HPV16 E6 oncoprotein inhibits apoptosis induced during serum-calcium differentiation of foreskin human keratinocytes

Transfection of human papillomavirus (HPV) 16 E6 oncogene into foreskin pri mary human keratinocytes (PHKs) causes the formation of colonies of viable cells resistant to serum-calcium differentiation. To define the stage of ke ratinocyte differentiation inhibited by E6, we examined the response of PHK s to serum and calcium with respect to parameters of both growth and differ entiation. The effect of HPV16 E6 was evaluated by infection with recombina nt retroviruses encoding the E6 protein. Results of these studies indicated that terminal differentiation of cultured foreskin keratinocytes, triggere d by serum and calcium, is a progressive process (2-3 weeks) that ends with cell death with characteristics of apoptosis. Human keratinocyte terminal differentiation was accompanied by time-related changes in the expression o f cellular proteins involved in the control pathways of apoptosis, includin g downregulation of Bcl-2 and p53 and upregulation of Bar, which coincided with the appearance of morphological signs of apoptosis. E6 expression did not override the differentiation-associated GI arrest or prevent the induct ion of squamous differentiation-specific markers, transglutaminase 1 and in volucrin. E8 expression led, however, to a significant reduction in cell st ratification and cell death by apoptosis, which correlated with prolonged e xpression of Bcl-2 and reduced elevation of Bar levels that occurred concom itant with a complete loss of p53. The data argue that E6 inhibits terminal differentiation of foreskin PHKs through inhibition of their differentiati on-induced apoptotic program. (C) 1999 Academic Press.