• RCAD is a Key Post-Translational Modification for the Proper Sorting of Digestive Enzymes and the Secretory Function of the Exocrine Pancreas

      Miller, Camille; College of Science and Mathematics; Sabbatini, Maria (2015-09-25)
      Introduction: Chronic alcohol consumption is one of the main cause of acute pancreatitis. An excess of alcohol causes an increase in free radicals formation and endoplasmic reticulum (ER) stress in pancreatic acinar cells. There are a number of ER proteins that participate in the correct folding/sorting of proteins. One of these proteins is the Ufm1 (Ubiquitin-fold modifier 1) conjugation system, which is a novel ubiquitin-like modification system and part of the unfolded protein response. Regulator of C53 and DDRGK1 (RCAD) has recently been identified as an important regulator of several signaling pathways, including the Ufm1 conjugation system. Endogenous RCAD forms a complex with two proteins, C53 and DDRGK1, and promotes Ufmylation of DDRGK1. Aim: To identify the role of RCAD for the proper sorting of pancreatic enzymes and the normal exocrine function of the pancreas. Methods: The relative expression of RCAD and DDRGK1 in alcoholic-treated rats compared with non-treated rats was evaluated by qPCR analysis. Adult male SD rats were fed the Lieber-DeCarli liquid diet containing ethanol (36 % of total calories) or an isocaloric substitution with maltose-dextrin ad lib for 8 weeks. To determine the functional relevance of RCAD in the exocrine pancreas, isolated pancreatic acini from WT and RCAD inducible conditional KO (CKO) mice were prepared. CCK-induced amylase secretion was measured. Results: Both RCAD and DDRGK1 transcript levels were up-regulated in alcoholic-treated rats. To generate RCAD CKO mice, we crossed RCADTrap-F/+mice with FLPo deleter mice B6(C3)-Tg(Pgk1-FLPo)10Sykr/J to remove the gene trap cassette. The floxed RCAD mice were crossed with ROSA26-CreERT2 mice (B6.129-Gt(ROSA)26Sorotm1(cre/ERT2) Tyj4/J in which CreERT2 was inserted into ROSA26 locus. The constitutive and CCK-inducible amylase secretion was much higher by isolated pancreatic acini prepared from RCAD CKO mice. Conclusion: RCAD is likely to be involved in a post-translational modification system for the proper sorting of amylase and normal pancreatic secretion. The significance of RCAD will provide an important insight into the pathophysiology of acute pancreatitis. Start Time: 34:06 End Time: 46:37