Upregulation of ΔNp63 transcript during immortalization of primary nasopharyngeal epithelial cells

Abstract

Mutation of the p53 gene is a common event in human cancer. However, p53 mutation is rarely observed in nasopharyngeal carcinoma which is a common cancer among Cantonese population living in Hong Kong and Southern China. Alternative ways of inactivation of function of the p53 gene may exist in nasopharyngeal carcinoma to facilitate its development. Recent study has shown that overexpression of the ΔNp63 isoform of the p63 gene could be detected in nasopharyngeal carcinoma. The ΔNp63 has been postulated to have dominant negative effect on function of p53 gene. Expression of ΔNp63 may functionally inactivate the p53 gene in nasopharyngeal carcinoma and plays a role in its development. Immortalization is a pre-requisite and fundamental property of human cancer cells. Molecular events occurring at the stage of cell immortalization may represent early events in human carcinogenesis. Our laboratory has been involved in the study of immortalization of human nasopharyngeal epithelial cells. We have examined the expression profile of ΔNp63 in human nasopharyngeal epithelial cells during immortalization. A panel of primary and immortalized nasopharyngeal epithelial cells at different stages of immortalization was used to examine the profile of ΔNp63 expression. Immortalization of nasopharyngeal epithelial cells was achieved by individual expression of telomerase, SV40T or HPV16E6E7. The immortalized nasopharyngeal epithelial cells are non-malignant in nature but harbor specific genetic alterations such as p16 and RASSF1A inactivation. Accurate quantitation of the transcripts of ΔNp63 was performed by real-time PCR. Low level of ΔNp63 transcript could be detected in all primary nasopharyngeal epithelial cells but not in senescing nasopharyngeal epithelial cells. Interestingly, marked increase in ΔNp63 expression (8 to 32 fold) was observed in all immortalized nasopharyngeal epithelial cells compared to primary nasopharyngeal epithelial cells suggesting a role in immortalization. The increase of ΔNp63 was also confirmed by western blotting using antibody against p63 and ΔNp63. The expression level of TAp63 remains low and unremarkable during immortalization of nasopharyngeal epithelial cells. Our results show that elevated expression of ΔNp63 is a common property of immortalized nasopharyngeal epithelial cells and nasopharyngeal carcinoma cells. The detection of elevated expression of ΔNp63 in immortalized but non-malignant nasopharyngeal epithelial cells show that it may represent an early event in premalignant nasopharyngeal epithelial cells. Our results support a role of ΔNp63 in the development of nasopharyngeal carcinoma.