Residual γH2AX foci as an indication of lethal DNA lesions
Tóm tắt
Evidence suggests that tumor cells exposed to some DNA damaging agents are more likely to die if they retain microscopically visible γH2AX foci that are known to mark sites of double-strand breaks. This appears to be true even after exposure to the alkylating agent MNNG that does not cause direct double-strand breaks but does produce γH2AX foci when damaged DNA undergoes replication. To examine this predictive ability further, SiHa human cervical carcinoma cells were exposed to 8 DNA damaging drugs (camptothecin, cisplatin, doxorubicin, etoposide, hydrogen peroxide, MNNG, temozolomide, and tirapazamine) and the fraction of cells that retained γH2AX foci 24 hours after a 30 or 60 min treatment was compared with the fraction of cells that lost clonogenicity. To determine if cells with residual repair foci are the cells that die, SiHa cervical cancer cells were stably transfected with a RAD51-GFP construct and live cell analysis was used to follow the fate of irradiated cells with RAD51-GFP foci. For all drugs regardless of their mechanism of interaction with DNA, close to a 1:1 correlation was observed between clonogenic surviving fraction and the fraction of cells that retained γH2AX foci 24 hours after treatment. Initial studies established that the fraction of cells that retained RAD51 foci after irradiation was similar to the fraction of cells that retained γH2AX foci and subsequently lost clonogenicity. Tracking individual irradiated live cells confirmed that SiHa cells with RAD51-GFP foci 24 hours after irradiation were more likely to die. Retention of DNA damage-induced γH2AX foci appears to be indicative of lethal DNA damage so that it may be possible to predict tumor cell killing by a wide variety of DNA damaging agents simply by scoring the fraction of cells that retain γH2AX foci.
Tài liệu tham khảo
Rogakou EP, Boon C, Redon C, Bonner WM: Megabase chromatin domains involved in DNA double-strand breaks In vivo. J Cell Biol. 1999, 146: 905-916. 10.1083/jcb.146.5.905.
Bonner WM, Redon CE, Dickey JS, Nakamura AJ, Sedelnikova OA, Solier S, Pommier Y: gammaH2AX and cancer. Nat Rev Cancer. 2008, 8: 957-967. 10.1038/nrc2523.
Kinner A, Wu W, Staudt C, Iliakis G: Gamma-H2AX in recognition and signaling of DNA double-strand breaks in the context of chromatin. Nucleic Acids Res. 2008, 36: 5678-5694. 10.1093/nar/gkn550.
Banath JP, MacPhail SH, Olive PL: Radiation sensitivity, H2AX phosphorylation, and kinetics of repair of DNA strand breaks in irradiated cervical cancer cell lines. Cancer Res. 2004, 64: 7144-7149. 10.1158/0008-5472.CAN-04-1433.
Klokov D, Macphail SM, Banath JP, Byrne JP, Olive PL: Phosphorylated histone H2AX in relation to cell survival in tumor cells and xenografts exposed to single and fractionated doses of X-rays. Radiother Oncol. 2006, 80: 223-229. 10.1016/j.radonc.2006.07.026.
Sak A, Stueben G, Groneberg M, Bocker W, Stuschke M: Targeting of Rad51-dependent homologous recombination: implications for the radiation sensitivity of human lung cancer cell lines. Br J Cancer. 2005, 92: 1089-1097. 10.1038/sj.bjc.6602457.
Henning W, Sturzbecher HW: Homologous recombination and cell cycle checkpoints: Rad51 in tumour progression and therapy resistance. Toxicology. 2003, 193: 91-109. 10.1016/S0300-483X(03)00291-9.
Bocker W, Iliakis G: Computational Methods for analysis of foci: validation for radiation-induced gamma-H2AX foci in human cells. Radiat Res. 2006, 165: 113-124. 10.1667/RR3486.1.
Kalocsay M, Hiller NJ, Jentsch S: Chromosome-wide Rad51 spreading and SUMO-H2A. Z-dependent chromosome fixation in response to a persistent DNA double-strand break. Mol Cell. 2009, 33: 335-343. 10.1016/j.molcel.2009.01.016.
Paull TT, Rogakou EP, Yamazaki V, Kirchgessner CU, Gellert M, Bonner WM: A critical role for histone H2AX in recruitment of repair factors to nuclear foci after DNA damage. Curr Biol. 2000, 10: 886-895. 10.1016/S0960-9822(00)00610-2.
Suzuki M, Suzuki K, Kodama S, Watanabe M: Phosphorylated histone H2AX foci persist on rejoined mitotic chromosomes in normal human diploid cells exposed to ionizing radiation. Radiat Res. 2006, 165: 269-276. 10.1667/RR3508.1.
Lobrich M, Kuhne M, Wetzel J, Rothkamm K: Joining of correct and incorrect DNA double-strand break ends in normal human and ataxia telangiectasia fibroblasts. Genes Chromosomes Cancer. 2000, 27: 59-68. 10.1002/(SICI)1098-2264(200001)27:1<59::AID-GCC8>3.0.CO;2-9.
Kato TA, Nagasawa H, Weil MM, Little JB, Bedford JS: Levels of gamma-H2AX Foci after low-dose-rate irradiation reveal a DNA DSB rejoining defect in cells from human ATM heterozygotes in two at families and in another apparently normal individual. Radiat Res. 2006, 166: 443-453. 10.1667/RR3604.1.
Limoli CL, Giedzinski E, Bonner WM, Cleaver JE: UV-induced replication arrest in the xeroderma pigmentosum variant leads to DNA double-strand breaks, gamma-H2AX formation, and Mre11 relocalization. Proc Natl Acad Sci USA. 2002, 99: 233-238. 10.1073/pnas.231611798.
Matsumoto M, Yaginuma K, Igarashi A, Imura M, Hasegawa M, Iwabuchi K, Date T, Mori T, Ishizaki K, Yamashita K, Inobe M, Matsunaga T: Perturbed gap-filling synthesis in nucleotide excision repair causes histone H2AX phosphorylation in human quiescent cells. J Cell Sci. 2007, 120: 1104-1112. 10.1242/jcs.03391.
Rogakou EP, Nieves-Neira W, Boon C, Pommier Y, Bonner WM: Initiation of DNA fragmentation during apoptosis induces phosphorylation of H2AX histone at serine 139. J Biol Chem. 2000, 275: 9390-9395. 10.1074/jbc.275.13.9390.
Tan TL, Essers J, Citterio E, Swagemakers SM, de Wit J, Benson FE, Hoeijmakers JH, Kanaar R: Mouse Rad54 affects DNA conformation and DNA-damage-induced Rad51 foci formation. Curr Biol. 1999, 9: 325-328. 10.1016/S0960-9822(99)80142-0.
Essers J, Houtsmuller AB, van VL, Paulusma C, Nigg AL, Pastink A, Vermeulen W, Hoeijmakers JH, Kanaar R: Nuclear dynamics of RAD52 group homologous recombination proteins in response to DNA damage. EMBO J. 2002, 21: 2030-2037. 10.1093/emboj/21.8.2030.
MacPhail SH, Banath JP, Yu TY, Chu EH, Lambur H, Olive PL: Expression of phosphorylated histone H2AX in cultured cell lines following exposure to X-rays. Int J Radiat Biol. 2003, 79: 351-358. 10.1080/0955300032000093128.
Olive PL, Banáth JP: Multicell spheroid response to drugs predicted with the comet assay. Cancer Res. 1997, 57: 5528-5533.
Olive PL, Banáth JP, MacPhail S: Lack of a correlation between radiosensitivity and DNA double-strand break induction or rejoining in six human tumor cell lines. Cancer Res. 1994, 54: 3939-3946.
Huang X, Traganos F, Darzynkiewicz Z: DNA Damage Induced by DNA Topoisomerase I- and Topoisomerase IIInhibitors Detected by Histone H2AX Phosphorylation in Relation to the Cell Cycle Phase and Apoptosis. Cell Cycle. 2003, 2: 614-619.
Koch K, Wrona A, Dikomey E, Borgmann K: Impact of homologous recombination on individual cellular radiosensitivity. Radiother Oncol. 2009, 90: 265-272. 10.1016/j.radonc.2008.07.028.
Banuelos CA, Banath JP, Kim JY, Aquino-Parsons C, Olive PL: {gamma}H2AX expression in tumors exposed to cisplatin and fractionated Irradiation. Clin Cancer Res. 2009, 15: 3344-3353. 10.1158/1078-0432.CCR-08-3114.
Olive PL, Johnston PJ: DNA damage from oxidants: influence of lesion complexity and chromatin organization. Oncol Res. 1997, 9: 287-294.
Stojic L, Cejka P, Jiricny J: High doses of SN1 type methylating agents activate DNA damage signaling cascades that are largely independent of mismatch repair. Cell Cycle. 2005, 4: 473-477.
Rydberg B: Radiation-induced heat-labile sites that convert into DNA double-strand breaks. Radiat Res. 2000, 153: 805-812. 10.1667/0033-7587(2000)153[0805:RIHLST]2.0.CO;2.
Stojic L, Mojas N, Cejka P, Di PM, Ferrari S, Marra G, Jiricny J: Mismatch repair-dependent G2 checkpoint induced by low doses of SN1 type methylating agents requires the ATR kinase. Genes Dev. 2004, 18: 1331-1344. 10.1101/gad.294404.
Stojic L, Cejka P, Jiricny J: High doses of SN1 type methylating agents activate DNA damage signaling cascades that are largely independent of mismatch repair. Cell Cycle. 2005, 4: 473-477.
Olive PL, Banath JP, Sinnott LT: Phosphorylated histone H2AX in spheroids, tumors, and tissues of mice exposed to etoposide and 3-amino-1,2,4-benzotriazine-1,3-dioxide. Cancer Res. 2004, 64: 5363-5369. 10.1158/0008-5472.CAN-04-0729.
Banath JP, Olive PL: Expression of phosphorylated histone H2AX as a surrogate of cell killing by drugs that create DNA double-strand breaks. Cancer Res. 2003, 63: 4347-4350.
Xie A, Puget N, Shim I, Odate S, Jarzyna I, Bassing CH, Alt FW, Scully R: Control of sister chromatid recombination by histone H2AX. Mol Cell. 2004, 16: 1017-1025. 10.1016/j.molcel.2004.12.007.
Bassing CH, Chua KF, Sekiguchi J, Suh H, Whitlow SR, Fleming JC, Monroe BC, Ciccone DN, Yan C, Vlasakova K, Livingston DM, Ferguson DO, Scully R, Alt FW: Increased ionizing radiation sensitivity and genomic instability in the absence of histone H2AX. Proc Natl Acad Sci USA. 2002, 99: 8173-8178. 10.1073/pnas.122228699.
Iakovlev VV, Pintilie M, Morrison A, Fyles AW, Hill RP, Hedley DW: Effect of distributional heterogeneity on the analysis of tumor hypoxia based on carbonic anhydrase IX. Lab Invest. 2007, 87: 1206-1217. 10.1038/labinvest.3700680.
Yoshikawa T, Kashino G, Ono K, Watanabe M: Phosphorylated H2AX foci in tumor cells have no correlation with their radiation sensitivities. J Radiat Res (Tokyo). 2009, 50: 151-160. 10.1269/jrr.08109.
MacPhail SH, Banath JP, Yu Y, Chu E, Olive PL: Cell cycle-dependent expression of phosphorylated histone H2AX: Reduced expression in unirradiated but not X-irradiated G(1)-phase cells. Radiat Res. 2003, 159: 759-767. 10.1667/RR3003.
The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1471-2407/10/4/prepub