Essential Role of Ubiquitin-Proteasome System in Normal Regulation of Insulin Secretion

Abstract

Insulin secretion from pancreatic ␤-cells occurs by sequential cellular processes, including glucose metabolism, electrical activity, Ca 2؉ entry, and regulated exocytosis. Abnormalities in any of these functions can impair insulin secretion. In the present study, we demonstrate that inhibition of proteasome activity severely reduces insulin secretion in the mouse pancreatic ␤-cell line MIN6-m9. Although no significant effects on glucose metabolism including ATP production were found in the presence of proteasome inhibitors, both glucose-and KCl-induced Ca 2؉ entry were drastically reduced. As Ca 2؉ -ionophore-induced insulin secretion was unaffected by proteasome inhibition, a defect in Ca 2؉ entry through voltage-dependent calcium channels (VDCCs) is the likely cause of the impaired insulin secretion. We found that the pore-forming ␣-subunit of VDCCs undergoes ubiquitination, which does not decrease but slightly increases expression of the ␣-subunit protein at the plasma membrane. However, electrophysiological analysis revealed that treatment with proteasome inhibitors results in a severe reduction in VDCC activity in MIN6-m9 cells, indicating that VDCC function is suppressed by proteasome inhibition. Furthermore, insulin secretion in isolated mouse pancreatic islets was also decreased by proteasome inhibition. These results demonstrate that the ubiquitin-proteasome system plays a critical role in insulin secretion by maintaining normal function of VDCCs.Insulin secretion from pancreatic ␤-cells is regulated by a variety of extracellular stimuli and intracellular signals (1-3). Regulated exocytosis of insulin-containing secretory granules is triggered by a rise in the intracellular (cytosolic) Ca 2ϩ concentration ([Ca 2ϩ ] i ) 3 (3). In pancreatic ␤-cells, elevation of [Ca 2ϩ ] i occurs by release of Ca 2ϩ from intracellular pools such as the endoplasmic reticulum (ER), mitochondria, and secretory granules and/or by influx of extracellular Ca 2ϩ mainly through voltage-dependent calcium channels (VDCCs) (4, 5). However, insulin secretion is induced by many secretagogues including glucose, sulfonylureas, and glucose-dependent potentiators such as glucose-dependent insulinotropic polypeptide, glucagon-like peptide-1, and pituitary adenylate cyclase-activating peptide, all of which depend critically on Ca 2ϩ entering the pancreatic ␤-cells through the VDCCs, as these secretions are abolished by specific VDCC channel blockers (4, 6, 7).We previously established two different mouse pancreatic ␤-cell sublines from MIN6 cells, which facilitate study of the mechanisms of impaired insulin secretion (8). One of them is MIN6-m9, a model of normal ␤-cells having insulin secretory properties similar to those of pancreatic islets, and the other is MIN6-m14, which exhibits abnormalities in glucose metabolism, ATP-sensitive potassium (K ATP ) channels, and VDCCs that impair insulin secretion (8). In the course of comparing these sublines, we found that several genes involved in the ubiquitin-proteasome system are down-regula...

Ubiquitin C-terminal hydrolase L1 is required for pancreatic beta cell survival and function in lipotoxic conditions

“…Emerging evidence points to the importance of UCHL1 in pancreatic beta cells [6,7,44], and UCHL1 was suggested to be a marker for newly formed pancreatic beta cells [20,39]. Notwithstanding previous correlative studies, our genetic loss-of-function data clearly show that UCHL1 is not required for normal mouse islet development and is dispensable under nonstressed conditions.…”

“…Multiple cellular functions and cell fate decisions are modulated by ubiquitination, a key reversible post-translational modification that can target proteins for degradation or modify their activity [3,4]. Emerging evidence implies essential roles for ubiquitination in both beta cell survival and insulin secretion [5][6][7][8]. The ubiquitin proteasome system and ubiquitin-mediated autophagy have been reported to be regulated by stress conditions, such as glucotoxicity and oxidative stress, resulting in decreased insulin secretion and beta cell apoptosis [6,7].…”

“…Emerging evidence implies essential roles for ubiquitination in both beta cell survival and insulin secretion [5][6][7][8]. The ubiquitin proteasome system and ubiquitin-mediated autophagy have been reported to be regulated by stress conditions, such as glucotoxicity and oxidative stress, resulting in decreased insulin secretion and beta cell apoptosis [6,7]. The roles of specific components of the ubiquitin proteasome system in beta cells are poorly understood.…”

Mitochondrial uncoupling protein 2 in pancreatic β‐cells

“…Whatever the mechanism, UCP2 does appear to be a regulator of the GSIS pathway, at least in INS-1E cells. Like other members of the GSIS pathway, UCP2 proteolysis appears to be proteasome mediated-the proteasome also regulates K ATP channels [47], voltage-dependent calcium channels [48] and proinsulin levels [49]. We can speculate that this multitarget proteasomal degradation system allows concerted regulation of the GSIS pathway under appropriate conditions.…”

Proteasome activity in experimental diabetes

“…A recent study involving the inappropriate degradation of insulin signalling molecules in experimental diabetes suggested that altered UPS might be one of the molecular mechanisms of insulin resistance [40]. Kawaguchi et al [41] have shown that UPS plays an essential role in the normal regulation of insulin secretion.…”