title: CCN2 is required for the TGF-β induced activation of Smad1-Erk1/2 signaling network.
abstract: Connective tissue growth factor (CCN2) is a multifunctional matricellular protein, which is frequently overexpressed during organ fibrosis. CCN2 is a mediator of the pro-fibrotic effects of TGF-β in cultured cells, but the specific function of CCN2 in the fibrotic process has not been elucidated. In this study we characterized the CCN2-dependent signaling pathways that are required for the TGF-β induced fibrogenic response. By depleting endogenous CCN2 we show that CCN2 is indispensable for the TGF-β-induced phosphorylation of Smad1 and Erk1/2, but it is unnecessary for the activation of Smad3. TGF-β stimulation triggered formation of the CCN2/β(3) integrin protein complexes and activation of Src signaling. Furthermore, we demonstrated that signaling through the α(v)β(3) integrin receptor and Src was required for the TGF-β induced Smad1 phosphorylation. Recombinant CCN2 activated Src and Erk1/2 signaling, and induced phosphorylation of Fli1, but was unable to stimulate Smad1 or Smad3 phosphorylation. Additional experiments were performed to investigate the role of CCN2 in collagen production. Consistent with the previous studies, blockade of CCN2 abrogated TGF-β-induced collagen mRNA and protein levels. Recombinant CCN2 potently stimulated collagen mRNA levels and upregulated activity of the COL1A2 promoter, however CCN2 was a weak inducer of collagen protein levels. CCN2 stimulation of collagen was dose-dependent with the lower doses (<50 ng/ml) having a stimulatory effect and higher doses having an inhibitory effect on collagen gene expression. In conclusion, our study defines a novel CCN2/α(v)β(3) integrin/Src/Smad1 axis that contributes to the pro-fibrotic TGF-β signaling and suggests that blockade of this pathway may be beneficial for the treatment of fibrosis.
authors.affiliation: The Arthritis Center, Boston University School of Medicine, Boston, Massachusetts, United States of America.
authors.forename: Sashidhar S
authors.initials: SS
authors.lastname: Nakerakanti
authors.forename: Andreea M
authors.initials: AM
authors.lastname: Bujor
authors.forename: Maria
authors.initials: M
authors.lastname: Trojanowska
mesh_terms: D003094:Collagen / Q000096:biosynthesis / Q000235:genetics; D055513:Connective Tissue Growth Factor / Q000378:metabolism*; D020405:Dermis / Q000166:cytology; D004789:Enzyme Activation / Q000187:drug effects; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000201:enzymology; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D007231:Infant, Newborn; D039302:Integrin alphaVbeta3 / Q000378:metabolism; D020935:MAP Kinase Signaling System / Q000187:drug effects*; D008297:Male; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism*; D008954:Models, Biological; D010766:Phosphorylation / Q000187:drug effects; D051898:Smad1 Protein / Q000378:metabolism*; D016212:Transforming Growth Factor beta / Q000494:pharmacology*; D015854:Up-Regulation / Q000187:drug effects; D019061:src-Family Kinases / Q000378:metabolism
chemical_list: C526317:CCN2 protein, human; D039302:Integrin alphaVbeta3; C497399:SMAD1 protein, human; D051898:Smad1 Protein; D016212:Transforming Growth Factor beta; D055513:Connective Tissue Growth Factor; D003094:Collagen; D019061:src-Family Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3