Protein kinase Dl mediates NF-kB activation induced by cholecystokinin and cholinergic signaling in pancreatic acinar cells

摘要

Protein kinase Dl mediates NF-kappaB activation induced by cholecystokinin and cholinergic signaling in pancreatic acinar cells. Am J Physiol Gastrointest Liver Phys-iol 295: G1190-G1201, 2008. First published October 9, 2008; .-The transcription factor NF-kappaBplays 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-kB activation induced by cholecystokinin-8 (CCK-8), tumor necrosis factor-alpha, and ethanol. However, the downstream participants in regulating NF-kB activation in exocrine pancreas remain poorly understood. Here, we demonstrate that protein kinase Dl (PKD1) is a key downstream target of PKCdelta and PKCe 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 auto-phosphorylation site (Ser916). The phosphorylation and activation of PKD1 correlated with NF-kappaB activity stimulated by CCK-8 or CCh, as measured by NF-kB 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. Con-versely, overexpression of PKDl resulted in augmentation of CCK-8-and CCh-stimulated NF-kappaB activation. Finally, the kinetics of PKDl and NF-kB activation during cerulein-induced rat pancreatitis showed that both PKDl and NF-kappaB activation were early events during acute pancreatitis and that their time courses of response were similar. Our results identify PKDl as a novel early conver-gent point for PKCdelta and epsilon in the signaling pathways mediating NF-kappaB activation in pancreatitis.