Rapid induction of senescence in human cervical carcinoma cells

Abstract

Expression of the bovine papillomavirus E2 regulatory protein in human cervical carcinoma cell lines repressed expression of the resident human papillomavirus E6 and E7 oncogenes and within a few days caused essentially all of the cells to synchronously display numerous phenotypic markers characteristic of cells undergoing replicative senescence. This process was accompanied by marked but in some cases transient alterations in the expression of cell cycle regulatory proteins and by decreased telomerase activity. We propose that the human papillomavirus E6 and E7 proteins actively prevent senescence from occurring in cervical carcinoma cells, and that once viral oncogene expression is extinguished, the senescence program is rapidly executed. Activation of endogenous senescence pathways in cancer cells may represent an alternative approach to treat human cancers. Cancer is thought to be caused by the stepwise accumulation of genetic changes, resulting in progressive phenotypic abnormality. Infection of cervical keratinocytes with a highrisk human papillomavirus (HPV) such as HPV18 is the initiating event in the great majority of cervical carcinomas (1). The HPV E6 and E7 proteins are continuously expressed in cervical carcinoma cells and seem to play a role in tumor development and in maintenance of the malignant phenotype (1). The role of the E6 and E7 proteins in the progression from normal keratinocytes to transformed cells has been studied in cell culture. Normal human keratinocytes can undergo a limited number of cell divisions in vitro and then stop proliferating, a phenomenon referred to as replicative senescence (2-4). Telomere length progressively shortens during cell passage (5, 6), and it has been proposed that once the length of one or more telomeres falls below a critical threshold, an active genetic program is mobilized that culminates in senescence (7,8). Cultured keratinocytes can escape senescence and become immortalized by inactivation of the retinoblastoma tumor suppressor pathway and activation of telomerase, the enzyme responsible for maintaining telomere length (9, 10). Expression of the HPV E6 and E7 proteins also can immortalize keratinocytes (11,12), an activity that is consistent with the known biochemical activities of these proteins. The E7 protein neutralizes the retinoblastoma tumor suppressor pathway, and the E6 protein stimulates telomerase activity in keratinocytes (1,6,13). Although the mechanistic basis for this effect on telomerase activity is not known, this activity seems crucial for immortalization because E6 mutants defective for telomerase activation are immortalization-defective (10). In addition, host-cell mutations are required for HPVimmortalized keratinocytes to become tumorigenic and to acquire other malignant characteristics, such as metastatic potential (9, 14 -20). The E6 and E7 proteins facilitate the acquisition of these mutations by causing accelerated degradation of p53 and p105 Rb , respectively, resulting in abrogation of DNA damage checkpoint control, elevat...

Inhibition of Bax activity is crucial for the antiapoptotic function of the human papillomavirus E6 oncoprotein

“…Whereas, we specifically target E6 by RNAi, treatment with IP10 resulted in the inhibition of both E6 and E7 expression. Yet, in contrast to studies indicating that combined E6 and E7 inhibition primarily results in senescence (Goodwin et al, 2000;Wells et al, 2000;Hall and Alexander, 2003), induction of apoptosis was reported. In addition, IP10 is likely to exert additional biological activities besides affecting E6/E7 expression.…”

“…E6 targets the PUMA/Bax Pathway M Vogt et al inhibition of E6 and E7. The combined inhibition of E6 and E7 primarily leads to growth arrest and senescence (Goodwin et al, 2000;Wells et al, 2000;Hall and Alexander, 2003), due to the release of pRb from the inhibitory influence of E7 (Psyrri et al, 2004). However, if growth arrest cannot be induced -due to the sustained inactivation of pRb-associated pathways by E7 -inhibition of E6 alone will result in Bax-induced apoptosis.…”

Silencing of human papillomavirus (HPV) E6/E7 oncogene expression affects both the contents and the amounts of extracellular microvesicles released from HPV‐positive cancer cells

“…58 HPVs interfere with both pathways through the activity of the viral E6 and E7 oncoproteins, 3 and reactivation of p53 and pRb activities after E6/E7 inhibition is linked to senescence induction. [35][36][37]57 This reconstitution of p53, as also observed under our experimental conditions, could affect exosome release as p53 can stimulate the TSAP6 and CHMP4C genes, which both enhance exosome formation by unknown mechanisms. [38][39][40][41] In line with this possibility, we found that the enhanced intracellular p53 levels on E6/E7 inhibition were linked to increased expression of both TSAP6 and CHMP4C in HeLa cells.…”

Human fibroblasts require the Rb family of tumor suppressors, but not p53, for PML-induced senescence

“…Our results suggest that a senescence response controlled by PML could be an important barrier to prevent the establishment of cervical carcinoma. Consistent with this idea, blocking the expression of E6 and E7 in cell lines derived from cervical carcinoma re-establishes the functions of p53 and Rb and induces senescence (Goodwin et al, 2000;Wells et al, 2000). Since PML is a critical regulator of senescence, studying the status of PML could be useful to predict the outcome of HPV infections of the cervix or the prognosis of premalignant cervical lesions.…”