Human papillomavirus 16 E6 expression disrupts the p53-mediated cellular response to DNA damage.

Abstract

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 malignant transformation, suggesting the requirement of additional cellular events. After DNA damage, normal mammalian cells exhibit G1 cell-cycle arrest and inhibition of replicative DNA synthesis. This mechanism, which requires wild-type p53, presumably allows cells to undertake DNA repair and avoid the fixation of mutations. We directly tested whether the normal response of cervical epithelial cells to DNA damage may be undermined by interactions between the E6 protein expressed by oncogenic 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 containing HPV16, HPV18, or mutant p53 alone. To test the effects of E6 alone 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 these 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 changes associated with cervical tumorigenesis.

Twenty years of p53 research: structural and functional aspects of the p53 protein

“…For example, 5-Gy ionizing irradiation produces p53-dependent G1 growth arrest in proliferating fibroblasts (Kastan et al, 1991). It is well established that wild-type p53 is required for the induction of G1 arrest in response to IR because cell lines engineered to lack p53 activity display an attenuated response (Kuerbitz et al, 1992;Kessis et al, 1993). In addition, embryonic ®broblasts from p53-null mice lose the G1 arrest response to IR .…”

Characterization of keratin and cell cycle protein expression in cell lines from squamous intraepithelial lesions progressing towards a malignant phenotype

“…Moreover, continuous transcription of HPV 33 E6-E7 suggests that p53 function is continuously affected. The effect of HPV E6 on the function of p53 is concentration dependent (Kessis et al, 1993;Lechner and Laimins, 1994), and an attempt was made to analyse the HPV E6 protein levels in the rafts of both cell lines, however the currently available antibodies were not sufficiently specific to warnant conclusions.…”

Human smooth muscle α-actin gene is a transcriptional target of the p53 tumor suppressor protein

“…The DNA-protein complexes were resolved on a 4% non-denaturing polyacrylamide gel in 0.5xTBE at 48C. The gels were dried and exposed to X-ray ®lm overnight 480 cells express mutant p53 (Kessis et al, 1993;Slichenmeyer et al, 1993;Kuerbitz et al, 1992;Kastan et al, 1991). These cells lack IR induction of p53 and the activation of the G1 check point.…”