NFI is an essential positive transcription factor for human papillomavirus type 16 early gene expression

Human papillomavirus type 16 (HPV16) is the primary etiologic agent for greater than 50% of all cervical carcinomas. Expression of the HPV16 E6 and E7 oncoproteins is under control of the upstream regulatory region (URR), which contains a myriad of transcription factor binding sites, including 7 half sites for NFI. These NFI binding sites were used as probes in electrophoretic mobility shift assays (EMSAs), and mutational analysis of individual and multiple NFI binding sites was performed in order to demonstrate the relative importance of particular NFI sites to URR activity. By using 5 NFI half sites as an enhancer, we were able to detect a 4-fold increase in URR activity. Our results define the role and relative contribution of NFI binding sites to the basal activity of the HPV16 promoter, and demonstrate that NFI binding sites can act independently to enhance HPV16 URR activity in immortalized keratinocytes.