Effects of Dexamethasone and Epidermal Growth Factor on Activity of Viral Promoter p97 and Expression of HPV-16 E6/E7 in Human Oral Keratinocytes Transformed with HPV-16 and Benzo(a)pyrene

Abstract

Primary human oral keratinocytes were previously transformed by transfection with clned human papilloavtrus type 16 (HPV-16) DNA and subsequent exposure tobenzo(a)pyrene, and an oral cancer cell line, CTHOK-16B-Bap, was established. To determine the effects of dexamethasone and epodermal growth facotr (RGF) on cell proliferation, the expression of HPV-16 E6/E7 and several proto-oncogenes, and activity of HPV-16 E6/E7 promotor, p97, the CTHOK-16B Bap cells were exposed to either dexamethasone and EGF alone or together. After incubation for 3 days, the degrees of both cell proliferation and the expression of HPV-16 E6/E7, EGF receptor (EGFR), c-myc, and c-fos fenes was determined, Dexamethasone and EGF, when added alone or together in the culture media, increased cell proliferation. Although dexamethasone did not affect the transctiptional level of HPV-16 E6/E7, EGFR, or c-myc in the cells, ot downregulated c-fos mRNA expression. On the other hand, EGF, alone or in confunction with dexzmethasone. down-regulated the transcriptional levels of HPV-16 E6/E7, c0myc, and c-fos genes, but it had little effect on the level if EGFR transctiption. These results suggest that the increaed proliferation of CTHOK-16B-BaP cells in the presence of dexamethasone and/or EGF may not depend on the extent of expression of such gense as HPV-16 E6/E7, EGFR, c-myc, and c-fos. In addition, the HPV-16 E6/E7 mRNA level was not changed and reduced by treatment with dexamethasone and EGF, respectively. Unexpectedly, however, dexamathasone and EGF enhanced the activity of the viral promoter p97 in CTHOK-16B-BaP line as analyzed by transient expression assays using the chloramphenicol α-cetyltransferase gene as a reporter. It appears that dominant regulatiory mechanisms presumably depending on the chromosomal integration site are able to override the respomse of the viral promoter to dexamethasone and EGF, Another EGF-responsive element (7454-7643 m) which acts as an enhancer may be located within the HPV-16 LCR portion.