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Alsuliam SM, Albadr NA, Alshammari GM, Almaiman SA, ElGasim Ahmed Yagoub A, Saleh A, Abdo Yahya M. Lepidium sativum alleviates diabetic nephropathy in a rat model by attenuating glucose levels, oxidative stress, and inflammation with concomitant suppression of TGF-β1. Saudi J Biol Sci 2023; 30:103720. [PMID: 37576066 PMCID: PMC10422013 DOI: 10.1016/j.sjbs.2023.103720] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/02/2023] [Revised: 06/15/2023] [Accepted: 06/23/2023] [Indexed: 08/15/2023] Open
Abstract
In this research, the treatment of diabetic nephropathy in rats induced by streptozotocin with L. sativium whole-plant aqueous extract was examined, and the mechanism of action was proposed. Adult male rats were grouped into: control, L. sativum, T1DM, and T1DM + L. sativum-treated groups. For 8 weeks, L. sativum S was given to rats at a final dose of 250 mg/kg. Treatment with L. sativum reduced the amount of fasting glucose, increased the amount of fasting insulin, and diminished the increase in hepatic and serum cholesterol, free fatty acid, and triglyceride levels. The level of serum LDL-c was reduced. At the level of the kidney, L. sativum reduced urine volume and albumin excretion and spiked creatinine excretion. It also attenuated the tubular damage in the rats' kidneys and reduced the amounts of major inflammatory markers, including nuclear factor-kappaα (NF-κB), tumor necrosis factor-α (TNF-α), and interleukin-6 (IL-6). Interestingly, L. sativium reduced the amount of mRNA transforming growth factor-β1 (TGF-β1), stimulated mRNA superoxide dismutase (SOD) and catalase (CAT), reduced lipid peroxide levels (MDA), and increased the glutathione (GSH), SOD, and CAT in the rat kidneys of the control and T1DM-treated group. In conclusion, L. sativum is a novel therapy against DN owing to its hypoglycemic effect, insulin-releasing, and antioxidant potential.
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Affiliation(s)
- Sarah M. Alsuliam
- Department of Food Science and Nutrition, College of Food and Agricultural Sciences, King Saud University, Riyadh 11451, Saudi Arabia
| | - Nawal A. Albadr
- Department of Food Science and Nutrition, College of Food and Agricultural Sciences, King Saud University, Riyadh 11451, Saudi Arabia
| | - Ghedeir M. Alshammari
- Department of Food Science and Nutrition, College of Food and Agricultural Sciences, King Saud University, Riyadh 11451, Saudi Arabia
| | - Salah A. Almaiman
- Department of Food Science and Nutrition, College of Food and Agricultural Sciences, King Saud University, Riyadh 11451, Saudi Arabia
| | - Abu ElGasim Ahmed Yagoub
- Department of Food Science and Nutrition, College of Food and Agricultural Sciences, King Saud University, Riyadh 11451, Saudi Arabia
| | - Ali Saleh
- Department of Food Science and Nutrition, College of Food and Agricultural Sciences, King Saud University, Riyadh 11451, Saudi Arabia
| | - Mohammed Abdo Yahya
- Department of Food Science and Nutrition, College of Food and Agricultural Sciences, King Saud University, Riyadh 11451, Saudi Arabia
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Jeon YG, Kim YY, Lee G, Kim JB. Physiological and pathological roles of lipogenesis. Nat Metab 2023; 5:735-759. [PMID: 37142787 DOI: 10.1038/s42255-023-00786-y] [Citation(s) in RCA: 84] [Impact Index Per Article: 42.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/18/2022] [Accepted: 03/15/2023] [Indexed: 05/06/2023]
Abstract
Lipids are essential metabolites, which function as energy sources, structural components and signalling mediators. Most cells are able to convert carbohydrates into fatty acids, which are often converted into neutral lipids for storage in the form of lipid droplets. Accumulating evidence suggests that lipogenesis plays a crucial role not only in metabolic tissues for systemic energy homoeostasis but also in immune and nervous systems for their proliferation, differentiation and even pathophysiological roles. Thus, excessive or insufficient lipogenesis is closely associated with aberrations in lipid homoeostasis, potentially leading to pathological consequences, such as dyslipidaemia, diabetes, fatty liver, autoimmune diseases, neurodegenerative diseases and cancers. For systemic energy homoeostasis, multiple enzymes involved in lipogenesis are tightly controlled by transcriptional and post-translational modifications. In this Review, we discuss recent findings regarding the regulatory mechanisms, physiological roles and pathological importance of lipogenesis in multiple tissues such as adipose tissue and the liver, as well as the immune and nervous systems. Furthermore, we briefly introduce the therapeutic implications of lipogenesis modulation.
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Affiliation(s)
- Yong Geun Jeon
- Center for Adipocyte Structure and Function, Institute of Molecular Biology and Genetics, School of Biological Sciences, Seoul National University, Seoul, South Korea
| | - Ye Young Kim
- Center for Adipocyte Structure and Function, Institute of Molecular Biology and Genetics, School of Biological Sciences, Seoul National University, Seoul, South Korea
| | - Gung Lee
- Center for Adipocyte Structure and Function, Institute of Molecular Biology and Genetics, School of Biological Sciences, Seoul National University, Seoul, South Korea
| | - Jae Bum Kim
- Center for Adipocyte Structure and Function, Institute of Molecular Biology and Genetics, School of Biological Sciences, Seoul National University, Seoul, South Korea.
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Shatoor AS, Al Humayed S, Almohiy HM. Astaxanthin attenuates hepatic steatosis in high-fat diet-fed rats by suppressing microRNA-21 via transactivation of nuclear factor erythroid 2-related factor 2. J Physiol Biochem 2021; 78:151-168. [PMID: 34651285 DOI: 10.1007/s13105-021-00850-9] [Citation(s) in RCA: 20] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/05/2021] [Accepted: 09/29/2021] [Indexed: 02/08/2023]
Abstract
This study examined whether astaxanthin (ASX) could alleviate hepatic steatosis in rats fed a high-fat diet (HFD) by modulating the nuclear factor erythroid 2-related factor 2 (Nrf2)/miR-21 axis. Rats (n = 8/group) were fed either a standard diet (3.8 kcal/g; 10% fat) or HFD (4.6 kcal/g; 40% fat) and treated orally with either the vehicle or ASX (6 mg/kg) daily for 8 days. Another group was fed HFD and treated with ASX and brusatol (an Nrf2 inhibitor) (2 mg/kg/twice per week/i.p.). ASX prevented the gain in body and liver weights and attenuated hepatic lipid accumulation in HFD-fed rats. In the control and HFD-fed rats, ASX did not affect food intake, serum free fatty acid (FFA) content, and glucose and insulin levels and tolerance. However, serum triglyceride (TG), cholesterol, and low-density lipoprotein-cholesterol levels; hepatic levels of TGs and FFAs; and hepatic levels of Srebp1, Srebp2, HMGCR, and fatty acid synthase mRNAs and miR-21 were reduced and the mRNA levels of Pparα were significantly increased in both the groups. These effects were associated with a reduction in the hepatic levels of reactive oxygen species, malondialdehyde, tumor necrosis factor-α, and interlukin-6 as well as an increase in superoxide dismutase levels, total glutathione content, and nuclear levels and activity of Nrf2. miR-21 levels were strongly correlated with the nuclear activity of Nrf2. Brusatol completely reversed the effects of ASX. In conclusion, ASX prevents hepatic steatosis mainly by transactivating Nrf2 and is associated with the suppression of miR-21 and Srebp1/2 and upregulation of Pparα expression.
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Affiliation(s)
- Abdullah S Shatoor
- Department of Medicine, Cardiology Section, College of Medicine, King Khalid University (KKU), Abha, Saudi Arabia.
| | - Suliman Al Humayed
- Department of Internal Medicine, College of Medicine, King Khalid University (KKU), Abha, Saudi Arabia
| | - Hussain M Almohiy
- Depatrtment of Radiology Science, College of Applied Medical Sciences, King Khalid University, Abha, Saudi Arabia
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Aldayel TS, Alshammari GM, Omar UM, Grace MH, Lila MA, Yahya MA. Hypoglycaemic, insulin releasing, and hepatoprotective effect of the aqueous extract of Aloe perryi Baker resin (Socotran Aloe) in streptozotocin-induced diabetic rats. JOURNAL OF TAIBAH UNIVERSITY FOR SCIENCE 2020. [DOI: 10.1080/16583655.2020.1855859] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 02/01/2023]
Affiliation(s)
- Tahany Saleh Aldayel
- Nutrition and Food Science, Department of Physical Sport Sciences, Princess Nourah Bint Abdulrahman University, Riyadh, Saudi Arabia
| | - Ghedeir M. Alshammari
- Department of Food Science and Nutrition, College of Food and Agricultural Science, King Saud University, Riyadh, Saudi Arabia
| | - Ulfat Mohammed Omar
- Department of Biochemistry, Faculty of Science, King Abdulaziz University; Immunology Unit, King Fahad Medical Research Center, Jeddah, Saudi Arabia
| | - Mary H. Grace
- Plants for Human Health Institute, Department of Food Bioprocessing and Nutrition Sciences, North Carolina State University, North Carolina Research Campus, Kannapolis, NC, USA
| | - Mary Ann Lila
- Plants for Human Health Institute, Department of Food Bioprocessing and Nutrition Sciences, North Carolina State University, North Carolina Research Campus, Kannapolis, NC, USA
| | - Mohammed A. Yahya
- Department of Food Science and Nutrition, College of Food and Agricultural Science, King Saud University, Riyadh, Saudi Arabia
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Daniel PV, Mondal P. Causative and Sanative dynamicity of ChREBP in Hepato-Metabolic disorders. Eur J Cell Biol 2020; 99:151128. [PMID: 33232883 DOI: 10.1016/j.ejcb.2020.151128] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/09/2020] [Revised: 10/22/2020] [Accepted: 10/28/2020] [Indexed: 12/12/2022] Open
Abstract
ChREBP is the master regulator of carbohydrate dependent glycolytic and lipogenic flux within metabolic tissues. It plays a vital role in hyper-calorific milieu by activating glycolysis, lipogenesis along with pentose phosphate shunt and glycogen synthesis, fostering immediate reduction in the systemic glycemic levels. Liver being the primary organ to sense disproportionate dietary intake and linked physiological stress, stimulates ChREBP to perform the aforementioned processes. Activated ChREBP also inhibits lipolysis and encourages proper disposal of excessive triglycerides into adipocytes from the liver ablating hepatic intracellular lipid trafficking. Chronic overeating or onset of positive energy balance, hyper-activates ChREBP and signals development, intensification of hepato-metabolic disorders, and allied discrepancies in the whole-body metabolic functioning. ChREBP thus gets negatively connotated as the primary regulator of hepatic disorders, owing to its inherent features as the primary glycemic sensor and the only transcription factor that can transduce glucose-dependent glycolytic and lipogenic signals. Through this review, we - try to recapitulate and emphasize on the sanative events coordinated by ChREBP in several pathophysiological states. In totality, we aim to uncouple the disease-causing aspects of ChREBP from its positive attributes evoked during a metabolic crisis, in hepato-metabolic diseases.
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Affiliation(s)
- P Vineeth Daniel
- School of Basic Sciences, Indian Institute of Technology Mandi, Mandi 175001, H.P, India.
| | - Prosenjit Mondal
- School of Basic Sciences, Indian Institute of Technology Mandi, Mandi 175001, H.P, India.
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Ghashghaeinia M, Köberle M, Mrowietz U, Bernhardt I. Proliferating tumor cells mimick glucose metabolism of mature human erythrocytes. Cell Cycle 2019; 18:1316-1334. [PMID: 31154896 PMCID: PMC6592250 DOI: 10.1080/15384101.2019.1618125] [Citation(s) in RCA: 32] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/10/2023] Open
Abstract
Mature human erythrocytes are dependent on anerobic glycolysis, i.e. catabolism (oxidation) of one glucose molecule to produce two ATP and two lactate molecules. Proliferating tumor cells mimick mature human erythrocytes to glycolytically generate two ATP molecules. They deliberately avoid or switch off their respiration, i.e. tricarboxylic acid (TCA) cycle and oxidative phosphorylation (OXPHOS) machinery and consequently dispense with the production of additional 36 ATP molecules from one glucose molecule. This phenomenon is named aerobic glycolysis or Warburg effect. The present review deals with the fate of a glucose molecule after entering a mature human erythrocyte or a proliferating tumor cell and describes why it is useful for a proliferating tumor cell to imitate a mature erythrocyte. Blood consisting of plasma and cellular components (99% of the cells are erythrocytes) may be regarded as a mobile organ, constantly exercising a direct interaction with other organs. Therefore, the use of drugs, which influences the biological activity of erythrocytes, has an immediate effect on the entire organism. Abbreviations: TCA: tricarboxylic acid cycle; OXPHOS: oxidative phosphorylation; GSH: reduced state of glutathione; NFκB: Nuclear factor of kappa B; PKB (Akt): protein kinase B; NOS: nitric oxide synthase; IgG: immune globulin G; H2S: hydrogen sulfide; slanDCs: Human 6-sulfo LacNAc-expressing dendritic cells; IL-8: interleukin-8; LPS: lipopolysaccharide; ROS: reactive oxygen species; PPP: pentose phosphate pathway; NADPH: nicotinamide adenine dinucleotide phosphate hydrogen; R5P: ribose-5-phophate; NAD: nicotinamide adenine dinucleotide; FAD: flavin adenine dinucleotide; O2●−: superoxide anion; G6P: glucose 6-phosphate; HbO2: Oxyhemoglobin; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; GAP: glyceraldehyde-3-phosphate; 1,3-BPG: 1,3-bis-phosphoglycerate; 2,3-BPG: 2,3-bisphosphoglycerte; PGAM1: phosphoglycerate mutase 1; 3-PG: 3-phosphoglycerate; 2-PG: 2-phosphoglycerate; MIPP1: Multiple inositol polyphosphate phosphatase; mTORC1: mammalian target of rapamycin complex 1; Ru5P: ribulose 5-phosphate; ox-PPP: oxidative branch of pentose phosphate pathway; PGK: phosphoglycerate kinase; IFN-γ: interferon-γ; LDH: lactate dehydrogenase; STAT3: signal transducer and activator of transcription 3; Rheb: Ras homolog enriched in Brain; H2O2: hydrogen peroxide; ROOH: lipid peroxide; SOD: superoxide dismutase; MRC: mitochondrial respiratory chain; MbFe2+-O2: methmyoglobin; RNR: ribonucleotide reductase; PRPP: phosphoribosylpyrophosphate; PPi: pyrophosphate; GSSG: oxidized state of glutathione; non-ox-PPP: non-oxidative branch of pentose phosphate pathway; RPI: ribose-5-phosphate isomerase; RPE: ribulose 5-phosphate 3-epimerase; X5P: xylulose 5-phosphate; TK: transketolase; TA: transaldolase; F6P: fructose-6-phosphate; AR2: aldose reductase 2; SD: sorbitol dehydrogenase; HK: hexokinase; MG: mehtylglyoxal; DHAP: dihydroxyacetone phosphate; TILs: tumor-infiltrating lymphocytes; MCTs: monocarboxylate transporters; pHi: intracellular pH; Hif-1α: hypoxia-induced factor 1; NHE1: sodium/H+ (Na+/H+) antiporter 1; V-ATPase: vacuolar-type proton ATPase; CAIX: carbonic anhydrase; CO2: carbon dioxide; HCO3−: bicarbonate; NBC: sodium/bicarbonate (Na+/HCO3−) symporter; pHe: extracellular pH; GLUT-1: glucose transporter 1; PGK-1: phosphoglycerate kinase 1
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Affiliation(s)
- Mehrdad Ghashghaeinia
- a Department of Dermatology , University Medical Center Schleswig-Holstein, Campus Kiel , Kiel , Germany
| | - Martin Köberle
- b Klinik und Poliklinik für Dermatologie und Allergologie, Fakultät für Medizin , Technische Universität München , Munich , Germany
| | - Ulrich Mrowietz
- a Department of Dermatology , University Medical Center Schleswig-Holstein, Campus Kiel , Kiel , Germany
| | - Ingolf Bernhardt
- c Laboratory of Biophysics, Faculty of Natural and Technical Sciences III , Saarland University , Saarbruecken , Germany
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Sae-Lee C, Moolsuwan K, Chan L, Poungvarin N. ChREBP Regulates Itself and Metabolic Genes Implicated in Lipid Accumulation in β-Cell Line. PLoS One 2016; 11:e0147411. [PMID: 26808438 PMCID: PMC4725739 DOI: 10.1371/journal.pone.0147411] [Citation(s) in RCA: 37] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/21/2015] [Accepted: 01/04/2016] [Indexed: 12/20/2022] Open
Abstract
Carbohydrate response element binding protein (ChREBP) is an important transcription factor that regulates a variety of glucose-responsive genes in hepatocytes. To date, only two natural isoforms, Chrebpα and Chrebpβ, have been identified. Although ChREBP is known to be expressed in pancreatic β cells, most of the glucose-responsive genes have never been verified as ChREBP targets in this organ. We aimed to explore the impact of ChREBP expression on regulating genes linked to accumulation of lipid droplets, a typical feature of β-cell glucotoxicity. We assessed gene expression in 832/13 cells overexpressing constitutively active ChREBP (caChREBP), truncated ChREBP with nearly identical amino acid sequence to Chrebpβ, or dominant negative ChREBP (dnChREBP). Among multiple ChREBP-controlled genes, ChREBP was sufficient and necessary for regulation of Eno1, Pklr, Mdh1, Me1, Pdha1, Acly, Acaca, Fasn, Elovl6, Gpd1, Cpt1a, Rgs16, Mid1ip1,Txnip, and Chrebpβ. Expression of Chrebpα and Srebp1c were not changed by caChREBP or dnChREBP. We identified functional ChREBP binding sequences that were located on the promoters of Chrebpβ and Rgs16. We also showed that Rgs16 overexpression lead to increased considerable amounts of lipids in 832/13 cells. This phenotype was accompanied by reduction of Cpt1a expression and slight induction of Fasn and Pklr gene in these cells. In summary, we conclude that Chrebpβ modulates its own expression, not that of Chrebpα; it also regulates the expression of several metabolic genes in β-cells without affecting SREBP-1c dependent regulation. We also demonstrate that Rgs16 is one of the ChREBP-controlled genes that potentiate accumulation of lipid droplets in β-cells.
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Affiliation(s)
- Chanachai Sae-Lee
- Clinical Molecular Pathology Laboratory, Department of Clinical Pathology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand
| | - Kanya Moolsuwan
- Clinical Molecular Pathology Laboratory, Department of Clinical Pathology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand
- Molecular Medicine Program, Multidisciplinary Unit, Faculty of Science, Mahidol University, Bangkok, Thailand
| | - Lawrence Chan
- Department of Medicine, Baylor College of Medicine, Houston, Texas, United States of America
| | - Naravat Poungvarin
- Clinical Molecular Pathology Laboratory, Department of Clinical Pathology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand
- * E-mail:
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Dentin R, Tomas-Cobos L, Foufelle F, Leopold J, Girard J, Postic C, Ferré P. Glucose 6-phosphate, rather than xylulose 5-phosphate, is required for the activation of ChREBP in response to glucose in the liver. J Hepatol 2012; 56:199-209. [PMID: 21835137 DOI: 10.1016/j.jhep.2011.07.019] [Citation(s) in RCA: 122] [Impact Index Per Article: 9.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/21/2010] [Revised: 07/07/2011] [Accepted: 07/11/2011] [Indexed: 12/04/2022]
Abstract
BACKGROUND & AIMS In liver, the glucose-responsive transcription factor ChREBP plays a critical role in converting excess carbohydrates into triglycerides through de novo lipogenesis. Although the importance of ChREBP in glucose sensing and hepatic energy utilization is strongly supported, the mechanism driving its activation in response to glucose in the liver is not fully understood. Indeed, the current model of ChREBP activation, which depends on Serine 196 and Threonine 666 dephosphorylation, phosphatase 2A (PP2A) activity, and xylulose 5-phosphate (X5P) as a signaling metabolite, has been challenged. METHODS We inhibited PP2A activity in HepG2 cells through the overexpression of SV40 small t antigen and addressed the importance of ChREBP dephosphorylation on Ser-196 using a phospho-specific antibody. To identify the exact nature of the metabolite signal required for ChREBP activity in liver, we focused on the importance of G6P synthesis in liver cells, through the modulation of glucose 6-phosphate dehydrogenase (G6PDH) activity, the rate-limiting enzyme of the pentose phosphate pathway in hepatocytes, and in HepG2 cells using both adenoviral and siRNA approaches. RESULTS In contrast to the current proposed model, our study reports that PP2A activity is dispensable for ChREBP activation in response to glucose and that dephosphorylation on Ser-196 is not sufficient to promote ChREBP nuclear translocation in the absence of a rise in glucose metabolism. By deciphering the respective roles of G6P and X5P as signaling metabolites, our study reveals that G6P produced by GK, but not X5P, is essential for both ChREBP nuclear translocation and transcriptional activity in response to glucose in liver cells. CONCLUSIONS Altogether, our study, by reporting that G6P is the glucose-signaling metabolite, challenges the PP2A/X5P-dependent model currently described for ChREBP activation in response to glucose in liver.
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Bennett KA, Forsyth L, Burchell A. Functional analysis of the 5' flanking region of the human G6PC3 gene: regulation of promoter activity by glucose, pyruvate, AMP kinase and the pentose phosphate pathway. Mol Genet Metab 2011; 103:254-61. [PMID: 21474354 DOI: 10.1016/j.ymgme.2011.03.015] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/06/2011] [Revised: 03/11/2011] [Accepted: 03/11/2011] [Indexed: 10/18/2022]
Abstract
G6PC3 is a widely expressed isoform of glucose-6-phosphatase, found in many foetal and adult tissues. Mutations in this gene cause developmental abnormalities and severe neutropenia due to abolition of glucose recycling between the cytoplasm and endoplasmic reticulum. Low G6PC3 expression as a result of promoter polymorphisms or dysregulation could produce similar outcomes. Here we investigated the regulation of human G6PC3 promoter activity. HeLa and H4IIE cells were transiently transfected with G6PC3 promoter coupled to the firefly luciferase gene, and promoter activity was measured by dual luciferase assay. Activity was highest in a 453 bp segment of the G6PC3 promoter, from -455 to -3 relative to the transcriptional start site. This promoter was unresponsive to glucostatic hormones. Its activity increased significantly between 1 and 5.5 mM glucose, and was not elevated further by glucose concentrations up to 25 mM. Pyruvate increased its activity, but β-hydroxybutyrate and sodium acetate did not. Promoter activity was reduced by inhibitors of hexokinase, glyceraldehyde phosphate dehydrogenase and the oxidative branch of the pentose phosphate pathway, but not by a transketolase inhibitor. Deletion of two adjacent Enhancer-boxes (-274 to -279 and -299 to -304) reduced promoter activity and abolished the glucose effect, suggesting they could function as a glucose response element. Deletion of an additional downstream 140 bp (-140 to -306) restored activity, but not the glucose response, suggesting the presence of repressor elements in this region. 5-Aminoimidazole-4-carboxamide 1-β-D-ribofuranoside (AICAR) reduced promoter activity, showing dependence on AMP-kinase. Regulation of the G6PC3 promoter is thus radically different to that of the hepatic isoform, G6PC. It is sensitive to carbohydrate, but not to fatty acid metabolites, and at much lower physiological concentrations. Based on these findings, we speculate that reduced G6PC3 expression could occur during hypoglycemic episodes in vivo, which are common in utero and in the postnatal period. If such episodes lower G6PC3 expression they could place the foetus or infant at risk of impaired immune function and development, and this possibility requires further examination both in vitro and in vivo.
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Affiliation(s)
- Kimberley Ann Bennett
- Maternal and Child Health Sciences, Ninewells Hospital and Medical School, University of Dundee, Dundee, DD1 9SY, UK.
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Amo K, Arai H, Uebanso T, Fukaya M, Koganei M, Sasaki H, Yamamoto H, Taketani Y, Takeda E. Effects of xylitol on metabolic parameters and visceral fat accumulation. J Clin Biochem Nutr 2011; 49:1-7. [PMID: 21765599 PMCID: PMC3128359 DOI: 10.3164/jcbn.10-111] [Citation(s) in RCA: 50] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/23/2010] [Accepted: 09/17/2010] [Indexed: 11/22/2022] Open
Abstract
Xylitol is widely used as a sweetener in foods and medications. Xylitol ingestion causes a small blood glucose rise, and it is commonly used as an alternative to high-energy supplements in diabetics. In previous studies, a xylitol metabolite, xylulose-5-phosphate, was shown to activate carbohydrate response element binding protein, and to promote lipogenic enzyme gene transcription in vitro; however, the effects of xylitol in vivo are not understood. Here we investigated the effects of dietary xylitol on lipid metabolism and visceral fat accumulation in rats fed a high-fat diet. Sprague-Dawley rats were fed a high-fat diet containing 0 g (control), 1.0 g/100 kcal (X1) or 2.0 g/100 kcal (X2) of xylitol. After the 8-week feeding period, visceral fat mass and plasma insulin and lipid concentrations were significantly lower in xylitol-fed rats than those in high-fat diet rats. Gene expression levels of ChREBP and lipogenic enzymes were higher, whereas the expression of sterol regulatory-element binding protein 1c was lower and fatty acid oxidation-related genes were significantly higher in the liver of xylitol-fed rats as compared with high-fat diet rats. In conclusion, intake of xylitol may be beneficial in preventing the development of obesity and metabolic abnormalities in rats with diet-induced obesity.
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Affiliation(s)
- Kikuko Amo
- Department of Clinical Nutrition, Institute of Health Biosciences, University of Tokushima Graduate School, Tokushima 770-8503, Japan
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Shimada M, Mochizuki K, Goda T. Feeding rats dietary resistant starch reduces both the binding of ChREBP and the acetylation of histones on the Thrsp gene in the jejunum. JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY 2011; 59:1464-1469. [PMID: 21244091 DOI: 10.1021/jf103111u] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 05/30/2023]
Abstract
We have previously reported that the thyroid hormone-responsive spot 14 protein (Thrsp) gene is expressed in rat jejunum. In this study, we found that jejunal mRNA and protein expressions of Thrsp were markedly reduced in rats fed a diet containing a high amount of resistant starch (RS), which is an indigestible starch, for 7 days, compared with those fed a regular starch diet. Furthermore, we found that the binding of carbohydrate response element binding protein (ChREBP), which is a key transcription factor for the Thrsp gene, and the acetylation of histones H3 and H4, which is one of the histone modifications for transactivation, on the Thrsp gene were reduced by feeding the RS diet. These results suggest that the reduction of jejunal Thrsp gene expression by feeding a diet rich in less-digestible starch is associated with decreases in the binding of ChREBP and the acetylation of histones on the gene.
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Affiliation(s)
- Masaya Shimada
- Department of Nutrition, Faculty of Health Sciences, Chiba Prefectural University of Health Sciences, Japan
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Abstract
Glucose metabolism represents a critical physiological program that not only provides energy to support cell proliferation, but also directly modulates signaling pathways of cell death. With the growing recognition of regulation of cell death by glucose metabolism, many techniques that can be applied in the study have been developed. This chapter discusses several protocols that aid in the analysis of glucose metabolism and cell death and the principles in practicing them under different conditions.
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Affiliation(s)
- Yuxing Zhao
- Department of Pharmacology, Duke University, Durham, North Carolina, USA
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Rauch MC, Ocampo ME, Bohle J, Amthauer R, Yáñez AJ, Rodríguez-Gil JE, Slebe JC, Reyes JG, Concha II. Hexose transporters GLUT1 and GLUT3 are colocalized with hexokinase I in caveolae microdomains of rat spermatogenic cells. J Cell Physiol 2006; 207:397-406. [PMID: 16419038 DOI: 10.1002/jcp.20582] [Citation(s) in RCA: 24] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/22/2022]
Abstract
Postmeiotic spermatogenic cells, but not meiotic spermatogenic cells respond differentially with glucose-induced changes in [Ca2+]i indicating a differential transport of glucose via facilitative hexose transporters (GLUTs) specifically distributed in the plasma membrane. Several studies have indicated that plasma membrane in mammalian cells is not homogeneously organized, but contains specific microdomains known as detergent-resistant membrane domains (DRMDs), lipid rafts or caveolae. The association of these domains and GLUTs isoforms has not been characterized in spermatogenic cells. We analyzed the expression and function of GLUT1 and GLUT3 in isolated spermatocytes and spermatids. The results showed that spermatogenic cells express both glucose transporters, with spermatids exhibiting a higher affinity glucose transport system. In addition, spermatogenic cells express caveolin-1, and glucose transporters colocalize with caveolin-1 in caveolin-enriched membrane fractions. Experiments in which the integrity of caveolae was disrupted by pretreatment with methyl-beta-cyclodextrin, indicated that the involvement of cholesterol-enriched plasma membrane microdomains were involved in the localization of GLUTs and uptake of 2-deoxyglucose. We also observed cofractionation of GLUT3 and caveolin-1 in low-buoyant density membranes together with their shift to higher densities after methyl-beta-cyclodextrin treatment. GLUT1 was found in all fractions isolated. Immunofluorescent studies indicated that caveolin-1, GLUT1, and hexokinase I colocalize in spermatocytes while caveolin-1, GLUT3, and hexokinase I colocalize in spermatids. These findings suggest the presence of hexose transporters in DRMDs, and further support a role for intact caveolae or cholesterol-enriched membrane microdomains in relation to glucose uptake and glucose phosphorylation. The results would also explain the different glucose-induced changes in [Ca2+]i in both cells.
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14
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Letexier D, Peroni O, Pinteur C, Beylot M. In vivo expression of carbohydrate responsive element binding protein in lean and obese rats. DIABETES & METABOLISM 2006; 31:558-66. [PMID: 16357804 DOI: 10.1016/s1262-3636(07)70231-8] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/18/2022]
Abstract
ChREBP (Carbohydrate response element binding protein) is considered to mediate the stimulatory effect of glucose on the expression of lipogenic genes. Its activity is stimulated by glucose. Less is known on the control of its expression. This expression could be controlled by nutritional (glucose, fatty acids) and hormonal (insulin) factors. We examined the in vivo nutritional control of ChREBP expression in liver and adipose tissue of Wistar rats. Compared respectively to the fed state and to a high carbohydrate diet, ChREBP mRNA concentrations were not modified by fasting or a high fat diet in rat liver and adipose tissue. FAS and ACC1 mRNA concentrations were on the contrary decreased as expected by fasting and high fat diets and these variations of FAS and ACC1 mRNA were positively related to those of SREBP-1c mRNA and protein, but not of ChREBP mRNA. Therefore i) ChREBP expression appears poorly responsive to modifications of nutritional condition, ii) modifications of the expression of ChREBP do not seem implicated in the physiological control of lipogenesis. To investigate the possible role of ChREBP in pathological situations we measured its mRNA concentrations in the liver and adipose tissue of obese Zucker rats. ChREBP expression was increased in the liver but not the adipose tissue of obese rats compared to their lean littermates. These results support a role of ChREBP in the development of hepatic steatosis and hypertriglyceridemia but not of obesity in this experimental model.
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Affiliation(s)
- D Letexier
- INSERM U499, IFR 62, Faculté RTH LAENNEC, University Claude Bernard-Lyon 1, rue G Paradin, 69008 Lyon, France
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15
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Dentin R, Benhamed F, Pégorier JP, Foufelle F, Viollet B, Vaulont S, Girard J, Postic C. Polyunsaturated fatty acids suppress glycolytic and lipogenic genes through the inhibition of ChREBP nuclear protein translocation. J Clin Invest 2005; 115:2843-54. [PMID: 16184193 PMCID: PMC1224299 DOI: 10.1172/jci25256] [Citation(s) in RCA: 224] [Impact Index Per Article: 11.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/04/2005] [Accepted: 07/19/2005] [Indexed: 12/19/2022] Open
Abstract
Dietary polyunsaturated fatty acids (PUFAs) are potent inhibitors of hepatic glycolysis and lipogenesis. Recently, carbohydrate-responsive element-binding protein (ChREBP) was implicated in the regulation by glucose of glycolytic and lipogenic genes, including those encoding L-pyruvate kinase (L-PK) and fatty acid synthase (FAS). The aim of our study was to assess the role of ChREBP in the control of L-PK and FAS gene expression by PUFAs. We demonstrated in mice, both in vivo and in vitro, that PUFAs [linoleate (C18:2), eicosapentanoic acid (C20:5), and docosahexaenoic acid (C22:6)] suppressed ChREBP activity by increasing ChREBP mRNA decay and by altering ChREBP translocation from the cytosol to the nucleus, independently of an activation of the AMP-activated protein kinase, previously shown to regulate ChREBP activity. In contrast, saturated [stearate (C18)] and monounsaturated fatty acids [oleate (C18:1)] had no effect. Since glucose metabolism via the pentose phosphate pathway is determinant for ChREBP nuclear translocation, the decrease in xylulose 5-phosphate concentrations caused by a PUFA diet favors a PUFA-mediated inhibition of ChREBP translocation. In addition, overexpression of a constitutive nuclear ChREBP isoform in cultured hepatocytes significantly reduced the PUFA inhibition of both L-PK and FAS gene expression. Our results demonstrate that the suppressive effect of PUFAs on these genes is primarily caused by an alteration of ChREBP nuclear translocation. In conclusion, we describe a novel mechanism to explain the inhibitory effect of PUFAs on the genes encoding L-PK and FAS and demonstrate that ChREBP is a pivotal transcription factor responsible for coordinating the PUFA suppression of glycolytic and lipogenic genes.
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Affiliation(s)
- Renaud Dentin
- Département d'Endocrinologie, Institut Cochin, INSERM U567 CNRS UMR8104, Université René Descartes, Paris, France
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16
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Im SS, Kang SY, Kim SY, Kim HI, Kim JW, Kim KS, Ahn YH. Glucose-stimulated upregulation of GLUT2 gene is mediated by sterol response element-binding protein-1c in the hepatocytes. Diabetes 2005; 54:1684-1691. [PMID: 15919789 DOI: 10.2337/diabetes.54.6.1684] [Citation(s) in RCA: 91] [Impact Index Per Article: 4.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/27/2023]
Abstract
GLUT2 is mainly expressed in the liver, beta-cells of the pancreas, and the basolateral membrane of kidney proximal tubules and plays an important role in glucose homeostasis in living organisms. The transcription of the GLUT2 gene is known to be upregulated in the liver during postprandial hyperglycemic states or in type 2 diabetes. However, a molecular mechanism by which glucose activates GLUT2 gene expression is not known. In this study, we report evidence that sterol response element-binding protein (SREBP)-1c plays a key role in glucose-stimulated GLUT2 gene expression. The GLUT2 promoter reporter is activated by SREBP-1c, and the activation is inhibited by a dominant-negative form of SREBP-1c (SREBP-1c DN). Adenoviral expression of SREBP-1c DN suppressed glucose-stimulated GLUT2 mRNA level in primary hepatocytes. An electrophoretic mobility shift assay and mutational analysis of the GLUT2 promoter revealed that SREBP-1c binds to the -84/-76 region of the GLUT2 promoter. Chromatin immunoprecipitation revealed that the binding of SREBP-1c to the -84/-76 region was increased by glucose concentration in a dose-dependent manner. These results indicate that SREBP-1c mediates glucose-stimulated GLUT2 gene expression in hepatocytes.
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Affiliation(s)
- Seung-Soon Im
- Department of Biochemistry and Molecular Biology, Yonsei University College of Medicine, 134 Shinchon-dong, Seodaemoon-gu, Seoul 120-752, Korea
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17
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Ranalletta M, Jiang H, Li J, Tsao TS, Stenbit AE, Yokoyama M, Katz EB, Charron MJ. Altered hepatic and muscle substrate utilization provoked by GLUT4 ablation. Diabetes 2005; 54:935-43. [PMID: 15793230 DOI: 10.2337/diabetes.54.4.935] [Citation(s) in RCA: 13] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/09/2023]
Abstract
Studies were conducted to explore altered substrate utilization and metabolism in GLUT4 null mice. Liver fatty acid synthase mRNA and fatty acid synthesis rates were dramatically increased in GLUT4 null mice compared with control mice and were supported by increased rates of the pentose phosphate pathway oxidative phase and sterol regulatory binding protein mRNA expression. Increased GLUT2 protein content, glucokinase mRNA, and glucose-6-phosphate in GLUT4 null mice may provide substrate for the enhanced fatty acid synthesis. Increased fatty acid synthesis, however, did not lead to hepatic triglyceride accumulation in GLUT4 null mice because of increased hepatic triglyceride secretion rates. GLUT4 null mice rapidly cleared orally administered olive oil, had reduced serum triglyceride concentrations in the fed and the fasted state, and increased skeletal muscle lipoprotein lipase when compared with controls. Oleate oxidation rates were increased in GLUT4 null skeletal muscle in association with mitochondrial hyperplasia/hypertrophy. This study demonstrated that GLUT4 null mice had increased hepatic glucose uptake and conversion into triglyceride for subsequent use by muscle. The ability of GLUT4 null mice to alter hepatic carbohydrate and lipid metabolism to provide proper nutrients for peripheral tissues may explain (in part) their ability to resist diabetes when fed a normal diet.
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Affiliation(s)
- Mollie Ranalletta
- Department of Biochemistry, Albert Einstein College of Medicine, 1300 Morris Park Ave., Bronx, NY 10462, USA
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18
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Abstract
The molecular mechanism of how hepatocytes maintain cholesterol homeostasis has become much more transparent with the discovery of sterol regulatory element binding proteins (SREBPs) in recent years. These membrane proteins are members of the basic helix-loop-helix-leucine zipper (bHLH-Zip) family of transcription factors. They activate the expression of at least 30 genes involved in the synthesis of cholesterol and lipids. SREBPs are synthesized as precursor proteins in the endoplasmic reticulum (ER), where they form a complex with another protein, SREBP cleavage activating protein (SCAP). The SCAP molecule contains a sterol sensory domain. In the presence of high cellular sterol concentrations SCAP confines SREBP to the ER. With low cellular concentrations, SCAP escorts SREBP to activation in the Golgi. There, SREBP undergoes two proteolytic cleavage steps to release the mature, biologically active transcription factor, nuclear SREBP (nSREBP). nSREBP translocates to the nucleus and binds to sterol response elements (SRE) in the promoter/enhancer regions of target genes. Additional transcription factors are required to activate transcription of these genes. Three different SREBPs are known, SREBPs-1a, -1c and -2. SREBP-1a and -1c are isoforms produced from a single gene by alternate splicing. SREBP-2 is encoded by a different gene and does not display any isoforms. It appears that SREBPs alone, in the sequence described above, can exert complete control over cholesterol synthesis, whereas many additional factors (hormones, cytokines, etc.) are required for complete control of lipid metabolism. Medicinal manipulation of the SREBP/SCAP system is expected to prove highly beneficial in the management of cholesterol-related disease.
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Affiliation(s)
- Lutz-W Weber
- Institute of Toxicology, GSF-National Research Center for Environment and Health, Munich, D-85758 Neuherberg, Germany.
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19
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Matsuzaka T, Shimano H, Yahagi N, Amemiya-Kudo M, Okazaki H, Tamura Y, Iizuka Y, Ohashi K, Tomita S, Sekiya M, Hasty A, Nakagawa Y, Sone H, Toyoshima H, Ishibashi S, Osuga JI, Yamada N. Insulin-independent induction of sterol regulatory element-binding protein-1c expression in the livers of streptozotocin-treated mice. Diabetes 2004; 53:560-9. [PMID: 14988238 DOI: 10.2337/diabetes.53.3.560] [Citation(s) in RCA: 140] [Impact Index Per Article: 6.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/17/2022]
Abstract
Insulin and glucose together have been previously shown to regulate hepatic sterol regulatory element-binding protein (SREBP)-1c expression. We sought to explore the nutritional regulation of lipogenesis through SREBP-1c induction in a setting where effects of sugars versus insulin could be distinguished. To do so, mice were insulin depleted by streptozotocin (STZ) administration and subjected to a fasting-refeeding protocol with glucose, fructose, or sucrose. Unexpectedly, the insulin-depleted mice exhibited a marked induction of SREBP-1c on all sugars, and this increase in SREBP-1c was even more dramatic than in the non-STZ-administered controls. The time course of changes in SREBP-1 induction varied depending on the type of sugars in both control and STZ-administered mice. Glucose refeeding gave a peak of SREBP-1c induction, whereas fructose refeeding caused slow and gradual increments, and sucrose refeeding fell between these two responses. Expression of various lipogenic enzymes were also gradually increased over time, irrespective of the types of sugars, with greater intensities in STZ-administered than in nontreated mice. In contrast, induction of hepatic glucokinase and suppression of phoshoenolpyruvate carboxykinase were insulin dependent in an early refed state. These data clearly demonstrate that nutritional regulation of SREBP-1c and lipogenic genes may be completely independent of insulin as long as sufficient carbohydrates are available.
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Affiliation(s)
- Takashi Matsuzaka
- Department of Internal Medicine, Institute of Clinical Medicine, University of Tsukuba, Ibaraki, Japan
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20
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Dentin R, Pégorier JP, Benhamed F, Foufelle F, Ferré P, Fauveau V, Magnuson MA, Girard J, Postic C. Hepatic glucokinase is required for the synergistic action of ChREBP and SREBP-1c on glycolytic and lipogenic gene expression. J Biol Chem 2004; 279:20314-26. [PMID: 14985368 DOI: 10.1074/jbc.m312475200] [Citation(s) in RCA: 344] [Impact Index Per Article: 16.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/12/2023] Open
Abstract
Hepatic glucokinase (GK) catalyzes the phosphorylation of glucose to glucose 6-phosphate (G6P), a step which is essential for glucose metabolism in liver as well as for the induction of glycolytic and lipogenic genes. The sterol regulatory element-binding protein-1c (SREBP-1c) has emerged as a major mediator of insulin action on hepatic gene expression, but the extent to which its transcriptional effect is caused by an increased glucose metabolism remains unclear. Through the use of hepatic GK knockout mice (hGK-KO) we have shown that the acute stimulation by glucose of l-pyruvate kinase (l-PK), fatty acid synthase (FAS), acetyl-CoA carboxylase (ACC), and Spot 14 genes requires GK expression. To determine whether the effect of SREBP-1c requires GK expression and subsequent glucose metabolism, a transcriptionally active form of SREBP-1c was overexpressed both in vivo and in primary cultures of control and hGK-KO hepatocytes. Our results demonstrate that the synergistic action of SREBP-1c and glucose metabolism via GK is necessary for the maximal induction of l-PK, ACC, FAS, and Spot 14 gene expression. Indeed, in hGK-KO hepatocytes overexpressing SREBP-1c, the effect of glucose on glycolytic and lipogenic genes is lost because of the impaired ability of these hepatocytes to efficiently metabolize glucose, despite a marked increase in low K(m) hexokinase activity. Our studies also reveal that the loss of glucose effect observed in hGK-KO hepatocytes is associated with a decreased in the carbohydrate responsive element-binding protein (ChREBP) gene expression, a transcription factor suggested to mediate glucose signaling in liver. Decreased ChREBP gene expression, achieved using small interfering RNA, results in a loss of glucose effect on endogenous glycolytic (l-PK) and lipogenic (FAS, ACC) gene expression, thereby demonstrating the direct implication of ChREBP in glucose action. Together these results support a model whereby both SREBP-1c and glucose metabolism, acting via ChREBP, are necessary for the dietary induction of glycolytic and lipogenic gene expression in liver.
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Affiliation(s)
- Renaud Dentin
- Département d'Endocrinologie, Institut Cochin, INSERM U567, CNRS UMR8104, Université René Descartes, 24 rue du Faubourg Saint Jacques, 75014 Paris, France
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21
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Tollet-Egnell P, Parini P, Ståhlberg N, Lönnstedt I, Lee NH, Rudling M, Flores-Morales A, Norstedt G. Growth hormone-mediated alteration of fuel metabolism in the aged rat as determined from transcript profiles. Physiol Genomics 2004; 16:261-7. [PMID: 14612592 DOI: 10.1152/physiolgenomics.00093.2002] [Citation(s) in RCA: 36] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022] Open
Abstract
Age-related changes in body composition and serum lipids resemble symptoms of adult-onset growth hormone (GH) deficiency. GH treatment has been shown to normalize these changes in both GH-deficient adult patients and elderly subjects. The aim of this study was to identify GH-responsive genes that might mediate positive effects of GH treatment on fuel metabolism and body composition. cDNA microarrays were used to analyze age- and GH-induced changes in gene expression patterns in male rats. Tissues analyzed were liver, adipose tissue, and skeletal muscle from animals on or off GH treatment. A value of 1.5 was chosen to denote differences (increased or decreased expression) in the level of mRNA expression. In the liver, 7.3% of the expressed genes were affected by age and 6.5% by GH. Similar values for the other tissues were 8.3% and 5.3% (fat), and 7.9% and 9.6% (muscle), respectively. Among the differentially expressed genes, we identified several that encode proteins involved in fuel metabolism. Old rats were shown to have induced expression of genes involved in hepatic glucose oxidation and lipid synthesis, whereas these pathways were reduced in adipose tissue. GH treatment induced the expression of genes for lipid oxidation in liver and for glucose oxidation in skeletal muscle. In adipose tissue, GH reduced the expression of genes involved in lipogenesis even further. Changes in transcript levels were reflected in serum in terms of altered lipid profiles. Serum levels of triglycerides, high-density lipoprotein (HDL) cholesterol, and total cholesterol were higher in the old animals than in the young and normalized by GH treatment.
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Affiliation(s)
- Petra Tollet-Egnell
- Department of Molecular Medicine, Karolinska Institutet, Karolinska Hospital, 171 76 Stockholm, Sweden
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22
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Leclerc I, da Silva Xavier G, Rutter GA. AMP- and stress-activated protein kinases: key regulators of glucose-dependent gene transcription in mammalian cells? PROGRESS IN NUCLEIC ACID RESEARCH AND MOLECULAR BIOLOGY 2003; 71:69-90. [PMID: 12102561 DOI: 10.1016/s0079-6603(02)71041-6] [Citation(s) in RCA: 13] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/31/2023]
Abstract
This article will discuss the role of two classes of serine/threonine protein kinases in the regulation of gene transcription in mammals. The first is AMP-activated protein kinase (AMPK), which is responsive to changes in the intracellular energy status. The second is the 'stress-activated" family of protein kinases, members of the mitogen-activated protein (MAP) kinase superfamily, whose regulation by a number of extracellular agents (including osmotic stresses, cytokines, and heat) is less well understood. Interest in these enzymes has grown in the past few years due to mounting evidence (both pharmacological and genetic) which has implicated them in the regulation of a number genes important in mammalian metabolism.
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Affiliation(s)
- Isabelle Leclerc
- Department of Biochemistry, School of Medical Sciences, University of Bristol, United Kingdom
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23
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Shimano H. Sterol regulatory element-binding protein family as global regulators of lipid synthetic genes in energy metabolism. VITAMINS AND HORMONES 2003; 65:167-94. [PMID: 12481547 DOI: 10.1016/s0083-6729(02)65064-2] [Citation(s) in RCA: 102] [Impact Index Per Article: 4.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Abstract
Sterol regulatory element-binding proteins (SREBPs) have been established as lipid synthetic transcription factors for cholesterol and fatty acid synthesis. SREBPs are synthesized as membrane-bound precursors with their N-terminal active portions entering the nucleus to activate target genes after proteolytic cleavage in a sterol-regulated manner. This cleavage step is regulated by a putative sterol-sensing molecule, SREBP-activating protein (SCAP), that forms a complex with SREBPs and traffics between the rough endoplasmic reticulum and Golgi. DNA cis-elements that SREBPs bind, originally identified as sterol-regulatory elements (SREs), now expands to a variety of SRE-like sequences and some of E-boxes, which makes SREBPs eligible to regulate a wide range of lipid genes. Animal experiments including transgenic and knockout mice suggest that three isoforms, SREBP-1a, -1c, and -2, have different roles in lipid synthesis. In differentiated tissues and organs, SREBP-1c is involved in fatty acid, whereas SREBP-2 plays a major role in regulation of cholesterol synthesis. SREBP-1a is expressed in growing cells, providing both cholesterol and fatty acids that are required for membrane synthesis. SREBP-1c seems to be a mediator for insulin/glucose signaling to lipogenesis, and could be involved in insulin resistance, remnant lipoproteins, and fatty livers. Future studies in this field will certainly focus on understanding the molecular mechanisms sensing cellular sterol and energy states leading to the activation of SREBP-mediated gene transcription.
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Affiliation(s)
- Hitoshi Shimano
- Department of Internal Medicine, Institute of Clinical Medicine, University of Tsukuba, Tsukuba, Ibaraki 305-8575, Japan
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24
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Foufelle F, Ferré P. New perspectives in the regulation of hepatic glycolytic and lipogenic genes by insulin and glucose: a role for the transcription factor sterol regulatory element binding protein-1c. Biochem J 2002; 366:377-91. [PMID: 12061893 PMCID: PMC1222807 DOI: 10.1042/bj20020430] [Citation(s) in RCA: 328] [Impact Index Per Article: 14.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/18/2002] [Revised: 05/27/2002] [Accepted: 06/13/2002] [Indexed: 02/07/2023]
Abstract
The regulation of hepatic glucose metabolism has a key role in whole-body energy metabolism, since the liver is able to store (glycogen synthesis, lipogenesis) and to produce (glycogenolysis, gluconeogenesis) glucose. These pathways are regulated at several levels, including a transcriptional level, since many of the metabolism-related genes are expressed according to the quantity and quality of nutrients. Recent advances have been made in the understanding of the regulation of hepatic glycolytic, lipogenic and gluconeogenic gene expression by pancreatic hormones, insulin and glucagon and glucose. Here we review the role of the transcription factors forkhead and sterol regulatory element binding protein-1c in the inductive and repressive effects of insulin on hepatic gene expression, and the pathway that leads from glucose to gene regulation with the recently discovered carbohydrate response element binding protein. We discuss how these transcription factors are integrated in a regulatory network that allows a fine tuning of hepatic glucose storage or production, and their potential importance in metabolic diseases.
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Affiliation(s)
- Fabienne Foufelle
- INSERM Unit 465, Centre de Recherches Biomédicales des Cordeliers, 15 rue de l'Ecole de Médecine, 75270 Paris Cedex 06, France.
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25
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Uyeda K, Yamashita H, Kawaguchi T. Carbohydrate responsive element-binding protein (ChREBP): a key regulator of glucose metabolism and fat storage. Biochem Pharmacol 2002; 63:2075-80. [PMID: 12110366 DOI: 10.1016/s0006-2952(02)01012-2] [Citation(s) in RCA: 148] [Impact Index Per Article: 6.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/25/2022]
Abstract
Feeding a high carbohydrate diet induces transcription of more than 15 genes involved in the metabolic conversion of glucose to fat. A new transcription factor binding to a glucose response element of the pyruvate kinase and lipogenesis enzyme genes was discovered recently. This factor, termed carbohydrate responsive element-binding protein (ChREBP), is activated in response to high glucose and up-regulates these genes. Cyclic AMP and a high fat diet inhibit ChREBP and slow down glucose utilization. ChREBP is able to control transcription of lipogenic enzyme genes in response to nutritional and hormonal inputs, and may play an important role in disease states such as diabetes, obesity, and hypertension.
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Affiliation(s)
- Kosaku Uyeda
- Department of Biochemistry, Veterans Affairs Medical Center and The University of Texas Southwestern Medical Center at Dallas, 75216, USA.
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26
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McInerney M, Serrano Rodriguez G, Pawlina W, Hurt CB, Fletcher BS, Laipis PJ, Frost SC. Glycogen phosphorylase is activated in response to glucose deprivation but is not responsible for enhanced glucose transport activity in 3T3-L1 adipocytes. BIOCHIMICA ET BIOPHYSICA ACTA 2002; 1570:53-62. [PMID: 11960689 DOI: 10.1016/s0304-4165(02)00154-x] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 10/27/2022]
Abstract
We have previously shown that glucose deprivation activates glucose transport in a time- and protein synthesis-dependent fashion in 3T3-L1 adipocytes, a mouse cell line. Coincident with this is loss of glycogen. Because glycogen phosphorylase (GP) is responsible for glycogen degradation, we have examined its regulation to determine the relationship between transport activation and glycogen turnover. We first cloned the adipose GP cDNA and found sequence similarity to rat and human liver GP. Because the mouse liver GP cDNA sequence was unavailable, we cloned this cDNA as well and showed 100% identity between mouse adipose and liver sequences. A 3.1 kb transcript was readily observed in total RNA isolated from mouse liver or adipose by Northern blot analysis but, surprisingly, not in either total or poly(A) selected RNA from 3T3-L1 adipocytes. To evaluate regulation in 3T3-L1 adipocytes, we amplified GP mRNA from total RNA using multiplex, semi-quantitative PCR but found that expression did not change in response to deprivation. GP protein levels did not change either. However, endogenous GP activity from glucose-deprived cells was significantly elevated relative to controls, due to an increase in the phosphorylated form of GP (GPa). Finally, we overexpressed GP to determine its direct influence on the glucose transport system. These results were negative, which suggests that the nutrient control of glucose transport and GP occurs independently despite kinetic similarities in transport activation and glycogen turnover.
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Affiliation(s)
- Melissa McInerney
- Department of Biochemistry and Molecular Biology, University of Florida College of Medicine, Box 100245, 1600 SW Archer Road, Gainesville, FL 32610, USA
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27
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Bandsma RH, Wiegman CH, Herling AW, Burger HJ, ter Harmsel A, Meijer AJ, Romijn JA, Reijngoud DJ, Kuipers F. Acute inhibition of glucose-6-phosphate translocator activity leads to increased de novo lipogenesis and development of hepatic steatosis without affecting VLDL production in rats. Diabetes 2001; 50:2591-7. [PMID: 11679439 DOI: 10.2337/diabetes.50.11.2591] [Citation(s) in RCA: 51] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/27/2023]
Abstract
Glucose-6-phosphatase (G6Pase) is a key enzyme in hepatic glucose metabolism. Altered G6Pase activity in glycogen storage disease and diabetic states is associated with disturbances in lipid metabolism. We studied the effects of acute inhibition of G6Pase activity on hepatic lipid metabolism in nonanesthetized rats. Rats were infused with an inhibitor of the glucose-6-phosphate (G6P) translocator (S4048, 30 mg. kg(-1). h(-1)) for 8 h. Simultaneously, [1-(13)C]acetate was administered for determination of de novo lipogenesis and fractional cholesterol synthesis rates by mass isotopomer distribution analysis. In a separate group of rats, Triton WR 1339 was injected for determination of hepatic VLDL-triglyceride production. S4048 infusion significantly decreased plasma glucose (-11%) and insulin (-48%) levels and increased hepatic G6P (201%) and glycogen (182%) contents. Hepatic triglyceride contents increased from 5.8 +/- 1.4 micromol/g liver in controls to 20.6 +/- 5.5 micromol/g liver in S4048-treated animals. De novo lipogenesis was increased >10-fold in S4048-treated rats, without changes in cholesterol synthesis rates. Hepatic mRNA levels of acetyl-CoA carboxylase and fatty acid synthase were markedly induced. Plasma triglyceride levels increased fourfold, but no differences in plasma cholesterol levels were seen. Surprisingly, hepatic VLDL-triglyceride secretion was not increased in S4048-treated rats. These studies demonstrate that inhibition of the G6Pase system leads to acute stimulation of fat synthesis and development of hepatic steatosis, without affecting hepatic cholesterol synthesis and VLDL secretion. The results emphasize the strong interactions that exist between hepatic carbohydrate and fat metabolism.
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Affiliation(s)
- R H Bandsma
- Groningen University Institute for Drug Exploration, Center for Liver, Digestive and Metabolic Diseases, Department of Pediatrics, Academic Hospital Groningen, Groningen, the Netherlands.
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Shimano H. Sterol regulatory element-binding proteins (SREBPs): transcriptional regulators of lipid synthetic genes. Prog Lipid Res 2001; 40:439-52. [PMID: 11591434 DOI: 10.1016/s0163-7827(01)00010-8] [Citation(s) in RCA: 568] [Impact Index Per Article: 23.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/30/2022]
Abstract
Roles of sterol regulatory element-binding proteins (SREBPs) have been established as lipid synthetic transcription factors especially for cholesterol and fatty acid synthesis. SREBPs have unique characteristics. Firstly, they are membrane-bound proteins and the N-terminal active portions enter nucleus to activate their target genes after proteolytic cleavage, which requires sterol-sensing molecule, SREBP-activating protein (SCAP) and is crucial for sterol-regulation. Secondly, they bind and activate sterol-regulatory (SREs) containing promoters as well as some E-boxes, which makes SREBPs eligible to regulate a wide range of lipid genes. Finally, three isoforms, SREBP-1a-1c, and have different roles in lipid synthesis. In vivo studies using transgenic and knockout mice suggest that SREBP-1 seems to be involved in energy metabolism including fatty acid and glucose/insulin metabolism, whereas SREBP-2 is specific to cholesterol synthesis. Future studies will be focused on understanding molecular mechanisms sensing cellular sterol and energy states where SREBPs are deeply involved.
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Affiliation(s)
- H Shimano
- Department of Internal Medicine, Institute of Clinical Medicine, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8575, Japan.
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29
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Massillon D. Regulation of the glucose-6-phosphatase gene by glucose occurs by transcriptional and post-transcriptional mechanisms. Differential effect of glucose and xylitol. J Biol Chem 2001; 276:4055-62. [PMID: 11087741 DOI: 10.1074/jbc.m007939200] [Citation(s) in RCA: 45] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
To understand how glucose regulates the expression of the glucose-6-phosphatase gene, the effect of glucose was studied in primary cultures of rat hepatocytes. Glucose-6-phosphatase mRNA levels increased about 10-fold when hepatocytes were incubated with 20 mm glucose. The rate of transcription of the glucose-6-phosphatase gene increased about 3-fold in hepatocytes incubated with glucose. The half-life of glucose-6-phosphatase mRNA was estimated to be 90 min in the absence of glucose and 3 h in its presence. Inhibition of the oxidative and the nonoxidative branches of the pentose phosphate pathway blocked the stimulation of glucose-6-phosphatase expression by glucose but not by xylitol or carbohydrates that enter the glycolytic/gluconeogenic pathways at the level of the triose phosphates. These results indicate that (i) the glucose induction of the mRNA for the catalytic unit of glucose-6-phosphatase occurs by transcriptional and post-transcriptional mechanisms and that (ii) xylitol and glucose increase the expression of this gene through different signaling pathways.
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Affiliation(s)
- D Massillon
- Department of Nutrition, Case Western Reserve University School of Medicine, Cleveland, Ohio 44106, USA
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30
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Affiliation(s)
- S Vaulont
- Institut Cochin de Génétique Moléculaire, U.129 INSERM, Université René Descartes, 75014 Paris, France.
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31
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Hasty AH, Shimano H, Yahagi N, Amemiya-Kudo M, Perrey S, Yoshikawa T, Osuga J, Okazaki H, Tamura Y, Iizuka Y, Shionoiri F, Ohashi K, Harada K, Gotoda T, Nagai R, Ishibashi S, Yamada N. Sterol regulatory element-binding protein-1 is regulated by glucose at the transcriptional level. J Biol Chem 2000; 275:31069-77. [PMID: 10913129 DOI: 10.1074/jbc.m003335200] [Citation(s) in RCA: 114] [Impact Index Per Article: 4.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
In vivo studies suggest that sterol regulatory element-binding protein (SREBP)-1 plays a key role in the up-regulation of lipogenic genes in the livers of animals that have consumed excess amounts of carbohydrates. In light of this, we sought to use an established mouse hepatocyte cell line, H2-35, to further define the mechanism by which glucose regulates nuclear SREBP-1 levels. First, we show that these cells transcribe high levels of SREBP-1c that are increased 4-fold upon differentiation from a prehepatocyte to a hepatocyte phenotype, making them an ideal cell culture model for the study of SREBP-1c induction. Second, we demonstrate that the presence of precursor and mature forms of SREBP-1 protein are positively regulated by medium glucose concentrations ranging from 5. 5 to 25 mm and are also regulated by insulin, with the amount of insulin in the fetal bovine serum being sufficient for maximal stimulation of SREBP-1 expression. Third, we show that the increase in SREBP-1 protein is due to an increase in SREBP-1 mRNA. Reporter gene analysis of the SREBP-1c promoter demonstrated a glucose-dependent induction of transcription. In contrast, expression of a fixed amount of the precursor form of SREBP-1c protein showed that glucose does not influence its cleavage. Fourth, we demonstrate that the glucose induction of SREBP could not be reproduced by fructose, xylose, or galactose nor by glucose analogs 2-deoxy glucose and 3-O-methyl glucopyranose. These data provide strong evidence for the induction of SREBP-1c mRNA by glucose leading to increased mature protein in the nucleus, thus providing a potential mechanism for the up-regulation of lipogenic genes by glucose in vivo.
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Affiliation(s)
- A H Hasty
- Department of Metabolic Diseasese, University of Tokyo, Tokyo 113-8655, Japan
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32
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Woods A, Azzout-Marniche D, Foretz M, Stein SC, Lemarchand P, Ferré P, Foufelle F, Carling D. Characterization of the role of AMP-activated protein kinase in the regulation of glucose-activated gene expression using constitutively active and dominant negative forms of the kinase. Mol Cell Biol 2000; 20:6704-11. [PMID: 10958668 PMCID: PMC86183 DOI: 10.1128/mcb.20.18.6704-6711.2000] [Citation(s) in RCA: 325] [Impact Index Per Article: 13.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/15/2022] Open
Abstract
In the liver, glucose induces the expression of a number of genes involved in glucose and lipid metabolism, e.g., those encoding L-type pyruvate kinase and fatty acid synthase. Recent evidence has indicated a role for the AMP-activated protein kinase (AMPK) in the inhibition of glucose-activated gene expression in hepatocytes. It remains unclear, however, whether AMPK is involved in the glucose induction of these genes. In order to study further the role of AMPK in regulating gene expression, we have generated two mutant forms of AMPK. One of these (alpha1(312)) acts as a constitutively active kinase, while the other (alpha1DN) acts as a dominant negative inhibitor of endogenous AMPK. We have used adenovirus-mediated gene transfer to express these mutants in primary rat hepatocytes in culture in order to determine their effect on AMPK activity and the transcription of glucose-activated genes. Expression of alpha1(312) increased AMPK activity in hepatocytes and blocked completely the induction of a number of glucose-activated genes in response to 25 mM glucose. This effect is similar to that observed following activation of AMPK by 5-amino-imidazolecarboxamide riboside. Expression of alpha1DN markedly inhibited both basal and stimulated activity of endogenous AMPK but had no effect on the transcription of glucose-activated genes. Our results suggest that AMPK is involved in the inhibition of glucose-activated gene expression but not in the induction pathway. This study demonstrates that the two mutants we have described will provide valuable tools for studying the wider physiological role of AMPK.
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Affiliation(s)
- A Woods
- Cellular Stress Group, MRC Clinical Sciences Centre, Imperial College School of Medicine, Hammersmith Hospital, London, United Kingdom
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33
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Sun Q, Sekar N, Goldwaser I, Gershonov E, Fridkin M, Shechter Y. Vanadate restores glucose 6-phosphate in diabetic rats: a mechanism to enhance glucose metabolism. Am J Physiol Endocrinol Metab 2000; 279:E403-10. [PMID: 10913041 DOI: 10.1152/ajpendo.2000.279.2.e403] [Citation(s) in RCA: 15] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 11/22/2022]
Abstract
Vanadate mimics the metabolic actions of insulin. In diabetic rodents, vanadate also sensitizes peripheral tissues to insulin. We have analyzed whether this latter effect is brought about by a mechanism other than the known insulinomimetic actions of vanadium in vitro. We report that the levels of glucose 6-phosphate (G-6-P) in adipose, liver, and muscle of streptozotocin-treated (STZ)-hyperglycemic rats are 77, 50, and 58% of those in healthy control rats, respectively. Normoglycemia was induced by vanadium or insulin therapy or by phlorizin. Vanadate fully restored G-6-P in all three insulin-responsive peripheral tissues. Insulin did not restore G-6-P in muscle, and phlorizin was ineffective in adipose and muscle. Incubation of diabetic adipose explants with glucose and vanadate in vitro increased lipogenic capacity three- to fourfold (half-maximally effective dose = 11 +/- 1 microM vanadate). Lipogenic capacity was elevated when a threshold level of approximately 7.5 +/- 0.3 nmol G-6-P/g tissue was reached. In summary, 1) chronic hyperglycemia largely reduces intracellular G-6-P in all three insulin-responsive tissues; 2) vanadate therapy restores this deficiency, but insulin therapy does not restore G-6-P in muscle tissue; 3) induction of normoglycemia per se (i.e., by phlorizin) restores G-6-P in liver only; and 4) glucose and vanadate together elevate G-6-P in adipose explants in vitro and significantly restore lipogenic capacity above the threshold of G-6-P level. We propose that hyperglycemia-associated decrease in peripheral G-6-P is a major factor responsible for peripheral resistance to insulin. The mechanism by which vanadate increases peripheral tissue capacity to metabolize glucose and to respond to the hormone involves elevation of this hexose phosphate metabolite and the cellular consequences of this elevated level of G-6-P.
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Affiliation(s)
- Q Sun
- Departments of Biological Chemistry, The Weizmann Institute of Science, Rehovot 76100, Israel
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34
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Bell ME, Bhatnagar S, Liang J, Soriano L, Nagy TR, Dallman MF. Voluntary sucrose ingestion, like corticosterone replacement, prevents the metabolic deficits of adrenalectomy. J Neuroendocrinol 2000; 12:461-70. [PMID: 10792586 DOI: 10.1046/j.1365-2826.2000.00488.x] [Citation(s) in RCA: 80] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 11/20/2022]
Abstract
We tested whether corticosterone replacement causes increased sucrose drinking in adrenalectomized (ADX) rats compared to sham-ADX (sham) rats. ADX rats given high doses of corticosterone drank as much sucrose as sham rats, whereas at three lower doses of corticosterone, drinking was similar between groups and was only approximately 40% of that ingested by shams. Compared to sham rats, ADX rats drinking saline, or saline and saccharin, gain weight more slowly, contain less white adipose tissue, and have higher sympathetic outflow as assessed by uncoupling protein content in brown adipose tissue. Allowing sucrose as well as saline to drink restored all of these variables to normal in ADX rats with no- or low-corticosterone. All endpoints from sucrose-drinking ADX rats with no-or low-corticosterone were indistinguishable from those in water-drinking shams. By contrast, sucrose-drinking ADX rats that were given high doses of corticosterone exhibited the usual catabolic effects of corticosterone on body weight gain and, unlike sucrose-drinking shams, were obese. We conclude that (i) high corticosterone stimulates the potability of sucrose and inhibits sympathetic stimulation of uncoupling protein; (ii) sucrose, without corticosterone, normalizes metabolic deficits in ADX rats probably through actions mediated both peripherally and by the central nervous system; and (iii) ADX rats have a distinct sucrose appetite.
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Affiliation(s)
- M E Bell
- Department of Physiology, University of California San Francisco, San Francisco, CA 94143-0444, USA
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35
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Wu H, MacFarlane WM, Tadayyon M, Arch JR, James RF, Docherty K. Insulin stimulates pancreatic-duodenal homoeobox factor-1 (PDX1) DNA-binding activity and insulin promoter activity in pancreatic beta cells. Biochem J 1999; 344 Pt 3:813-8. [PMID: 10585868 PMCID: PMC1220703] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/14/2023]
Abstract
Pancreatic-duodenal homoeobox factor-1 (PDX1) is a homoeodomain transcription factor that plays an important role in linking glucose metabolism in pancreatic beta cells to the regulation of insulin gene transcription. Our previous results indicated that glucose activates PDX1 DNA-binding activity and insulin promoter activity via a stress-activated signalling pathway involving phosphatidylinositol 3-kinase (PtdIns 3-kinase) and stress-activated protein kinase 2 (SAPK2/p38). The present study was undertaken to determine the effects of other metabolizable and non-metabolizable nutrients. The results indicate that non-metabolizable nutrients, with the exception of 2-deoxyglucose, had no effect. Metabolizable nutrients that could stimulate calcium uptake and insulin release were shown to activate both PDX1 and the insulin promoter. The possible role of insulin acting via an autoregulatory loop was therefore examined. Insulin was shown to potently activate PDX1 DNA-binding activity and insulin promoter activity. The effects of insulin were inhibited by the PtdIns 3-kinase inhibitors wortmannin and LY294002 and by the SAPK2 inhibitor SB203580, suggesting that its effects were mediated via activation of PtdIns 3-kinase and SAPK2. Further support for the insulin-mediated activation of SAPK2 came from the observation that both glucose and insulin stimulated the phosphorylation of SAPK2. These results suggest that both glucose and insulin stimulate PDX1 DNA-binding activity and insulin promoter activity via a pathway involving PtdIns 3-kinase and SAPK2.
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Affiliation(s)
- H Wu
- Department of Molecular Biology, University of Aberdeen, Institute of Medical Sciences, Foresterhill, Aberdeen AB25 2ZD, U.K
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36
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Cournarie F, Azzout-Marniche D, Foretz M, Guichard C, Ferre P, Foufelle F. The inhibitory effect of glucose on phosphoenolpyruvate carboxykinase gene expression in cultured hepatocytes is transcriptional and requires glucose metabolism. FEBS Lett 1999; 460:527-32. [PMID: 10556529 DOI: 10.1016/s0014-5793(99)01407-6] [Citation(s) in RCA: 27] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022]
Abstract
Phosphoenolpyruvate carboxykinase (PEPCK) is the rate-limiting enzyme of gluconeogenesis in the liver. PEPCK gene expression is controlled at the transcriptional level and is mainly regulated by hormones that are involved in glucose homeostasis. In this study, we have investigated the role of glucose on PEPCK gene expression in cultured hepatocytes. We demonstrate that glucose counteracts the stimulatory effect of glucocorticoids and cAMP on PEPCK expression. Glucose must be metabolized through glucokinase to have its inhibitory effect. The effect of glucose is mainly transcriptional and the region responsible for glucose inhibition is localized in the first 490 bp of the promoter.
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Affiliation(s)
- F Cournarie
- U465 INSERM, Institut Biomédical des Cordeliers (Université Paris 6), 15 rue de l'Ecole de Médecine, F-75270, Paris, France
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37
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Abstract
Fatty acid synthase (EC 2.3.1.85) is an enzyme involved in the lipogenic pathway allowing fatty acid synthesis from glucose. Glucose up-regulates the transcription of the fatty acid synthase gene in both adipocytes and hepatocytes, with insulin having only an indirect role. The signal metabolite could be glucose-6-phosphate rather than glucose itself. The glucose response element of the fatty acid synthase gene has not yet been precisely identified, although a -2 kb region of the fatty acid synthase promoter is sufficient to confer nutritional responsiveness to a reporter gene. ADD1/SREBP1, a b-HLH-LZ transcription factor belonging to the sterol regulatory element-binding protein family might be involved in the transduction of the glucose effect. Finally, the stimulatory effect of glucose on the expression of the fatty acid synthase gene is inhibited by the activation of AMP-activated protein kinase. Interestingly enough, AMP-activated protein kinase is structurally and functionally related to the yeast SNF1 protein kinase complex which is essential for the transcriptional activation of glucose-repressed genes in Saccharomyces cerevisiae.
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Affiliation(s)
- P Ferré
- U465 INSERM, Centre Biomédical des Cordeliers, Paris, France.
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38
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Foretz M, Foufelle F, Ferré P. Polyunsaturated fatty acids inhibit fatty acid synthase and spot-14-protein gene expression in cultured rat hepatocytes by a peroxidative mechanism. Biochem J 1999; 341 ( Pt 2):371-6. [PMID: 10393095 PMCID: PMC1220369] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/13/2023]
Abstract
In vivo, polyunsaturated fatty acids (PUFA) inhibit the expression of hepatic genes related to the lipogenic process such as fatty acid synthase and spot-14-protein (S14) genes. In vitro studies have suggested that this was a direct transcriptional effect of PUFA. In hepatocytes, the inhibition of the lipogenic rate by PUFA is not specific, but is linked to a cytotoxic effect due to peroxidative mechanisms. We have investigated whether peroxidation could also explain the inhibitory effect of PUFA on gene expression. Rat hepatocytes were cultured for 24 h with mono-unsaturated or PUFA. PUFA inhibited the expression of fatty acid synthase and S14 genes, and this inhibition was directly related to the number of unsaturations. However, the beta-actin and albumin mRNA concentrations were also affected by the most unsaturated fatty acids, suggesting a non-specific effect of PUFA on gene expression. Measurement of lactate dehydrogenase released into the medium indicated a cytotoxicity of PUFA. This was associated with their peroxidation as evaluated by the presence of thiobarbituric acid-reactive substances in the culture medium. The addition of high concentrations of antioxidants abolished lipid peroxidation and lactate dehydrogenase leakage and completely reversed the inhibitory effect of PUFA on gene expression. This suggests (i) that the results obtained previously in cultured hepatocytes in the presence of low concentrations of antioxidants must be interpretated cautiously and (ii) that in vivo, the inhibitory effect of PUFA on lipogenesis-related genes could be indirect through hormonal or metabolic changes or that their effect on gene expression is somehow linked to peroxidative mechanisms.
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Affiliation(s)
- M Foretz
- INSERM Unit 465, Centre Biomédical des Cordeliers, 15 rue de l'Ecole de Médecine, F-75270 Paris Cédex 06, France
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39
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Wang Y, Lee-Kwon W, Martindale JL, Adams L, Heller P, Egan JM, Bernier M. Modulation of CCAAT/enhancer-binding protein-alpha gene expression by metabolic signals in rodent adipocytes. Endocrinology 1999; 140:2938-47. [PMID: 10385384 DOI: 10.1210/endo.140.7.6793] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/22/2023]
Abstract
The transcription factor CCAAT/enhancer-binding protein-alpha (C/EBPalpha) is a positive modulator of transcription for several adipocyte-specific genes that play a role in energy metabolism. However, there is little information available regarding the regulation of its expression by metabolic signals. Exposure to insulin for 5-24 h attenuated C/EBPalpha expression when 3T3-L1 adipocytes were incubated in 24 mM glucose, but not in 5.7 mM glucose. Nuclear run-on transcription assays indicated a transcriptional repression of C/EBPalpha gene, but not that of C/EBPbeta. Glucosamine, a product of the hexosamine pathway, in the presence of low glucose mimicked high glucose's ability to reduce C/EBPalpha messenger RNA expression in insulin-treated cells. Similar results were obtained with xylitol, an activator of the pentose phosphate pathway. There was no correlation between the accumulation of hexosamine pathway metabolites (e.g. UDP-N-acetylhexosamines) and/or changes in intracellular protein glycosylation with the ability of high glucose, glucosamine, or xylitol to down-regulate C/EBPalpha gene expression. None of these treatments caused a reduction in intracellular ATP levels. Stable transfection of 3T3-L1 cells with the 5'-flanking 468-bp sequence of the mouse C/EBPalpha gene fused to luciferase demonstrated that promoter activity was also reduced by these nutrients. Of interest, treatment of rats with glucose or glucosamine led to a reduction in C/EBPalpha messenger RNA levels in epididymal, but not omental, fat. Taken together, these results suggest that metabolic signals serve to down-regulate C/EBPalpha expression both in vitro and in vivo.
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Affiliation(s)
- Y Wang
- Laboratory of Clinical Investigation, National Institute on Aging, National Institutes of Health, Baltimore, Maryland 21224-6825, USA
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40
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Abstract
The transition of nonfailing to failing cardiac hypertrophy cannot be prevented by current drug regimens. This investigation examined whether possible drug targets have remained unexplored because they do not result in acute improvement of heart function. Of major importance, in this respect, is an inadequate performance of the sarco(endo)plasmic reticulum Ca(2+)-ATPase (SERCA2). In the present approach, binding sequences within the proximal promoter of SERCA2 are described which may be useful in the development of drugs (i.e., transcriptional modulators) that interfere selectively with the transcription of genes of the cardiomyocyte. The proximal promoter region of the SERCA2 genes has a thyroid response element, 9 potential Sp1-binding sites (5'-GGGCGG-3', 5'-CCGCCC-3' and 5'-GGGAGG-3'), and an E-box motif (5'-CACATG-3'), which may function as glucose response elements. This region also has 2 putative fatty-acid response elements (5'-GGGGGA-3'). It is proposed that the beneficial effects of the camitine palmitoyltransferase-1 inhibitor etomoxir arise from a shift in fuel metabolism involving glucose response elements and/or peroxisomal proliferator-activated receptors. Although the relative contribution of these DNA regulatory elements remains to be defined, it appears that they provide the driving force that prevents the decrease in transcriptional activity of the SERCA2 gene in the hypertrophic heart. It is further concluded that etomoxir represents a member of a novel class of transcriptional modulators that improve function of hypertrophied hearts with unimpeded blood flow by modulating gene expression of the cardiomyocyte.
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Affiliation(s)
- A Zarain-Herzberg
- Departamento de Bioquímica, Facultad de Medicina, Universidad Nacional Autónoma de México, Mexico City
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41
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Foretz M, Pacot C, Dugail I, Lemarchand P, Guichard C, Le Lièpvre X, Berthelier-Lubrano C, Spiegelman B, Kim JB, Ferré P, Foufelle F. ADD1/SREBP-1c is required in the activation of hepatic lipogenic gene expression by glucose. Mol Cell Biol 1999; 19:3760-8. [PMID: 10207099 PMCID: PMC84202 DOI: 10.1128/mcb.19.5.3760] [Citation(s) in RCA: 414] [Impact Index Per Article: 15.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
The transcription of genes encoding proteins involved in the hepatic synthesis of lipids from glucose is strongly stimulated by carbohydrate feeding. It is now well established that in the liver, glucose is the main activator of the expression of this group of genes, with insulin having only a permissive role. While ADD1/SREBP-1 has been implicated in lipogenic gene expression through temporal association with food intake and ectopic gain-of-function experiments, no genetic evidence for a requirement for this factor in glucose-mediated gene expression has been established. We show here that the transcription of ADD1/SREBP-1c in primary cultures of hepatocytes is controlled positively by insulin and negatively by glucagon and cyclic AMP, establishing a link between this transcription factor and carbohydrate availability. Using adenovirus-mediated transfection of a powerful dominant negative form of ADD1/SREBP-1c in rat hepatocytes, we demonstrate that this factor is absolutely necessary for the stimulation by glucose of L-pyruvate kinase, fatty acid synthase, S14, and acetyl coenzyme A carboxylase gene expression. These results demonstrate that ADD1/SREBP-1c plays a crucial role in mediating the expression of lipogenic genes induced by glucose and insulin.
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Affiliation(s)
- M Foretz
- U465 INSERM, Institut Biomédical des Cordeliers, 75270 Paris Cedex 06, France
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42
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Yamada K, Tanaka T, Noguchi T. Characterization and purification of carbohydrate response element-binding protein of the rat L-type pyruvate kinase gene promoter. Biochem Biophys Res Commun 1999; 257:44-9. [PMID: 10092507 DOI: 10.1006/bbrc.1999.0410] [Citation(s) in RCA: 16] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022]
Abstract
The L-III transcriptional regulatory element of the rat pyruvate kinase L gene is located between -170 and -150 base pairs upstream of the hepatocyte-specific transcription initiation site. As the L-III element is not only necessary for cell type-specific expression but also for transcriptional stimulation by carbohydrates, it is also referred to as a carbohydrate-response element. Electrophoretic mobility shift assays using rat liver nuclear extract showed that L-III element-binding protein (L-IIIBP) was observed as multiple bands. These bands disappeared when the nuclear extract was preincubated at 60 degrees C for 5 min and were competed with unlabeled L-III oligonucleotide but not with unlabeled adenovirus major late promoter E box oligonucleotide. In addition, these bands were not affected in the presence of antiserum against upstream stimulating factor (USF). Thus, we conclude that L-IIIBP is different from USF. Then, heat-labile L-IIIBP was purified from rat liver nuclear extracts. Purified L-IIIBP exhibited two bands on sodium dodecyl sulfate/polyacrylamide gel electrophoresis by silver staining. Ultraviolet crosslinking experiment showed that both bands had binding activity to the L-III oligonucleotide.
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Affiliation(s)
- K Yamada
- Department of Biochemistry, Fukui Medical University, Shimoaizuki, Matsuoka, Fukui, 910-1193, Japan
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43
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Barbosa-Tessmann IP, Pineda VL, Nick HS, Schuster SM, Kilberg MS. Transcriptional regulation of the human asparagine synthetase gene by carbohydrate availability. Biochem J 1999; 339 ( Pt 1):151-8. [PMID: 10085239 PMCID: PMC1220139] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/11/2023]
Abstract
Transcription of the asparagine synthetase (AS) gene is induced by amino acid deprivation. The present data illustrate that this gene is also under transcriptional control by carbohydrate availability. Incubation of human HepG2 hepatoma cells in glucose-free medium resulted in an increased AS mRNA content, reaching a maximum of about 14-fold over control cells after approx. 12 h. Extracellular glucose caused the repression of the content of AS mRNA in a concentration-dependent manner, with a k1/2 (concentration causing a half-maximal repression) of 1 mM. Fructose, galactose, mannose, 2-deoxyglucose and xylitol were found to maintain the mRNA content of both AS and the glucose-regulated protein GRP78 in a state of repression, whereas 3-O-methylglucose did not. Incubation in either histidine-free or glucose-free medium also resulted in adaptive regulation of the AS gene in BNL-CL.2 mouse hepatocytes, rat C6 glioma cells and human MOLT4 lymphocytes, in addition to HepG2 cells. In contrast, the steady-state mRNA content of GRP78 was unaffected by amino acid availability. Transient transfection assays using a reporter gene construct documented that glucose deprivation increases AS gene transcription via elements within the proximal 3 kbp of the AS promoter. These results illustrate that human AS gene transcription is induced following glucose limitation of the cells.
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Affiliation(s)
- I P Barbosa-Tessmann
- Department of Biochemistry and Molecular Biology, University of Florida College of Medicine, Box 100245, JHMHC Gainesville, FL 32610-0245, USA
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44
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Roder K, Wolf SS, Sickinger S, Schweizer M. FIRE3 in the promoter of the rat fatty acid synthase (FAS) gene binds the ubiquitous transcription factors CBF and USF but does not mediate an insulin response in a rat hepatoma cell line. EUROPEAN JOURNAL OF BIOCHEMISTRY 1999; 260:743-51. [PMID: 10103003 DOI: 10.1046/j.1432-1327.1999.00216.x] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/20/2022]
Abstract
Several putative insulin-responsive elements (IRE) in the fatty acid synthase (FAS) promoter have been identified and shown to be functional in adipocytes and hepatocytes. Here we report on the insulin-responsiveness in the rat hepatoma cell line H4IIE of four cis-elements in the FAS promoter: the FAS insulin-responsive elements, FIRE2 and FIRE3; the inverted CCAAT element, ICE; and the insulin/glucose-binding element, designated hepatic FIRE element, hFIRE, originally identified in rat hepatocytes. Using electrophoretic mobility shift assay (EMSA) competition experiments together with supershifts and in vitro transcription/translation we show that FIRE3 (-68/-58) binds not only the upstream stimulatory factors USF-1/USF-2 but also the CCAAT-binding factor CBF, also known as the nuclear factor Y, NF-Y. The putative IRE FIRE2, which shows sequence similarity to FIRE3, is located between -267 and -249. Gel retardation experiments indicate that USF-1 and USF-2 also bind to this element, which contains an imperfect E-box motif. Using the same approach we have shown that hFIRE binds the stimulatory proteins Sp1 and Sp3 in addition to CBF. Transient transfection experiments using FAS promoter constructs deleted for FIRE2 and FIRE3 demonstrate that neither of these elements mediates the insulin response of the FAS promoter in the rat hepatoma cell line H4IIE, however, ICE at -103/-87 is responsible for mediating the effect of the insulin antagonist cAMP. The hFIRE element located at -57/-34, in spite of its role in the glucose/insulin response in primary rat hepatocytes, is apparently not involved in the insulin regulation of the rat FAS promoter in H4IIE cells. The fact that the topology of the promoters of the FAS genes in rat, human, goose and chicken is conserved regarding CBF-binding sites and USF-binding sites implies an important role for these ubiquitously expressed transcription factors in the regulation of the FAS promoter.
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Affiliation(s)
- K Roder
- Genetics and Microbiology Department, Institute of Food Research, Norwich, UK
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Casado M, Vallet VS, Kahn A, Vaulont S. Essential role in vivo of upstream stimulatory factors for a normal dietary response of the fatty acid synthase gene in the liver. J Biol Chem 1999; 274:2009-13. [PMID: 9890958 DOI: 10.1074/jbc.274.4.2009] [Citation(s) in RCA: 109] [Impact Index Per Article: 4.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
In the liver, transcription of several genes encoding lipogenic and glycolytic enzymes, in particular the gene for fatty acid synthase (FAS), is known to be stimulated by dietary carbohydrates. The molecular dissection of the FAS promoter pointed out the critical role of an E box motif, located at position -65 with respect to the start site of transcription, in mediating the glucose- and insulin-dependent regulation of the gene. Upstream stimulatory factors (USF1 and USF2) and sterol response element binding protein 1 (SREBP1) were shown to be able to interact in vitro with this E box. However, to date, the relative contributions of USFs and SREBP1 ex vivo remain controversial. To gain insight into the specific roles of these factors in vivo, we have analyzed the glucose responsiveness of hepatic FAS gene expression in USF1 and USF2 knock-out mice. In both types of mouse lines, defective in either USF1 or USF2, induction of the FAS gene by refeeding a carbohydrate-rich diet was severely delayed, whereas expression of SREBP1 was almost normal and insulin response unchanged. Therefore, USF transactivators, and especially USF1/USF2 heterodimers, seem to be essential to sustain the dietary induction of the FAS gene in the liver.
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Affiliation(s)
- M Casado
- Institut Cochin de Génétique Moléculaire, U.129 INSERM Unité de Recherches en Physiologie et Pathologie Génétiques et Moléculaires, 75014 Paris, France
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Hasegawa J, Osatomi K, Wu RF, Uyeda K. A novel factor binding to the glucose response elements of liver pyruvate kinase and fatty acid synthase genes. J Biol Chem 1999; 274:1100-7. [PMID: 9873057 DOI: 10.1074/jbc.274.2.1100] [Citation(s) in RCA: 48] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
Transcription of the liver type pyruvate kinase and lipogenesis enzyme genes is induced by high carbohydrate in liver. We have found a novel protein factor in rat liver nuclei that binds to the glucose response element (CACGTG motifs) of the pyruvate kinase gene (Liu, Z. , Thompson, K. S., and Towle, H. C. (1993) J. Biol. Chem. 268, 12787-12795) and the "insulin response element" of fatty acid synthase gene. The amounts of this DNA-binding protein, termed "glucose response element binding protein" (GRBP) in the nuclear extract, were increased in liver by a high carbohydrate diet and decreased by starvation, high fat, and high protein diet. GRBP also occurs in cytosols of liver and is dependent on carbohydrate. Both the nuclear and the cytosolic GRBP showed similar properties, except the former was more resistant to thermal inactivation than the latter. Kinetics of glucose activation of the cytosolic GRBP in a primary culture of hepatocytes indicated that a half-maximum activation was achieved after 6 h, and glucose concentration required for the maximum activation of the GRBP was approximately 12 mM. Dibutyryl-cAMP, okadaic acid, and forskolin inhibited glucose activation of both GRBP and liver pyruvate kinase transcription. These results suggested that GRBP may be a factor that recognizes the glucose response motif site and may be involved in mediating carbohydrate response of the pyruvate kinase gene.
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Affiliation(s)
- J Hasegawa
- Research and Development, Dallas Veterans Affairs Medical Center and Department of Biochemistry, University of Texas Southwestern Medical Center, Dallas, Texas 75216, USA
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Yamada K, Noguchi T. Nutrient and hormonal regulation of pyruvate kinase gene expression. Biochem J 1999; 337 ( Pt 1):1-11. [PMID: 9854017 PMCID: PMC1219928] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/09/2023]
Abstract
Mammalian pyruvate kinase (PK), a key glycolytic enzyme, has two genes named PKL and PKM, which produce the L- and R-type isoenzymes by means of alternative promoters, and the M1-and M2-types by mutually exclusive alternative splicing respectively. The expression of these genes is tissue-specific and under developmental, dietary and hormonal control. The L-type isoenzyme (L-PK) gene contains multiple regulatory elements necessary for regulation in the 5' flanking region, up to position -170. Both L-II and L-III elements are required for stimulation of L-PK gene transcription by carbohydrates such as glucose and fructose, although the L-III element is itself responsive to carbohydrates. The L-II element is also responsible for the gene regulation by polyunsaturated fatty acids. Nuclear factor-1 proteins and hepatocyte nuclear factor 4, which bind to the L-II element, may also be involved in carbohydrate and polyunsaturated fatty acid regulation of the L-PK gene respectively. However, the L-III-element-binding protein that is involved in carbohydrate regulation remains to be clarified, although involvement by an upstream stimulating factor has been proposed. Available evidence suggests that the carbohydrate signalling pathway to the L-PK gene includes a glucose metabolite, possibly glucose 6-phosphate or xylulose 5-phosphate, as well as phosphorylation and dephosphorylation mechanisms. In addition, at least five regulatory elements have been identified in the 5' flanking region of the PKM gene up to position -279. Sp1-family proteins bind to two proximal elements, but the binding of proteins to other elements have not yet been clarified. Glucose may stimulate the transcription of the PKM gene via hexosamine derivatives. Sp1 may be involved in this regulation via its dephosphorylation, although the carbohydrate response element has not been determined precisely in the PKM gene. Thus glucose stimulates transcription of the PKM gene by the mechanism which is probably different from the L-PK gene.
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Affiliation(s)
- K Yamada
- Department of Biochemistry, Fukui Medical University, Shimoaizuki, Matsuoka, Fukui 910-1193, Japan
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Plee-Gautier E, Aggerbeck M, Beurton F, Antoine B, Grimal H, Barouki R, Forest C. Identification of an adipocyte-specific negative glucose response region in the cytosolic aspartate aminotransferase gene. Endocrinology 1998; 139:4936-44. [PMID: 9832431 DOI: 10.1210/endo.139.12.6342] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 11/19/2022]
Abstract
Cytosolic aspartate aminotransferase (cAspAT) participates in gluconeogenesis in the liver and is expected to exert a glyceroneogenic function in the adipose tissue when the supply of glucose is limited. Here we demonstrate that adipose cAspAT messenger RNA (mRNA) is increased when rats are fed a low carbohydrate diet. In the 3T3-F442A, BFC-1 adipocyte cell lines and differentiated adipocytes in primary culture, a 24 h glucose deprivation induces approximately a 4-fold increase in cytosolic AspAT (cAspAT) mRNA, whereas mitochondrial AspAT mRNA remains unchanged. cAspAT activity is also increased in a weaker but reproducible manner. Addition of glucose within a physiological range of concentrations reverses the increase of cAspAT mRNA in 8 h (EC50 = 1.25 g/liter). Such a regulation requires protein synthesis and is specific for adipocytes differentiated in culture. It does not occur in Fao or H4IIE hepatoma cells, in C2 muscle cells, or in 293 kidney cells. 2-deoxyglucose mimicks glucose, while 3-orthomethyl-glucose has no effect, suggesting that glucose-6-phosphate is the effector. cAspAT mRNA stability is not affected by glucose deprivation. To ascertain the transcriptional nature of the glucose effect, we have stably transfected 3T3-F442A adipoblasts with constructs containing the chloramphenicol acetyltransferase reporter gene under the control of either 5'-deletions of the cAspAT gene promoter or internal fragments in an heterologous context. We demonstrate that a glucose response element(s) is present in the region between -1838 and -1702 bp relative to the translation start site. In this region, three DNA sequences bind nuclear proteins from adipocytes as shown by footprinting experiments. Our results indicate that cAspAT gene transcription is repressed by glucose selectively in adipocytes.
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Affiliation(s)
- E Plee-Gautier
- Centre de Recherche sur l'Endocrinologie Moléculaire et le Développement, Centre National de la Recherche Scientifique, Meudon, France
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Tan J, Yang HS, Patel MS. Regulation of mammalian pyruvate dehydrogenase alpha subunit gene expression by glucose in HepG2 cells. Biochem J 1998; 336 ( Pt 1):49-56. [PMID: 9806883 PMCID: PMC1219840 DOI: 10.1042/bj3360049] [Citation(s) in RCA: 11] [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
We report the effect of glucose on the expression of the gene encoding the pyruvate dehydrogenase (E1) alpha subunit (E1alpha) in human hepatoma (HepG2) cells. Total pyruvate dehydrogenase complex activity as well as the levels of protein and mRNA of the E1alpha subunit were significantly increased in HepG2 cells cultured in medium containing 16.7 mM glucose compared with 1.0 mM glucose for a period of 4 weeks. The level of E1alpha mRNA was elevated approx. 2-fold in HepG2 cells cultured for 24 h in medium containing 16.7 mM glucose compared with 1 mM glucose. This effect was specific to glucose and independent of insulin. Nuclear run-on assays and promoter analysis indicate that the glucose-induced increases in the levels of E1alpha mRNA in HepG2 cells are due to increased transcription of the human E1alpha (PDHA1) gene. Mutational analysis of the E1alpha promoter region has identified two regions, from -78 to -73 bp (CCCCTG) and from -8 to -3 bp (GCGGTG), that are responsible for the effect of glucose on promoter activity; the former exhibits a larger effect. These two sequences represent new variations of the carbohydrate-response element that has been identified in other genes. The stimulation of E1alpha promoter activity by glucose was abolished by okadaic acid at 100 nM but not at 5 nM, suggesting that glucose-mediated regulation of pyruvate dehydrogenase complex E1alpha gene transcription involves a phosphorylation/dephosphorylation mechanism, possibly involving protein phosphatase-1.
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Affiliation(s)
- J Tan
- Department of Biochemistry, School of Medicine and Biomedical Sciences, State University of New York at Buffalo, 140 Farber Hall, 3435 Main Street, Buffalo, NY 14214, USA
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Abstract
Successfully igniting the yeast glycolytic flux during the transition from gluconeogenic to fermentative growth seems to be a matter of balance and coordination between a multitude of events. The contours of the sugar sensing and signalling pathways that regulate this transition are only beginning to emerge.
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Affiliation(s)
- P Gonçalves
- Seccão Autónoma de Biotecnologia, Faculdade de Ciências e Tecnologia, Universidade Nova de Lisboa, Monte de Caparica, Portugal
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