title: The autoantigen DNA topoisomerase I interacts with chemokine receptor 7 and exerts cytokine-like effects on dermal fibroblasts.
abstract: OBJECTIVE
  Previous studies have demonstrated that, once released into the extracellular environment, the systemic sclerosis (SSc)-associated autoantigen DNA topoisomerase I (topo I) binds specifically to the surface of fibroblasts via an unknown receptor. We extended these results by identifying topo I-mediated cellular effects and characterizing the specific target of topo I on fibroblast surfaces.
  
  
  METHODS
  Purified topo I was used to investigate intracellular signaling pathway activation and tested for cell migration. To demonstrate the expression of specific chemokine receptors on fibroblasts, we performed immunoblotting and flow cytometry. To evaluate the direct interaction between chemokine receptor and topo I, a protein-protein based enzyme-linked immunosorbent assay (ELISA) was used. Finally, topo I coupled to the fluorochrome phycoerythrin (PE) was used to investigate competition of topo I specific binding on fibroblast surfaces with chemokine ligand.
  
  
  RESULTS
  Topo I stimulated the phosphorylation of phospholipase Cγ1, c-Raf, ERK-1/2, and p38 MAPK, intracellular signaling pathways that stimulated fibroblast migration via a G(αi)  protein-coupled receptor. CCR7 was found to interact directly with topo I. Furthermore, its ligand, CCL21, competed in vitro for this interaction and in vivo with the binding of PE-coupled topo I to fibroblast surfaces.
  
  
  CONCLUSIONS
  These new roles of topo I in fibroblast physiology and the identification of its target on the cell surface demonstrate that topo I is a bifunctional autoantigen and open up new perspectives of study in the field of SSc-associated anti-topo I autoantibodies.
authors.affiliation: Laboratory for Research in Autoimmunity, Notre-Dame Hospital, Centre Hospitalier de l'Université de Montréal, and Université de Montréal, Montréal, Québec, Canada.
authors.forename: Julie
authors.initials: J
authors.lastname: Arcand
authors.forename: Geneviève
authors.initials: G
authors.lastname: Robitaille
authors.forename: Martial
authors.initials: M
authors.lastname: Koenig
authors.forename: Jean-Luc
authors.initials: JL
authors.lastname: Senécal
authors.forename: Yves
authors.initials: Y
authors.lastname: Raymond
mesh_terms: D001323:Autoantibodies / Q000276:immunology; D001324:Autoantigens / Q000276:immunology; D002465:Cell Movement / Q000187:drug effects; D002478:Cells, Cultured; D054421:Chemokine CCL21 / Q000276:immunology / Q000378:metabolism; D004264:DNA Topoisomerases, Type I / Q000276:immunology / Q000378:metabolism / Q000494:pharmacology*; D020405:Dermis / Q000166:cytology; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D010766:Phosphorylation; D011485:Protein Binding; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism; D054400:Receptors, CCR7 / Q000187:drug effects* / Q000276:immunology / Q000378:metabolism; D012595:Scleroderma, Systemic; D015398:Signal Transduction; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
chemical_list: D001323:Autoantibodies; D001324:Autoantigens; C515478:CCL21 protein, human; C515370:CCR7 protein, human; D054421:Chemokine CCL21; D054400:Receptors, CCR7; D019908:Proto-Oncogene Proteins c-raf; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D048051:p38 Mitogen-Activated Protein Kinases; D004264:DNA Topoisomerases, Type I; C528909:TOP1 protein, human