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Mol Cell Biol. Mar 1994; 14(3): 1815–1823.
PMCID: PMC358539

DNA strand breaks: the DNA template alterations that trigger p53-dependent DNA damage response pathways.


The tumor suppressor protein p53 serves as a critical regulator of a G1 cell cycle checkpoint and of apoptosis following exposure of cells to DNA-damaging agents. The mechanism by which DNA-damaging agents elevate p53 protein levels to trigger G1/S arrest or cell death remains to be elucidated. In fact, whether damage to the DNA template itself participates in transducing the signal leading to p53 induction has not yet been demonstrated. We exposed human cell lines containing wild-type p53 alleles to several different DNA-damaging agents and found that agents which rapidly induce DNA strand breaks, such as ionizing radiation, bleomycin, and DNA topoisomerase-targeted drugs, rapidly triggered p53 protein elevations. In addition, we determined that camptothecin-stimulated trapping of topoisomerase I-DNA complexes was not sufficient to elevate p53 protein levels; rather, replication-associated DNA strand breaks were required. Furthermore, treatment of cells with the antimetabolite N(phosphonoacetyl)-L-aspartate (PALA) did not cause rapid p53 protein increases but resulted in delayed increases in p53 protein levels temporally correlated with the appearance of DNA strand breaks. Finally, we concluded that DNA strand breaks were sufficient for initiating p53-dependent signal transduction after finding that introduction of nucleases into cells by electroporation stimulated rapid p53 protein elevations. While DNA strand breaks appeared to be capable of triggering p53 induction, DNA lesions other than strand breaks did not. Exposure of normal cells and excision repair-deficient xeroderma pigmentosum cells to low doses of UV light, under conditions in which thymine dimers appear but DNA replication-associated strand breaks were prevented, resulted in p53 induction attributable to DNA strand breaks associated with excision repair. Our data indicate that DNA strand breaks are sufficient and probably necessary for p53 induction in cells with wild-type p53 alleles exposed to DNA-damaging agents.

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  • Almon E, Goldfinger N, Kapon A, Schwartz D, Levine AJ, Rotter V. Testicular tissue-specific expression of the p53 suppressor gene. Dev Biol. 1993 Mar;156(1):107–116. [PubMed]
  • Andreassen PR, Margolis RL. 2-Aminopurine overrides multiple cell cycle checkpoints in BHK cells. Proc Natl Acad Sci U S A. 1992 Mar 15;89(6):2272–2276. [PMC free article] [PubMed]
  • Avemann K, Knippers R, Koller T, Sogo JM. Camptothecin, a specific inhibitor of type I DNA topoisomerase, induces DNA breakage at replication forks. Mol Cell Biol. 1988 Aug;8(8):3026–3034. [PMC free article] [PubMed]
  • Benjamin RC, Gill DM. Poly(ADP-ribose) synthesis in vitro programmed by damaged DNA. A comparison of DNA molecules containing different types of strand breaks. J Biol Chem. 1980 Nov 10;255(21):10502–10508. [PubMed]
  • Borek C, Ong A, Morgan WF, Cleaver JE. Morphological transformation of 10T1/2 mouse embryo cells can be initiated by DNA double-strand breaks alone. Mol Carcinog. 1991;4(3):243–247. [PubMed]
  • Bryant PE. Enzymatic restriction of mammalian cell DNA using Pvu II and Bam H1: evidence for the double-strand break origin of chromosomal aberrations. Int J Radiat Biol Relat Stud Phys Chem Med. 1984 Jul;46(1):57–65. [PubMed]
  • Bryant PE. Use of restriction endonucleases to study relationships between DNA double-strand breaks, chromosomal aberrations and other end-points in mammalian cells. Int J Radiat Biol. 1988 Dec;54(6):869–890. [PubMed]
  • Bryant PE, Riches AC. Oncogenic transformation of murine C3H 10T1/2 cells resulting from DNA double-strand breaks induced by a restriction endonuclease. Br J Cancer. 1989 Dec;60(6):852–854. [PMC free article] [PubMed]
  • Busse PM, Bose SK, Jones RW, Tolmach LJ. The action of caffeine on X-irradiated HeLa cells. II. Synergistic lethality. Radiat Res. 1977 Sep;71(3):666–677. [PubMed]
  • Chen GL, Yang L, Rowe TC, Halligan BD, Tewey KM, Liu LF. Nonintercalative antitumor drugs interfere with the breakage-reunion reaction of mammalian DNA topoisomerase II. J Biol Chem. 1984 Nov 10;259(21):13560–13566. [PubMed]
  • Chung HW, Phillips JW, Winegar RA, Preston RJ, Morgan WF. Modulation of restriction enzyme-induced damage by chemicals that interfere with cellular responses to DNA damage: a cytogenetic and pulsed-field gel analysis. Radiat Res. 1991 Jan;125(1):107–113. [PubMed]
  • Clarke AR, Purdie CA, Harrison DJ, Morris RG, Bird CC, Hooper ML, Wyllie AH. Thymocyte apoptosis induced by p53-dependent and independent pathways. Nature. 1993 Apr 29;362(6423):849–852. [PubMed]
  • D'Arpa P, Beardmore C, Liu LF. Involvement of nucleic acid synthesis in cell killing mechanisms of topoisomerase poisons. Cancer Res. 1990 Nov 1;50(21):6919–6924. [PubMed]
  • Devary Y, Gottlieb RA, Smeal T, Karin M. The mammalian ultraviolet response is triggered by activation of Src tyrosine kinases. Cell. 1992 Dec 24;71(7):1081–1091. [PubMed]
  • Donehower LA, Harvey M, Slagle BL, McArthur MJ, Montgomery CA, Jr, Butel JS, Bradley A. Mice deficient for p53 are developmentally normal but susceptible to spontaneous tumours. Nature. 1992 Mar 19;356(6366):215–221. [PubMed]
  • Dunn WC, Regan JD. Inhibition of DNA excision repair in human cells by arabinofuranosyl cytosine: effect on normal and xeroderma pigmentosum cells. Mol Pharmacol. 1979 Mar;15(2):367–374. [PubMed]
  • Elia MC, DeLuca JG, Bradley MO. Significance and measurement of DNA double strand breaks in mammalian cells. Pharmacol Ther. 1991;51(3):291–327. [PubMed]
  • Fornace AJ., Jr Mammalian genes induced by radiation; activation of genes associated with growth control. Annu Rev Genet. 1992;26:507–526. [PubMed]
  • Fornace AJ, Jr, Nebert DW, Hollander MC, Luethy JD, Papathanasiou M, Fargnoli J, Holbrook NJ. Mammalian genes coordinately regulated by growth arrest signals and DNA-damaging agents. Mol Cell Biol. 1989 Oct;9(10):4196–4203. [PMC free article] [PubMed]
  • Fornace AJ, Jr, Seres DS. Detection of DNA single-strand breaks during the repair of UV damage in xeroderma pigmentosum cells. Radiat Res. 1983 Jan;93(1):107–111. [PubMed]
  • Fritsche M, Haessler C, Brandner G. Induction of nuclear accumulation of the tumor-suppressor protein p53 by DNA-damaging agents. Oncogene. 1993 Feb;8(2):307–318. [PubMed]
  • George J, Castellazzi M, Buttin G. Prophage induction and cell division in E. coli. III. Mutations sfiA and sfiB restore division in tif and lon strains and permit the expression of mutator properties of tif. Mol Gen Genet. 1975 Oct 22;140(4):309–332. [PubMed]
  • Giaccia AJ, MacLaren RA, Denko N, Nicolaou D, Stamato TD. Increased sensitivity to killing by restriction enzymes in the XR-1 DNA double-strand break repair-deficient mutant. Mutat Res. 1990 Jul;236(1):67–76. [PubMed]
  • Gottlieb TM, Jackson SP. The DNA-dependent protein kinase: requirement for DNA ends and association with Ku antigen. Cell. 1993 Jan 15;72(1):131–142. [PubMed]
  • Hall PA, McKee PH, Menage HD, Dover R, Lane DP. High levels of p53 protein in UV-irradiated normal human skin. Oncogene. 1993 Jan;8(1):203–207. [PubMed]
  • Hartwell LH, Weinert TA. Checkpoints: controls that ensure the order of cell cycle events. Science. 1989 Nov 3;246(4930):629–634. [PubMed]
  • Horoszewicz JS, Leong SS, Kawinski E, Karr JP, Rosenthal H, Chu TM, Mirand EA, Murphy GP. LNCaP model of human prostatic carcinoma. Cancer Res. 1983 Apr;43(4):1809–1818. [PubMed]
  • Hsiang YH, Lihou MG, Liu LF. Arrest of replication forks by drug-stabilized topoisomerase I-DNA cleavable complexes as a mechanism of cell killing by camptothecin. Cancer Res. 1989 Sep 15;49(18):5077–5082. [PubMed]
  • Huisman O, D'Ari R. An inducible DNA replication-cell division coupling mechanism in E. coli. Nature. 1981 Apr 30;290(5809):797–799. [PubMed]
  • Isaacs WB, Carter BS, Ewing CM. Wild-type p53 suppresses growth of human prostate cancer cells containing mutant p53 alleles. Cancer Res. 1991 Sep 1;51(17):4716–4720. [PubMed]
  • Jeggo PA. Studies on mammalian mutants defective in rejoining double-strand breaks in DNA. Mutat Res. 1990 Jul;239(1):1–16. [PubMed]
  • Kastan MB, Onyekwere O, Sidransky D, Vogelstein B, Craig RW. Participation of p53 protein in the cellular response to DNA damage. Cancer Res. 1991 Dec 1;51(23 Pt 1):6304–6311. [PubMed]
  • Kaufmann WK, Boyer JC, Estabrooks LL, Wilson SJ. Inhibition of replicon initiation in human cells following stabilization of topoisomerase-DNA cleavable complexes. Mol Cell Biol. 1991 Jul;11(7):3711–3718. [PMC free article] [PubMed]
  • Kessis TD, Slebos RJ, Nelson WG, Kastan MB, Plunkett BS, Han SM, Lorincz AT, Hedrick L, Cho KR. Human papillomavirus 16 E6 expression disrupts the p53-mediated cellular response to DNA damage. Proc Natl Acad Sci U S A. 1993 May 1;90(9):3988–3992. [PMC free article] [PubMed]
  • Kuerbitz SJ, Plunkett BS, Walsh WV, Kastan MB. Wild-type p53 is a cell cycle checkpoint determinant following irradiation. Proc Natl Acad Sci U S A. 1992 Aug 15;89(16):7491–7495. [PMC free article] [PubMed]
  • Lau CC, Pardee AB. Mechanism by which caffeine potentiates lethality of nitrogen mustard. Proc Natl Acad Sci U S A. 1982 May;79(9):2942–2946. [PMC free article] [PubMed]
  • Li JC, Kaminskas E. Accumulation of DNA strand breaks and methotrexate cytotoxicity. Proc Natl Acad Sci U S A. 1984 Sep;81(18):5694–5698. [PMC free article] [PubMed]
  • Liu LF. DNA topoisomerase poisons as antitumor drugs. Annu Rev Biochem. 1989;58:351–375. [PubMed]
  • Liu LF, Rowe TC, Yang L, Tewey KM, Chen GL. Cleavage of DNA by mammalian DNA topoisomerase II. J Biol Chem. 1983 Dec 25;258(24):15365–15370. [PubMed]
  • Livingstone LR, White A, Sprouse J, Livanos E, Jacks T, Tlsty TD. Altered cell cycle arrest and gene amplification potential accompany loss of wild-type p53. Cell. 1992 Sep 18;70(6):923–935. [PubMed]
  • Lowe SW, Schmitt EM, Smith SW, Osborne BA, Jacks T. p53 is required for radiation-induced apoptosis in mouse thymocytes. Nature. 1993 Apr 29;362(6423):847–849. [PubMed]
  • Luchnik AN, Hisamutdinov TA, Georgiev GP. Inhibition of transcription in eukaryotic cells by X-irradiation: relation to the loss of topological constraint in closed DNA loops. Nucleic Acids Res. 1988 Jun 10;16(11):5175–5190. [PMC free article] [PubMed]
  • Malkin D, Li FP, Strong LC, Fraumeni JF, Jr, Nelson CE, Kim DH, Kassel J, Gryka MA, Bischoff FZ, Tainsky MA, et al. Germ line p53 mutations in a familial syndrome of breast cancer, sarcomas, and other neoplasms. Science. 1990 Nov 30;250(4985):1233–1238. [PubMed]
  • Maltzman W, Czyzyk L. UV irradiation stimulates levels of p53 cellular tumor antigen in nontransformed mouse cells. Mol Cell Biol. 1984 Sep;4(9):1689–1694. [PMC free article] [PubMed]
  • Morgan WF, Yates BL, Rufer JT, Abella Columna E, Valcarcel ER, Phillips JW. Chromosomal aberration induction in CHO cells by combined exposure to restriction enzymes and X-rays. Int J Radiat Biol. 1991 Oct;60(4):627–634. [PubMed]
  • O'Connor PM, Ferris DK, White GA, Pines J, Hunter T, Longo DL, Kohn KW. Relationships between cdc2 kinase, DNA cross-linking, and cell cycle perturbations induced by nitrogen mustard. Cell Growth Differ. 1992 Jan;3(1):43–52. [PubMed]
  • Robbins JH, Kraemer KH, Lutzner MA, Festoff BW, Coon HG. Xeroderma pigmentosum. An inherited diseases with sun sensitivity, multiple cutaneous neoplasms, and abnormal DNA repair. Ann Intern Med. 1974 Feb;80(2):221–248. [PubMed]
  • Ryan AJ, Squires S, Strutt HL, Johnson RT. Camptothecin cytotoxicity in mammalian cells is associated with the induction of persistent double strand breaks in replicating DNA. Nucleic Acids Res. 1991 Jun 25;19(12):3295–3300. [PMC free article] [PubMed]
  • Schwartz D, Goldfinger N, Rotter V. Expression of p53 protein in spermatogenesis is confined to the tetraploid pachytene primary spermatocytes. Oncogene. 1993 Jun;8(6):1487–1494. [PubMed]
  • Schwartz JL. Monofunctional alkylating agent-induced S-phase-dependent DNA damage. Mutat Res. 1989 Apr;216(2):111–118. [PubMed]
  • Shero JH, Bordwell B, Rothfield NF, Earnshaw WC. High titers of autoantibodies to topoisomerase I (Scl-70) in sera from scleroderma patients. Science. 1986 Feb 14;231(4739):737–740. [PubMed]
  • Slichenmyer WJ, Nelson WG, Slebos RJ, Kastan MB. Loss of a p53-associated G1 checkpoint does not decrease cell survival following DNA damage. Cancer Res. 1993 Sep 15;53(18):4164–4168. [PubMed]
  • Srivastava S, Zou ZQ, Pirollo K, Blattner W, Chang EH. Germ-line transmission of a mutated p53 gene in a cancer-prone family with Li-Fraumeni syndrome. Nature. 1990 Dec 20;348(6303):747–749. [PubMed]
  • Tewey KM, Rowe TC, Yang L, Halligan BD, Liu LF. Adriamycin-induced DNA damage mediated by mammalian DNA topoisomerase II. Science. 1984 Oct 26;226(4673):466–468. [PubMed]
  • Tishler RB, Calderwood SK, Coleman CN, Price BD. Increases in sequence specific DNA binding by p53 following treatment with chemotherapeutic and DNA damaging agents. Cancer Res. 1993 May 15;53(10 Suppl):2212–2216. [PubMed]
  • Tolmach LJ, Jones RW, Busse PM. The action of caffeine on X-irradiated HeLa cells. I. Delayed inhibition of DNA synthesis. Radiat Res. 1977 Sep;71(3):653–665. [PubMed]
  • Wang TC, Smith KC. Postreplication repair in ultraviolet-irradiated human fibroblasts: formation and repair of DNA double-strand breaks. Carcinogenesis. 1986 Mar;7(3):389–392. [PubMed]
  • Walters RA, Gurley LR, Tobey RA. Effects of caffeine on radiation-induced phenomena associated with cell-cycle traverse of mammalian cells. Biophys J. 1974 Feb;14(2):99–118. [PMC free article] [PubMed]
  • Weinert TA, Hartwell LH. The RAD9 gene controls the cell cycle response to DNA damage in Saccharomyces cerevisiae. Science. 1988 Jul 15;241(4863):317–322. [PubMed]
  • Weinert TA, Hartwell LH. Characterization of RAD9 of Saccharomyces cerevisiae and evidence that its function acts posttranslationally in cell cycle arrest after DNA damage. Mol Cell Biol. 1990 Dec;10(12):6554–6564. [PMC free article] [PubMed]
  • Winegar RA, Phillips JW, Youngblom JH, Morgan WF. Cell electroporation is a highly efficient method for introducing restriction endonucleases into cells. Mutat Res. 1989 Jan-Feb;225(1-2):49–53. [PubMed]
  • Witkin EM. The radiation sensitivity of Escherichia coli B: a hypothesis relating filament formation and prophage induction. Proc Natl Acad Sci U S A. 1967 May;57(5):1275–1279. [PMC free article] [PubMed]
  • Yin Y, Tainsky MA, Bischoff FZ, Strong LC, Wahl GM. Wild-type p53 restores cell cycle control and inhibits gene amplification in cells with mutant p53 alleles. Cell. 1992 Sep 18;70(6):937–948. [PubMed]
  • Zhan Q, Carrier F, Fornace AJ., Jr Induction of cellular p53 activity by DNA-damaging agents and growth arrest. Mol Cell Biol. 1993 Jul;13(7):4242–4250. [PMC free article] [PubMed]
  • Zajac-Kaye M, Ts'o PO. DNAase I encapsulated in liposomes can induce neoplastic transformation of Syrian hamster embryo cells in culture. Cell. 1984 Dec;39(3 Pt 2):427–437. [PubMed]

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