Epidermal growth factor induction of human papillomavirus type 16 E6/E7 MRNA in tumor cells involves two AP-1 binding sites in the viral enhancer.

The early genes E6 and E7 from human papillomaviruses (HPVs) play a key role in the development of cervical cancer. Modulation of E6 and E7 gene expression may alter tumour progression; therefore, modifiers of viral transcription such as hormones or growth factors are potential risk factors in cancer development. We have analysed the effects of epidermal growth factor (EGF) on E6/E7 mRNA from human papillomavirus type 16 (HPV-16) by Northern blot in two cell lines, SiHa cervical carcinoma cells, and HPK IA, an HPV-16-immortalized keratinocyte cell line. E6/E7 mRNA is EGF-inducible in SiHa cells, with the earliest response after 2 h. In contrast, in HPK IA cells no increase in E6/E7 RNA is observed, suggesting a differential EGF response of viral transcription in tumour cells compared with keratinocytes. We demonstrate that the cell type-specific HPV-16 enhancer is a target of EGF-induced signals, as its activity is amplified by EGF in SiHa cell transfections. However, when transfected into HPK IA keratinocytes, the viral enhancer shows no EGF response. The enhancer contains two binding sites for the transcription factor AP-1, a potential mediator of the EGF signalling cascade. Enhancer subfragments with single AP-1 binding sites are also EGF-responsive in SiHa cells. Mutating either AP-1 site in the complete enhancer decreases the EGF response, whereas a double mutation causes a complete loss of EGF regulation, suggesting that the EGF induction of HPV-16 early transcription requires AP-1 activation. We conclude that alterations of EGF responsiveness that increase viral oncogene expression may contribute to cervical cancer progression.