FIG. 8.

E2-mediated repression of the HPV-11 E6 promoter can be alleviated by forming a promoter-bound TBP (or TFIID)-TFIIB-pol II-TFIIF complex. (A) TBP as the TATA-binding factor for PIC assembly. In vitro transcription was performed as outlined at the bottom by preincubating transcriptional templates with TBP (T), TBP-TFIIB (T/B), TBP-TFIIB-pol II (T/B/II), or TBP-TFIIB-pol II-TFIIF (T/B/II/F), in different reaction tubes at 30°C for 30 min. The remaining components and NTPs were then added, in the absence (−) or presence (+) of HPV-11 E2, to initiate transcription. The reactions were processed as described in Materials and Methods. Regular E2-mediated reactions with all four protein components but without a two-step incubation were also conducted for comparison (lanes 1 and 2 and lanes 11 and 12). (B) TFIID as the TATA-binding factor for PIC assembly. In vitro transcription was performed as outlined in panel A, except that TFIID (D) was used as the TATA-binding factor.