Proteasome inhibition alters glucose-stimulated (pro) insulin secretion and turnover in pancreatic β-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 [ H-3] tyrosine, decreased significantly by 90% in islets and 71% in MIN6 cells pretreated with the proteasome inhibitor lactacystin ( 10 mu M) for 2 h. To follow the fate of newly synthesized ( pro) insulin, islets were pulse- labeled with [H-3] tyrosine ( 40 mu Ci) for 20 min and chased +/- lactacystin ( 10 mu M) for up to 4 h. The release of newly synthesized ( pro) insulin ([H-3] 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 [H-3] 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 [H-3] 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.