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)

The hormone leptin is expressed and secreted by the adipose tissue and impacts on the central nervous system. Leptin is involved in the regulation of energy balance, satiety, and body composition. The lack of active leptin results in obesity, high food intake, hyperglycemia, and hyperinsulinemia. We present data supporting effects of leptin on the endocrine pancreas. We found the leptin receptor to be expressed in insulin- and glucagon-secretin cells derived from mouse, hamster, and rat pancreas. In the isolated perfused rat pancreas leptin is a potent inhibitor of basal and glucose-induced insulin secretion, especially during the first phase of the insulin response. At isolated mouse islets and insulin-secreting INS-1 cells leptin reduced promptly and persistently the intracellular Ca2+ levels. Cytoplasmic Ca2+ oscillation amplitude was decreased and the oscillation frequency increased. These findings suggest functional active receptors for leptin on insulin-secreting B-cells. Therefore, leptin is a metabolic hormone and not only a signal to the brain indicating filled fat stores. Our data suggest that leptin is also a signal back to the endocrine pancreas that no more insulin is required to replenish fat stores. Thus, an "adipo-insular axis" operating with two arms exists: insulin and glucagon are signals to the adipocyte. This releases leptin, which could be the mediator of the respective feedback to the pancreas. A defective leptin suppression of insulin secretion could contribute to hyperinsulinemia and disturbances of glucose metabolism.
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PMID:Leptin: a potent inhibitor of insulin secretion. 939 72

Proglucagon is processed differentially in pancreatic alpha-cells and intestinal endocrine L cells to release either glucagon or glucagon-like peptide-1-(7-36amide) (tGLP-1), two peptide hormones with opposing biological actions. Previous studies have demonstrated that the prohormone convertase PC2 is responsible for the processing of proglucagon to glucagon, and have suggested that the related endoprotease PC3 is involved in the formation of tGLP-1. To understand better the biosynthetic pathway of tGLP-1, proglucagon processing was studied in the mouse pituitary cell line AtT-20, a cell line that mimics the intestinal pathway of proglucagon processing and in the rat insulinoma cell line INS-1. In both of these cell lines, proglucagon was initially cleaved to glicentin and the major proglucagon fragment (MPGF) at the interdomain site Lys70-Arg71. In both cell lines, MPGF was cleaved successively at the monobasic site Arg77 and then at the dibasic site Arg109-Arg110, thus releasing tGLP-1, the cleavages being less extensive in INS-1 cells. Glicentin was completely processed to glucagon in INS-1 cells, but was partially converted to oxyntomodulin and very low levels of glucagon in AtT-20 cells in the face of generation of tGLP-1. Adenovirus-mediated co-expression of PC3 and proglucagon in GH4C1 cells (normally expressing no PC2 or PC3) resulted in the formation of tGLP-1, glicentin, and oxyntomodulin, but no glucagon. When expressed in alphaTC1-6 (transformed pancreatic alpha-cells) or in rat primary pancreatic alpha-cells in culture, PC3 converted MPGF to tGLP-1. Finally, GLP-1-(1-37) was cleaved to tGLP-1 in vitro by purified recombinant PC3. Taken together, these results indicate that PC3 has the same specificity as the convertase that is responsible for the processing of proglucagon to tGLP-1, glicentin and oxyntomodulin in the intestinal L cell, and it is concluded that this enzyme is thus able to act alone in this processing pathway.
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PMID:Role of the prohormone convertase PC3 in the processing of proglucagon to glucagon-like peptide 1. 940 57

Leptin controls feeding behavior and insulin secretion from pancreatic beta-cells. Insulin stimulates the production of leptin, thereby establishing an adipoinsular axis. Earlier we identified leptin receptors on pancreatic beta-cells and showed leptin-mediated inhibition of insulin secretion by activation of ATP-sensitive potassium channels. Here we examine transcriptional effects of leptin on the promoter of the rat insulin I gene in rodent beta-cells. A fall in levels of preproinsulin mRNA is detected in vivo in islets of ob/ob mice 24 h after a single injection of leptin, in isolated ob/ob islets treated with leptin in vitro and in the beta-cell line INS-1 on leptin exposure when preproinsulin mRNA expression is stimulated by 25 mM glucose or 10 nM glucagon-like peptide 1. Under these conditions, transcriptional activity of -410 bp of the rat insulin I promoter is inhibited by leptin, whereas transactivation of a 5'-deleted promoter (-307 bp) is not. The -307 sequence contains the known glucose-responsive control elements (E2:A3/4). Constitutive activation of ATP-sensitive potassium channels by diazoxide does not alter leptin inhibition of preproinsulin mRNA levels. Distinct protein-DNA complexes appear on the rat insulin I promoter sequences located between -307 and -410 with nuclear extracts from ob/ob islets in response to leptin, including a signal transducer and activator of transcription (STAT)5b binding site. These results indicate that leptin inhibits transcription of the preproinsulin gene by altering transcription factor binding to sequences upstream from the elements (307 bp) that confer glucose responsivity to the rat insulin I gene promoter. Thus leptin exerts inhibitory effects on both insulin secretion and insulin gene expression in pancreatic beta-cells, but by different cellular mechanisms.
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PMID:Leptin inhibits insulin gene transcription and reverses hyperinsulinemia in leptin-deficient ob/ob mice. 989 92

Activation of adenylyl cyclase by Gs-coupled receptors for insulinotropic hormones such as glucagon-like peptide-1 and pituitary adenylate cyclase-activating polypeptide plays a critical role in stimulating glucose-induced insulin secretion. Despite this important role of insulinotropic hormones in the regulation of insulin secretion, little is known about which of the multiple subtypes of adenylyl cyclase are expressed in beta-cells. Here we report the use of PCR primers designed to amplify all subtypes of adenylyl cyclase from cDNA prepared from human and rat islets and from insulin-secreting beta-cell lines. PCR products were cloned and sequenced to identify the subtypes of adenylyl cyclase amplified. Adenylyl cyclase types V and VI, known to couple to Galphas and Gbetagamma in the cAMP signaling pathway, account for all subtypes identified in human islets and INS-1 cells and the majority of subtypes in rat islets and HIT-T15 cells. These findings indicate that pancreatic beta-cells are particularly well suited to transmit signals via Gs-coupled receptors such as that for glucagon-like peptide-1.
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PMID:Expression of adenylyl cyclase subtypes in pancreatic beta-cells. 992 Aug 5

Glucagon-like peptide 1 (7-36)amide (GLP-1) is an insulinotropic intestinal peptide hormone with a potential role as antidiabetogenic therapeutic agent. It mediates a potentiation of glucose-induced insulin secretion, by activation of adenylate cyclase and subsequent elevation of cytosolic free calcium, [Ca2+]cyt. We investigated the role of protein kinase A (PKA) in GLP-1 signal transduction, using isolated mouse islets as well as the differentiated beta-cell line INS-1. Two specific inhibitors of PKA, (Rp)-adenosine cyclic 3',5'-phosporothioate (Rp-cAMPS, up to 3 mM) and KT5720 (up to 10 microM), did not inhibit the GLP-1-induced [Ca2+]cyt elevation. Another PKA inhibitor, H-89, reduced the [Ca2+]cyt elevation only when applied at high concentrations (10-40 microM), higher than sufficient for PKA inhibition in many cell types. Furthermore, at these concentrations, H-89 also inhibited presumably PKA-independent processes such as glucose-induced [Ca2+]cyt elevations and intracellular calcium storage. This suggests a PKA-independent action of H-89. Similarly to H-89, the potent but unselective protein kinase inhibitor staurosporine inhibited the GLP-1-induced [Ca2+]cyt elevation only at high concentrations, at which it also inhibited glucose-induced [Ca2+]cyt elevations. The same observations as with GLP-1 were made when adenylate cyclase was stimulated with forskolin, for selective examination of signal transduction downstream of receptor and G protein. Our results suggest that the GLP-1-induced [Ca2+]cyt elevation is mediated independently of PKA and thus belongs to the yet-little-characterized ensemble of effects that are mediated by binding of cAMP to other target proteins.
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PMID:Glucagon-like peptide 1 elevates cytosolic calcium in pancreatic beta-cells independently of protein kinase A. 1046 60

The effect of leptin on insulin secretion is controversial due to conflicting results in the literature. In the present study, we incubated insulin-producing rat insulinoma INS-1 cells for 60 min and examined the effects of recombinant murine leptin (20 nmol/l). We found that leptin (0.1-100 nmol/l) did not affect the insulin response to glucose (1-20 mmol/l). However, when cells were incubated with agents that increase the intracellular content of cAMP, i.e., glucagon-like peptide-1 (100 nmol/l), pituitary adenylate cyclase activating polypeptide (100 nmol/l), forskolin (2.5 micromol/l), dibutyryl-cAMP (1 mmol/l), or 3-isobutyl-1-methylxanthine (100 micromol/l), leptin significantly reduced insulin secretion (by 34-58%, P < 0.05-0.001). In contrast, when insulin secretion was stimulated by the cholinergic agonist carbachol (100 micromol/l) or the phorbol ester 12-O-tetradecanoylphorbol 13-acetate (1 micromol/l), both of which activate protein kinase C, leptin was without effect. We conclude that leptin inhibits insulin secretion from INS-1 cells under conditions in which intracellular cAMP is increased. This suggests that the cAMP-protein kinase A signal transduction pathway is a target for leptin to inhibit insulin secretion in insulin-producing cells.
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PMID:Leptin inhibits insulin secretion induced by cellular cAMP in a pancreatic B cell line (INS-1 cells). 1051 32

Glucose controls long-term processes in the pancreatic beta-cell such as metabolic enzymes gene expression, cell growth, and apoptosis. Such control is likely mediated via the expression of immediate-early response genes since several of these genes including c-fos are strongly induced by glucose in the beta-cell line INS-1, provided costimulation with cAMP-raising glucoincretin hormones. This study addresses the mechanism of c-fos gene activation by glucose. Glucose in the presence of chlorophenylthio-cAMP generated a low threefold induction of the c-fos/basic luciferase reporter gene, which includes only the c-fos promoter. In contrast, the c-fos/intron construct containing the first intron in addition to promoter elements showed a pronounced 16-fold induction, comparable to the increased c-fos mRNA accumulation. Similar observations were made with glucose in combination with the glucoincretins glucagon-like peptide 1, glucose-dependent insulinotropic polypeptide, and pituitary adenylyl cyclase-activating peptide 38. Deletion of a 119 bp region in intron 1 that includes a transcriptional arrest site did not affect the inductive process. In contrast, a 534 bp deletion comprising a major part of the intron reduced the induction by 75%. At the promoter level, mutating the cAMP response element reduced by more than 60% the transcriptional activation whereas mutating the serum response element had no effect. Inhibitors of protein kinase A and Ca(2+)/calmodulin-dependent protein kinases each reduced by 50% the reporter gene activation and together fully prevented the glucose-glucoincretin effect. In conclusion, the strong induction of c-fos by glucose and glucoincretins results from Ca(2+) and cAMP signaling pathways addressing both the CRE in the promoter and essential response element(s) in the first intron that are unrelated to the transcription arrest site.
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PMID:Essentiality of intron control in the induction of c-fos by glucose and glucoincretin peptides in INS-1 beta-cells. 1062 87

The biochemical mechanisms involved in regulation of insulin secretion are not completely understood. The rat INS-1 cell line has been used to gain insight in this area because it secretes insulin in response to glucose concentrations in the physiological range. However, the magnitude of the response is far less than that seen in freshly isolated rat islets. In the current study, we have stably transfected INS-1 cells with a plasmid containing the human proinsulin gene. After antibiotic selection and clonal expansion, 67% of the resultant clones were found to be poorly responsive to glucose in terms of insulin secretion (< or =2-fold stimulation by 15 mmol/l compared with 3 mmol/l glucose), 17% of the clones were moderately responsive (2- to 5-fold stimulation), and 16% were strongly responsive (5- to 13-fold stimulation). The differences in responsiveness could not be ascribed to differences in insulin content. Detailed analysis of one of the strongly responsive lines (832/13) revealed that its potent response to glucose (average of 10-fold) was stable over 66 population doublings (approximately 7.5 months of tissue culture) with half-maximal stimulation at 6 mmol/l glucose. Furthermore, in the presence of 15 mmol/l glucose, insulin secretion was potentiated significantly by 100 pmol/l isobutylmethylxanthine (320%), 1 mmol/l oleate/palmitate (77%), and 50 nmol/l glucagon-like peptide 1 (60%), whereas carbachol had no effect. Glucose-stimulated insulin secretion was also potentiated by the sulfonylurea tolbutamide (threefold at 3 mmol/l glucose and 50% at 15 mmol/l glucose) and was abolished by diazoxide, which demonstrates the operation of the ATP-sensitive K+ channel (K(ATP)) in 832/13 cells. Moreover, when the K(ATP) channel was bypassed by incubation of cells in depolarizing K+ (35 mmol/l), insulin secretion was more effectively stimulated by glucose in 832/13 cells than in parental INS-1 cells, which demonstrates the presence of a K(ATP) channel-independent pathway of glucose sensing. We conclude that clonal selection of INS-1 cells allows isolation of cell lines that exhibit markedly enhanced and stable responsiveness to glucose and several of its known potentiators. These lines may be attractive new vehicles for studies of beta-cell function.
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PMID:Isolation of INS-1-derived cell lines with robust ATP-sensitive K+ channel-dependent and -independent glucose-stimulated insulin secretion. 1086 64

Hepatocyte nuclear factors 3 (HNF-3 alpha, -3 beta and -3 gamma) belong to an evolutionarily conserved family of transcription factors that are critical for diverse biological processes such as development, differentiation and metabolism. Gene expression studies have shown that HNF3 proteins are critical regulators of the early-onset type 2 diabetes genes HNF-1 alpha, HNF-4 alpha and IPF-1/PDX-1 (MODY3, 1 and 4, respectively) and of glucagon transcription and pancreatic alpha-cell function. In this study, we investigated whether genetic variation in the genes encoding HNF-3 alpha, HNF-3 beta and HNF-3 gamma predisposes humans to hyperglycemic or hypoglycemic syndromes. In addition, we report the cloning and partial nucleotide sequence of the human HNF-3 alpha, -3 beta and -3 gamma genes. Mutation screening included 96 subjects with type 2 diabetes mellitus, as well as one family with persistent neonatal hypoglycemia. No functional mutations were detected in the coding sequences of the three HNF-3 genes. Our results suggest that mutations in HNF-3 genes are not a common cause of type 2 diabetes mellitus. The data provided will facilitate genetic studies in other populations and will advance our understanding of the role HNF-3 plays in the development of diabetes mellitus and other metabolic disorders of glucose homeostasis.
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PMID:The human HNF-3 genes: cloning, partial sequence and mutation screening in patients with impaired glucose homeostasis. 1089 56

IA-2, a member of the protein tyrosine phosphatase family, represents a major target autoantigen in type 1 diabetes. To study the regulation of IA-2 gene expression, we used INS-1 insulinoma cells to analyze beta-cell signal transduction pathways as well as the effect of metabolic and hormonal factors involved in the regulation of the insulin secretory pathway. Quantitative competitive reverse transcriptase-polymerase chain reaction revealed that an increase of cellular cAMP mediated by forskolin (10 micromol/l, 24 h) or 3-isobutyl-1-methylxanthine (100 micromol/l, 24 h) induced maximal stimulation of IA-2 mRNA levels (451 +/- 85 and 338 +/- 86% compared with basal conditions; P < 0.001). In contrast, activation of protein kinase C (PKC) by short-term treatment with phorbol 12-myristate 13-acetate (PMA) (1 micromol/l, 6 h) did not alter IA-2 expression, whereas depletion of PKC by prolonged culturing (24 h) exerted a significant inhibition (57 +/- 24%; P < 0.05). cAMP-dependent upregulation was confirmed by the findings that glucagon (10 micromol/l, 24-48 h) increased levels of IA-2 mRNA (190 +/- 35%; P < 0.05), whereas short-term incubation with high glucose concentration showed no effect. However, prolonged incubation in high glucose (21 mmol/l) induced a time- and dose-dependent increase of IA-2 mRNA expression, reaching maximal values after 144 h (285 +/- 68%; P < 0.05). These studies demonstrate that stimuli of insulin secretion that operate by activation of adenylate cyclase generating cAMP significantly increase IA-2 gene expression. In contrast, activation of PKC by high glucose concentration or PMA exerted no effect, suggesting that IA-2 gene expression is not simply coupled to insulin secretion, but may be involved in the fine regulation of beta-cell function. These findings may be important to clarify the function of IA-2 in beta-cells and elucidate mechanisms involved in the induction of autoimmunity to IA-2.
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PMID:Regulation of the diabetes-associated autoantigen IA-2 in INS-1 pancreatic beta-cells. 1090 70


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