Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P01275 (glucagon)
 26,492 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

GLP-1 (glucagon-like peptide-1) is a potent insulin secretagogue released from L cells in the intestine. The regulation of GLP-1 secretion has been described both in vivo and in vitro in several animal species, but data from human cellular models are lacking. For this purpose, factors and cell-signaling pathways regulating GLP-1 secretion were investigated in the NCI-H716 human intestinal cell line. After differentiation, these cells homogeneously produced 16.8 pmol GLP-1/mg protein with a basal release of 4.2% during a 2-h incubation period. Nutrients, such as palmitic acid, oleic acid, and meat hydrolysate, stimulated GLP-1 secretion in a dose-dependent manner, as did the cholinergic agonist carbachol and the neuromediator gastrin-releasing peptide. Along with stimulating GLP-1 release, gastrin-releasing peptide, like ionomycin, increased intracellular calcium levels. Activators of PKA and PKC were able to increase GLP-1 secretion in NCI-H716 cells. However, neither PKA activators nor meat hydrolysate increased proglucagon mRNA levels. These findings indicate that the NCI-H716 cell line constitutes a unique model to study the cellular mechanism of GLP-1 secretion in humans and suggest potential interspecies divergence in the regulation of proglucagon gene expression in enteroendocrine cells.
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PMID:A human cellular model for studying the regulation of glucagon-like peptide-1 secretion. 1156 18

A full biphasic insulin response is the most sensitive index for well-coupled beta-cell signal transduction. While first-phase insulin response is extremely sensitive to potentiating and inhibiting modulations, full expression of second-phase response requires near maximally activated beta-cell fuel metabolism. In the isolated rat pancreas, accelerated calcium entry or activation of protein kinase (PK)-A or PKC result in no insulin response in the absence of fuel metabolism. At submaximal levels of beta-cell fuel secretagogue, arginine (which promotes calcium entry) or glucagon (which activates PKA) produces a small first-phase insulin response but minimal or no second-phase response; carbachol (which activates PKC and promotes calcium entry) generates biphasic insulin response in the presence of minimal fuel (3.3 mmol/l glucose). Glucagon produces full biphasic response in the presence of 10.0 mmol/l glucose, whereas arginine requires near-maximal stimulatory glucose (16.7 mmol) to produce full biphasic insulin response. Thus, PKA and PKC signal pathways potentiate primary signals generated by fuel secretagogues to induce full biphasic insulin response, while calcium recruitment alone is insufficient to potentiate primary signals generated at low levels of fuel secretagogue. We suggest that three families of PKs (calmodulin-dependent PK [CaMK], PKA, and PKC) function as distal amplifiers for stimulus-secretion coupling signals originating from fuel metabolism, as well as from incretins acting through membrane receptors, adenylate cyclase, and phospholipase C. Several isoenzymes of PKA and PKC are present in pancreatic beta-cells, but the specific function of most is still undefined. Each PK isoenzyme is activated and subsequently phosphorylates its specific effector protein by binding to a highly specific anchoring protein. Some diabetes-related beta-cell derangements may be linked to abnormal function of one or more PK isoenzymes. Identification and characterization of the specific function of the individual PK isoenzymes may provide the tool to improve the insulin response of the diabetic patient.
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PMID:Beta-cell protein kinases and the dynamics of the insulin response to glucose. 1181 61

The mammalian insulin gene is exclusively expressed in the beta cells of the endocrine pancreas. Two decades of intensive physiological and biochemical studies have led to the identification of regulatory sequence motifs along the insulin promoter and to the isolation of transcription factors which interact to activate gene transcription. The majority of the islet-restricted (BETA2, PDX-1, RIP3b1-Act/C1) and ubiquitous (E2A, HEB) insulin-binding proteins have been characterized. Transcriptional regulation results not only from specific combinations of these activators through DNA-protein and protein-protein interactions, but also from their relative nuclear concentrations, generating a cooperativity and transcriptional synergism unique to the insulin gene. Their DNA binding activity and their transactivating potency can be modified in response to nutrients (glucose, NEFA) or hormonal stimuli (insulin, leptin, glucagon like peptide-1, growth hormone, prolactin) through kinase-dependent signalling pathways (PI3-K, p38MAPK, PKA, CaMK) modulating their affinities for DNA and/or for each other. From the overview of the research presented, it is clear that much more study is required to fully comprehend the mechanisms involved in the regulated-expression of the insulin gene in the beta cell to prevent its impairment in diabetes.
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PMID:Regulation of insulin gene transcription. 1191 36

Signal transduction properties of exendin-4 (Ex-4) underlying its ability to stimulate rat insulin I gene promoter (RIP1) activity were assessed in the pancreatic beta-cell line INS-1. Ex-4 acted via glucagon-like peptide-1 receptors to stimulate RIP1 in a glucose-dependent manner, as measured in cells transfected with a -410-bp RIP1-luciferase construct (RIP1-Luc). The action of Ex-4 was independent of cAMP and PKA because it was not blocked by cotransfection with dominant-negative G alpha(s), was unaffected by pretreatment with the membrane-permeant cAMP antagonist 8-Br-Rp-cAMPS, and remained apparent after treatment with PKA inhibitors H-89 or KT 5720. Similarly, cotransfection with a dominant-negative isoform of the type-2 cAMP-regulated guanine nucleotide exchange factor (Epac2) failed to alter the response to Ex-4. Ro 31-8220, a serine/threonine protein kinase inhibitor that targets PKC as as well as the 90-kDa ribosomal S6 kinase (RSK) and mitogen- and stress-activated protein kinase (MSK) family of cAMP response element-binding protein (CREB) kinases, blocked the stimulatory action of Ex-4 at RIP1-Luc. However, selective inhibition of PKC using K-252c, prolonged exposure to phorbol 1,2-myristate-13-acetate, or cotransfection with dominant-negative atypical PKC-zeta, was without effect. A-CREB, a dominant-negative inhibitor of basic region-leucine zipper transcription factors (bZIPs) related in structure to CREB, inhibited the action of Ex-4 at RIP1-Luc, whereas A-ATF-2 was ineffective. Similarly, introduction of deletions at the RIP1 cAMP response element (CRE), or truncation of RIP1 to remove the CRE, nearly abolished the action of Ex-4. Inactivating mutations introduced at the A4/A3 elements, binding sites for the glucose-regulated homeodomain transcription factor PDX-1, did not diminish the response to Ex-4, although a marked reduction of basal promoter activity was observed. The glucose-dependent stimulation of RIP1-Luc by Ex-4 was reproduced using a synthetic reporter (RIP1-CRE-Luc) incorporating multimerized CREs of the RIP1 nonpalindromic sequence 5'-TGACGTCC-3'. It is concluded that the bZIP and CRE-mediated stimulation of RIP1 by Ex-4 explains, at least in part, how this insulinotropic hormone facilitates transcriptional activity of the rat insulin I gene.
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PMID:Exendin-4 as a stimulator of rat insulin I gene promoter activity via bZIP/CRE interactions sensitive to serine/threonine protein kinase inhibitor Ro 31-8220. 1202 Nov 95

Activation of protein kinase A (cAMP-dependent protein kinase; PKA) triggers insulin secretion in the beta-cell. Adenylate cyclase toxin (ACT), a bacterial exotoxin with adenylate cyclase activity, and forskolin, an activator of adenylate cyclase, both dose-dependently increased insulin secretion in the presence, but not the absence, of glucose in insulin-secreting betaTC3 cells. The stimulation of cAMP release by either agent was dose-dependent but glucose-independent. Omission of extracellular Ca(2+) totally abolished the effects of ACT on insulin secretion and cytosolic cAMP accumulation. ACT and forskolin caused rapid and dramatic increases in cytosolic Ca(2+), which were blocked by nifedipine and the omission of extracellular Ca(2+). Omission of glucose completely blocked the effects of forskolin and partially blocked the effects of ACT on cytosolic Ca(2+). PKA alpha, beta and gamma catalytic subunits (Calpha, Cbeta and Cgamma respectively) were identified in betaTC6 cells by confocal microscopy. Glucose and glucagon-like polypeptide-1 (GLP-1) caused translocation of Calpha to the nucleus and of Cbeta to the plasma membrane and the nucleus, but did not affect the distribution of Cgamma. In conclusion, glucose and GLP-1 amplify insulin secretion via cAMP production and PKAbeta activation.
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PMID:Protein kinase A translocation and insulin secretion in pancreatic beta-cells: studies with adenylate cyclase toxin from Bordetella pertussis. 1218 Sep 8

Isolated rat islets were exposed to cAMP-elevating agents and/or nutrients. Insulin exocytosis subsequently triggered by a depolarizing concentration of K(+) or a stimulatory concentration of glucose was employed as an index of time-dependent potentiation (TDP). Stimulatory concentrations (>or=5.5 mM) of glucose caused TDP, and 6 micro M forskolin (an activator of adenylyl cyclase) significantly enhanced it (3.1-fold at most). Forskolin produced an 8.0-fold increase in islet cell cAMP; however, it returned to the baseline after washout by the time of stimulation of exocytosis. Two millimoles of dibutyryl cAMP (a cAMP analog), 0.1 mM isobutylmethylxanthine (a phosphodiesterase inhibitor), and 100 nM glucagon-like peptide-1 (an incretin hormone) also enhanced glucoseinduced TDP. The time-dependent effect of cAMP was not attenuated by protein kinase A inhibitors (200 micro M adenosine 3',5'-cyclic monophosphothioate, Rp isomer, and 10 micro M H89). Although glucose-induced TDP was attenuated by NaN(3) (a mitochondrial poison) and cerulenin (an inhibitor of protein acylation), cAMP enhancement of it was unaffected by these agents. In conclusion, cAMP time-dependently stimulates insulin exocytosis, provided the extracellular glucose concentration is equivalent to or higher than ambient plasma levels. Protein kinase A, mitochondrial metabolism, and protein acylation are not involved in this cAMP action. Incretin stimulation of insulin exocytosis may occur in part via this mechanism.
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PMID:Time-dependent stimulation of insulin exocytosis by 3',5'-cyclic adenosine monophosphate in the rat islet beta-cell. 1239 13

The activity of glycogen phosphorylase (GPase) in the active a-form (GPase a) is dependent on the hydration state of hepatocytes. We establish that GPase a catalysis in catfish (Ameiurus nebulosus) hepatocytes is a function of medium osmolarity and that a linear relationship exists between GPase a activity and osmolarity between 254 mosmol l(-1) and 478 mosmol l(-1). Exposure of isolated hepatocytes to hyperosmotic media increases enzyme activity up to 7-fold, indicative of covalent phosphorylation. GPase activation associated with cell shrinkage peaks within 10 min of exposure. The average degree of activation (2.7-fold-increase of GPase a) is only slightly less than in hepatocytes exposed to glucagon (3.1-fold-increase) under isosmotic conditions; with glucagon, the maximum is reached within 2 min. Phosphorylation status remains elevated during the entire 40 min experimental period; cells do not undergo regulatory volume increase (RVI) during this period and do not regain pre-exposure volume. We interpret the increased GPase a activity as an inherent response to hyperosmotic stress, likely brought about by molecular crowding. Activation of the enzyme results in increased glucose production from endogenous glycogen. Glucose is not retained in the liver cells, but may act as an oxidative substrate in extrahepatic tissues for the increased metabolic demand of ion regulation. Protein kinase A or intracellular Ca(2+) make apparently small contributions to the activation of GPase, leaving us to speculate on alternate routes of enzyme activation. Conversely, hepatocyte swelling in hyposmotic medium leads to significant decreases in GPase a activity and curtailed glucose output. A minimum is attained in 10 min, and pre-insult rates are re-established within 40 min, somewhat lagging behind readjustment in cell volume by regulatory volume decrease (RVD). We conclude that cell swelling and subsequent RVD do not signify stress to the cells and metabolic demand may be decreased under cell swelling conditions. Alteration of GPase phosphorylation with extracellular osmolarity appears to be a general phenomenon, since we also find it in hepatocytes of another freshwater catfish (Clarias batrachus) and a marine scorpaenid (Sebastes caurinus).
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PMID:Cell volume affects glycogen phosphorylase activity in fish hepatocytes. 1289 65

Glucagon-like peptide-1 (GLP-1) elevates the intracellular free calcium concentration ([Ca2+]i) and insulin secretion in a Na+-dependent manner. To investigate a possible role of Na ion in the action of GLP-1 on pancreatic islet cells, we measured the glucose-and GLP-1-induced intracellular Na+ concentration ([Na+]i), [Ca2+]i, and insulin secretion in hamster islet cells in various concentrations of Na+. The [Na+]i and [Ca2+]i were monitored in islet cells loaded with sodium-binding benzofuran isophthalate and fura 2, respectively. In the presence of 135 mM Na+ and 8 mM glucose, GLP-1 (10 nM) strongly increased the [Na+]i, [Ca2+]i, and insulin secretion. In the presence of 13.5 mM Na+, both glucose and GLP-1 increased neither the [Na+]i nor the [Ca2+]i. In a Na+-free medium, GLP-1 and glucose did not increase the [Na+]i. SQ-22536, an inhibitor of adenylate cyclase, and H-89, an inhibitor of PKA, incompletely inhibited the response. In the presence of both 8 mM glucose and H-89, 8-pCPT-2'-O-Me-cAMP, a PKA-independent cAMP analog, increased the insulin secretion and the [Na+]i. Therefore, we conclude that GLP-1 increases the cAMP level via activation of adenylate cyclase, which augments the membrane Na+ permeability through PKA-dependent and PKA-independent mechanisms, thereby increasing the [Ca2+]i and promoting insulin secretion from hamster islet cells.
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PMID:Glucagon-like peptide-1 induces a cAMP-dependent increase of [Na+]i associated with insulin secretion in pancreatic beta-cells. 1453 75

cAMP has previously been shown to promote cell survival in a variety of cell types, but the downstream signaling pathway(s) of this antiapoptotic effect is unclear. Thus the role of cAMP signaling through PKA and cAMP-regulated guanine nucleotide exchange factors (cAMP-GEFs) in cAMP's antiapoptotic action was investigated in the present study. cAMP's protective effect against bile acid-, Fas ligand-, and TNF-alpha-induced apoptosis in rat hepatocytes was largely unaffected by the selective PKA inhibitor, Rp-8-(4-chlorophenylthio)-cAMP (Rp-cAMP). In contrast, a novel cAMP analog, 8-(4-chlorophenylthio)-2'-O-methyl (CPT-2-Me)-cAMP, which activated cAMP-GEFs in hepatocytes without activating PKA, protected hepatocytes against apoptosis induced by bile acids, Fas ligand, and TNF-alpha. The role of cAMP-GEF and PKA on activation of Akt, a kinase implicated in cAMP survival signaling, was investigated. Inhibition of PKA with RP-cAMP had no effect on cAMP-mediated Akt phosphorylation, whereas CPT-2-Me-cAMP, which did not activate PKA, induced phosphatidylinositol 3-kinase (PI3-kinase)-dependent activation of Akt. Pretreatment of hepatocytes with the PI3-kinase inhibitor, Ly-294002, prevented CPT-2-Me-cAMP's protective effect against bile acid and Fas ligand, but not TNF-alpha-mediated apoptosis. Glucagon, CPT-cAMP, and CPT-2-Me-cAMP all activated Rap 1, a downstream effector of cAMP-GEF. These results suggest that a PKA-independent cAMP/cAMP-GEF/Rap pathway exists in hepatocytes and that activation of cAMP-GEFs promotes Akt phosphorylation and hepatocyte survival. Thus a cAMP/cAMP-GEF/Rap/PI3-kinase/Akt signaling pathway may confer protection against bile acid- and Fas-induced apoptosis in hepatocytes.
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PMID:Activation of cAMP-guanine exchange factor confers PKA-independent protection from hepatocyte apoptosis. 1504 79

We have examined the expression and activity of inducible nitric oxide synthase (iNOS) and the activity of neuronal constitutive NOS (ncNOS) in isolated rat pancreatic islets, stimulated by a "hyperglycaemic" concentration of glucose, and whether the NOS activities could be modulated by activation of the cyclic AMP/protein kinase A (cyclic AMP/PKA) system in relation to the insulin secretory process. Here, we show that glucose stimulation (20 mmol/l) induces iNOS and increases ncNOS activity. No iNOS is detectable at basal glucose levels (3.3 mmol/l). The addition of glucagon-like-peptide 1 (GLP-1) or dibutyryl-cAMP to islets incubated with 20 mmol/l glucose results in a marked suppression of iNOS expression and activity, a reduction in ncNOS activity and increased insulin release. The GLP-1-induced suppression of glucose-stimulated iNOS activity and expression and its stimulation of insulin release is, at least in part, PKA dependent, since the PKA inhibitor H-89 reverses the effects of GLP-1. These observations have been confirmed by confocal microscopy showing the glucose-stimulated expression of iNOS, its suppression by GLP-1 and its reversion by H-89 in beta-cells. We have also found that the NO scavenger cPTIO and the NOS inhibitor L-NAME potentiate the insulin response to glucose, again suggesting that NO is a negative modulator of glucose-stimulated insulin release. We conclude that the induction of iNOS and the increase in ncNOS activity caused by glucose in rat islets is suppressed by the cyclic AMP/PKA system. The inhibition of iNOS expression by the GLP-1/cyclic AMP/PKA pathway might possibly be of therapeutic potential in NO-mediated beta-cell dysfunction and destruction.
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PMID:Glucose stimulates the expression and activities of nitric oxide synthases in incubated rat islets: an effect counteracted by GLP-1 through the cyclic AMP/PKA pathway. 1555 23


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