Home LiteratureArticle Details
PMID: 21765476 Published · ppublish English Journal Article Research Support, N.I.H., Extramural Research Support, N.I.H., Intramural Research Support, Non-U.S. Gov't

CHEK2 genomic and proteomic analyses reveal genetic inactivation or endogenous activation across the 60 cell lines of the US National Cancer Institute.

Oncogene ·Vol. 31 ·No. 4 ·2012-01-26 ·Pages 403-18

Zoppoli G, Solier S, Reinhold WC, Liu H, Connelly JW, Monks A, Shoemaker RH, Abaan OD, Davis SR, Meltzer PS, Doroshow JH, Pommier Y

Abstract

CHEK2 encodes a serine/threonine kinase (Chk2) activated by ATM in response to DNA double-strand breaks. On the one hand, CHEK2 has been described as a tumor suppressor with proapoptotic, cell-cycle checkpoint and mitotic functions. On the other hand, Chk2 is also commonly activated (phosphorylated at T68) in cancers and precancerous lesions. Here, we report an extensive characterization of CHEK2 across the panel of 60 established cancer cell lines from the NCI Anticancer Screen (the NCI-60) using genomic and proteomic analyses, including exon-specific mRNA expression, DNA copy-number variation (CNV) by aCGH, exome sequencing, as well as western blot analyses for total and activated (pT68-Chk2) Chk2. We show that the high heterogeneity of Chk2 levels in cancer cells is primarily due to its inactivation (owing to low gene expression, alternative splicing, point mutations, copy-number alterations and premature truncation) or reduction of protein levels. Moreover, we observe that a significant percentage of cancer cells (12% of the NCI-60 and HeLa cells) show high endogenous Chk2 activation, which is always associated with p53 inactivation, and which is accompanied by downregulation of the Fanconi anemia and homologous recombination pathways. We also report the presence of activated Chk2 (pT68-Chk2) along with histone γ-H2AX in centrosomes.

MeSH Terms
Ataxia Telangiectasia Mutated Proteins Cell Cycle Proteins/physiology Cell Line, Tumor Checkpoint Kinase 2 Chromosomal Instability DNA Damage DNA-Binding Proteins/physiology Exons Fanconi Anemia/genetics Gene Expression Regulation, Neoplastic Gene Silencing Genomics Humans Neoplasms/genetics Phosphorylation Point Mutation Protein Serine-Threonine Kinases/analysis,genetics,physiology Proteomics RNA, Messenger/analysis Recombination, Genetic Tumor Suppressor Protein p53/physiology Tumor Suppressor Proteins/physiology
Chemicals
Cell Cycle Proteins DNA-Binding Proteins RNA, Messenger Tumor Suppressor Protein p53 Tumor Suppressor Proteins Checkpoint Kinase 2 ATM protein, human Ataxia Telangiectasia Mutated Proteins CHEK2 protein, human Protein Serine-Threonine Kinases
Authors & Affiliations
12 authors, click to expand affiliations / ORCID
Zoppoli G
Laboratory of Molecular Pharmacology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD, USA. gabriele.zoppoli@unige.it
Solier S
Reinhold W C
Liu H
Connelly J W
Monks A
Shoemaker R H
Abaan O D
Davis S R
Meltzer P S
Doroshow J H
Pommier Y
References (84)
84 references, click to expand
  1. Aberrations of the Chk2 tumour suppressor in advanced urinary bladder cancer.
    Oncogene. 2004 Nov 4;23(52):8545-51 PMID: 15361851
  2. The cell cycle checkpoint kinase Chk2 is a negative regulator of mitotic catastrophe.
    Oncogene. 2004 May 27;23(25):4353-61 PMID: 15048074
  3. Targeting chk2 kinase: molecular interaction maps and therapeutic rationale.
    Curr Pharm Des. 2005;11(22):2855-72 PMID: 16101442
  4. Heterozygous germ line hCHK2 mutations in Li-Fraumeni syndrome.
    Science. 1999 Dec 24;286(5449):2528-31 PMID: 10617473
  5. Cellular inhibition of checkpoint kinase 2 (Chk2) and potentiation of camptothecins and radiation by the novel Chk2 inhibitor PV1019 [7-nitro-1H-indole-2-carboxylic acid {4-[1-(guanidinohydrazone)-ethyl]-phenyl}-amide].
    J Pharmacol Exp Ther. 2009 Dec;331(3):816-26 PMID: 19741151
  6. Chk2 splice variants express a dominant-negative effect on the wild-type Chk2 kinase activity.
    Biochim Biophys Acta. 2010 Mar;1803(3):386-95 PMID: 20080130
  7. Centrosomal localization of DNA damage checkpoint proteins.
    J Cell Biochem. 2007 May 15;101(2):451-65 PMID: 17171639
  8. Apoptosis associated with deregulated E2F activity is dependent on E2F1 and Atm/Nbs1/Chk2.
    Mol Cell Biol. 2004 Apr;24(7):2968-77 PMID: 15024084
  9. Threonine 68 phosphorylation by ataxia telangiectasia mutated is required for efficient activation of Chk2 in response to ionizing radiation.
    Cancer Res. 2000 Nov 1;60(21):5934-6 PMID: 11085506
  10. p53 C-terminal phosphorylation by CHK1 and CHK2 participates in the regulation of DNA-damage-induced C-terminal acetylation.
    Mol Biol Cell. 2005 Apr;16(4):1684-95 PMID: 15659650
  11. The CHEK2 gene and inherited breast cancer susceptibility.
    Oncogene. 2006 Sep 25;25(43):5912-9 PMID: 16998506
  12. GammaH2AX and cancer.
    Nat Rev Cancer. 2008 Dec;8(12):957-67 PMID: 19005492
  13. A role for the Tip60 histone acetyltransferase in the acetylation and activation of ATM.
    Proc Natl Acad Sci U S A. 2005 Sep 13;102(37):13182-7 PMID: 16141325
  14. Promyelocytic leukemia activates Chk2 by mediating Chk2 autophosphorylation.
    J Biol Chem. 2006 Sep 8;281(36):26645-54 PMID: 16835227
  15. Destabilization of CHK2 by a missense mutation associated with Li-Fraumeni Syndrome.
    Cancer Res. 2001 Nov 15;61(22):8062-7 PMID: 11719428
  16. SIFT: Predicting amino acid changes that affect protein function.
    Nucleic Acids Res. 2003 Jul 1;31(13):3812-4 PMID: 12824425
  17. The apoptotic ring: a novel entity with phosphorylated histones H2AX and H2B and activated DNA damage response kinases.
    Cell Cycle. 2009 Jun 15;8(12):1853-9 PMID: 19448405
  18. Regulation of the Chk2 protein kinase by oligomerization-mediated cis- and trans-phosphorylation.
    Mol Cancer Res. 2003 Jun;1(8):598-609 PMID: 12805407
  19. Topoisomerase I levels in the NCI-60 cancer cell line panel determined by validated ELISA and microarray analysis and correlation with indenoisoquinoline sensitivity.
    Mol Cancer Ther. 2009 Jul;8(7):1878-84 PMID: 19584232
  20. GammaH2AX in cancer cells: a potential biomarker for cancer diagnostics, prediction and recurrence.
    Cell Cycle. 2006 Dec;5(24):2909-13 PMID: 17172873
  21. Replication stress and oxidative damage contribute to aberrant constitutive activation of DNA damage signalling in human gliomas.
    Oncogene. 2010 Sep 9;29(36):5095-102 PMID: 20581868
  22. Identification and characterization of a novel and specific inhibitor of the ataxia-telangiectasia mutated kinase ATM.
    Cancer Res. 2004 Dec 15;64(24):9152-9 PMID: 15604286
  23. Chk2 tumour suppressor protein in human spermatogenesis and testicular germ-cell tumours.
    Oncogene. 2001 Sep 13;20(41):5897-902 PMID: 11593395
  24. TRANSFAC and its module TRANSCompel: transcriptional gene regulation in eukaryotes.
    Nucleic Acids Res. 2006 Jan 1;34(Database issue):D108-10 PMID: 16381825
  25. Concomitant inactivation of p53 and Chk2 in breast cancer.
    Oncogene. 2002 Feb 21;21(9):1316-24 PMID: 11857075
  26. Regulation of CHK2 by DNA-dependent protein kinase.
    J Biol Chem. 2005 Mar 25;280(12):12041-50 PMID: 15668230
  27. Transcript and protein expression profiles of the NCI-60 cancer cell panel: an integromic microarray study.
    Mol Cancer Ther. 2007 Mar;6(3):820-32 PMID: 17339364
  28. 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):3988-92 PMID: 8387205
  29. ATM activation is accompanied with earlier stages of prostate tumorigenesis.
    Biochim Biophys Acta. 2006 Oct;1763(10):1090-7 PMID: 16997395
  30. The CHK2-BRCA1 tumour suppressor pathway ensures chromosomal stability in human somatic cells.
    Nat Cell Biol. 2010 May;12(5):492-9 PMID: 20364141
  31. Kinetics of histone H2AX phosphorylation and Chk2 activation in A549 cells treated with topotecan and mitoxantrone in relation to the cell cycle phase.
    Cytometry A. 2008 Jun;73(6):480-9 PMID: 18459160
  32. DNA damage-activated kinase Chk2 is independent of proliferation or differentiation yet correlates with tissue biology.
    Cancer Res. 2001 Jul 1;61(13):4990-3 PMID: 11431331
  33. Phosphorylation of HuR by Chk2 regulates SIRT1 expression.
    Mol Cell. 2007 Feb 23;25(4):543-57 PMID: 17317627
  34. Cytometric assessment of DNA damage by exogenous and endogenous oxidants reports aging-related processes.
    Cytometry A. 2007 Nov;71(11):905-14 PMID: 17879239
  35. CHK2 kinase--a busy messenger.
    Nat Rev Mol Cell Biol. 2001 Dec;2(12):877-86 PMID: 11733767
  36. DNA damage response as a candidate anti-cancer barrier in early human tumorigenesis.
    Nature. 2005 Apr 14;434(7035):864-70 PMID: 15829956
  37. A gene expression database for the molecular pharmacology of cancer.
    Nat Genet. 2000 Mar;24(3):236-44 PMID: 10700175
  38. Proteomic profiling of the NCI-60 cancer cell lines using new high-density reverse-phase lysate microarrays.
    Proc Natl Acad Sci U S A. 2003 Nov 25;100(24):14229-34 PMID: 14623978
  39. Genome-wide mRNA and microRNA profiling of the NCI 60 cell-line screen and comparison of FdUMP[10] with fluorouracil, floxuridine, and topoisomerase 1 poisons.
    Mol Cancer Ther. 2010 Dec;9(12):3105-14 PMID: 21159603
  40. Analysis of Food and Drug Administration-approved anticancer agents in the NCI60 panel of human tumor cell lines.
    Mol Cancer Ther. 2010 May;9(5):1451-60 PMID: 20442306
  41. The Chk2 protein kinase.
    DNA Repair (Amst). 2004 Aug-Sep;3(8-9):1039-47 PMID: 15279791
  42. Phosphorylation of threonine 68 promotes oligomerization and autophosphorylation of the Chk2 protein kinase via the forkhead-associated domain.
    J Biol Chem. 2002 May 31;277(22):19389-95 PMID: 11901158
  43. E2F1 uses the ATM signaling pathway to induce p53 and Chk2 phosphorylation and apoptosis.
    Mol Cancer Res. 2004 Apr;2(4):203-14 PMID: 15140942
  44. The checkpoint kinases Chk1 and Chk2 regulate the functional associations between hBRCA2 and Rad51 in response to DNA damage.
    Oncogene. 2008 Jun 26;27(28):3977-85 PMID: 18317453
  45. Karyotypic complexity of the NCI-60 drug-screening panel.
    Cancer Res. 2003 Dec 15;63(24):8634-47 PMID: 14695175
  46. Linkage of ATM to cell cycle regulation by the Chk2 protein kinase.
    Science. 1998 Dec 4;282(5395):1893-7 PMID: 9836640
  47. Human DNA damage checkpoints and their relevance to soft tissue sarcoma.
    Pathol Int. 2004 Jan;54(1):26-31 PMID: 14674991
  48. Epigenetic inactivation of checkpoint kinase 2 gene in non-small cell lung cancer and its relationship with clinicopathological features.
    Lung Cancer. 2009 Aug;65(2):247-50 PMID: 19362748
  49. Death receptor-induced activation of the Chk2- and histone H2AX-associated DNA damage response pathways.
    Mol Cell Biol. 2009 Jan;29(1):68-82 PMID: 18955500
  50. Alternative splicing and mutation status of CHEK2 in stage III breast cancer.
    Oncogene. 2004 Nov 4;23(52):8535-44 PMID: 15361853
  51. Chk2 activates E2F-1 in response to DNA damage.
    Nat Cell Biol. 2003 May;5(5):401-9 PMID: 12717439
  52. Cooperative functions of Chk1 and Chk2 reduce tumour susceptibility in vivo.
    EMBO J. 2010 Oct 20;29(20):3558-70 PMID: 20834228
  53. Presence and expression of human papillomavirus sequences in human cervical carcinoma cell lines.
    Am J Pathol. 1985 Jun;119(3):361-6 PMID: 2990217
  54. The NCI60 human tumour cell line anticancer drug screen.
    Nat Rev Cancer. 2006 Oct;6(10):813-23 PMID: 16990858
  55. Identification of a highly potent and selective DNA-dependent protein kinase (DNA-PK) inhibitor (NU7441) by screening of chromenone libraries.
    Bioorg Med Chem Lett. 2004 Dec 20;14(24):6083-7 PMID: 15546735
  56. Integrating data on DNA copy number with gene expression levels and drug sensitivities in the NCI-60 cell line panel.
    Mol Cancer Ther. 2006 Apr;5(4):853-67 PMID: 16648555
  57. Antisense inhibition of Chk2/hCds1 expression attenuates DNA damage-induced S and G2 checkpoints and enhances apoptotic activity in HEK-293 cells.
    FEBS Lett. 2001 Sep 7;505(1):7-12 PMID: 11557032
  58. Chk2 is a tumor suppressor that regulates apoptosis in both an ataxia telangiectasia mutated (ATM)-dependent and an ATM-independent manner.
    Mol Cell Biol. 2002 Sep;22(18):6521-32 PMID: 12192050
  59. The DNA damage response: ten years after.
    Mol Cell. 2007 Dec 14;28(5):739-45 PMID: 18082599
  60. Complex engagement of DNA damage response pathways in human cancer and in lung tumor progression.
    Carcinogenesis. 2007 Oct;28(10):2082-8 PMID: 17522062
  61. 53BP1 functions in an ATM-dependent checkpoint pathway that is constitutively activated in human cancer.
    Nat Cell Biol. 2002 Dec;4(12):998-1002 PMID: 12447382
  62. DNA damage-induced activation of p53 by the checkpoint kinase Chk2.
    Science. 2000 Mar 10;287(5459):1824-7 PMID: 10710310
  63. Exon array analyses across the NCI-60 reveal potential regulation of TOP1 by transcription pausing at guanosine quartets in the first intron.
    Cancer Res. 2010 Mar 15;70(6):2191-203 PMID: 20215517
  64. Polo-like kinase 1 and Chk2 interact and co-localize to centrosomes and the midbody.
    J Biol Chem. 2003 Mar 7;278(10):8468-75 PMID: 12493754
  65. Detection of in vitro kinase generated protein phosphorylation sites using gamma[18O4]-ATP and mass spectrometry.
    Anal Chem. 2007 Oct 15;79(20):7603-10 PMID: 17877366
  66. ATM-Chk2-p53 activation prevents tumorigenesis at an expense of organ homeostasis upon Brca1 deficiency.
    EMBO J. 2006 May 17;25(10):2167-77 PMID: 16675955
  67. Chk2/hCds1 functions as a DNA damage checkpoint in G(1) by stabilizing p53.
    Genes Dev. 2000 Feb 1;14(3):278-88 PMID: 10673500
  68. Chk1 and Chk2 kinases in checkpoint control and cancer.
    Cancer Cell. 2003 May;3(5):421-9 PMID: 12781359
  69. Low-penetrance susceptibility to breast cancer due to CHEK2(*)1100delC in noncarriers of BRCA1 or BRCA2 mutations.
    Nat Genet. 2002 May;31(1):55-9 PMID: 11967536
  70. Inhibition of poly(ADP-ribose) polymerase enhances cell death and improves tumor growth delay in irradiated lung cancer models.
    Clin Cancer Res. 2007 May 15;13(10):3033-42 PMID: 17505006
  71. The ATM-Chk2-Cdc25A checkpoint pathway guards against radioresistant DNA synthesis.
    Nature. 2001 Apr 12;410(6830):842-7 PMID: 11298456
  72. Ataxia telangiectasia-mutated phosphorylates Chk2 in vivo and in vitro.
    Proc Natl Acad Sci U S A. 2000 Sep 12;97(19):10389-94 PMID: 10973490
  73. CHK2 kinase: cancer susceptibility and cancer therapy - two sides of the same coin?
    Nat Rev Cancer. 2007 Dec;7(12):925-36 PMID: 18004398
  74. The predictive value of molecular markers (p53, EGFR, ATM, CHK2) in multimodally treated squamous cell carcinoma of the oesophagus.
    Br J Cancer. 2007 Nov 19;97(10):1404-8 PMID: 17940507
  75. DNA damage checkpoint kinase Chk2 triggers replicative senescence.
    EMBO J. 2004 Jul 7;23(13):2554-63 PMID: 15192702
  76. Chk2 molecular interaction map and rationale for Chk2 inhibitors.
    Clin Cancer Res. 2006 May 1;12(9):2657-61 PMID: 16675556
  77. Chk2-deficient mice exhibit radioresistance and defective p53-mediated transcription.
    EMBO J. 2002 Oct 1;21(19):5195-205 PMID: 12356735
  78. Expression of checkpoint kinase 2 in breast carcinomas: correlation with key regulators of tumor cell proliferation, angiogenesis, and survival.
    Histol Histopathol. 2006 Apr;21(4):373-82 PMID: 16437383
  79. p14ARF activates a Tip60-dependent and p53-independent ATM/ATR/CHK pathway in response to genotoxic stress.
    Mol Cell Biol. 2006 Jun;26(11):4339-50 PMID: 16705183
  80. Activated checkpoint kinase 2 provides a survival signal for tumor cells.
    Cancer Res. 2006 Dec 15;66(24):11576-9 PMID: 17178848
  81. Chk2 oligomerization studied by phosphopeptide ligation: implications for regulation and phosphodependent interactions.
    J Biol Chem. 2008 Dec 19;283(51):36019-30 PMID: 18948271
  82. Mutation analysis of 24 known cancer genes in the NCI-60 cell line set.
    Mol Cancer Ther. 2006 Nov;5(11):2606-12 PMID: 17088437
  83. Checkpoint kinase 2 (Chk2) monomers or dimers phosphorylate Cdc25C after DNA damage regardless of threonine 68 phosphorylation.
    J Biol Chem. 2002 Dec 13;277(50):48418-26 PMID: 12386164
  84. Activation of the DNA damage checkpoint and genomic instability in human precancerous lesions.
    Nature. 2005 Apr 14;434(7035):907-13 PMID: 15829965
Article Info
Journal
Oncogene
Abbr.
Oncogene
ISSN
1476-5594
DOI
10.1038/onc.2011.283
Email
gabriele.zoppoli@unige.it
Published
2012-01-26
Epub
2011-00-18
Pages
403-18
Language
English
Region
England
NLM ID
8711562
PMCID
PMC7489305
Subset
IM
Grants
Intramural NIH HHS · Z01 BC006150 · United States
Intramural NIH HHS · Z99 CA999999 · United States
Intramural NIH HHS · ZIC BC011497 · United States
Corrections
ErratumIn
-
External Links
PubMed source DOI: 10.1038/onc.2011.283 PMC Full Text View journal details
Analysis Services
Analysis Services

Contact

No. 2 Wenbo Road, Zhangqiu District, Jinan, Shandong

Qilu Normal University · Genelibs Bioinformatics Lab

750 Shunhua Rd, Jinan

2F, Bldg F, University Science Park

Tel: 0531-88819269

WeChat Official Account

Follow our WeChat subscription account for real-time updates and the latest in medical and biological research.


Business Email

E-mail: product@genelibs.com

Copyright © 2026.Genelibs Jinan Honghui Biotechnology Co., Ltd. 鲁ICP备20007159号-2