CN-NFAT-Rcan1 signaling axis in cholecystokinin (CKK)-induced pancreatic growth.

摘要

An increase in circulating CCK induced by feeding protease inhibitors leads to growth of the exocrine pancreas, which is mediated at the acinar cell by the phosphatase calcineurin (CN). Here we examined activation of CN-regulated Nuclear Factor of Activated T-cells (NFATs) and the role of NFATs as transcriptional mediators of CN.;Western blots and confocal imaging using isolated mouse pancreatic acini showed that CCK dose-dependently stimulated NFAT translocation into the nucleus. This shift correlated with CCK-induced activation of an NFAT-luciferase reporter and was blocked by CN inhibitors FK506, Cyclosporine A and overexpression of endogenous protein inhibitor of CN. Parallel changes took place in vivo. Pancreatic growth led to an increase in nuclear NFATs and subsequent activation of NFAT-luciferase in the pancreas, but not liver. These occurred in CN-dependent fashion as they were blocked by FK506.;Activation of NFATs, well-known transcription factors, led us to examine genome-wide, CN-dependent transcriptional events. FK506 blocked expression of 38 genes, as confirmed by qPCR. The CN-dependent genes were linked to growth-related processes, while their promoters were enriched in NFAT and NFAT/API sites. Multiple novel NFAT targets were validated by ChIP. One of these, a CN feedback inhibitor Rcan1 (Regulator of Calcineurin 1), was induced >50 fold during 1--8h course of pancreatic growth and strongly inhibited (>99%) by FK506. We overexpressed Rcan1 in an inducible, acinar cell-specific fashion. Rcan1 inhibited CN-NFAT signaling, as shown by NFAT-luciferase reporter and qPCR. Most notably, the increase in exocrine pancreas size, protein/DNA content and acinar proliferation were blocked in Rcan1 expressing mice. In sum, our results document CCK-mediated and CN-dependent activation of NFATs. We profile adaptive pancreatic growth and delineate NFAT-regulated genes. This leads us to identify Rcan1 as an important new regulator of pancreatic growth and firmly establish that CN-NFAT signaling is required for this response.