GLP-1 and GIP receptors signal through distinct ß-arrestin 2-dependent pathways to regulate pancreatic ß cell function.

Glucagon-like peptide 1 (GLP-1R) and glucose-dependent insulinotropic polypeptide (GIPR) receptors are G-protein-coupled receptors involved in glucose homeostasis. Diabetogenic conditions decrease ß-arrestin 2 (ARRB2) levels in human islets. In mouse ß cells, ARRB2 dampens insulin secretion by partially uncoupling cyclic AMP (cAMP)/protein kinase A (PKA) signaling at physiological doses of GLP-1, whereas at pharmacological doses, the activation of extracellular signal-related kinase (ERK)/cAMP-responsive element-binding protein (CREB) requires ARRB2. In contrast, GIP-potentiated insulin secretion needs ARRB2 in mouse and human islets. The GIPR-ARRB2 axis is not involved in cAMP/PKA or ERK signaling but does mediate GIP-induced F-actin depolymerization. Finally, the dual GLP-1/GIP agonist tirzepatide does not require ARRB2 for the potentiation of insulin secretion. Thus, ARRB2 plays distinct roles in regulating GLP-1R and GIPR signaling, and we highlight (1) its role in the physiological context and the possible functional consequences of its decreased expression in pathological situations such as diabetes and (2) the importance of assessing the signaling pathways engaged by the agonists (biased/dual) for therapeutic purposes.

The nuclear receptor REV-ERBα is implicated in the alteration of ß-cell autophagy and survival under diabetogenic conditions.

Pancreatic ß-cell failure in type 2 diabetes mellitus (T2DM) is associated with impaired regulation of autophagy which controls ß-cell development, function, and survival through clearance of misfolded proteins and damaged organelles. However, the mechanisms responsible for defective autophagy in T2DM ß-cells remain unknown. Since recent studies identified circadian clock transcriptional repressor REV-ERBα as a novel regulator of autophagy in cancer, in this study we set out to test whether REV-ERBα-mediated inhibition of autophagy contributes to the ß-cell failure in T2DM. Our study provides evidence that common diabetogenic stressors (e.g., glucotoxicity and cytokine-mediated inflammation) augment ß-cell REV-ERBα expression and impair ß-cell autophagy and survival. Notably, pharmacological activation of REV-ERBα was shown to phenocopy effects of diabetogenic stressors on the ß-cell through inhibition of autophagic flux, survival, and insulin secretion. In contrast, negative modulation of REV-ERBα was shown to provide partial protection from inflammation and glucotoxicity-induced ß-cell failure. Finally, using bioinformatic approaches, we provide further supporting evidence for augmented REV-ERBα activity in T2DM human islets associated with impaired transcriptional regulation of autophagy and protein degradation pathways. In conclusion, our study reveals a previously unexplored causative relationship between REV-ERBα expression, inhibition of autophagy, and ß-cell failure in T2DM.

Mechanisms of Beta-Cell Apoptosis in Type 2 Diabetes-Prone Situations and Potential Protection by GLP-1-Based Therapies.

Type 2 diabetes (T2D) is characterized by chronic hyperglycemia secondary to the decline of functional beta-cells and is usually accompanied by a reduced sensitivity to insulin. Whereas altered beta-cell function plays a key role in T2D onset, a decreased beta-cell mass was also reported to contribute to the pathophysiology of this metabolic disease. The decreased beta-cell mass in T2D is, at least in part, attributed to beta-cell apoptosis that is triggered by diabetogenic situations such as amyloid deposits, lipotoxicity and glucotoxicity. In this review, we discussed the molecular mechanisms involved in pancreatic beta-cell apoptosis under such diabetes-prone situations. Finally, we considered the molecular signaling pathways recruited by glucagon-like peptide-1-based therapies to potentially protect beta-cells from death under diabetogenic situations.

Methods to Study Roles of ß-Arrestins in the Regulation of Pancreatic ß-Cell Function.

Novel findings reveal important functional roles for ß-arrestin 1 and ß-arrestin 2 in the regulation of insulin secretion, ß-cell survival, and ß-cell mass plasticity not only by glucose but also by G-protein-coupled receptors, such as the glucagon-like peptide-1 (GLP-1) and the pituitary adenylate cyclase-activating polypeptide (PACAP) receptors or GPR40, or tyrosine kinase receptors, such as the insulin receptor. Here, we describe experimental protocols to knock down ß-arrestins by small interference RNA, to follow subcellular localization of ß-arrestins in the cytosol and nucleus of the insulinoma INS-1E rat pancreatic ß-cell line, and to analyze ß-arrestin protein expression by Western blot using INS-1E cells and isolated mouse or human pancreatic islets. We also provide details on how to genotype ß-arrestin 2 knockout (Arrb2-/-) mice and to evaluate ß-arrestin-mediated roles in ß-cell mass plasticity and ß-cell signaling using immunocytochemistry on pancreatic sections or on primary dispersed ß-cells from wild-type mice and Arrb2-/- mice.

Targeting protein misfolding to protect pancreatic beta-cells in type 2 diabetes.

The islet in type 2 diabetes is characterized by beta-cell dysfunction and deficit, increased beta-cell apoptosis and amyloid deposits that derived from islet amyloid polypeptide (IAPP). In species such as humans that are vulnerable to developing type 2 diabetes, IAPP has the propensity to form toxic oligomers that contribute to beta-cell dysfunction and apoptosis, defining type 2 diabetes as a protein misfolding disorder. In this report, we review mechanisms known to contribute to protein misfolding and formation of toxic oligomers, and the deleterious consequences of these oligomers on beta-cell function and survival. Finally, we will consider approaches to prevent protein misfolding and formation of toxic oligomers as potential novel therapeutic targets for type 2 diabetes and other protein misfolding diseases.

Proteasomal degradation of the histone acetyl transferase p300 contributes to beta-cell injury in a diabetes environment.

In type 2 diabetes, amyloid oligomers, chronic hyperglycemia, lipotoxicity, and pro-inflammatory cytokines are detrimental to beta-cells, causing apoptosis and impaired insulin secretion. The histone acetyl transferase p300, involved in remodeling of chromatin structure by epigenetic mechanisms, is a key ubiquitous activator of the transcriptional machinery. In this study, we report that loss of p300 acetyl transferase activity and expression leads to beta-cell apoptosis, and most importantly, that stress situations known to be associated with diabetes alter p300 levels and functional integrity. We found that proteasomal degradation is the mechanism subserving p300 loss in beta-cells exposed to hyperglycemia or pro-inflammatory cytokines. We also report that melatonin, a hormone produced in the pineal gland and known to play key roles in beta-cell health, preserves p300 levels altered by these toxic conditions. Collectively, these data imply an important role for p300 in the pathophysiology of diabetes.

ERK1 is dispensable for mouse pancreatic beta cell function but is necessary for glucose-induced full activation of MSK1 and CREB.

AIMS/HYPOTHESIS: Insufficient insulin secretion from pancreatic beta cells, which is associated with a decrease in beta cell mass, is a characteristic of type 2 diabetes. Extracellular signal-related kinase 1 and 2 (ERK1/2) inhibition in beta cells has been reported to affect insulin secretion, gene transcription and survival, although whether ERK1 and ERK2 play distinct roles is unknown. The aim of this study was to assess the individual roles of ERK1 and ERK2 in beta cells using ERK1 (also known as Mapk3)-knockout mice (Erk1 -/- mice) and pharmacological approaches. METHODS: NAD(P)H, free cytosolic Ca2+ concentration and insulin secretion were determined in islets. ERK1 and ERK2 subplasmalemmal translocation and activity was monitored using total internal reflection fluorescence microscopy. ERK1/2, mitogen and stress-activated kinase1 (MSK1) and cAMP-responsive element-binding protein (CREB) activation were evaluated by western blot and/or immunocytochemistry. The islet mass was determined from pancreatic sections. RESULTS: Glucose induced rapid subplasmalemmal recruitment of ERK1 and ERK2. When both ERK1 and ERK2 were inhibited simultaneously, the rapid transient peak of the first phase of glucose-induced insulin secretion was reduced by 40% (p < 0.01), although ERK1 did not appear to be involved in this process. By contrast, ERK1 was required for glucose-induced full activation of several targets involved in beta cell survival; MSK1 and CREB were less active in Erk1 -/- mouse beta cells (p < 0.01) compared with Erk1 +/+ mouse beta cells, and their phosphorylation could only be restored when ERK1 was re-expressed and not when ERK2 was overexpressed. Finally, the islet mass of Erk1 -/- mice was slightly increased in young animals (4-month-old mice) vs Erk1 +/+ mice (section occupied by islets [mean ± SEM]: 0.74% ± 0.03% vs 0.62% ± 0.04%; p < 0.05), while older mice (10 months old) were less prone to age-associated pancreatic peri-insulitis (infiltrated islets [mean ± SEM]: 7.51% ± 1.34% vs 2.03% ± 0.51%; p < 0.001). CONCLUSIONS/INTERPRETATION: ERK1 and ERK2 play specific roles in beta cells. ERK2 cannot always compensate for the lack of ERK1 but the absence of a clear-cut phenotype in Erk1 -/- mice shows that ERK1 is dispensable in normal conditions.

ß Cell-specific increased expression of calpastatin prevents diabetes induced by islet amyloid polypeptide toxicity.

The islet in type 2 diabetes (T2D) shares many features of the brain in protein misfolding diseases. There is a deficit of ß cells with islet amyloid derived from islet amyloid polypeptide (IAPP), a protein coexpressed with insulin. Small intracellular membrane-permeant oligomers, the most toxic form of IAPP, are more frequent in ß cells of patients with T2D and rodents expressing human IAPP. ß Cells in T2D, and affected cells in neurodegenerative diseases, share a comparable pattern of molecular pathology, including endoplasmic reticulum stress, mitochondrial dysfunction, attenuation of autophagy, and calpain hyperactivation. While this adverse functional cascade in response to toxic oligomers is well described, the sequence of events and how best to intervene is unknown. We hypothesized that calpain hyperactivation is a proximal event and tested this in vivo by ß cell-specific suppression of calpain hyperactivation with calpastatin overexpression in human IAPP transgenic mice. ß Cell-specific calpastatin overexpression was remarkably protective against ß cell dysfunction and loss and diabetes onset. The critical autophagy/lysosomal pathway for ß cell viability was protected with calpain suppression, consistent with findings in models of neurodegenerative diseases. We conclude that suppression of calpain hyperactivation is a potentially beneficial disease-modifying strategy for protein misfolding diseases, including T2D.

Activation of Melatonin Signaling Promotes ß-Cell Survival and Function.

Type 2 diabetes mellitus (T2DM) is characterized by pancreatic islet failure due to loss of ß-cell secretory function and mass. Studies have identified a link between a variance in the gene encoding melatonin (MT) receptor 2, T2DM, and impaired insulin secretion. This genetic linkage raises the question whether MT signaling plays a role in regulation of ß-cell function and survival in T2DM. To address this postulate, we used INS 832/13 cells to test whether activation of MT signaling attenuates proteotoxicity-induced ß-cell apoptosis and through which molecular mechanism. We also used nondiabetic and T2DM human islets to test the potential of MT signaling to attenuate deleterious effects of glucotoxicity and T2DM on ß-cell function. MT signaling in ß-cells (with duration designed to mimic typical nightly exposure) significantly enhanced activation of the cAMP-dependent signal transduction pathway and attenuated proteotoxicity-induced ß-cell apoptosis evidenced by reduced caspase-3 cleavage (∼40%), decreased activation of stress-activated protein kinase/Jun-amino-terminal kinase (∼50%) and diminished oxidative stress response. Activation of MT signaling in human islets was shown to restore glucose-stimulated insulin secretion in islets exposed to chronic hyperglycemia as well as in T2DM islets. Our data suggest that ß-cell MT signaling is important for the regulation of ß-cell survival and function and implies a preventative and therapeutic potential for preservation of ß-cell mass and function in T2DM.

Insulin-degrading enzyme inhibition, a novel therapy for type 2 diabetes?

The insulin-degrading enzyme (IDE) has been identified as a type 2 diabetes and Alzheimer's disease susceptibility gene, though its physiological function remains unclear. Maianti et al. (2014) now propose that an IDE inhibitor may be a promising therapeutic strategy for type 2 diabetes.

Autophagy defends pancreatic ß cells from human islet amyloid polypeptide-induced toxicity.

Type 2 diabetes (T2D) is characterized by a deficiency in ß cell mass, increased ß cell apoptosis, and extracellular accumulation of islet amyloid derived from islet amyloid polypeptide (IAPP), which ß cells coexpress with insulin. IAPP expression is increased in the context of insulin resistance, the major risk factor for developing T2D. Human IAPP is potentially toxic, especially as membrane-permeant oligomers, which have been observed to accumulate within ß cells of patients with T2D and rodents expressing human IAPP. Here, we determined that ß cell IAPP content is regulated by autophagy through p62-dependent lysosomal degradation. Induction of high levels of human IAPP in mouse ß cells resulted in accumulation of this amyloidogenic protein as relatively inert fibrils within cytosolic p62-positive inclusions, which temporarily averts formation of toxic oligomers. Mice hemizygous for transgenic expression of human IAPP did not develop diabetes; however, loss of ß cell-specific autophagy in these animals induced diabetes, which was attributable to accumulation of toxic human IAPP oligomers and loss of ß cell mass. In human IAPP-expressing mice that lack ß cell autophagy, increased oxidative damage and loss of an antioxidant-protective pathway appeared to contribute to increased ß cell apoptosis. These findings indicate that autophagy/lysosomal degradation defends ß cells against proteotoxicity induced by oligomerization-prone human IAPP.

UCHL1 deficiency exacerbates human islet amyloid polypeptide toxicity in ß-cells: evidence of interplay between the ubiquitin/proteasome system and autophagy.

The islet in type 2 diabetes mellitus (T2DM) is characterized by a deficit in ß-cells and increased ß-cell apoptosis attributable at least in part to intracellular toxic oligomers of IAPP (islet amyloid polypeptide). ß-cells of individuals with T2DM are also characterized by accumulation of polyubiquitinated proteins and deficiency in the deubiquitinating enzyme UCHL1 (ubiquitin carboxyl-terminal esterase L1 [ubiquitin thiolesterase]), accounting for a dysfunctional ubiquitin/proteasome system. In the present study, we used mouse genetics to elucidate in vivo whether a partial deficit in UCHL1 enhances the vulnerability of ß-cells to human-IAPP (hIAPP) toxicity, and thus accelerates diabetes onset. We further investigated whether a genetically induced deficit in UCHL1 function in ß-cells exacerbates hIAPP-induced alteration of the autophagy pathway in vivo. We report that a deficit in UCHL1 accelerated the onset of diabetes in hIAPP transgenic mice, due to a decrease in ß-cell mass caused by increased ß-cell apoptosis. We report that UCHL1 dysfunction aggravated the hIAPP-induced defect in the autophagy/lysosomal pathway, illustrated by the marked accumulation of autophagosomes and cytoplasmic inclusions positive for SQSTM1/p62 and polyubiquitinated proteins with lysine 63-specific ubiquitin chains. Collectively, this study shows that defective UCHL1 function may be an early contributor to vulnerability of pancreatic ß-cells for protein misfolding and proteotoxicity, hallmark defects in islets of T2DM. Also, given that deficiency in UCHL1 exacerbated the defective autophagy/lysosomal degradation characteristic of hIAPP proteotoxicity, we demonstrate a previously unrecognized role of UCHL1 in the function of the autophagy/lysosomal pathway in ß-cells.

ß-Cell failure in type 2 diabetes: a case of asking too much of too few?

The islet in type 2 diabetes (T2DM) is characterized by a deficit in ß-cells, increased ß-cell apoptosis, and extracellular amyloid deposits derived from islet amyloid polypeptide (IAPP). In the absence of longitudinal studies, it is unknown if the low ß-cell mass in T2DM precedes diabetes onset (is a risk factor for diabetes) or develops as a consequence of the disease process. Although insulin resistance is a risk factor for T2DM, most individuals who are insulin resistant do not develop diabetes. By inference, an increased ß-cell workload results in T2DM in some but not all individuals. We propose that the extent of the ß-cell mass that develops during childhood may underlie subsequent successful or failed adaptation to insulin resistance in later life. We propose that a low innate ß-cell mass in the face of subsequent insulin resistance may expose ß-cells to a burden of insulin and IAPP biosynthetic demand that exceeds the cellular capacity for protein folding and trafficking. If this threshold is crossed, intracellular toxic IAPP membrane permeant oligomers (cylindrins) may form, compromising ß-cell function and inducing ß-cell apoptosis.

Roles and regulation of the transcription factor CREB in pancreatic ß -cells.

The preservation of a functional pancreatic ß-cell mass has become a major point of research in type 2 diabetes (T2D) and the future therapies of T2D notably aim at protecting the ß-cell from dysfunction and apoptotic death. ß-cell proliferation, survival and insulin secretion are regulated by crucial transcription factors which are activated by signalling pathways engaged by nutrients, G-protein coupled receptors or tyrosine kinase receptors. Among these factors, the cAMP-responsive element-binding protein (CREB) has emerged as a key transcriptional element for the maintenance of an efficient glucose sensing, insulin exocytosis, insulin gene transcription and ß-cell survival. CREB activates the transcription of target genes within the ß-cells in response to a diverse array of stimuli including glucose, incretin hormones such as the glucagon-like peptide-1 (GLP-1) or the gastric inhibitory polypeptide (GIP), the pituitary adenylate cyclase-activating polypeptide (PACAP), or growth factors such as the insulin like growth factor-1 (IGF-1). All these stimuli phosphorylate CREB at a particular residue, serine 133, which is required for CREB-mediated transcription. However, the molecular mechanisms by which CREB activates gene transcription in ß-cells vary according to the nature of the stimulus. These mechanisms involve different protein kinases, scaffold proteins and cofactors which allow CREB to specifically regulate the expression of crucial genes such as insulin, BCL-2, cyclin D1, cyclin A2 or IRS-2. In this review, we summarize the signalling pathways that lead to CREB phosphorylation in ß-cells and the molecular features of each signalling pathway that rise specificity at the level of CREB activation and regulation.

Cyclin-dependent kinase 5 promotes pancreatic ß-cell survival via Fak-Akt signaling pathways.

OBJECTIVE: Cyclin-dependent kinase 5 (CDK5) regulatory subunit-associated protein 1-like 1 has recently been linked to type 2 diabetes by genome-wide association studies. While CDK5 and its regulatory protein p35 are both expressed and display enzymatic activity in pancreatic ß-cells, their precise role in the ß-cell remains unknown. Because type 2 diabetes is characterized by a deficit in ß-cell mass and increased ß-cell apoptosis, we investigated the role of CDK5 in ß-cell survival. RESEARCH DESIGN AND METHODS: We used INS 832/13 cells, rat islets isolated from wild-type or human islet amyloid polypeptide (h-IAPP) transgenic rats, and pancreatic tissue from rats and humans with and without type 2 diabetes and investigated the effect of CDK5/p35 inhibition (by small interfering RNA or by chemical inhibition) as well as CDK5/p35 overexpression on ß-cell vulnerability to apoptosis. RESULTS: CDK5 inhibition led to increased ß-cell apoptosis. To identify the mechanisms involved, we examined the phosphorylation state of focal adhesion kinase (Fak)(Ser732), a known target of CDK5. Following CDK5 inhibition, the phosphorylation of Fak(Ser732) decreased with resulting attenuation of phosphatidylinositol 3-kinase (PI3K)/Akt survival pathway. Conversely, CDK5 overexpression increased Fak(Ser732) phosphorylation and protected ß-cells against apoptosis induced by the inhibition of the ß-1 integrin signaling pathway. Also, Fak(Ser732) phosphorylation was less abundant in ß-cells in both h-IAPP transgenic rats and humans with type 2 diabetes. CONCLUSIONS: This study shows that by regulating Fak phosphorylation and subsequently PI3K/Akt survival pathway, CDK5 plays a previously unrecognized role in promoting ß-cell survival.

ß-cell dysfunctional ERAD/ubiquitin/proteasome system in type 2 diabetes mediated by islet amyloid polypeptide-induced UCH-L1 deficiency.

OBJECTIVE: The islet in type 2 diabetes is characterized by ß-cell apoptosis, ß-cell endoplasmic reticulum stress, and islet amyloid deposits derived from islet amyloid polypeptide (IAPP). Toxic oligomers of IAPP form intracellularly in ß-cells in humans with type 2 diabetes, suggesting impaired clearance of misfolded proteins. In this study, we investigated whether human-IAPP (h-IAPP) disrupts the endoplasmic reticulum-associated degradation/ubiquitin/proteasome system. RESEARCH DESIGN AND METHODS: We used pancreatic tissue from humans with and without type 2 diabetes, isolated islets from h-IAPP transgenic rats, isolated human islets, and INS 832/13 cells transduced with adenoviruses expressing either h-IAPP or a comparable expression of rodent-IAPP. Immunofluorescence and Western blotting were used to detect polyubiquitinated proteins and ubiquitin carboxyl-terminal hydrolase L1 (UCH-L1) protein levels. Proteasome activity was measured in isolated rat and human islets. UCH-L1 was knocked down by small-interfering RNA in INS 832/13 cells and apoptosis was evaluated. RESULTS: We report accumulation of polyubiquinated proteins and UCH-L1 deficiency in ß-cells of humans with type 2 diabetes. These findings were reproduced by expression of oligomeric h-IAPP but not soluble rat-IAPP. Downregulation of UCH-L1 expression and activity to reproduce that caused by h-IAPP in ß-cells induced endoplasmic reticulum stress leading to apoptosis. CONCLUSIONS: Our results indicate that defective protein degradation in ß-cells in type 2 diabetes can, at least in part, be attributed to misfolded h-IAPP leading to UCH-L1 deficiency, which in turn further compromises ß-cell viability.

The effect of curcumin on human islet amyloid polypeptide misfolding and toxicity.

Type 2 diabetes involves aberrant misfolding of human islet amyloid polypeptide (h-IAPP) and resultant pancreatic amyloid deposits. Curcumin, a biphenolic small molecule, has offered potential benefits in other protein misfolding diseases, such as Alzheimer's disease. Our aim was to investigate whether curcumin alters h-IAPP misfolding and protects from cellular toxicity at physiologically relevant concentrations. The effect of curcumin on h-IAPP misfolding in vitro was investigated by electron paramagnetic resonance spectroscopy, ThT fluorescence and electron microscopy. Our in vitro studies revealed that curcumin significantly reduces h-IAPP fibril formation and aggregates formed in the presence of curcumin display alternative morphology and structure. We then tested a potential protective effect of curcumin against h-IAPP toxicity on ß-cells. Micromolar concentrations of curcumin partially protect INS cells from exogenous IAPP toxicity. This protective effect, however, is limited to a narrow concentration range, as curcumin becomes cytotoxic at micromolar concentrations. In different models of endogenous over-expression of h-IAPP (INS cells and h-IAPP transgenic rat islets), curcumin failed to protect ß-cells from h-IAPP-induced apoptosis. While curcumin has the ability to inhibit amyloid formation, the present data suggest that, without further modification, it is unlikely to be therapeutically useful in protection of ß-cells in type 2 diabetes.

Calcium-activated calpain-2 is a mediator of beta cell dysfunction and apoptosis in type 2 diabetes.

The islet in type 2 diabetes (T2DM) and the brain in neurodegenerative diseases share progressive cell dysfunction, increased apoptosis, and accumulation of locally expressed amyloidogenic proteins (islet amyloid polypeptide (IAPP) in T2DM). Excessive activation of the Ca(2+)-sensitive protease calpain-2 has been implicated as a mediator of oligomer-induced cell death and dysfunction in neurodegenerative diseases. To establish if human IAPP toxicity is mediated by a comparable mechanism, we overexpressed human IAPP in rat insulinoma cells and freshly isolated human islets. Pancreas was also obtained at autopsy from humans with T2DM and nondiabetic controls. We report that overexpression of human IAPP leads to the formation of toxic oligomers and increases beta cell apoptosis mediated by increased cytosolic Ca(2+) and hyperactivation of calpain-2. Cleavage of alpha-spectrin, a marker of calpain hyperactivation, is increased in beta cells in T2DM. We conclude that overactivation of Ca(2+)-calpain pathways contributes to beta cell dysfunction and apoptosis in T2DM.

GLP-1 mediates antiapoptotic effect by phosphorylating Bad through a beta-arrestin 1-mediated ERK1/2 activation in pancreatic beta-cells.

Strategies based on activating GLP-1 receptor (GLP-1R) are intensively developed for the treatment of type 2 diabetes. The exhaustive knowledge of the signaling pathways linked to activated GLP-1R within the beta-cells is of major importance. In beta-cells, GLP-1 activates the ERK1/2 cascade by diverse pathways dependent on either Galpha(s)/cAMP/cAMP-dependent protein kinase (PKA) or beta-arrestin 1, a scaffold protein. Using pharmacological inhibitors, beta-arrestin 1 small interfering RNA, and islets isolated from beta-arrestin 1 knock-out mice, we demonstrate that GLP-1 stimulates ERK1/2 by two temporally distinct pathways. The PKA-dependent pathway mediates rapid and transient ERK1/2 phosphorylation that leads to nuclear translocation of the activated kinases. In contrast, the beta-arrestin 1-dependent pathway produces a late ERK1/2 activity that is restricted to the beta-cell cytoplasm. We further observe that GLP-1 phosphorylates the cytoplasmic proapoptotic protein Bad at Ser-112 but not at Ser-155. We find that the beta-arrestin 1-dependent ERK1/2 activation engaged by GLP-1 mediates the Ser-112 phosphorylation of Bad, through p90RSK activation, allowing the association of Bad with the scaffold protein 14-3-3, leading to its inactivation. beta-Arrestin 1 is further found to mediate the antiapoptotic effect of GLP-1 in beta-cells through the ERK1/2-p90RSK-phosphorylation of Bad. This new regulatory mechanism engaged by activated GLP-1R involving a beta-arrestin 1-dependent spatiotemporal regulation of the ERK1/2-p90RSK activity is now suspected to participate in the protection of beta-cells against apoptosis. Such signaling mechanism may serve as a prototype to generate new therapeutic GLP-1R ligands.