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C600071200v1
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Papers In Press, published online ahead of print March 29, 2006
J. Biol. Chem, 10.1074/jbc.C600071200
Submitted on March 22, 2006
Revised on March 29, 2006
Accepted on March 29, 2006

REV3 and REV1 play major roles in recombination-independent repair of DNA interstrand cross-links mediated by monoubiquitinated PCNA

Xi Shen, Sohee Jun, Lindsey E. O'Neal, Eiichiro Sonoda, Mats Bemark, Julian E. Sale, and Lei Li

Experimental Radiation Oncology, The University of Texas M. D. Anderson Cancer Center, Houston, TX 77030

Corresponding Author: Leili{at}mdanderson.org

DNA interstrand cross-links (ICLs) are the most cytotoxic lesions to eukaryotic genome and are repaired by both homologous recombination-dependent and -independent mechanisms. To better understand the role of lesion bypass polymerases in ICL repair, we investigated recombination-independent repair of ICLs in REV3 and REV1 deletion mutants constructed in avian DT40 cells and mouse embryonic fibroblast cells. Our results showed that Rev3 plays a major role in recombination-independent ICL repair, which may account for the extreme sensitivity of REV3 mutants to cross-linking agents. This result raised the possibility that the NER gap synthesis, when encountering an adducted base present in the ICL repair intermediate, can lead to recruitment of Rev3, analogous to the recruitment of Pol eta during replicative synthesis. Indeed, the monoubiquitination-defective PCNA mutant exhibits impaired recombination-independent ICL repair as well as drastically reduced mutation rate, indicating that the PCNA switch is utilized to enable lesion bypass during DNA repair synthesis. Analyses of a REV1 deletion mutant also revealed a significant reduction in recombination-independent ICL repair, suggesting that Rev1 cooperates with Rev3 in recombination-independent ICL repair. Moreover, deletion of REV3 or REV1 significantly altered the spectrum of mutations resulting from ICL repair, further confirming their involvement in mutagenic repair of ICLs.


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