JNK1 Deficient Insulin-Producing Cells Are Protected against Interleukin-1β-Induced Apoptosis Associated with Abrogated Myc Expression (original) (raw)
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JNK1 Protects against Glucolipotoxicity-Mediated Beta-Cell Apoptosis
PLoS ONE, 2014
Pancreatic b-cell dysfunction is central to type 2 diabetes pathogenesis. Prolonged elevated levels of circulating free-fatty acids and hyperglycemia, also termed glucolipotoxicity, mediate b-cell dysfunction and apoptosis associated with increased c-Jun N-terminal Kinase (JNK) activity. Endoplasmic reticulum (ER) and oxidative stress are elicited by palmitate and high glucose concentrations further potentiating JNK activity. Our aim was to determine the role of the JNK subtypes JNK1, JNK2 and JNK3 in palmitate and high glucose-induced b-cell apoptosis. We established insulin-producing INS1 cell lines stably expressing JNK subtype specific shRNAs to understand the differential roles of the individual JNK isoforms. JNK activity was increased after 3 h of palmitate and high glucose exposure associated with increased expression of ER and mitochondrial stress markers. JNK1 shRNA expressing INS1 cells showed increased apoptosis and cleaved caspase 9 and 3 compared to nonsense shRNA expressing control INS1 cells when exposed to palmitate and high glucose associated with increased CHOP expression, ROS formation and Puma mRNA expression. JNK2 shRNA expressing INS1 cells did not affect palmitate and high glucose induced apoptosis or ER stress markers, but increased Puma mRNA expression compared to nonsense shRNA expressing INS1 cells. Finally, JNK3 shRNA expressing INS1 cells did not induce apoptosis compared to nonsense shRNA expressing INS1 cells when exposed to palmitate and high glucose but showed increased caspase 9 and 3 cleavage associated with increased DP5 and Puma mRNA expression. These data suggest that JNK1 protects against palmitate and high glucose-induced b-cell apoptosis associated with reduced ER and mitochondrial stress.
JNK3 is abundant in insulin-secreting cells and protects against cytokine-induced apoptosis
Diabetologia, 2009
Aims/hypothesis In insulin-secreting cells, activation of the c-Jun NH 2 -terminal kinase (JNK) pathway triggers apoptosis. Whereas JNK1 and JNK2 are ubiquitously produced, JNK3 has been described exclusively in neurons. This report aims to characterise the expression and role in apoptosis of the three JNK isoforms in insulin-secreting cells exposed to cytokines. Methods Sections of human and mouse pancreases were used for immunohistochemistry studies with isoformspecific anti-JNK antibodies. Human, pig, mouse and rat pancreatic islets were isolated by enzymatic digestion and RNA or protein extracts were prepared. RNA and protein levels were determined by quantitative RT-PCR and western blotting respectively, using JNK-isoform-specific primers and isoform-specific antibodies; activities of the three JNK isoforms were determined by kinase assays following quantitative immunoprecipitation/depletion of JNK3. JNK silencing was performed with small interfering RNAs and apoptotic rates were determined in INS-1E cells by scoring cells displaying pycnotic nuclei. Results JNK3 and JNK2 mRNAs are the predominant isoforms expressed in human pancreatic islets. JNK3 is nuclear while JNK2 is also cytoplasmic. In INS-1E cells, JNK3 knockdown increases c-Jun levels and caspase-3 cleavage and sensitises cells to cytokine-induced apoptosis; in contrast, JNK1 or JNK2 knockdown is protective. Conclusions/interpretation In insulin-secreting cells, JNK3 plays an active role in preserving pancreatic beta cell mass from cytokine attacks. The specific localisation of JNK3 in the nucleus, its recruitment by cytokines, and its effects on key transcription factors such as c-Jun, indicate that JNK3 is certainly an important player in the transcriptional control of genes expressed in insulin-secreting cells.
Cytokine, 2003
The stress-activated protein kinase c-Jun NH 2-terminal kinase (JNK) is a central signal for interleukin-1b (IL-1b)-induced apoptosis in insulin-producing b-cells. The cell-permeable peptide inhibitor of JNK (JNKI1), that introduces the JNK binding domain (JBD) of the scaffold protein islet-brain 1 (IB1) inside cells, effectively prevents b-cell death caused by this cytokine. To define the molecular targets of JNK involved in cytokine-induced b-cell apoptosis we investigated whether JNKI1 or stable expression of JBD affected the expression of selected pro-and anti-apoptotic genes induced in rat (RIN-5AH-T 2 B) and mouse (bTC3) insulinoma cells exposed to IL-1b. Inhibition of JNK significantly reduced phosphorylation of the specific JNK substrate c-Jun (p < 0:05), IL-1b-induced apoptosis (p < 0:001), and IL-1b-mediated c-fos gene expression. However, neither JNKI1 nor JBD did influence IL-1b-induced NO synthesis or iNOS expression or the transcription of the genes encoding mitochondrial manganese superoxide dismutase (MnSOD), catalase (CAT), glutathione peroxidase (GPx), glutathione-S-transferase q (GSTq), heat shock protein (HSP) 70, IL-1b-converting enzyme (ICE), caspase-3, apoptosis-inducing factor (AIF), Bcl-2 or Bcl-x L. We suggest that the anti-apoptotic effect of JNK inhibition by JBD is independent of the transcription of major pro-and anti-apoptotic genes, but may be exerted at the translational or posttranslational level.
Molecular biology of the cell, 2013
Elevated interleukin-1β (IL-1β) induces apoptosis in pancreatic β-cells through endoplasmic reticulum (ER) stress induction and subsequent c-jun-N-terminal kinase 1/2 (JNK1/2) activation. In earlier work we showed that JNK1/2 activation is initiated before ER stress and apoptotic induction in response to IL-1β. However, the detailed regulatory mechanisms are not completely understood. Because the ER is the organelle responsible for Ca(2+) handling and storage, here we examine the effects of IL-1β on cellular Ca(2+) movement and mitochondrial dysfunction and evaluate the role of JNK1/2. Our results show that in RINm5F cells and human primary β-cells, IL-1β alters mitochondrial membrane potential, mitochondrial permeability transition pore opening, ATP content, and reactive oxygen species production and these alterations are preceded by ER Ca(2+) release via IP3R channels and mitochondrial Ca(2+) uptake. All these events are prevented by JNK1/2 small interfering RNA (siRNA), indicatin...
PLoS ONE, 2012
We have recently shown that silencing of the brain/islet specific c-Jun N-terminal Kinase3 (JNK3) isoform enhances both basal and cytokine-induced beta-cell apoptosis, whereas silencing of JNK1 or JNK2 has opposite effects. While it is known that JNK1 or JNK2 may promote apoptosis by inhibiting the activity of the pro-survival Akt pathway, the effect of JNK3 on Akt has not been documented. This study aims to determine the involvement of individual JNKs and specifically JNK3 in the regulation of the Akt signaling pathway in insulin-secreting cells. JNK3 silencing strongly decreases Insulin Receptor Substrate 2 (IRS2) protein expression, and blocks Akt2 but not Akt1 activation by insulin, while the silencing of JNK1 or JNK2 activates both Akt1 and Akt2. Concomitantly, the silencing of JNK1 or JNK2, but not of JNK3, potently phosphorylates the glycogen synthase kinase3 (GSK3b). JNK3 silencing also decreases the activity of the transcription factor Forkhead BoxO3A (FoxO3A) that is known to control IRS2 expression, in addition to increasing c-Jun levels that are known to inhibit insulin gene expression. In conclusion, we propose that JNK1/2 on one hand and JNK3 on the other hand, have opposite effects on insulin-signaling in insulin-secreting cells; JNK3 protects beta-cells from apoptosis and dysfunction mainly through maintenance of a normal IRS2 to Akt2 signaling pathway. It seems that JNK3 mediates its effects mainly at the transcriptional level, while JNK1 or JNK2 appear to mediate their pro-apoptotic effect in the cytoplasm.
IB1 Reduces Cytokine-induced Apoptosis of Insulin-secreting Cells
Journal of Biological Chemistry, 2000
IB1/JIP-1 is a scaffold protein that interacts with upstream components of the c-Jun N-terminal kinase (JNK) signaling pathway. IB1 is expressed at high levels in pancreatic  cells and may therefore exert a tight control on signaling events mediated by JNK in these cells. Activation of JNK by interleukin 1 (IL-1) or by the upstream JNK constitutive activator ⌬MEKK1 promoted apoptosis in two pancreatic  cell lines and decreased IB1 content by 50 -60%. To study the functional consequences of the reduced IB1 content in  cell lines, we used an insulin-secreting cell line expressing an inducible IB1 antisense RNA that lead to a 38% IB1 decrease. Reducing IB1 levels in these cells increased phosphorylation of c-Jun and increased the apoptotic rate in presence of IL-1. Nitric oxide production was not stimulated by expression of the IB1 antisense RNA. Complementary experiments indicated that overexpression of IB1 in insulin-producing cells prevented JNKmediated activation of the transcription factors c-Jun, ATF2, and Elk1 and decreased IL-1-and ⌬MEKK1-induced apoptosis. These data indicate that IB1 plays an anti-apoptotic function in insulin-producing cells probably by controlling the activity of the JNK signaling pathway.
Diabetes, 2003
We previously reported that interleukin-1β (IL-1β) alone does not cause apoptosis of β-cells, whereas when combined with γ-interferon (IFN-γ) and tumor necrosis factor-α (TNF-α), it exerts a distinct apoptotic effect. Studies in β-cell lines indicated that IL-1β reduced expression of islet brain (IB)-1/JNK interacting protein (JIP)-1, a JNK scaffold protein with antiapoptotic action. We examined whether variations in IB1/JIP-1 expression in purified primary β-cells affect their susceptibility to cytokine-induced apoptosis. Exposure to IL-1β for 24 h decreased cellular IB1/JIP-1 content by 66 ± 17%; this IL-1β effect was maintained in the presence of TNF-α + IFN-γ, which did not influence IB1/JIP-1 levels by themselves. Addition of IL-1β to TNF-α + IFN-γ increased apoptosis from 20 ± 2% to 59 ± 5%. A similar increase in TNF-α + IFN-γ-induced apoptosis was produced by adenoviral expression of antisense IB1/JIP-1 and was not further enhanced by addition of IL-1β, indicating that IL-1β-...
Proceedings of the National Academy of Sciences, 2005
The c-Jun NH2-terminal kinase isoform (JNK) 1 is implicated in type 2 diabetes. However, a potential role for the JNK2 protein kinase in diabetes has not been established. Here, we demonstrate that JNK2 may play an important role in type 1 (insulin-dependent) diabetes that is caused by autoimmune destruction of  cells. Studies of nonobese diabetic mice demonstrated that disruption of the Mapk9 gene (which encodes the JNK2 protein kinase) decreased destructive insulitis and reduced disease progression to diabetes. CD4 ؉ T cells from JNK2-deficient nonobese diabetic mice produced less IFN-␥ but significantly increased amounts of IL-4 and IL-5, indicating polarization toward the Th2 phenotype. This role of JNK2 to control the Th1͞Th2 balance of the immune response represents a mechanism of protection against autoimmune diabetes. We conclude that JNK protein kinases may have important roles in diabetes, including functions of JNK1 in type 2 diabetes and JNK2 in type 1 diabetes.
Functional in vivo interactions between JNK1 and JNK2 isoforms in obesity and insulin resistance
Proceedings of the National Academy of Sciences, 2006
The c-Jun N-terminal kinases (JNKs) are key regulators of inflammation and interfere with insulin action in cultured cells and whole animals. Obesity increases total JNK activity, and JNK1, but not JNK2, deficiency results in reduced adiposity and improved insulin sensitivity. Interestingly, a higher-than-normal level of JNK activation is observed in Jnk2 ؊/؊ mice, particularly in the liver, indicating an interaction between the isoforms that might have masked the metabolic activity of JNK2 in isolated mutant mice. To address the role of the JNK2 isoform in metabolic homeostasis, we intercrossed Jnk1 ؊/؊ and Jnk2 ؊/؊ mice and examined body weight and glucose metabolism in the resulting mutant allele combinations. Among all of the viable genotypes examined, we observed only reduced body weight and increased insulin sensitivity in Jnk1 ؊/؊ and Jnk1 ؉/؊ Jnk2 ؊/؊ mice. These two groups of mice also exhibited reduced total JNK activity and cytokine expression in liver tissue compared with all other genotypes examined. These data indicate that the JNK2 isoform is also involved in metabolic regulation, but its function is not obvious when JNK1 is fully expressed because of regulatory crosstalk between the two isoforms.
Diabetes, 2000
To characterize the differentiation events that selectively target insulin-producing cells to interleukin (IL)-1-induced apoptosis, we studied IL-1 signaling via mitogen-activated protein kinase (MAPK) and stressactivated protein kinase in 2 pancreatic endocrine cell lines. We studied the glucagon-secreting AN-glu cell line and the insulin and the islet amyloid polypeptideproducing -cell line (AN-ins cells), which is derived by stable transfection of AN-glu cells with the transcription factor pancreatic duodenal homeobox factor-1. AN-ins cells were more sensitive to the cytotoxic action of IL-1. This increased sensitivity was not associated with a more pronounced IL-1-induced nitric oxide production in AN-ins cells, but it correlated with a more marked activation of the 3 MAPKs extracellular signal-regulated kinases (ERKs)-1/2, c-Jun NH 2 -terminal kinase (JNK), and p38 MAPK (p38). This led to increased phosphorylation of the transcription factors c-Jun, Elk-1, and ATF2 and of heat shock protein 25. Inhibition of ERK-1/2 and p38 did not prevent but aggravated IL-1-induced cell death. In contrast, inhibition of JNK by transfection with the dominant negative inhibitor of the JNK-binding domain prevented apoptosis in both cell types. Cell death could be elicited by overexpressing the catalytic domain of MAPK kinase kinase 1, a specific activator of JNK and nuclear factor-B, which does not recruit ERK-1/2 or p38. Coactivation of ERK-1/2 with JNK did not prevent apoptosis. In conclusion, increased MAPK signaling in response to IL-1 may represent a novel molecular marker of -cell differentiation. JNK inhibition represents an effective means of preventing IL-1-activated -cell destruction.