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Jiang FX, Morahan G. Insulin-secreting β cells require a post-genomic concept. World J Diabetes 2016; 7:198-208. [PMID: 27226815 PMCID: PMC4873311 DOI: 10.4239/wjd.v7.i10.198] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/28/2015] [Accepted: 03/18/2016] [Indexed: 02/05/2023] Open
Abstract
Pancreatic insulin-secreting β cells are essential in maintaining normal glucose homeostasis accomplished by highly specialized transcription of insulin gene, of which occupies up to 40% their transcriptome. Deficiency of these cells causes diabetes mellitus, a global public health problem. Although tremendous endeavors have been made to generate insulin-secreting cells from human pluripotent stem cells (i.e., primitive cells capable of giving rise to all cell types in the body), a regenerative therapy to diabetes has not yet been established. Furthermore, the nomenclature of β cells has become inconsistent, confusing and controversial due to the lack of standardized positive controls of developmental stage-matched in vivo cells. In order to minimize this negative impact and facilitate critical research in this field, a post-genomic concept of pancreatic β cells might be helpful. In this review article, we will briefly describe how β cells were discovered and islet lineage is developed that may help understand the cause of nomenclatural controversy, suggest a post-genomic definition and finally provide a conclusive remark on future research of this pivotal cell.
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2
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Kaneto H, Matsuoka TA. Role of pancreatic transcription factors in maintenance of mature β-cell function. Int J Mol Sci 2015; 16:6281-97. [PMID: 25794287 PMCID: PMC4394532 DOI: 10.3390/ijms16036281] [Citation(s) in RCA: 50] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/16/2014] [Revised: 02/10/2015] [Accepted: 02/16/2015] [Indexed: 12/12/2022] Open
Abstract
A variety of pancreatic transcription factors including PDX-1 and MafA play crucial roles in the pancreas and function for the maintenance of mature β-cell function. However, when β-cells are chronically exposed to hyperglycemia, expression and/or activities of such transcription factors are reduced, which leads to deterioration of β-cell function. These phenomena are well known as β-cell glucose toxicity in practical medicine as well as in the islet biology research area. Here we describe the possible mechanism for β-cell glucose toxicity found in type 2 diabetes. It is likely that reduced expression levels of PDX-1 and MafA lead to suppression of insulin biosynthesis and secretion. In addition, expression levels of incretin receptors (GLP-1 and GIP receptors) in β-cells are decreased, which likely contributes to the impaired incretin effects found in diabetes. Taken together, down-regulation of insulin gene transcription factors and incretin receptors explains, at least in part, the molecular mechanism for β-cell glucose toxicity.
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Affiliation(s)
- Hideaki Kaneto
- Department of Diabetes, Endocrinology and Metabolism, Kawasaki Medical School, 577, Matsushima, Kurashiki 701-0192, Japan.
| | - Taka-aki Matsuoka
- Department of Metabolic Medicine, Osaka University Graduate School of Medicine, Osaka 565-0871, Japan.
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3
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Abstract
Type 2 diabetes is characterized by pancreatic β-cell dysfunction and insulin resistance, and the number of patients has markedly increased worldwide. In the diabetic state, hyperglycemia per se and subsequent induction of oxidative stress decrease insulin biosynthesis and secretion, leading to the aggravation of Type 2 diabetes. In addition, there is substantial reduction in expression and/or activities of several insulin gene transcription factors. This process is known as β-cell glucose toxicity, which is often observed under diabetic conditions. Taken together, it is likely that oxidative stress explains, at least in part, the molecular mechanism for β-cell glucose toxicity, which is often observed in Type 2 diabetes.
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4
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Han SI, Yasuda K, Kataoka K. ATF2 interacts with beta-cell-enriched transcription factors, MafA, Pdx1, and beta2, and activates insulin gene transcription. J Biol Chem 2011; 286:10449-56. [PMID: 21278380 DOI: 10.1074/jbc.m110.209510] [Citation(s) in RCA: 26] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
Pancreatic β-cell-restricted expression of insulin is established through several critical cis-regulatory elements located in the insulin gene promoter region. The principal cis elements are A-boxes, E1, and C1/RIPE3b. The β-cell-enriched transcription factors Pdx1 and Beta2 bind to the A-boxes and E1 element, respectively. A β-cell-specific trans-acting factor binding to C1/RIPE3b (termed RIPE3b1 activator) was detected by electrophoretic mobility shift assay and has been identified as MafA, a member of the Maf family of basic leucine zipper (bZip) proteins. Here, ATF2, a member of the ATF/CREB family of basic leucine zipper proteins, was identified as a component of the RIPE3b1 activator. ATF2 alone was unable to bind to the C1/RIPE3b element but acquired binding capacity upon complex formation with MafA. ATF2 also interacted with Pdx1 and Beta2, and co-expression of ATF2, MafA, Pdx1, and Beta2 resulted in a synergistic activation of the insulin promoter. Immunohistochemical analysis of mouse pancreas tissue sections showed that ATF2 is enriched in islet endocrine cells, including β-cells. RNAi-mediated knockdown of MafA or ATF2 in the MIN6 β-cell line resulted in a significant decrease in endogenous levels of insulin mRNA. These data indicate that ATF2 is an essential component of the positive regulators of the insulin gene expression.
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Affiliation(s)
- Song-iee Han
- Laboratory of Molecular and Developmental Biology, Graduate School of Biological Science, Nara Institute of Science and Technology, Ikoma 630-0192, Japan
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5
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Wong WPS, Tiano JP, Liu S, Hewitt SC, Le May C, Dalle S, Katzenellenbogen JA, Katzenellenbogen BS, Korach KS, Mauvais-Jarvis F. Extranuclear estrogen receptor-alpha stimulates NeuroD1 binding to the insulin promoter and favors insulin synthesis. Proc Natl Acad Sci U S A 2010; 107:13057-62. [PMID: 20616010 PMCID: PMC2919966 DOI: 10.1073/pnas.0914501107] [Citation(s) in RCA: 115] [Impact Index Per Article: 7.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/18/2022] Open
Abstract
Estrogen receptors (ERs) protect pancreatic islet survival in mice through rapid extranuclear actions. ERalpha also enhances insulin synthesis in cultured islets. Whether ERalpha stimulates insulin synthesis in vivo and, if so, through which mechanism(s) remain largely unknown. To address these issues, we generated a pancreas-specific ERalpha knockout mouse (PERalpha KO(-/-)) using the Cre-loxP strategy and used a combination of genetic and pharmacologic tools in cultured islets and beta cells. Whereas 17beta-estradiol (E2) treatment up-regulates pancreatic insulin gene and protein content in control ERalpha lox/lox mice, these E2 effects are abolished in PERalpha KO(-/-) mice. We find that E2-activated ERalpha increases insulin synthesis by enhancing glucose stimulation of the insulin promoter activity. Using a knock-in mouse with a mutated ERalpha eliminating binding to the estrogen response elements (EREs), we show that E2 stimulation of insulin synthesis is independent of the ERE. We find that the extranuclear ERalpha interacts with the tyrosine kinase Src, which activates extracellular signal-regulated kinases(1/2), to increase nuclear localization and binding to the insulin promoter of the transcription factor NeuroD1. This study supports the importance of ERalpha in beta cells as a regulator of insulin synthesis in vivo.
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Affiliation(s)
| | - Joseph P. Tiano
- Division of Endocrinology, Metabolism and Molecular Medicine and
| | - Suhuan Liu
- Division of Endocrinology, Metabolism and Molecular Medicine and
- Comprehensive Center on Obesity, Department of Medicine, Northwestern University School of Medicine, Chicago, IL 60611
| | - Sylvia C. Hewitt
- National Institute of Environmental Health Sciences, Research Triangle Park, NC 27709
| | - Cedric Le May
- Division of Endocrinology, Metabolism and Molecular Medicine and
| | - Stéphane Dalle
- Institut National de la Santé et de la Recherche Médicale U661, Institut de Génomique Fonctionnelle, Montpellier 34094, France; and
| | | | | | - Kenneth S. Korach
- National Institute of Environmental Health Sciences, Research Triangle Park, NC 27709
| | - Franck Mauvais-Jarvis
- Division of Endocrinology, Metabolism and Molecular Medicine and
- Comprehensive Center on Obesity, Department of Medicine, Northwestern University School of Medicine, Chicago, IL 60611
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6
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Wilson LM, Wong SHK, Yu N, Geras-Raaka E, Raaka BM, Gershengorn MC. Insulin but not glucagon gene is silenced in human pancreas-derived mesenchymal stem cells. Stem Cells 2010; 27:2703-11. [PMID: 19785038 DOI: 10.1002/stem.229] [Citation(s) in RCA: 14] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/09/2023]
Abstract
We previously characterized human islet-derived precursor cells (hIPCs) as a specific type of mesenchymal stem cell capable of differentiating to insulin (INS)- and glucagon (GCG)-expressing cells. However, during proliferative expansion, INS transcript becomes undetectable and then cannot be induced, a phenomenon consistent with silencing of the INS gene. We explored this possibility by determining whether ectopic expression of transcription factors known to induce transcription of this gene in beta cells, pancreatic and duodenal homeobox factor 1 (Pdx1), V-maf musculoaponeurotic fibrosarcoma oncogene homolog A (Mafa), and neurogenic differentiation 1 (Neurod1), would activate INS gene expression in long-term hIPC cultures. Coexpression of all three transcription factors had little effect on INS mRNA levels but unexpectedly increased GCG mRNA at least 100,000-fold. In contrast to the endogenous promoter, an exogenous rat INS promoter was activated by expression of Pdx1 and Mafa in hIPCs. Chromatin immunoprecipitation (ChIP) assays using antibodies directed at posttranslationally modified histones show that regions of the INS and GCG genes have similar levels of activation-associated modifications but the INS gene has higher levels of repression-associated modifications. Furthermore, the INS gene was found to be less accessible to micrococcal nuclease digestion than the GCG gene. Lastly, ChIP assays show that exogenously expressed Pdx1 and Mafa bind at very low levels to the INS promoter and at 20- to 25-fold higher levels to the GCG promoter in hIPCs. We conclude that the INS gene in hIPCs is modified epigenetically ("silenced") so that it is resistant to activation by transcription factors.
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Affiliation(s)
- Leah M Wilson
- Clinical Endocrinology Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland 20892-8029, USA
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7
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Kaneto H, Katakami N, Matsuhisa M, Matsuoka TA. Role of reactive oxygen species in the progression of type 2 diabetes and atherosclerosis. Mediators Inflamm 2010; 2010:453892. [PMID: 20182627 PMCID: PMC2825658 DOI: 10.1155/2010/453892] [Citation(s) in RCA: 362] [Impact Index Per Article: 24.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/28/2009] [Accepted: 11/13/2009] [Indexed: 02/06/2023] Open
Abstract
Type 2 diabetes is the most prevalent and serious metabolic disease all over the world, and its hallmarks are pancreatic beta-cell dysfunction and insulin resistance. Under diabetic conditions, chronic hyperglycemia and subsequent augmentation of reactive oxygen species (ROS) deteriorate beta-cell function and increase insulin resistance which leads to the aggravation of type 2 diabetes. In addition, chronic hyperglycemia and ROS are also involved in the development of atherosclerosis which is often observed under diabetic conditions. Taken together, it is likely that ROS play an important role in the development of type 2 diabetes and atherosclerosis.
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Affiliation(s)
- Hideaki Kaneto
- Department of Metabolic Medicine, Graduate School of Medicine, Osaka University, 2-2 Yamadaoka, Suita, Osaka 565-0871, Japan.
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8
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Harmon JS, Bogdani M, Parazzoli SD, Mak SSM, Oseid EA, Berghmans M, Leboeuf RC, Robertson RP. beta-Cell-specific overexpression of glutathione peroxidase preserves intranuclear MafA and reverses diabetes in db/db mice. Endocrinology 2009; 150:4855-62. [PMID: 19819955 PMCID: PMC2775976 DOI: 10.1210/en.2009-0708] [Citation(s) in RCA: 148] [Impact Index Per Article: 9.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 11/19/2022]
Abstract
Chronic hyperglycemia causes oxidative stress, which contributes to damage in various tissues and cells, including pancreatic beta-cells. The expression levels of antioxidant enzymes in the islet are low compared with other tissues, rendering the beta-cell more susceptible to damage caused by hyperglycemia. The aim of this study was to investigate whether increasing levels of endogenous glutathione peroxidase-1 (GPx-1), specifically in beta-cells, can protect them against the adverse effects of chronic hyperglycemia and assess mechanisms that may be involved. C57BLKS/J mice overexpressing the antioxidant enzyme GPx-1 only in pancreatic beta-cells were generated. The biological effectiveness of the overexpressed GPx-1 transgene was documented when beta-cells of transgenic mice were protected from streptozotocin. The transgene was then introgressed into the beta-cells of db/db mice. Without use of hypoglycemic agents, hyperglycemia in db/db-GPx(+) mice was initially ameliorated compared with db/db-GPx(-) animals and then substantially reversed by 20 wk of age. beta-Cell volume and insulin granulation and immunostaining were greater in db/db-GPx(+) animals compared with db/db-GPx(-) animals. Importantly, the loss of intranuclear musculoaponeurotic fibrosarcoma oncogene homolog A (MafA) that was observed in nontransgenic db/db mice was prevented by GPx-1 overexpression, making this a likely mechanism for the improved glycemic control. These studies demonstrate that enhancement of intrinsic antioxidant defenses of the beta-cell protects it against deterioration during hyperglycemia.
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Affiliation(s)
- Jamie S Harmon
- Pacific Northwest Diabetes Research Institute, 720 Broadway, Seattle, Washington 98122, USA
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9
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Vanderford NL, Cantrell JEL, Popa GJ, Ozcan S. Multiple kinases regulate mafA expression in the pancreatic beta cell line MIN6. Arch Biochem Biophys 2008; 480:138-42. [PMID: 18948074 DOI: 10.1016/j.abb.2008.10.001] [Citation(s) in RCA: 13] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/20/2008] [Revised: 10/03/2008] [Accepted: 10/04/2008] [Indexed: 01/13/2023]
Abstract
MafA is a basic leucine zipper transcription factor expressed within the beta cells of the pancreas and is required to maintain normal glucose homeostasis as it is involved in various aspects of beta cell biology. MafA protein levels are known to increase in response to high glucose through mechanisms that have yet to be fully characterized. We investigated whether discrete intracellular signaling events control mafA expression. We found that the general kinase inhibitor staurosporine induces mafA expression without altering the stability of the protein. Inhibition of the MAP-kinase JNK mimics the effects of staurosporine on the expression of mafA. Calmodulin kinase and calcium signaling are also important in stimulating mafA expression by high glucose. However, staurosporine, JNK, and calmodulin kinase have different effects on the induction of insulin expression. These data reveal that MafA levels are tightly controlled by the coordinated action of multiple kinase pathways.
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Affiliation(s)
- Nathan L Vanderford
- Department of Molecular and Cellular Biochemistry, University of Kentucky, College of Medicine, 741 South Limestone Street, Lexington, KY 40536, USA
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10
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Han SI, Aramata S, Yasuda K, Kataoka K. MafA stability in pancreatic beta cells is regulated by glucose and is dependent on its constitutive phosphorylation at multiple sites by glycogen synthase kinase 3. Mol Cell Biol 2007; 27:6593-605. [PMID: 17682063 PMCID: PMC2099218 DOI: 10.1128/mcb.01573-06] [Citation(s) in RCA: 61] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Regulation of insulin gene expression by glucose in pancreatic beta cells is largely dependent on a cis-regulatory element, termed RIPE3b/C1, in the insulin gene promoter. MafA, a member of the Maf family of basic leucine zipper (bZip) proteins, is a beta-cell-specific transcriptional activator that binds to the C1 element. Based on increased C1-binding activity, MafA protein levels appear to be up-regulated in response to glucose, but the underlying molecular mechanism for this is not well understood. In this study, we show evidence supporting that the amino-terminal region of MafA is phosphorylated at multiple sites by glycogen synthase kinase 3 (GSK3) in beta cells. Mutational analysis of MafA and pharmacological inhibition of GSK3 in MIN6 beta cells strongly suggest that the rate of MafA protein degradation is regulated by glucose, that MafA is constitutively phosphorylated by GSK3, and that phosphorylation is a prerequisite for rapid degradation of MafA under low-glucose conditions. Our data suggest a new glucose-sensing signaling pathway in islet beta cells that regulates insulin gene expression through the regulation of MafA protein stability.
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Affiliation(s)
- Song-Iee Han
- Laboratory of Molecular and Developmental Biology, Graduate School of Biological Science, Nara Institute of Science and Technology, Ikoma, Japan
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11
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Ren J, Jin P, Wang E, Liu E, Harlan DM, Li X, Stroncek DF. Pancreatic islet cell therapy for type I diabetes: understanding the effects of glucose stimulation on islets in order to produce better islets for transplantation. J Transl Med 2007; 5:1. [PMID: 17201925 PMCID: PMC1769476 DOI: 10.1186/1479-5876-5-1] [Citation(s) in RCA: 31] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/05/2006] [Accepted: 01/03/2007] [Indexed: 01/28/2023] Open
Abstract
While insulin replacement remains the cornerstone treatment for type I diabetes mellitus (T1DM), the transplantation of pancreatic islets of Langerhans has the potential to become an important alternative. And yet, islet transplant therapy is limited by several factors, including far too few donor pancreases. Attempts to expand mature islets or to produce islets from stem cells are far from clinical application. The production and expansion of the insulin-producing cells within the islet (so called beta cells), or even creating cells that secrete insulin under appropriate physiological control, has proven difficult. The difficulty is explained, in part, because insulin synthesis and release is complex, unique, and not entirely characterized. Understanding beta-cell function at the molecular level will likely facilitate the development of techniques to manufacture beta-cells from stem cells. We will review islet transplantation, as well as the mechanisms underlying insulin transcription, translation and glucose stimulated insulin release.
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Affiliation(s)
- Jiaqiang Ren
- Department of Transfusion Medicine, Clinical Center, National Institutes of Health, Bethesda, MD, 20892, USA
| | - Ping Jin
- Department of Transfusion Medicine, Clinical Center, National Institutes of Health, Bethesda, MD, 20892, USA
| | - Ena Wang
- Department of Transfusion Medicine, Clinical Center, National Institutes of Health, Bethesda, MD, 20892, USA
| | - Eric Liu
- National Institute of Diabetes, Digestive and Kidney Disease, National Institutes of Health, Bethesda, MD, 20892, USA
| | - David M Harlan
- National Institute of Diabetes, Digestive and Kidney Disease, National Institutes of Health, Bethesda, MD, 20892, USA
| | - Xin Li
- Department of Transfusion Medicine, Clinical Center, National Institutes of Health, Bethesda, MD, 20892, USA
| | - David F Stroncek
- Department of Transfusion Medicine, Clinical Center, National Institutes of Health, Bethesda, MD, 20892, USA
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12
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Nishimura W, Salameh T, Kondo T, Sharma A. Regulation of insulin gene expression by overlapping DNA-binding elements. Biochem J 2006; 392:181-9. [PMID: 16050808 PMCID: PMC1317677 DOI: 10.1042/bj20050970] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/17/2022]
Abstract
The transcription factor MafA/RIPE3b1 is an important regulator of insulin gene expression. MafA binds to the insulin enhancer element RIPE3b (C1-A2), now designated as insulin MARE (Maf response element). The insulin MARE element shares an overlapping DNA-binding region with another insulin enhancer element A2. A2.2, a beta-cell-specific activator, like the MARE-binding factor MafA, binds to the overlapping A2 element. Our previous results demonstrated that two nucleotides in the overlapping region are required for the binding of both factors. Surprisingly, instead of interfering with each other's binding activity, the MafA and the A2-binding factors co-operatively activated insulin gene expression. To understand the molecular mechanisms responsible for this functional co-operation, we have determined the nucleotides essential for the binding of the A2.2 factor. Using this information, we have constructed non-overlapping DNA-binding elements and their derivatives, and subsequently analysed the effect of these modifications on insulin gene expression. Our results demonstrate that the overlapping binding site is essential for maximal insulin gene expression. Furthermore, the overlapping organization is critical for MafA-mediated transcriptional activation, but has a minor effect on the activity of A2-binding factors. Interestingly, the binding affinities of both MafA and A2.2 to the overlapping or non-overlapping binding sites were not significantly different, implying that the overlapping binding organization may increase the activation potential of MafA by physical/functional interactions with A2-binding factors. Thus our results demonstrate a novel mechanism for the regulation of MafA activity, and in turn beta-cell function, by altering expression and/or binding of the A2.2 factor. Our results further suggest that the major downstream targets of MafA will in addition to the MARE element have a binding site for the A2.2 factor.
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Affiliation(s)
- Wataru Nishimura
- *Section of Islet Transplantation and Cell Biology, Joslin Diabetes Center, Harvard Medical School, Boston, MA 02215, U.S.A
- †Department of Medicine, Harvard Medical School, Boston, MA 02215, U.S.A
| | - Therese Salameh
- *Section of Islet Transplantation and Cell Biology, Joslin Diabetes Center, Harvard Medical School, Boston, MA 02215, U.S.A
| | - Takuma Kondo
- *Section of Islet Transplantation and Cell Biology, Joslin Diabetes Center, Harvard Medical School, Boston, MA 02215, U.S.A
- †Department of Medicine, Harvard Medical School, Boston, MA 02215, U.S.A
| | - Arun Sharma
- *Section of Islet Transplantation and Cell Biology, Joslin Diabetes Center, Harvard Medical School, Boston, MA 02215, U.S.A
- †Department of Medicine, Harvard Medical School, Boston, MA 02215, U.S.A
- To whom correspondence should be addressed, at Research Division, Joslin Diabetes Center, One Joslin Place, Boston, MA 02215, U.S.A. (email )
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13
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Hagman DK, Hays LB, Parazzoli SD, Poitout V. Palmitate inhibits insulin gene expression by altering PDX-1 nuclear localization and reducing MafA expression in isolated rat islets of Langerhans. J Biol Chem 2005; 280:32413-8. [PMID: 15944145 PMCID: PMC1361267 DOI: 10.1074/jbc.m506000200] [Citation(s) in RCA: 162] [Impact Index Per Article: 8.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
Abnormalities in lipid metabolism have been proposed as contributing factors to both defective insulin secretion from the pancreatic beta cell and peripheral insulin resistance in type 2 diabetes. Previously, we have shown that prolonged exposure of isolated rat islets of Langerhans to excessive fatty acid levels impairs insulin gene transcription. This study was designed to assess whether palmitate alters the expression and binding activity of the key regulatory factors pancreas-duodenum homeobox-1 (PDX-1), MafA, and Beta2, which respectively bind to the A3, C1, and E1 elements in the proximal region of the insulin promoter. Nuclear extracts of isolated rat islets cultured with 0.5 mm palmitate exhibited reduced binding activity to the A3 and C1 elements but not the E1 element. Palmitate did not affect the overall expression of PDX-1 but reduced its nuclear localization. In contrast, palmitate blocked the stimulation of MafA mRNA and protein expression by glucose. Combined adenovirus-mediated overexpression of PDX-1 and MafA in islets completely prevented the inhibition of insulin gene expression by palmitate. These results demonstrate that prolonged exposure of islets to palmitate inhibits insulin gene transcription by impairing nuclear localization of PDX-1 and cellular expression of MafA.
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Affiliation(s)
- Derek K. Hagman
- From the Pacific Northwest Research Institute, Seattle, Washington 98122 and the
| | - Lori B. Hays
- From the Pacific Northwest Research Institute, Seattle, Washington 98122 and the
| | - Susan D. Parazzoli
- From the Pacific Northwest Research Institute, Seattle, Washington 98122 and the
| | - Vincent Poitout
- From the Pacific Northwest Research Institute, Seattle, Washington 98122 and the
- Department of Medicine, University of Washington, Seattle, Washington 98195
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14
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Matsuoka TA, Artner I, Henderson E, Means A, Sander M, Stein R. The MafA transcription factor appears to be responsible for tissue-specific expression of insulin. Proc Natl Acad Sci U S A 2004; 101:2930-3. [PMID: 14973194 PMCID: PMC365722 DOI: 10.1073/pnas.0306233101] [Citation(s) in RCA: 213] [Impact Index Per Article: 10.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022] Open
Abstract
Insulin gene expression is regulated by several islet-enriched transcription factors. However, MafA is the only beta cell-specific activator. Here, we show that MafA selectively induces endogenous insulin transcription in non-beta cells. MafA was also first detected in the insulin-producing cells formed during the second and predominant phase of beta cell differentiation, and absent in the few insulin-positive cells found in Nkx6.1(-/-) pancreata, which lack the majority of second-phase beta cells. These results demonstrate that MafA is a potent insulin activator that is likely to function downstream of Nkx6.1 during islet insulin-producing cell development.
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Affiliation(s)
- Taka-aki Matsuoka
- Department of Molecular Physiology and Biophysics, Vanderbilt University School of Medicine, 723 Light Hall, Nashville, TN 37232, USA
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15
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Leibowitz G, Ferber S, Edlund H, Gross DJ, Cerasi E, Melloul D, Kaiser N. IPF1/PDX1 deficiency and beta-cell dysfunction in Psammomys obesus, an animal With type 2 diabetes. Diabetes 2001; 50:1799-806. [PMID: 11473041 DOI: 10.2337/diabetes.50.8.1799] [Citation(s) in RCA: 50] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 11/13/2022]
Abstract
The homeodomain transcription factor IPF1/PDX1 is required in beta-cells for efficient expression of insulin, glucose transporter 2, and prohormone convertases 1/3 and 2. Psammomys obesus, a model of diet-responsive type 2 diabetes, shows markedly depleted insulin stores when given a high-energy (HE) diet. Despite hyperglycemia, insulin mRNA levels initially remained unchanged and then decreased gradually to 15% of the basal level by 3 weeks. Moreover, insulin gene expression was not increased when isolated P. obesus islets were exposed to elevated glucose concentrations. Consistent with these observations, no functional Ipf1/Pdx1 gene product was detected in islets of newborn or adult P. obesus using immunostaining, Western blot, DNA binding, and reverse transcriptase-polymerase chain reaction analyses. Other beta-cell transcription factors (e.g., ISL-1, Nkx2.2, and Nkx6.1) were expressed in P. obesus islets, and the DNA binding activity of the insulin transcription factors RIPE3b1-Act and IEF1 was intact. Ipf1/Pdx1 gene transfer to isolated P. obesus islets normalized the defect in glucose-stimulated insulin gene expression and prevented the rapid depletion of insulin content after exposure to high glucose. Taken together, these results suggest that the inability of P. obesus islets to adapt to dietary overload, with depletion of insulin content as a consequence, results from IPF1/PDX1 deficiency. However, because not all animals become hyperglycemic on HE diet, additional factors may be important for the development of diabetes in this animal model.
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Affiliation(s)
- G Leibowitz
- Department of Endocrinology and Metabolism, Hebrew University- Hadassah Medical Center, Jerusalem.
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16
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Kaytor EN, Qian J, Towle HC, Olson LK. An indirect role for upstream stimulatory factor in glucose-mediated induction of pyruvate kinase and S14 gene expression. Mol Cell Biochem 2000; 210:13-21. [PMID: 10976753 DOI: 10.1023/a:1007006429041] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/12/2022]
Abstract
Transcription of the L-type pyruvate kinase (L-PK) and S14 genes is induced in hepatocytes in response to increased glucose metabolism. The regulatory sequences of these genes responsible for induction by glucose have been mapped to related E-box containing motifs in the promoters. Similarly, L-PK promoter activity is stimulated in a differentiated pancreatic beta-cell line, INS-1, in response to elevated glucose. By mutational analysis, we demonstrate that the sequence requirements for glucose induction in the INS-1 cell are identical to those observed in the hepatocyte, suggesting that the same transcriptional factor(s) is responsible for regulation of L-PK expression in the two cell types. One nuclear factor that binds to the glucose regulatory sequences of both of these genes is the Upstream Stimulatory Factor (USF), a ubiquitous E-box binding protein. Mice deleted for the USF2 gene display a severely delayed response to carbohydrate feeding (Vallet et al. [26]). This observation, however, does not differentiate between a direct and an indirect role for USF in the process. To gain further insight into the possible involvement of USF in glucose signaling, we have used a recombinant adenoviral construct that expresses a dominant negative form of USF. This dominant negative can dimerize with endogenous USF and is shown to inhibit DNA binding of USF in hepatocytes and INS-1 cells. However, expression of the dominant negative USF did not block the ability of glucose to stimulate L-PK or S14 gene expression in hepatocytes or L-PK promoter activity in INS-1 cells. We conclude that USF does not act by binding to the glucose regulatory sequences of the S14 or L-PK genes and the role of USF in the process of glucose induction is indirect.
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Affiliation(s)
- E N Kaytor
- Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis 55455, USA
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Leibiger B, Moede T, Schwarz T, Brown GR, Köhler M, Leibiger IB, Berggren PO. Short-term regulation of insulin gene transcription by glucose. Proc Natl Acad Sci U S A 1998; 95:9307-12. [PMID: 9689076 PMCID: PMC21334 DOI: 10.1073/pnas.95.16.9307] [Citation(s) in RCA: 83] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/08/2023] Open
Abstract
Whereas short-term regulation of insulin biosynthesis at the level of translation is well accepted, glucose-dependent transcriptional control is still believed to be a long-term effect occurring after more than 2 hr of glucose stimulation. Because pancreatic beta cells are exposed to elevated glucose levels for minutes rather than hours after food uptake, we hypothesized the existence of a short-term transcriptional control. By studying the dynamics of newly synthesized (prepro)insulin RNA and by employing on-line monitoring of gene expression in single, insulin-producing cells, we were able to provide convincing evidence that insulin gene transcription indeed is affected by glucose within minutes. Exposure of insulinoma cells and isolated pancreatic islets to elevated glucose for only 15 min resulted in a 2- to 5-fold elevation in (prepro)insulin mRNA levels within 60-90 min. Similarly, insulin promoter-driven green fluorescent protein expression in single insulin-producing cells was significantly enhanced after transient glucose stimulation. Thus, short-term signaling, such as that involved in insulin secretion, also may regulate insulin gene transcription.
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Affiliation(s)
- B Leibiger
- The Rolf Luft Center for Diabetes Research, Department of Molecular Medicine, Karolinska Institute, S-171 76 Stockholm, Sweden
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Marshak S, Totary H, Cerasi E, Melloul D. Purification of the beta-cell glucose-sensitive factor that transactivates the insulin gene differentially in normal and transformed islet cells. Proc Natl Acad Sci U S A 1996; 93:15057-62. [PMID: 8986763 PMCID: PMC26355 DOI: 10.1073/pnas.93.26.15057] [Citation(s) in RCA: 127] [Impact Index Per Article: 4.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/03/2023] Open
Abstract
The beta cell-specific glucose-sensitive factor (GSF), which binds the A3 motif of the rat I and human insulin promoters, is modulated by extracellular glucose. A single mutation in the GSF binding site of the human insulin promoter abolishes the stimulation by high glucose only in normal islets, supporting the suggested physiological role of GSF in the glucose-regulated expression of the insulin gene. GSF binding activity was observed in all insulin-producing cells. We have therefore purified this activity from the rat insulinoma RIN and found that a single polypeptide of 45 kDa was responsible for DNA binding. Its amino acid sequence, determined by microsequencing, provided direct evidence that GSF corresponds to insulin promoter factor 1 (IPF-1; also known as PDX-1) and that, in addition to its essential roles in development and differentiation of pancreatic islets and in beta cell-specific gene expression, it functions as mediator of the glucose effect on insulin gene transcription in differentiated beta cells. The human cDNA coding for GSF/IPF-1 has been cloned, its cell and tissue distribution is described. Its expression in the glucagon-producing cell line alpha TC1 transactivates the wild-type human insulin promoter more efficiently than the mutated construct. It is demonstrated that high levels of ectopic GSF/IPF-1 inhibit the expression of the human insulin gene in normal islets, but not in transformed beta TC1 cells. These results suggest the existence of a control mechanism, such as requirement for a coactivator of GSF/IPF-1, which may be present in limiting amounts in normal as opposed to transformed beta cells.
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Affiliation(s)
- S Marshak
- Department of Endocrinology and Metabolism, Hebrew University Hadassah Medical Center, Jerusalem, Israel
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