View Abstract

A4358 - PTP-Alpha Promotes TGF-Beta Profibrotic Responses Via Activation of Src
Author Block: Y. Aschner1, M. R. Nelson2, M. I. Wong2, H. Roybal2, R. Holly2, G. P. Downey3; 1Pulmonary Sciences and Critical Care Medicine, University of Colorado Denver, Aurora, CO, United States, 2Academic Affairs, National Jewish Health, Denver, CO, United States, 3Natl Jewish Health, Denver, CO, United States.
Rationale: The pathogenesis of pulmonary fibrosis remains incompletely understood and current treatments are not curative and are poorly tolerated. While the role of tyrosine kinases is recognized in the pathogenesis of lung tissue fibrosis, the contribution of protein tyrosine phosphatases is less well characterized. We have previously reported that Protein Tyrosine Phosphatase-alpha (PTPα) regulates TGF-β dependent pro-fibrotic signaling in lung fibroblasts. Mice globally deficient in PTPα are protected from experimental models of pulmonary fibrosis, and this phenotype is recapitulated in transgenic mice with fibroblast-specific but not Alveolar Type II-specific deletion of Ptpra (the gene encoding PTPα), suggesting that PTPα promotes pro-fibrotic signaling via fibroblast-mediated pathways. Herein, we sought to determine the molecular mechanisms underlying Ptpra control of TGF-β signaling pathways.
Methods: Ptpra-/- mice and C57Bl/6 wild-type controls were treated with single-dose bleomycin by intra-tracheal instillation for 21 days to induce tissue fibrosis. Murine embryonic fibroblasts (MEF) derived from wild type (WT) and Ptpra-/- mice were studied in culture using TGF-β stimulation assays. Downstream fibrotic responses and markers of TGF-β signal transduction were assessed by Western blot analysis, RT-PCR, and immunofluorescence microscopy.
Results: The best known function of PTPα is to dephosphorylate and thereby activate Src (at Tyrosine residue 527). We hypothesize that PTPα acts indirectly to promote fibrogenesis through the TGF-β signaling cascade via activation of Src. Stimulation of WT MEFs with TGF-β results in activation of Src as evidenced by dephosphorylation of the C-terminal inhibitory Tyr527 and increased levels of phosphorylation on Tyr416. By contrast, TGF-β-induced Src activation was markedly attenuated in Ptpra-/- MEFs. Src activation was markedly reduced in lung tissue of Ptpra-/- mice treated with bleomycin, compared to WT mice as assessed by immunostaining with antibodies to non-phospho-Tyr527-Src. Finally, in human IPF lung tissue we observed an increase in the amount of active (non-phospho-Tyr527-Src) compared to normal controls, as assessed by western blot analysis.
Conclusions: We conclude that PTPα promotes pro-fibrotic signaling in response to TGF-β stimulation and that this action is indirect through activation of Src (by de-phosphorylation of the inhibitory Tyr 527 residue). We speculate that activated Src phosphorylates TGF-β Receptor II, driving downstream signaling though canonical and non-canonical TGF-β pathways, leading to tissue fibrosis.