Activation of protein kinase D1 in mast cells in response to innate, adaptive, and growth factor signals.
Little is known about the serine/threonine kinase protein kinase D (PKD)1 in mast cells. We sought to define ligands that activate PKD1 in mast cells and to begin to address the contributions of this enzyme to mast cell activation induced by diverse agonists. Mouse bone marrow-derived mast cells (BMMC) contained both PKD1 mRNA and immunoreactive PKD1 protein. Activation of BMMC through TLR2, Kit, or FcepsilonRI with Pam(3)CSK(4) (palmitoyl-3-cysteine-serine-lysine-4), stem cell factor (SCF), and cross-linked IgE, respectively, induced activation of PKD1, as determined by immunochemical detection of autophosphorylation. Activation of PKD1 was inhibited by the combined PKD1 and protein kinase C (PKC) inhibitor Gö 6976 but not by broad-spectrum PKC inhibitors, including bisindolylmaleimide (Bim) I. Pam(3)CSK(4) and SCF also induced phosphorylation of heat shock protein 27, a known substrate of PKD1, which was also inhibited by Gö 6976 but not Bim I in BMMC. This pattern also extended to activation-induced increases in mRNA encoding the chemokine CCL2 (MCP-1) and release of the protein. In contrast, both pharmacologic agents inhibited exocytosis of beta-hexosaminidase induced by SCF or cross-linked IgE. Our findings establish that stimuli representing innate, adaptive, and growth factor pathways activate PKD1 in mast cells. In contrast with certain other cell types, activation of PKD1 in BMMC is largely independent of PKC activation. Furthermore, our findings also indicate that PKD1 preferentially influences transcription-dependent production of CCL2, whereas PKC predominantly regulates the rapid exocytosis of preformed secretory granule mediators.
Murphy
Thomas R
TR
Department of Medicine, Harvard Medical School, and Division of Rheumatology, Immunology and Allergy, Brigham and Women's Hospital, Boston, MA 02115, USA.
Legere
Henry J
HJ
Katz
Howard R
HR
D000222:Adaptation, Physiological / Q000276:immunology*; D000818:Animals; D001665:Binding Sites; D001854:Bone Marrow Cells / Q000166:cytology / Q000187:drug effects / Q000276:immunology; D002227:Carbazoles / Q000494:pharmacology; D018932:Chemokine CCL2 / Q000187:drug effects / Q000235:genetics / Q000276:immunology; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D005089:Exocytosis / Q000187:drug effects; D007113:Immunity, Innate / Q000276:immunology; D007073:Immunoglobulin E / Q000276:immunology; D007211:Indoles / Q000494:pharmacology; D008024:Ligands; D055666:Lipopeptides; D008301:Maleimides / Q000494:pharmacology; D008407:Mast Cells / Q000187:drug effects / Q000276:immunology*; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D010455:Peptides / Q000494:pharmacology; D010766:Phosphorylation; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000276:immunology; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D015398:Signal Transduction / Q000276:immunology*; D019089:Stem Cell Factor / Q000276:immunology* / Q000494:pharmacology; D013329:Structure-Activity Relationship
D002227:Carbazoles; C491768:Ccl2 protein, mouse; D018932:Chemokine CCL2; D007211:Indoles; D008024:Ligands; D055666:Lipopeptides; D008301:Maleimides; C508075:Pam(3)CSK(4) peptide; D010455:Peptides; D012333:RNA, Messenger; D019089:Stem Cell Factor; C081021:Go 6976; D007073:Immunoglobulin E; C089212:protein kinase D; D011493:Protein Kinase C; C070515:bisindolylmaleimide I