The co-activators CBP and p300 are important for normal cell differentiation and cell cycle progression and are the targets for viral proteins that dysregulate these cellular processes. We show here that the E6 protein from the oncogenic human papillomavirus type 16 (HPV-16) binds to three regions (C/H1, C/H3 and the C-terminus) of both CBP and p300. The interaction of E6 with CBP/p300 was direct and independent of proteins known to bind the co-activators, such as p53. The E6 protein from low-risk HPV type 6 did not interact with C/H3 or the C-terminus but associated with the C/H1 domain at 50% of the level of HPV-16. HPV-16 E6 inhibited the intrinsic transcriptional activity of CBP/p300 and decreased the ability of p300 to activate p53-and NF-κB-responsive promoter elements. Interestingly, some mutations in HPV-16 E6 abrogated C/H3-E6 interactions, but did not alter the ability of E6 to associate with the C/H1 domain, suggesting that these modified proteins could be used to delineate the functional significance of the C/H1 and C/H3 domains of CBP/p300.

The p63 transcription factor (TP63) is critical in development, growth and differentiation of stratifying epithelia. This is highlighted by the severity of congenital abnormalities caused by TP63 mutations in humans, the dramatic phenotypes in knockout mice and de-regulation of TP63 expression in neoplasia altering the tumour suppressive roles of the TP53 family. In order to define the normal role played by TP63 and provide the basis for better understanding how this network is perturbed in disease, we used chromatin immunoprecipitation combined with massively parallel sequencing (ChIP-seq) to identify >7500 high-confidence TP63-binding regions across the entire genome, in primary human neonatal foreskin keratinocytes (HFKs). Using integrative strategies, we demonstrate that only a subset of these sites are bound by TP53 in response to DNA damage. We identify a role for TP63 in transcriptional regulation of multiple genes genetically linked to cleft palate and identify AP-2alpha (TFAP2A) as a co-regulator of a subset of these genes. We further demonstrate that AP-2gamma (TFAP2C) can bind a subset of these regions and that acute depletion of either TFAP2A or TFAP2C alone is sufficient to reduce terminal differentiation of organotypic epidermal skin equivalents, indicating overlapping physiological functions with TP63.

The E7 gene product of human papillomavirus type 16 (HPV16) binds to the retinoblastoma gene product (pRb) and dissociates pRb‐E2F complexes. However, the observation that the ability of E7 to bind pRb is not required for the HPV16‐induced immortalization of primary keratinocytes prompted a search for other cellular factors bound by E7. Using a glutathione‐S‐transferase (GST) fusion protein system, we show that E7 complexes with AP1 transcription factors including c‐Jun, JunB, JunD and c‐Fos. The ability of E7 to complex with c‐Jun in vivo is demonstrated by co‐immunoprecipitation and the yeast two‐hybrid system. An analysis of E7 point mutants in the GST system indicates that the E7 zinc‐finger motif, but not the pRb binding domain, is involved in these interactions. Using c‐Jun deletion mutants, E7 binding maps between amino acids 224 and 286 of c‐Jun. E7 trans‐activates c‐Jun‐induced transcription from a Jun responsive promoter, and this activity correlates with the ability of E7 mutants to bind Jun proteins. Finally, a transcriptionally inactive c‐Jun deletion, which can bind E7, interferes with the E7‐induced transformation of rat embryo fibroblasts in cooperation with an activated ras, indicating that the Jun‐E7 interaction is physiologically relevant and that Jun factors may be targeted in the E7 transformation pathway.