HUMAN PAPILLOMAVIRUS-16 E6 EXPRESSION DISRUPTS THE P53-MEDIATED CELLULAR-RESPONSE TO DNA DAMAGE

Infection with certain types of human papillomaviruses (HPV) is highly associated with carcinomas of the human uterine cervix. However, HPV infection alone does not appear to be sufficient for the process of ma lignant transformation, suggesting the requirement of additional cellu lar events. After DNA damage, normal mammalian cells exhibit G1 cell-c ycle arrest and inhibition of replicative DNA synthesis. This mechanis m, which requires wild-type p53, presumably allows cells to undertake DNA repair and avoid the fixation of mutations. We directly tested whe ther the normal response of cervical epithelial cells to DNA damage ma y be undermined by interactions between the E6 protein expressed by on cogenic HPV types and wild-type p53. We treated primary keratinocytes with the DNA-damaging agent actinomycin D and demonstrated inhibition of replicative DNA synthesis and a significant increase in p53 protein levels. In contrast, inhibition of DNA synthesis and increases in p53 protein did not occur after actinomycin D treatment of keratinocytes immortalized with HPV16 E6/E7 or in cervical carcinoma cell lines cont aining HPV16, HPV18, or mutant p53 alone. To test the effects of E6 al one on the cellular response to DNA damage, HPV16 E6 was expressed in the carcinoma cell line RKO, resulting in undetectable baseline levels of p53 protein and loss of the G1 arrest that normally occurs in thes e cells after DNA damage. These findings demonstrate that oncogenic E6 can disrupt an important cellular response to DNA damage mediated by p53 and may contribute to the subsequent accumulation of genetic chang es associated with cervical tumorigenesis.