Bombesin, vasopressin, endothelin, bradykinin, and platelet-derived growth factor rapidly activate protein kinase D through a protein kinase C-dependent signal transduction pathway.
Protein kinase D (PKD) is a serine/threonine protein kinase that is activated by phorbol esters via protein kinase C in intact cells. To assess the physiological significance of this putative pathway, we examined the regulation of PKD in living cells by mitogenic regulatory peptides and by platelet-derived growth factors (PDGF). Our results demonstrate that bombesin rapidly induces PKD activation in Swiss 3T3 cells, as shown by autophosphorylation and syntide-2 phosphorylation assays. Maximum PKD activation (14-fold above base-line levels) was obtained 90 s after bombesin stimulation. Bombesin also induced PKD activation in Rat-1 cells stably transfected with the bombesin/gastrin releasing peptide (GRP) receptor and in COS-7 cells transiently co-transfected with PKD and bombesin/GRP receptor expression constructs. No inducible kinase activity was demonstrated when COS-7 cells were transfected with a kinase-deficient PKD mutant. Bombesin-mediated PKD activation was prevented by treatment of Swiss 3T3 cells with the protein kinase C inhibitors GF 1092030X and Ro 31-8220. In contrast, these compounds did not inhibit PKD activity when added directly in vitro. Vasopressin, endothelin, and bradykinin also activated PKD in Swiss 3T3 cells through a PKC-dependent pathway. Platelet-derived growth factor-stimulated PKD activation in Swiss 3T3 cells and in porcine aortic endothelial cells stably transfected with PDGF-beta receptors. Treatment with GF 1092030X or Ro 31-8220 inhibited PKD activation induced by PDGF. Thus, our results indicate that PKD is activated by multiple signaling peptides through a protein kinase C-dependent signal transduction pathway in a variety of cell types.
Zugaza
J L
JL
Growth Regulation Laboratory, Imperial Cancer Research Fund, P.O. Box 123, 44 Lincoln's Inn Fields, London WC2A 3PX, United Kingdom.
Waldron
R T
RT
Sinnett-Smith
J
J
Rozengurt
E
E
D016475:3T3 Cells; D000818:Animals; D001839:Bombesin / Q000494:pharmacology; D001920:Bradykinin / Q000494:pharmacology; D019556:COS Cells; D016232:Endothelins / Q000494:pharmacology; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D051379:Mice; D009479:Neuropeptides / Q000494:pharmacology*; D010766:Phosphorylation; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects*; D014667:Vasopressins / Q000494:pharmacology
D016232:Endothelins; D004791:Enzyme Inhibitors; D009479:Neuropeptides; D010982:Platelet-Derived Growth Factor; D014667:Vasopressins; C089212:protein kinase D; D017346:Protein Serine-Threonine Kinases; D011493:Protein Kinase C; D001839:Bombesin; D001920:Bradykinin