Jul 23, 2005

Expression of mutated cationic trypsinogen reduces cellular viability in AR4-2J cells

Biochemical and Biophysical Research Communications
Sebastian GaiserHans Bödeker

Abstract

Mutations in the human cationic trypsinogen are associated with hereditary pancreatitis. The cDNA coding for human cationic trypsinogen was subcloned into the expression vector pcDNA3. The mutations R122H, N29I, A16V, D22G, and K23R were introduced by site directed mutagenesis. We constructed an expression vector coding for active trypsin by subcloning the cDNA of trypsin lacking the coding region for the trypsin activating peptide behind an appropriate signal peptide. Expression of protein was verified by Western blot and measurement of enzymatic activity. AR4-2J cells were transiently transfected with the different expression vectors and cell viability and intracellular caspase-3 activity were quantified. In contrast to wild-type trypsinogen, expression of active trypsin and mutated trypsinogens reduced cell viability of AR4-2J cells. Expression of trypsin and R122H trypsinogen induced caspase-3 activity. Acinar cells might react to intracellular trypsin activity by triggering apoptosis.

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Citations

Mentioned in this Paper

Establishment and Maintenance of Localization
Pathogenic Aspects
Transfection
Tissue Membrane
Aequorea victoria
Pathogenesis
Apoptosis, Intrinsic Pathway
Trypsin Inhibitors
expression vector
Caspase-3

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Apoptosis

Apoptosis is a specific process that leads to programmed cell death through the activation of an evolutionary conserved intracellular pathway leading to pathognomic cellular changes distinct from cellular necrosis