Background & aims: Cholecystokinin (CCK) stimulates protein synthesis in pancreatic acini at the translational level, although the signaling mechanisms involved remain uncharacterized. Two intermediates controlling translation are p70 S6 kinase and PHAS-I. We previously showed that CCK activates p70 S6K in pancreatic acini through phosphatidylinositol 3-kinase (PI 3K). In the present study we investigated the role of PI 3K, p70 S6K, and PHAS-I in mediating CCK-stimulated protein synthesis.
Methods: Protein synthesis was measured by [35S]methionine incorporation into pancreatic protein using acini from rats with streptozotocin-induced diabetes. p70 S6 K activity was measured. PHAS-I was identified by Western analysis. PHAS-I/eIF-4E association was measured as the amount of PHAS-I recovered after purification of translation factor eIF-4E by 7-methyl guanosine triphosphate-Sepharose.
Results: Rapamycin and PI 3K inhibitors, wortmannin and LY294002, blocked CCK-stimulated p70 S6K activity. Rapamycin inhibited basal protein synthesis and blocked the increase to all CCK concentrations. Wortmannin and LY294002 dose-dependently inhibited basal and CCK-stimulated protein synthesis and also blocked insulin-stimulated protein synthesis. CCK dose-dependently increased PHAS-I phosphorylation via a rapamycin- and LY294002-sensitive pathway and decreased the amount of PHAS-I associated with eIF-4E. Rapamycin and LY294002 eliminated this effect of CCK.
Conclusions: CCK stimulation of protein synthesis in pancreatic acini is sensitive to rapamycin and PI 3K inhibitors and involves PHAS-I phosphorylation and its association with eIF-4E.