Src mediates cytokine-stimulated gene expression in airway myocytes through ERK MAPK
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Src mediates cytokine-stimulated gene expression in airway myocytes through ERK MAPK Cherie A Singer1*, Beata Lontay1, Helmut Unruh2, Andrew J Halayko3 and William T Gerthoffer1,4
Abstract The p38 and extracellular signal-regulated kinases (ERK) mitogen-activated protein kinases (MAPK) participate in cytokine-stimulated inflammatory gene expression in airway smooth muscle cells. The following study was undertaken to determine whether Src tyrosine kinases are signaling intermediaries upstream of cytokine-stimulated MAPK activation and gene expression. Treating human airway myocytes with interleukin (IL)-1b, tumor necrosis factor (TNF) a and interferon (IFN) g caused a rapid 1.8-fold increase in Src family tyrosine kinase activity within 1 minute that remained 2.3 to 2.7 fold above basal conditions for 15 minutes. This activity was blocked by addition of 30 μM PP1, a pyrimidine inhibitor specific for Src family tyrosine kinases, in immune-complex assays to confirm that this stimulus activates Src tyrosine kinase. Addition of PP1 also blocked cytokine-stimulated expression of IL-1b, IL-6 and IL-8, while decreasing phosphorylation of ERK, but not p38 MAPK. Since this inflammatory stimulus may activate additional inflammatory signaling pathways downstream of Src, we tested the effects of PP1 on phosphorylation of signal transducers and activators of transcription (STAT). PP1 had no effect on cytokinestimulated STAT 1 or STAT 3 phosphorylation. These results demonstrate that Src tyrosine kinases participate in the regulation of IL-1b, IL-6 and IL-8 expression and that these effects of Src are mediated through activation of ERK MAPK and not p38 MAPK or STAT1/STAT3 phosphorylation. Findings Our laboratory has examined signaling pathways regulating secretion of inflammatory mediators by human airway smooth muscle cells. The synthesis and secretion of Th1/Th2 cytokines, along with CC and C-X-C chemokines, chemotactic proteins, peptide growth factors and their receptors can be induced in these myocytes by exposure to, among others, interleukin (IL)-1b, tumor necrosis factor (TNF) a, interferon (IFN) g, or transforming growth factor b [1,2] and contributes to inflammatory airway disease. In previous studies, we used a complementary DNA expression array to analyze expression of inflammatory mediators following treatment with a pro-inflammatory stimulus consisting of IL-1b, TNFa and IFNg and established that this stimulus induces expression of multiple inflammatory mediators including IL-1b, IL-6, and IL-8 [3]. Pharmacological inhibitors of mitogen-activated protein kinase (MAPK) activation were used to further demonstrate that both * Correspondence: [email protected] 1 University of Nevada School of Medicine, Department of Pharmacology Reno, NV 89557, USA Full list of author information is available at the end of the article
p38 and ERK MAPK are upstream mediators of IL-6 and IL-8 expression, while ERK MAPK alone was involved in mediating IL-1b expression. Src tyrosine kinases are one of t
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