Protein kinase D1 mediates NF-kappaB activation induced by cholecystokinin and cholinergic signaling in pancreatic acinar cells.
The transcription factor NF-kappaB plays a critical role in inflammatory and cell death responses during acute pancreatitis. Previous studies in our laboratory demonstrated that protein kinase C (PKC) isoforms PKCdelta and epsilon are key regulators of NF-kappaB activation induced by cholecystokinin-8 (CCK-8), tumor necrosis factor-alpha, and ethanol. However, the downstream participants in regulating NF-kappaB activation in exocrine pancreas remain poorly understood. Here, we demonstrate that protein kinase D1 (PKD1) is a key downstream target of PKCdelta and PKCepsilon in pancreatic acinar cells stimulated by two major secretagogues, CCK-8 and the cholinergic agonist carbachol (CCh), and that PKD1 is necessary for NF-kappaB activation induced by CCK-8 and CCh. Both CCK-8 and CCh dose dependently induced a rapid and striking activation of PKD1 in rat pancreatic acinar cells, as measured by in vitro kinase assay and by phosphorylation at PKD1 activation loop (Ser744/748) or autophosphorylation site (Ser916). The phosphorylation and activation of PKD1 correlated with NF-kappaB activity stimulated by CCK-8 or CCh, as measured by NF-kappaB DNA binding. Either inhibition of PKCdelta or epsilon by isoform-specific inhibitory peptides, genetic deletion of PKCdelta and epsilon in pancreatic acinar cells, or knockdown of PKD1 by using small interfering RNAs in AR42J cells resulted in a marked decrease in PKD1 and NF-kappaB activation stimulated by CCK-8 or CCh. Conversely, overexpression of PKD1 resulted in augmentation of CCK-8- and CCh-stimulated NF-kappaB activation. Finally, the kinetics of PKD1 and NF-kappaB activation during cerulein-induced rat pancreatitis showed that both PKD1 and NF-kappaB activation were early events during acute pancreatitis and that their time courses of response were similar. Our results identify PKD1 as a novel early convergent point for PKCdelta and epsilon in the signaling pathways mediating NF-kappaB activation in pancreatitis.
Veterans Affairs Greater Los Angeles Healthcare System, West Los Angeles VA Healthcare Center, Los Angeles, CA 90073, USA. jzyuan@ucla.edu
Jingzhen
J
Yuan
Aurelia
A
Lugea
Ling
L
Zheng
Ilya
I
Gukovsky
Mouad
M
Edderkaoui
Enrique
E
Rozengurt
Stephen J
SJ
Pandol
D000818:Animals; D002217:Carbachol / Q000494:pharmacology*; D002108:Ceruletide; D002766:Cholecystokinin / Q000502:physiology*; D018679:Cholinergic Agonists / Q000494:pharmacology*; D051379:Mice; D016328:NF-kappa B / Q000187:drug effects / Q000378:metabolism*; D046790:Pancreas, Exocrine / Q000166:cytology* / Q000502:physiology; D010195:Pancreatitis / Q000139:chemically induced / Q000503:physiopathology; D011493:Protein Kinase C; D051745:Protein Kinase C-delta / Q000235:genetics / Q000502:physiology; D051744:Protein Kinase C-epsilon / Q000235:genetics / Q000502:physiology; D011494:Protein Kinases / Q000502:physiology*; D051381:Rats
D018679:Cholinergic Agonists; D016328:NF-kappa B; D002108:Ceruletide; D002217:Carbachol; D002766:Cholecystokinin; D011494:Protein Kinases; C089212:protein kinase D; D011493:Protein Kinase C; D051745:Protein Kinase C-delta; D051744:Protein Kinase C-epsilon