Human Papillomavirus Type 16 E6 and E7 Cause Polyploidy in Human Keratinocytes and Up-Regulation of G2-M-phase Proteins

Human papillomavirus type 16 proteins E6 and E7 have been shown to cause centrosome amplification and lagging chromosomes during mitosis. These abnormalities during mitosis can result in missegregation of the chromosomes, leading to chromosomal instability. Genomic instability is thought to be an essential part of the conversion of a normal cell to a cancer cell. We now show that E6 and E7 together cause polyploidy in primary human keratinocytes soon after these genes are introduced into the cells. Polyploidy seems to result from a spindle checkpoint failure arising from abrogation of the normal functions of p53 and retinoblastoma family members by E6 and E7, respectively. In addition, E6 and E7 cause deregulation of cellular genes such as Plk1, Aurora-A, cdk1, and Nek2, which are known to control the G2-M-phase transition and the ordered progression through mitosis.

Induction of tetraploidy through loss of p53 and upregulation of Plk1 by Human Papillomavirus type-16 E6

Cancer cells are insensitive to many signals that inhibit growth of untransformed cells. Here, we show that primary human epithelial cells expressing human papillomavirus (HPV) type-16 E6/E7 bypass arrest caused by the DNA-damaging drug adriamycin and become tetraploid. To determine the contribution of E6 in the context of E7 to the resistance of arrest and induction of tetraploidy, we used an E6 mutant unable to degrade p53 or RNAi targeting p53 for knockdown. The E6 mutant fails to generate tetraploidy; however, the presence of E7 is sufficient to bypass arrest while the p53 RNAi permits both arrest insensitivity and tetraploidy. We published previously that polo-like kinase 1 (Plk1) is upregulated in E6/E7-expressing cells. We observe here that abnormal expression of Plk1 protein correlates with tetraploidy. Using the p53 binding-defective mutant of E6 and p53 RNAi, we show that p53 represses Plk1, suggesting that loss of p53 results in tetraploidy through upregulation of Plk1. Consistent with this hypothesis, overexpression of Plk1 in cells generates tetraploidy but does not confer resistance to arrest. These results support a model for transformation caused by HPV-16 where bypass of arrest and tetraploidy are separable consequences of p53 loss with Plk1 required only for the latter effect.