Mutations of the human papillomavirus type 16 E7 gene that affect transformation, transactivation and phosphorylation by the E7 protein

The human papillomavirus type 16 (HPV-16) E7 gene cooperates with an activated ras oncogene to transform primary rodent cells and is important in the immortal-ization of cervical keratinocytes. We have generated a series of point mutations within the E7 gene and show that mutation of residues erine 31 and serine 71 affect the phosphorylation f the E7 protein, but do not alter its ability to cooperate with ras. Further mutations which alter cysteine residues in a-Cys-X-X-Cys- motif decrease transformation markedly, although they do not abolish it entirely. All the mutations generated displayed a decreased ability to transactivate the adenovirus E2 promoter. These results show that neither phosphorylation of E7 nor its ability to transactivate are required for transformation by E7. Human genital papillomaviruses (HPVs) are frequently associated with cervical dysplasias and neoplasias. HPV types 16, 18, 31 and 33 are commonly found in high grade lesions and carcinomas, whereas types 6 and 11 are typically found in condylomata and low grade cervical intraepithelial lesions (Diirst et al., 1983; Gissmann et al., 1983). In the majority of cervical carcinomas the HPV DNA is integrated into the cellular genome, integration usually occurring within the viral E1 or E2 open reading frames (ORF) and is often accompanied by deletions in this region (Schwarz et al., 1983; Schneider-Gadicke & Schwarz, 1986). The viral early region encoding the E6 and E70RF is selectively retained (Schwarz et al., 1983; Schneider-Gadicke & Schwarz, 1986) and expressed in cervical cancer-derived cell lines (Smotkin & Wettstein, 1986; Androphy et al., 1987; Banks et al., 1987). Transformation of primary rodent epithelial cells by genital HPVs requires a functional E7 gene to cooperate with the activated ras (Phelps et al.