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Differential stability of the DNA-activated protein kinase catalytic subunit mRNA in human glioma cells

Oncogene volume 18, pages 1361–1368 (1999)Cite this article

Abstract

DNA-dependent protein kinase (DNA-PK) functions in double-strand break repair and immunoglobulin [V(D)J] recombination. We previously established a radiation-sensitive human cell line, M059J, derived from a malignant glioma, which lacks the catalytic subunit (DNA-PKcs) of the DNA-PK multiprotein complex. Although previous Northern blot analysis failed to detect the DNA-PKcs transcript in these cells, we show here through quantitative studies that the transcript is present, albeit at greatly reduced (∼20x) levels. Sequencing revealed no genetic alteration in either the promoter region, the kinase domain, or the 3′ untranslated region of the DNA-PKcs gene to account for the reduced transcript levels. Nuclear run-on transcription assays indicated that the rate of DNA-PKcs transcription in M059J and DNA-PKcs proficient cell lines was similar, but the stability of the DNA-PKcs message in the M059J cell line was drastically (∼20x) reduced. Furthermore, M059J cells lack an alternately spliced DNA-PKcs transcript that accounts for a minor (5 – 20%) proportion of the DNA-PKcs message in all other cell lines tested. Thus, alterations in DNA-PKcs mRNA stability and/or the lack of the alternate mRNA may result in the loss of DNA-PKcs activity. This finding has important implications as DNA-PKcs activity is essential to cells repairing damage induced by radiation or radiomimetric agents.

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Acknowledgements

We would like to thank Dr Roseline Godbout and Dr Susan Lees-Miller for helpful discussions and critically reading the manuscript. This work has been supported by grants from the National Cancer Institute of Canada and the Medical Research Council. CWA is supported by the Office of Health and Environmental Research of the US Department of Energy and by US Public Health grant GM52825.

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Authors and Affiliations

  1. Experimental Oncology, Cross Cancer Institute, University of Alberta, Edmonton, Alberta, T6G 1Z2, Canada

    Anne M Galloway, Charlotte A Spencer & M Joan Allalunis-Turner

  2. Department of Oncology, University of Alberta, Edmonton, Alberta, T6G 1Z2, Canada

    Charlotte A Spencer & M Joan Allalunis-Turner

  3. Biology Department, Brookhaven National Laboratory, Upton, New York, 11973, USA

    Carl W Anderson

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  1. Anne M Galloway
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Galloway, A., Spencer, C., Anderson, C. et al. Differential stability of the DNA-activated protein kinase catalytic subunit mRNA in human glioma cells. Oncogene 18, 1361–1368 (1999). https://doi.org/10.1038/sj.onc.1202433

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