Selective expression of death receptor 4 and induction of apoptosis by HPV E6

Choi, Eunah; Koh, Hyonseok; Shin, Soonah; Kim, Kunhong

Abstract

Human papillomaviruses, especially type 16 and 18, have been known to be closely associated with human cervix cancer. The E6 and E7 proteins encoded by these viruses disrupt normal cell cycle regulation. The E7 protein interacts with pRb and abrogates its tumor-suppressive activity, while the E6 protein cooperates with E6AP, a ubiquitin E3 ligase, to tzrget p53 tumor suppressor protein for ubiquitin-dependent degradation. Telomerase activity is detected in more than 90% of immortalized and cancer cells but absent in most normal somatic cells and E6 oncoprotein could activate telomerase. In this experiment, we hypothesized that the hTERT promoter can be used for tumor-specific expression of transgenes to induce selective cancer cell death in HPV E6 positive cells. First, we tested whether hTERT core promoter is active only in cancer cells and whether the presence of E6 protein could potentiate the activation of the promoter by using luciferase construct and we found that hTERT promoter is active only in cancer cells, and the presence of E6 protein could augment hTERT promoter activity. Then, we constructed mammalian expression vector to express TRAIL-death Receptor (DR) 4 driven by hTERT promoter to test whether DR4 expression is induced by the promoter in the presence of E6 oncoprotein and we found that DR4 expression was induced. Finally, we tested whether the induction of DR4 could contribute to cancer cell death by observing caspase 3 activation and we found that in the presence of E6, caspase 3 activation is more evident than in the absence of 6. Taken together, our results reveal that the proper use of hTERT promoter could be useful and powerful for inducing selective cancer cell (especially for E6 positive cancer cells) death with effective death inducing transgene.