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First published online 1 September 2005
doi: 10.1242/jcs.02544
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Research Article |
1 Department of Molecular Genetics, Institute of Development, Aging and Cancer, Tohoku University, Seiryomachi 4-1, Sendai 980-8575, Japan
2 Department of Genome Repair Dynamics, Radiation Biology Center, Kyoto University, Kyoto 606-8501, Japan
3 Experimental Immunology Branch, NIH, National Cancer Institute, Bethesda, MD 20892-1360, USA
4 GeneCare Research Institute, 200 Kajiwara, Kamakura, Kanagawa 247-0063, Japan
5 Department of Pathology, University of Washington, Seattle, WA 98195-7470, USA
* Author for correspondence (e-mail: ayasui{at}idac.tohoku.ac.jp)
Accepted 6 June 2005
Werner syndrome is an autosomal recessive accelerated-aging disorder caused by a defect in the WRN gene, which encodes a member of the RecQ family of DNA helicases with an exonuclease activity. In vitro experiments have suggested that WRN functions in several DNA repair processes, but the actual functions of WRN in living cells remain unknown. Here, we analyzed the kinetics of the intranuclear mobilization of WRN protein in response to a variety of types of DNA damage produced locally in the nucleus of human cells. A striking accumulation of WRN was observed at laser-induced double-strand breaks, but not at single-strand breaks or oxidative base damage. The accumulation of WRN at double-strand breaks was rapid, persisted for many hours, and occurred in the absence of several known interacting proteins including polymerase ß, poly(ADP-ribose) polymerase 1 (PARP1), Ku80, DNA-dependent protein kinase (DNA-PKcs), NBS1 and histone H2AX. Abolition of helicase activity or deletion of the exonuclease domain had no effect on accumulation, whereas the presence of the HRDC (helicase and RNaseD C-terminal) domain was necessary and sufficient for the accumulation. Our data suggest that WRN functions mainly at DNA double-strand breaks and structures resembling double-strand breaks in living cells, and that an autonomous accumulation through the HRDC domain is the initial response of WRN to the double-strand breaks.
Key words: Werner protein, Double-strand breaks, Laser irradiation, Damage accumulation, HRDC domain
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