Proteasome inhibition alters glucose-stimulated (pro)insulin secretion and turnover in pancreatic {beta}-cells.
Metabolic labeling studies were conducted in freshly isolated mouse islets and a beta-cell line (MIN6) to examine the effects of proteasome inhibition on glucose-stimulated (pro)insulin synthesis and secretion. Glucose-stimulated (pro)insulin synthesis, as determined by the incorporation of [(3)H]tyrosine, decreased significantly by 90% in islets and 71% in MIN6 cells pretreated with the proteasome inhibitor lactacystin (10 microM) for 2 h. To follow the fate of newly synthesized (pro)insulin, islets were pulse-labeled with [(3)H]tyrosine (40 microCi) for 20 min and chased +/- lactacystin (10 microM) for up to 4 h. The release of newly synthesized (pro)insulin ([(3)H]tyrosine-labeled) was similar between lactacystin-treated and control islets despite a 51% decrease (p <0.05) in total immunoreactive (pro)insulin secretion by lactacystin-treated islets. The specific radioactivity of [(3)H]tyrosine-labeled (pro)insulin in the extracellular medium of lactacystin-treated islets (0.52 +/- 0.16 cpm/microunits) was 2-fold greater relative to control islets (0.25 +/- 0.06 cpm/microunits). Induction of the unfolded protein response by lactacystin, as evidenced by the up-regulation of endoplasmic reticulum (ER) chaperones (GRP78/BiP, GRP94, protein disulfide isomerase) and induction of the stress-inducible transcription factor C/EBP-homologous protein/GADD153 (CHOP/GADD153), likely contributed to the release of newly synthesized (pro)insulin to relieve ER stress. The present data indicate proteasome inhibition did not prevent, but increased (p <0.05), the intracellular degradation of [(3)H]tyrosine-labeled (pro-)insulin from 8 to 24% in islets. Collectively, these data indicate beta-cells may balance glucose-stimulated (pro)insulin synthesis and secretion with the activity of the proteasome to regulate protein concentrations in the ER.
Kitiphongspattana
Kajorn
K
Division of Nutritional Sciences, Department of Animal Sciences, Institute for Genomic Biology, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801, USA.
Mathews
Clayton E
CE
Leiter
Edward H
EH
Gaskins
H Rex
HR
D000111:Acetylcysteine / Q000031:analogs & derivatives* / Q000494:pharmacology; D000818:Animals; D000091342:Endoplasmic Reticulum Chaperone BiP; D005260:Female; D005455:Fluorescent Antibody Technique; D005947:Glucose / Q000378:metabolism*; D007515:Islets of Langerhans / Q000378:metabolism*; D051379:Mice; D018613:Microscopy, Confocal; D025821:Polyubiquitin / Q000378:metabolism; D011384:Proinsulin / Q000096:biosynthesis / Q000378:metabolism*; D061988:Proteasome Inhibitors
D000091342:Endoplasmic Reticulum Chaperone BiP; C000717248:Hspa5 protein, mouse; D061988:Proteasome Inhibitors; D025821:Polyubiquitin; C067713:lactacystin; D011384:Proinsulin; D005947:Glucose; D000111:Acetylcysteine