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Mutant K-ras enhances apoptosis in embryonic stem cells in combination with DNA damage and is associated with increased levels of p19ARF

Oncogene volume 20, pages 2144–2152 (2001)Cite this article

Abstract

The roles of K-ras in neoplasia are not entirely understood, although there is evidence that K-ras affects susceptibility to apoptosis, modulating survival of DNA damaged cells which would otherwise be eliminated. In this study, we investigated the effects of mutant K-ras on apoptosis in vitro following DNA damage. To avoid complications resulting from mutations in other cancer-related genes and from the presence of endogenous K-ras, we derived K-ras null embryonic stem cells. Expression of either wild-type or mutant K-ras was reconstructed by stable plasmid transfection. The cell lines were treated with etoposide, cisplatin and UV radiation and apoptosis measured flow cytometrically. Mutant K-ras potentiated the effect of etoposide-derived DNA damage by increasing apoptosis, whereas absence of K-ras had the opposite effect. This pattern was similar but less marked with cisplatin, whereas UV yielded no difference in apoptosis with regard to K-ras status, suggesting that the effect of K-ras on apoptosis is dependent on the nature of the DNA damage. To investigate possible mechanisms, we examined the expression of p19ARF mRNA by RT–PCR. Cells expressing mutant K-ras produced elevated levels of p19ARF mRNA, which could link K-ras status with p53 expression and hence susceptibility to DNA damage-induced apoptosis.

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Acknowledgements

We are grateful to the Cancer Research Campaign, the Association for International Cancer Research, the Scottish Hospital Endowment Research Trust and the Royal Society for supporting this work.

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Authors and Affiliations

  1. Sir Alastair Currie CRC Laboratories, Molecular Medicine Centre, University of Edinburgh, Crewe Road, Edinburgh, EH4 2XU, UK

    David G Brooks, Roberta M James, Charles E Patek, James Williamson & Mark J Arends

  2. Department of Pathology, University of Cambridge, Addenbrooke's Hospital, Hills Road, Cambridge, CB2 2QQ, UK

    David G Brooks & Mark J Arends

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  1. David G Brooks
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Brooks, D., James, R., Patek, C. et al. Mutant K-ras enhances apoptosis in embryonic stem cells in combination with DNA damage and is associated with increased levels of p19ARF. Oncogene 20, 2144–2152 (2001). https://doi.org/10.1038/sj.onc.1204309

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