Nucleic Acids Research Advance Access published online on July 24, 2008
Nucleic Acids Research, doi:10.1093/nar/gkn450
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Molecular Biology |
DNA repair of clustered lesions in mammalian cells: involvement of non-homologous end-joining
Department of Molecular and Cellular Physiology, Louisiana Health Sciences Center, Shreveport, LA 71130, USA
*To whom correspondence should be addressed. Tel: 318 675 4213; Fax: 318 675 4217; Email: lclary{at}lsuhsc.edu
Received December 19, 2007. Revised June 23, 2008. Accepted June 26, 2008.
Clustered lesions are defined as 
two lesions within 20 bps and are generated in DNA by ionizing radiation. In vitro studies and work in bacteria have shown that attempted repair of two closely opposed lesions can result in the formation of double strand breaks (DSBs). Since mammalian cells can repair DSBs by non-homologous end-joining (NHEJ), we hypothesized that NHEJ would repair DSBs formed during the removal of clustered tetrahydrofurans (furans). However, two opposing furans situated 2, 5 or 12 bps apart in a firefly luciferase reporter plasmid caused a decrease in luciferase activity in wild-type, Ku80 or DNA-PKcs-deficient cells, indicating the generation of DSBs. Loss of luciferase activity was maximal at 5 bps apart and studies using siRNA implicate the major AP endonuclease in the initial cleavage. Since NHEJ-deficient cells had equivalent luciferase activity to their isogenic wild-type cells, NHEJ was not involved in accurate repair of clustered lesions. However, quantitation and examination of re-isolated DNA showed that damage-containing plasmids were inaccurately repaired by Ku80-dependent, as well as Ku80-independent mechanisms. This work indicates that not even NHEJ can completely prevent the conversion of clustered lesions to potentially lethal DSBs, so demonstrating the biological relevance of ionizing radiation-induced clustered damage.
CiteULike 
Connotea 
Del.icio.us What's this?
This article has been cited by other articles:
|
|
 |
|
  S. Bellon, N. Shikazono, S. Cunniffe, M. Lomax, and P. O'Neill Processing of thymine glycol in a clustered DNA damage site: mutagenic or cytotoxic Nucleic Acids Res., July 1, 2009; 37(13): 4430 - 4440. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. G. Kozmin, Y. Sedletska, A. Reynaud-Angelin, D. Gasparutto, and E. Sage The formation of double-strand breaks at multiply damaged sites is driven by the kinetics of excision/incision at base damage in eukaryotic cells Nucleic Acids Res., April 1, 2009; 37(6): 1767 - 1777. [Abstract] [Full Text] [PDF]
|
|