p53-Mediated DNA Repair Responses to UV Radiation: Studies of Mouse Cells Lacking p53, p21, and/or gadd45 Genes

doi:10.1128/MCB.20.10.3705-3714.2000

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Molecular and Cellular Biology, May 2000, p. 3705-3714, Vol. 20, No. 10
0270-7306/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

p53-Mediated DNA Repair Responses to UV Radiation: Studies of Mouse Cells Lacking p53, p21, and/or gadd45 Genes

Martin L. Smith,¹^,^* James M. Ford,²^, M. Christine Hollander,¹ Rachel A. Bortnick,¹ Sally A. Amundson,¹ Young R. Seo,³ Chu-Xia Deng,⁴ Philip C. Hanawalt,² and Albert J. Fornace Jr.¹

Division of Basic Science, National Cancer Institute,¹ and National Institute of Diabetes, Digestive, and Kidney Diseases,⁴ National Institutes of Health, Bethesda, Maryland 20892; Department of Biological Sciences, Stanford University, Stanford, California 94305²; and Indiana University Cancer Center/Walther Oncology Center and Department of Microbiology, Indiana University School of Medicine, Indianapolis, Indiana 46202³

Received 14 January 2000/Returned for modification 17 February 2000/Accepted 22 February 2000

Human cells lacking functional p53 exhibit a partial deficiency in nucleotide excision repair (NER), the pathway for repairof UV-induced DNA damage. The global genomic repair (GGR) subpathwayof NER, but not transcription-coupled repair (TCR), is mainlyaffected by p53 loss or inactivation. We have utilized mouse embryofibroblasts (MEFs) lacking p53 genes or downstream effector genesof the p53 pathway, gadd45 (Gadd45a) or p21 (Cdkn1a), as wellas MEFs lacking both gadd45 and p21 genes to address the potentialcontribution of these downstream effectors to p53-associated DNArepair. Loss of p53 or gadd45 had a pronounced effect on GGR,while p21 loss had only a marginal effect, determined by measurementsof repair synthesis (unscheduled DNA synthesis), by immunoassaysto detect removal of UV photoproducts from genomic DNA, and byassays determining strand-specific removal of CPDs from the mousedhfr gene. Taken together, the evidence suggests a role for Gadd45,but relatively little role for p21, in DNA repair responses toUV radiation. Recent evidence suggests that Gadd45 binds to UV-damagedchromatin and may affect lesion accessibility. MEFs lacking p53or gadd45 genes exhibited decreased colony-forming ability afterUV radiation and cisplatin compared to wild-type MEFs, indicatingtheir sensitivity to DNA damage. We provide evidence that Gadd45affects chromatin remodelling of templates concurrent with DNArepair, thus indicating that Gadd45 may participate in the couplingbetween chromatin assembly and DNArepair.

^* Corresponding author. Present address: Indiana University Cancer Center, Department of Microbiology and Walther Oncology Center,R4-155, Indiana University School of Medicine, Indianapolis, IN46202.

Present address: Departments of Medicine and Genetics, Division of Oncology, Stanford University School of Medicine, Stanford,CA94305.

Molecular and Cellular Biology, May 2000, p. 3705-3714, Vol. 20, No. 10
0270-7306/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

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