The E6 oncoprotein of human papillomaviruses (HPV) promotes oncogenesis by inactivating tumor suppressor protein p53 i. e. it binds to and enhances the degradation of p53. To study whether inactivation of p53 is solely responsible for E6-induced oncogenesis, we constructed several plasmid vectors expressing wild-type (wt) or mutant (mt) E6 proteins. RKO cells that express wt p53 were stably transfected with these plasmids and challenged with DNA damaging agents. The level of p53 was significantly increased by DNA damaging agents in control cells and cells transfected with plasmids expressing mt E6 that do not bind to p53. As expected, p53 did not increase in cells transfected with plasmids expressing mt E6 that do bind to p53. To investigate the oncogenic effect of these various E6 proteins, we determined the mutation frequency of the hprt locus in control cells and cells expressing different E6 proteins. We found that cells expressing wt E6 and mt E6 (capable or incapable of binding to p53) showed notable increases in the mutation frequency at hprt locus compared with that of control cells. The elevation of mutation frequency in cells expressing mt E6 was similar to that in cells expressing wt E6. These data indicate that E6-induced mutagenicity is induced not only via p53 inactivation, but also via p53-independent pathways.