The E7 protein of human papillomavirus type 16 sensitizes primary human keratinocytes to apoptosis

Abstract

The`high risk' human papillomaviruses are associated with the development of anogenital carcinomas and their E6 and E7 genes possess immortalizing and transforming functions in several in vitro culture systems. Recently the E6 gene has also been shown to enhance the apoptosis of human mammary epithelial cells. To determine the apoptotic activity of these oncogenes in the natural host cell, we infected genital keratinocytes with retroviruses expressing either HPV-16 E6, E7, or both the E6 and E7 (E6/7) genes. Apoptosis was quantitated under normal growth conditions or when induced by tumor necrosis factor a/cycloheximide or sulfur mustard. In contrast to previous ®ndings with mammary epithelial cells, the E6 gene did not signi®cantly augment either spontaneous or induced apoptosis. E6 also did not suppress apoptosis in normal keratinocytes (despite dramatically reducing their p53 levels), suggesting that p53-independent events mediated this eect. In contrast, E7 increased both spontaneous and induced apoptosis as well as the cellular levels of p53 and p21 protein. Interestingly, coexpression of E6 abrogated E7-facilitated apoptosis by tumor necrosis factor a nearly completely, but had only a minor protective eect on sulfur mustard induced apoptosis in these cells, demonstrating at least in part the p53-dependence and -independence of these two apoptotic pathways. Finally, our results indicate that the apoptosis of normal and E7-expressing keratinocytes is dierentially aected by E6 expression and that E7, when unaccompanied by E6, sensitizes keratinocytes to apoptosis.

The HPV E7 oncoprotein inhibits tumor necrosis factor α-mediated apoptosis in normal human fibroblasts

“…Apoptosis is induced more rapidly and e ciently by TNF R1 than Fas antibody in IMR-90 cells (Figure 1d). This may also explain the di erences between this study and a published report by Richard Schlegel's laboratory that provided evidence for increased TNF-mediated apoptosis in E7-expressing keratinocytes (Stoppler et al, 1998). We have obtained similar results in keratinocytes: TNF/ cycloheximide treatment induces a relatively low level of apoptosis and the apoptotic response is much delayed compared to ®broblasts (J Basile et al, 2001, manuscript in preparation).…”

The human papillomavirus (HPV) 16 E6 oncoprotein leads to an increase in gene expression of the angiogenic switch molecule FGF-BP in non-immortalized human keratinocytes

“…This demonstrates that the expression of E6 alone, without further cellular changes, is su cient to upregulate the FGF-BP promoter in non-transformed primary human keratinocytes during their senescent growth phase. It has already been described that E7-expressing primary keratinocytes possess an increased p53 steady state level in comparison to non-HPV oncogeneexpressing keratinocytes as well as to E6-expressing keratinocytes in which p53 levels are further reduced (Demers et al, 1994;StoÈ ppler et al, 1998). As previously shown, the up-regulated p53 steady state level in E7-expressing cells led to an increase in the promoter activity of a p53-dependent promoter.…”

The human papillomavirus 16 E6 protein can either protect or further sensitize cells to TNF: effect of dose

“…In one paper in which the entire HPV 18 sequence was present, the authors found that it conferred resistance to TNF. 71 E7, on the other hand, has been reported both to protect 72 and to sensitize 73 human fibroblasts or keratinocytes, respectively, from TNF. Our own results with the CaSki and SiHa cells (Figure 1) are consistent with the E6 dose-dependent response we have worked out in greater molecular detail in the U2OS cells, and it may be that dosage effects are important for viral proteins other than E6.…”