = no FLAG E2 DNA; WT = 1. Values are expressed as means ± SEM. *, P ≤ 0.05 by two-tailed t test compared to control. (B) HeLa cells were transfected with FLAG-HPV-31 E1 and FLAG-WT or Y138 E2 mutants. After 48 h, the ChIP assays were performed with rat anti-E1 antibodies. HPV-18 LCR DNA was amplified using RT-PCR, and values were normalized to those of input DNA. No E1 = 1. Values are expressed as means ± SEM. *, P ≤ 0.05 by two-tailed t test compared to no E1 group. No statistically significant difference was found between WT and Y138 mutant groups. E1 only (n = 6), no E1 (n = 12), E1+ WT (n = 11), E1 + Y138F (n = 12), and E1 + Y138E (n = 12). (C) HeLa cells were transfected with WT and Y138 E2 mutants. After 48 h, RNA was isolated and cDNA was synthesized. Real-time PCR was used to measure HPV-18 E6 cDNA and normalized to actin cDNA.