Brd4-independent transcriptional repression function of the papillomavirus E2 proteins

The papillomavirus E2 protein is a critical viral regulatory protein with transcription, DNA replication, and genome maintenance functions. We have previously identified the cellular bromodomain protein Brd4 as a major E2-interacting protein and established that it participates in tethering bovine papillornavirus type I E2 and viral genomes to host cell mitotic chromosomes. We have also shown that Brd4 mediates E2-dependent transcriptional activation, which is strongly inhibited by the disruption of E2/Brd4 binding as well as by short hairpin RNA (shRNA) knockdown of Brd4 expression levels. Since several mutants harboring single amino acid substitutions within the E2 transactivation domain that are defective for both transcriptional transactivation and Brd4 binding are also defective for transcriptional repression, we examined the role of Brd4 in E2 repression of the human papillomavirus E6/E7 promoter. Surprisingly, in a variety of in vivo assays, including transcription reporter assays, HeLa cell proliferation and colony reduction assays, and Northern blot analyses, neither blocking of the binding of E2 to Brd4 nor shRNA knockdown of Brd4 affected the E2 repression function. Our study provides evidence for a Brd4-independent mechanism of E2-mediated repression and suggests that different cellular factors must be involved in E2-mediated transcriptional activation and repression functions.