Figure 3. MiR-184 expression is down-regulated by E6 oncoprotein at transcription level via decreased the binding activity of p53 onto the miR-184 promoter due to p53 degradation by E6.

(A) TL-1 and SiHa cells were treated with miR- 184 promoter reporter plasmid (−839/+1), wild-type p53 expression vector (5 μg), shp53 (5 μg), and/or shE6 shRNA (5 μg). (B) TL-10 and C33A were treated with the miR-184 promoter reporter plasmid (−839/+1), shp53 shRNA (5 μg), wild-type p53 (5 μg), and/or E6 expression vectors (5 μg). The cell lysates were separated by SDS-PAGE for the evaluation of protein expression by specific antibodies using western blotting. A ChIP assay was performed to evaluate the binding ability of p53 onto the miR-184 promoter. The products were amplified by PCR. Luciferase reporter assay was performed to evaluate the reporter activity of miR-184 promoter (−839/+1). The miR-184 level was determined by real-time PCR.