GENOMIC INSTABILITY AND TELOMERASE ACTIVITY IN HUMAN BRONCHIAL EPITHELIAL-CELLS DURING IMMORTALIZATION BY HUMAN PAPILLOMAVIRUS-16 E6 AND E7GENES

Human papilloma virus types 16 and 18 contribute to the development of cervical carcinomas in which the E6 and E7 genes are frequently retai ned and expressed in the tumors. Our study explored the ability of the E6 and/or E7 genes to immortalize normal human bronchial epithelial ( NHBE) cells and to reactivate telomerase expression in these cells. We have introduced the human papillomavirus type 16 E6 or E7 genes alone or in combination (E6/E7) into NHBE cells using the retroviral constr uct pLXSN. Cells expressing either the E6 or the E7 oncoproteins alone displayed an increased colony-forming efficiency and a slightly exten ded in vitro life span before entering a crisis, from which immortaliz ed cell lines were not obtained. Telomerase activity was not detected in cells expressing either E6 or E7 individually. Cells expressing the E6/E7 oncoproteins in combination had a substantially increased life span before entering crisis. A subpopulation of these cells escaped fr om crisis and achieved 130 population doublings, suggesting immortaliz ation. Telomerase activity was detected in these postcrisis cells, but was not detected prior to crisis. In addition, karyotypic analysis sh owed evidence of genomic instability in mass cultures as well as clone s expressing E6, E7, or E6/E7. Abnormalities included numerous monosom ies and trisomies, chromatid gaps and breaks, double minutes, and aber rant chromosomes. These results demonstrate that expression of E6 and/ or E7 is sufficient to induce genomic instability and an extended life span to NHBE cells, but the presence of both E6 and E7, along with at least one additional genetic or epigenetic event achieved during cris is, was required for reactivation of telomerase and the immortalizatio n in this human cell type. (C) 1997 Academic Press.