Dexamethasone up-regulates the inhibitory adaptor protein Dok-1 and suppresses downstream activation of the mitogen-activated protein kinase pathway in antigen-stimulated RBL-2H3 mast cells.
The glucocorticoid dexamethasone suppresses antigen-induced degranulation, cytokine production, and intermediate signaling events in RBL-2H3 mast cells, although the exact mechanisms are uncertain. By microarray analysis, we discovered that expression of the inhibitory adaptor protein, downstream of tyrosine kinase (Dok)-1, was up-regulated 4-fold in dexamethasone-treated RBL-2H3 cells. The up-regulation was apparent with as little as 1 to 10 nM dexamethasone. Treatment with dexamethasone also enhanced tyrosine phosphorylation of Dok-1, augmented recruitment of Ras GTPase-activating protein (RasGAP) by Dok-1, and inhibited activation of the mitogen-activated protein (MAP) kinase pathway in antigen-stimulated cells. The same effects were obtained by transient overexpression of Dok-1 but not by overexpression of Dok-1 that was mutated in RasGAP-binding domain. The negative regulatory role of Dok-1 was further validated by the expression of small interfering RNA directed against Dok-1, which enhanced activation of MAP kinase and subsequent release of arachidonic acid and tumor necrosis factor-alpha. These findings identify Dok-1 as mediator of the antiallergic actions of dexamethasone and as a negative regulator of the MAP kinase pathway and downstream release of inflammatory mediators.
Hiragun
Takaaki
T
Laboratory of Molecular Immunology, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892-1760, USA.
Peng
Ze
Z
Beaven
Michael A
MA
D000818:Animals; D000941:Antigens / Q000494:pharmacology; D016718:Arachidonic Acid / Q000378:metabolism; D001483:Base Sequence; D045744:Cell Line, Tumor; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000235:genetics*; D003907:Dexamethasone / Q000494:pharmacology*; D005786:Gene Expression Regulation / Q000187:drug effects*; D007700:Kinetics; D020935:MAP Kinase Signaling System / Q000502:physiology; D008407:Mast Cells; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors*; D020411:Oligonucleotide Array Sequence Analysis; D010750:Phosphoproteins / Q000235:genetics*; D034741:RNA, Small Interfering / Q000235:genetics; D016601:RNA-Binding Proteins / Q000235:genetics*; D051381:Rats; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
D000941:Antigens; D017931:DNA Primers; D004268:DNA-Binding Proteins; C499185:Dok1 protein, rat; D010750:Phosphoproteins; D034741:RNA, Small Interfering; D016601:RNA-Binding Proteins; D014409:Tumor Necrosis Factor-alpha; D016718:Arachidonic Acid; D003907:Dexamethasone; D020928:Mitogen-Activated Protein Kinases