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Absence of cancer–associated changes in human fibroblasts immortalized with telomerase

Abstract

The ectopic expression of telomerase1,2 in normal human cells results in an extended lifespan3,4, indicating that telomere shortening regulates the timing of cellular senescence. As telomerase expression is a hallmark of cancer, we investigated the long–term effects of forced expression of human telomerase catalytic component (hTERT) in normal human fibroblasts. In vitro growth requirements, cell–cycle checkpoints and karyotypic stability in telomerase–expressing cells are similar to those of untransfected controls. In addition, co–expression of telomerase, the viral oncoproteins HPV16 E6/E7 (which inactivate p53 and pRB) and oncogenic HRAS does not result in growth in soft agar. Thus, although ectopic expression of telomerase in human fibroblasts is sufficient for immortalization, it does not result in changes typically associated with malignant transformation.

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Figure 1: Growth rates and relative telomerase activity in fibroblasts transfected with TERT.
Figure 2: The retinoblastoma protein phosphorylation status in normal and telomerase–expressing fibroblasts.
Figure 3: Effects of UV irradiation on the expression of p53 and p21Cip1/Waf1 protein in TERT–expressing fibroblast clones, B52 and G6, measured by western–blot analysis.
Figure 4: Oncogenic HRAS induces a senescence–like phenotype in telomerase–expressing fibroblasts.
Figure 5: Lack of anchorage–independent growth of telomerase–expressing fibroblasts expressing HPV16 E6/E7 and oncogenic HRAS.

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References

  1. Nakamura, T.M. et al. Telomerase catalytic subunit homologs from fission yeast and human. Science 277, 955– 959 (1997).

    Article  CAS  Google Scholar 

  2. Meyerson, M. et al. hEST2, the putative human telomerase catalytic subunit gene, is up–regulated in tumor cells and during immortalization. Cell 90, 785–795 ( 1997).

    Article  CAS  Google Scholar 

  3. Bodnar, A.G. et al. Extension of life–span by introduction of telomerase into normal human cells. Science 279, 349 –352 (1998).

    Article  CAS  Google Scholar 

  4. Vaziri, H. & Benchimol, S. Reconstitution of telomerase activity in normal human cells leads to elongation of telomeres and extended replicative life span. Curr. Biol. 8, 279– 282 (1998).

    Article  CAS  Google Scholar 

  5. Shay, J.W., Wright, W.E. & Werbin, H. Defining the molecular mechanisms of human cell immortalization. Biochim. Biophys. Acta 1072, 1– 7 (1991).

    CAS  PubMed  Google Scholar 

  6. Abercrombie, M. Contact inhibition in tissue culture. In Vitro 6, 128–142 (1970).

    Article  CAS  Google Scholar 

  7. Scher, C.D. & Todaro, G.J. Selective growth of human neoplastic cells in medium lacking serum growth factor. Exp. Cell Res. 68, 479–481 (1971).

    Article  CAS  Google Scholar 

  8. Buchovitch, K., Duffy, L.A. & Harlow, E. The retinoblastoma protein is phosphorylated during specific phases of the cell cycle. Cell 58, 1097–1105 (1989).

    Article  Google Scholar 

  9. DeCaprio, J.A. et al. The product of the retinoblastoma susceptibility gene has properties of a cell cycle regulatory element. Cell 58, 1085–1095 (1989).

    Article  CAS  Google Scholar 

  10. Kastan, M.B., Onyekwere, O., Sidransky, D., Vogelstein, B. & Craig, R.W. Participation of p53 protein in the cellular response to DNA damage. Cancer Res. 51 , 6304–6311 (1991).

    CAS  Google Scholar 

  11. Levine, A.J., Momand, J. & Finlay, C.A. The p53 tumor suppressor gene. Nature 351, 453–456 (1991).

    Article  CAS  Google Scholar 

  12. DiLeonardo, A., Linke, S.P., Clarkin, K. & Wahl, G.M. DNA damage triggers a prolonged p53–dependent G1 arrest and long term induction of Cip1 in normal human fibroblasts. Genes Dev. 8, 2540–2551 (1994).

    Article  CAS  Google Scholar 

  13. Brown, J.P., Wei, W. & Sedivy, J.M. Bypass of senescence after disruption of p21Cip1/Waf1 gene in normal diploid human fibroblasts. Science 277, 831–834 (1997).

    Article  CAS  Google Scholar 

  14. White, A.E., Livanos, E.M. & Tlsty, T.D. Differential disruption of genomic integrity and cell cycle regulation in normal human fibroblasts by the HPV oncoproteins. Genes Dev. 8, 666–677 ( 1994).

    Article  CAS  Google Scholar 

  15. Serrano, M., Lin, A.W., McCurrach, M.E., Beach, D. & Lowe, S.W. Oncogenic ras provokes premature cell senescence associated with accumulation of p53 and p16INK4a. Cell 88, 593–602 (1997).

    Article  CAS  Google Scholar 

  16. Dimri, G.P. et al. A biomarker that identifies senscent human cells in culture and in aging skin in vivo. Proc. Natl Acad. Sci. USA 92, 9363–9367 (1995).

    Article  CAS  Google Scholar 

  17. Land, H., Parada, L.F. & Weinberg, R.A. Tumorigenic conversion of primary embryo fibroblasts requires at least two cooperating oncogenes. Nature 304, 596–602 (1983).

    Article  CAS  Google Scholar 

  18. Newbold, R.F. & Overell, R.W. Fibroblast immortality is a prerequisite for transformation by EJ c–Ha–ras oncogene. Nature 304, 648–651 ( 1983).

    Article  CAS  Google Scholar 

  19. Sager, R. Senescence as a mode of tumor suppression. Environ. Health Perspect. 93, 59–62 ( 1991).

    Article  CAS  Google Scholar 

  20. Nowell, P.C. & Croce, C.M. Cytogenetics of neoplasia. in Development and Recognition of the Transformed Cell (eds Greene, M.I. & Hamaoka, T.) 1–19 (Plenum, New York, 1987).

    Google Scholar 

  21. Honda, T. et al. Spontaneous immortalization of cultured skin fibroblasts obtained from a high–dose atomic bomb survivor. Mutat. Res. 354, 15–26 ( 1996).

    Article  Google Scholar 

  22. Saksela, E. & Moorhead, P.S. Aneuploidy in the degenerative phase of serial cultivation of human cell strains. Proc. Natl Acad. Sci. USA 50, 390–395 ( 1963).

    Article  CAS  Google Scholar 

  23. Benn, P.A. Specific chromosome abberations in senescent fibroblast cell lines derived from human embryos. Am. J. Hum. Genet. 28, 465–473 (1976).

    CAS  PubMed  PubMed Central  Google Scholar 

  24. Halbert, C.L., Demers, G.W. & Galloway, D.A. The E6 and E7 genes of human papillomavirus type 6 have weak immortalizing activity in human epithelial cells. J. Virol. 66, 2125–2134 ( 1992).

    CAS  PubMed  PubMed Central  Google Scholar 

  25. Holt, S.E., Gollahon, L.S., Willingham, T., Barbosa, M.S. & Shay, J.W. p53 levels in human mammary epithelial cells expressing wild–type and mutant human papilloma virus type 16 (HPV–16) E6 proteins: relationship to reactivation of telomerase and immortalization. Inter. J. Oncol. 8, 263 –270 (1996).

    CAS  Google Scholar 

  26. Shay, J.W., Tomlinson, G., Piatyszek, M.A. & Gollahon, L.S. Spontaneous in vitro immortalization of breast epithelial cells from a patient with Li–Fraumeni syndrome. Mol. Cell. Biol. 15, 425–432 (1995).

    Article  CAS  Google Scholar 

  27. Holt, S.E., Aisner, D.L., Shay, J.W. & Wright, W.E. Lack of cell cycle regulation of telomerase activity in human cells. Proc. Natl Acad. Sci. USA 94, 10687–10692 (1997).

    Article  CAS  Google Scholar 

  28. Clark, G.J., Cox, A.D., Graham, S.M. & Der, C.J. in Methods in Enzymology (eds Balch, W.E., Der, C.J. & Hall, A.) 395– 412 (Academic Press, San Diego, 1995).

    Google Scholar 

  29. Gustashaw K.M. in The AGT Cytogenetics Laboratory Manual (eds Barch, M.J., Knutsen, T. & Spurbeck, J.L.) 259–324 (Lippincott–Raven, Philadelphia, 1997).

    Google Scholar 

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Acknowledgements

We acknowledge J. Rohde, M. Liao, D. Cohen, J. Doolittle, S. Donovan and S. Patel for excellent technical assistance. This work was supported in part by the National Institutes of Aging (AG07992), the National Cancer Institute (CA71443) and Geron Corporation. C.P.M. is supported by The Robert Wood Johnson Foundation Minority Medical Faculty Development Program. J.W.S. is an Ellison Medical Foundation Senior Scholar. S.E.H. was supported by a postdoctoral fellowship from the National Institute of Aging.

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Correspondence to Jerry W. Shay.

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Morales, C., Holt, S., Ouellette, M. et al. Absence of cancer–associated changes in human fibroblasts immortalized with telomerase. Nat Genet 21, 115–118 (1999). https://doi.org/10.1038/5063

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