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Alternative recombination pathways in UV-irradiated XP variant cells

Oncogene volume 24, pages 3708–3714 (2005)Cite this article

Abstract

XP variant (XP-V) cells lack the damage-specific polymerase η and exhibit prolonged replication arrest after UV irradiation due to impaired bypass of UV photoproducts. To analyse the outcome of the arrested replication forks, homologous recombination (HR, Rad51 events) and fork breakage (Rad50 events) were assayed by immunofluorescent detection of foci-positive cells. Within 1 h of irradiation, XP-V cells showed more Rad51-positive cells than normal cells, while neither cell type showed an increase in Rad50 foci. Beyond 1 h, the frequency of Rad51-positive cells reached similar levels in both cell types, then declined at higher UV doses. At these later times, Rad50-positive cells increased with dose and to a greater extent in XP-V cells. Few cells were simultaneously positive for both sets of foci, suggesting a mutually exclusive recruitment of recombination proteins, or that these pathways operate at different stages during S phase. Analysis of cells containing a vector of tandemly arranged enhanced green fluorescent protein genes also showed that UV-induced HR was higher in XP-V cells. These results suggest that cells make an early commitment to HR and that at later times a subset of arrested forks degrade into double-strand breaks, two alternative pathways that are greater in XP-V cells.

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Acknowledgements

The work described here was supported by the following grants: American Cancer Society RPG-00-036-01-CNE and NASA NAG2-1632 (to CLL); the National Institutes of Environmental Health Sciences grant 1 RO1 ES 8061 and the Ellison Senior Scholar Award (to JEC). We are also grateful to the XP Society, Poughkeepsie, NY, for their continued support and encouragement to one of us (JEC). We thank Dr William Hyun, Molecular Diagnostics, UCSF Cancer Center, for assistance in immunofluorescence.

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  1. Radiation Oncology Research Laboratory, Department of Radiation Oncology, University of California, 1855 Folsom St., MCB-200, San Francisco, CA, 94103-0806, USA

    Charles L Limoli & E Giedzinski

  2. Auerback Melanoma Laboratory, UCSF Cancer Center, University of California, Room N431, Box 0808, San Francisco, CA, 94143-0808, USA

    J E Cleaver

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  1. Charles L Limoli
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  2. E Giedzinski
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Limoli, C., Giedzinski, E. & Cleaver, J. Alternative recombination pathways in UV-irradiated XP variant cells. Oncogene 24, 3708–3714 (2005). https://doi.org/10.1038/sj.onc.1208515

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