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Nucleic Acids Res. 2011 Aug;39(15):6500-10. doi: 10.1093/nar/gkr257. Epub 2011 Apr 29.

Restoration of G1 chemo/radioresistance and double-strand-break repair proficiency by wild-type but not endonuclease-deficient Artemis.

Author information

1
Department of Pharmacology and Toxicology, Massey Cancer Center, Virginia Commonwealth University, Richmond, VA 23298, USA.

Abstract

Deficiency in Artemis is associated with lack of V(D)J recombination, sensitivity to radiation and radiomimetic drugs, and failure to repair a subset of DNA double-strand breaks (DSBs). Artemis harbors an endonuclease activity that trims both 5'- and 3'-ends of DSBs. To examine whether endonucleolytic trimming of terminally blocked DSBs by Artemis is a biologically relevant function, Artemis-deficient fibroblasts were stably complemented with either wild-type Artemis or an endonuclease-deficient D165N mutant. Wild-type Artemis completely restored resistance to γ-rays, bleomycin and neocarzinostatin, and also restored DSB-repair proficiency in G0/G1 phase as measured by pulsed-field gel electrophoresis and repair focus resolution. In contrast, cells expressing the D165N mutant, even at very high levels, remained as chemo/radiosensitive and repair deficient as the parental cells, as evidenced by persistent γ-H2AX, 53BP1 and Mre11 foci that slowly increased in size and ultimately became juxtaposed with promyelocytic leukemia protein nuclear bodies. In normal fibroblasts, overexpression of wild-type Artemis increased radioresistance, while D165N overexpression conferred partial repair deficiency following high-dose radiation. Restoration of chemo/radioresistance by wild-type, but not D165N Artemis suggests that the lack of endonucleolytic trimming of DNA ends is the principal cause of sensitivity to double-strand cleaving agents in Artemis-deficient cells.

PMID:
21531702
PMCID:
PMC3159448
DOI:
10.1093/nar/gkr257
[Indexed for MEDLINE]
Free PMC Article

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