Intracellular Autoactivation of Human Cationic Trypsinogen Mutants Causes Reduced Trypsinogen Secretion and Acinar Cell Death

Abstract

Mutations in the activation peptide of human cationic trypsinogen have been found in patients with chronic pancreatitis. Previous biochemical studies demonstrated that mutations p.D19A, p.D22G, and p.K23R strongly stimulate trypsinogen autoactivation. In the present study, we characterized the cell biological effects of these mutants using human embryonic kidney 293T and AR42J rat acinar cells. We found that relative to wild-type trypsinogen, secretion of the mutants from transfected cells was markedly decreased. This apparent secretion defect was completely rescued by inhibition of autoactivation via (1)

Expression of XBP1s in bone marrow stromal cells is critical for myeloma cell growth and osteoclast formation

“…24 The human BM stromal cell lines KM101 cells were cultured in ␣-MEM (Invitrogen) supplemented with 10% FBS and P/S. IRE1␣ ϩ/ϩ , IRE1␣ Ϫ/Ϫ , XBP1 ϩ/ϩ , and XBP1 Ϫ/Ϫ mouse embryonic fibroblast (MEF) cells were kindly provided by Dr Randy Kaufman (University of Michigan, Ann Arbor, MI) and maintained in DMEM supplemented with 10% FBS, L-glutamine, P/S, and additional essential and nonessential amino acids (Invitrogen).…”

Endoplasmic Reticulum Stress Is Chronically Activated in Chronic Pancreatitis

“…Our data suggest that sustained ER stress resulting from misfolding of mutated trypsinogens may be one such mechanism. Such a possibility was recognized in the carboxypeptidase A1 mutations recently recognized in hereditary chronic pancreatitis (45) and is further supported by CP-associated mutations confirming ER stress induction by expression of some mutations (of PRSS1 (41,44), activation peptide of PRSS1 (42), and CTRC (43)) in HEK or rat acinar cell lines. .…”

“…Accumulation of unfolded and misfolded proteins and alteration of ER Ca 2ϩ are well recognized inducers of ER stress in various cells (8,40). Although Ca 2ϩ responses are important for ER stress in caerulein-induced pancreatic injury, alteration of secretion and accumulation of misfolded proteins may be another mechanism (but not necessarily independent of pathologic Ca 2ϩ signaling) of ER stress as has been demonstrated recently in in vitro studies of several CP-associated mutations in the trypsin system (41)(42)(43)(44)(45). In addition to hereditary mutations, alcohol has been shown to cause ER stress in the pancreas (46).…”

Etiology and Risk Factors of Acute and Chronic Pancreatitis

“…Other PRSS1 variants, such as the activation peptide mutants p.D22G, p.K23R, and p.K23_I24insIDK, are clinically rare and tend to associate primarily with sporadic idiopathic disease. These mutants all exhibit reduced secretion because trypsinogen becomes intracellularly autoactivated and degraded [51-53]. It appears likely that inside the endoplasmic reticulum active trypsin is sensed as a misfolded state of trypsinogen, which prompts degradation.…”