title: Priming and potentiation of DNA damage response by fibronectin in human colon cancer cells and tumor-derived myofibroblasts.
abstract: We have previously shown that the genotoxin-induced apoptosis in mouse embryo fibroblasts was enhanced by the extracellular matrix protein fibronectin (FN). In the present study, we tested the hypothesis that FN regulates the DNA damage response (DDR) signaling pathways in HCT116 (p53-wt) and HT29 (p53-mut) human colon cancer cells and tumor-derived myofibroblasts. DNA damage recognition mechanisms were analyzed by immunofluorescence staining, cell cycle analysis and immunoblotting addressed at specific molecular sensors and executors involved in the DDR pathways. The results show that FN, but not collagen type IV or Matrigel, initiates and potentiates the DDR to the anticancer drug cisplatin in an α5 integrin and cell cycle-dependent manner (S and G2/M phases) in human colon cancer cells. Accordingly, we demonstrate that adhesion of HCT116 cells to FN upregulated the expression of α5 integrin FN receptors at the cell surface. These FN-induced DDR pathways include the concerted phosphorylation of histone H2AX on Ser139 detected as nuclear foci (γ-H2AX, 15 and 25 kDa forms), of ataxia telangiectasia mutated (ATM-Ser1981), checkpoint kinase 2 (CHK2-Thr68, 62 and 67 kDa) and p53-Ser15. These FN-induced γ-H2AX signals were interrupted or attenuated by selective inhibitors acting on the DDR pathway kinases, including wortmannin (targeting the phosphatidylinositol-3-kinase-related protein kinases, PIKK), KU55933 (ATM), NU7026 (DNA-dependent protein kinase catalytic subunit, DNA-PK-cs) and SP600125 (JNK2, stress activated protein kinase/c-Jun N-terminal kinase-2). Adhesion to FN also potentiated the γ-H2AX signals and the cytotoxic effects of cisplatin in human colon tumor-derived myofibroblasts. These data provide evidence that FN promotes DNA damage recognition and chemosensitization to cisplatin via the potentiation of the DNA damage signaling responses in human colon cancer cells and tumor-derived myofibroblasts.
authors.affiliation: Laboratory of Experimental Cancerology, Ghent University Hospital, Ghent, Belgium.
authors.forename: Olivier
authors.initials: O
authors.lastname: DE Wever
authors.affiliation: CNRS UMR 7098, Paris.|Université Pierre-et-Marie-Curie (UPMC) Paris-6, Paris, France.
authors.forename: Joëlle
authors.initials: J
authors.lastname: Sobczak-Thépot
authors.affiliation: Unité de Glycobiologie Structurale et Fonctionnelle UMR USTL/CNRS no. 8576 - IFR147, Villeneuve-d'Ascq, France.
authors.forename: Anne-Sophie
authors.initials: AS
authors.lastname: Vercoutter-Edouart
authors.affiliation: Unité de Glycobiologie Structurale et Fonctionnelle UMR USTL/CNRS no. 8576 - IFR147, Villeneuve-d'Ascq, France.
authors.forename: Jean-Claude
authors.initials: JC
authors.lastname: Michalski
authors.affiliation: INSERM U673 and U938, Laboratory of Cancer Biology and Therapeutics, Centre de recherche Saint-Antoine.
authors.forename: Radia
authors.initials: R
authors.lastname: Ouelaa-Benslama
authors.affiliation: Moores UCSD Cancer Center, UCSD School of Medicine, La Jolla, CA, USA.
authors.forename: Dwayne G
authors.initials: DG
authors.lastname: Stupack
authors.affiliation: Laboratory of Experimental Cancerology, Ghent University Hospital, Ghent, Belgium.
authors.forename: Marc
authors.initials: M
authors.lastname: Bracke
authors.affiliation: Moores UCSD Cancer Center, UCSD School of Medicine, La Jolla, CA, USA.
authors.forename: Jean Y J
authors.initials: JYJ
authors.lastname: Wang
authors.affiliation: INSERM U673 and U938, Laboratory of Cancer Biology and Therapeutics, Centre de recherche Saint-Antoine.|Université Pierre-et-Marie-Curie (UPMC) Paris-6, Paris, France.
authors.forename: Christian
authors.initials: C
authors.lastname: Gespach
authors.affiliation: INSERM U673 and U938, Laboratory of Cancer Biology and Therapeutics, Centre de recherche Saint-Antoine.|Université Pierre-et-Marie-Curie (UPMC) Paris-6, Paris, France.
authors.forename: Shahin
authors.initials: S
authors.lastname: Emami
mesh_terms: D002448:Cell Adhesion / Q000187:drug effects; D002453:Cell Cycle / Q000187:drug effects; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D064447:Checkpoint Kinase 2; D002945:Cisplatin / Q000494:pharmacology; D003110:Colonic Neoplasms / Q000235:genetics* / Q000378:metabolism*; D003432:Cross-Linking Reagents / Q000494:pharmacology; D004249:DNA Damage / Q000187:drug effects* / Q000235:genetics*; D005353:Fibronectins / Q000378:metabolism* / Q000494:pharmacology*; D045325:HCT116 Cells; D019073:HT29 Cells; D006657:Histones / Q000378:metabolism; D006801:Humans; D058628:Myofibroblasts / Q000378:metabolism* / Q000473:pathology; D010766:Phosphorylation / Q000187:drug effects; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D017469:Receptors, Fibronectin / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects
chemical_list: D003432:Cross-Linking Reagents; D005353:Fibronectins; C468783:H2AX protein, human; D006657:Histones; D017469:Receptors, Fibronectin; D064447:Checkpoint Kinase 2; C578634:CHEK2 protein, human; C578633:Chek2 protein, mouse; D017346:Protein Serine-Threonine Kinases; D002945:Cisplatin