Aryl Hydrocarbon Receptor Nuclear Translocator/Hypoxia-inducible Factor-1β Plays a Critical Role in Maintaining Glucose-stimulated Anaplerosis and Insulin Release from Pancreatic β-Cells

• DOI: 10.1074/jbc.m110.149062
• Publication Date: 2010-11-09T05:39:10Z
• Authors: Renjitha Pillai; Peter Huypens; Mei Huang; Stephanie Schaefer; Tanya Sheinin; Shawn D. Wettig; Jamie W. Joseph
• Keywords: Pancreatic Islets; Hypoxia-Inducible Factors

The metabolic pathways that are involved in regulating insulin secretion from pancreatic β-cells still incompletely understood. One potential regulator of the phenotype is transcription factor aryl hydrocarbon receptor nuclear translocator (ARNT)/hypoxia-inducible (HIF)-1β. ARNT/HIF-1β levels profoundly reduced islets obtained type 2 diabetic patients. However, no study to date has investigated key release lack ARNT/HIF-1β. In this study, we confirm siRNA-mediated knockdown inhibits glucose-stimulated secretion. We next consequence loss knockdown. demonstrate with expression exhibit a 31% reduction glycolytic flux without significant changes glucose oxidation or ATP:ADP ratio. Metabolic profiling treated siRNAs against gene revealed glycolysis, anaplerosis, and glucose-induced fatty acid production were down-regulated, all events addition, both first second phase significantly after Together, our data suggest an important role for it may play critical maintaining normal competence β-cells. IntroductionThe ability β-cell maintain homeostasis critically depends on existence functional sensor operates within physiologic range concentrations (1Matschinsky F.M. Diabetes. 1996; 45: 223-241Crossref PubMed Scopus (0) Google Scholar, 2Matschinsky Nat. Rev. Drug Discov. 2009; 8: 399-416Crossref (328) Scholar). glucose-phosphorylating enzyme glucokinase (GK) 3The abbreviations used are: GK, glucokinase; ARNT/HIF-1β, translocator/hypoxia-inducible factor-1β; GSIS, secretion; LC-CoA, long-chain acyl CoA; PC, pyruvate carboxylase; PDH, dehydrogenase; DIC, dicarboxylate carrier; CIC, citrate BisTris, 2-[bis(2-hydroxyethyl)amino]-2-(hydroxymethyl)propane-1,3-diol; PPP, pentose phosphate pathway; MEc, cytosolic malic enzyme; OGC, α-ketoglutarate carrier. been identified as rate-limiting step metabolism, allowing adapt rate accordance circulating (2Matschinsky 3German M.S. Proc. Natl. Acad. Sci. U.S.A. 1993; 90: 1781-1785Crossref (181) Downstream metabolism leads elevation ratio point where promotes closure KATP channels, resulting depolarization plasma membrane opening voltage-gated calcium enter cytosol promote exocytosis (4Prentki M. Matschinsky Physiol. 1987; 67: 1185-1248Crossref 5Ashcroft Rorsman P. Prog. Biophys. Mol. Biol. 1989; 54: 87-143Crossref (945) 6MacDonald P.E. Joseph J.W. Philos. Trans. R. Soc. Lond. B 2005; 360: 2211-2225Crossref (253) This so-called “KATP channel-dependent pathway” appears be particularly acute release, whereas more sustained requires -independent (7Henquin J.C. Nenquin Ravier M.A. Szollosi A. Diabetes Obes. Metab. 11: 168-179Crossref (69) 8Henquin Jonas Gilon Eur. J. Clin. Invest. 2003; 33: 742-750Crossref (151) 9Maechler Kennedy E.D. Pozzan T. Wollheim C.B. EMBO 1997; 16: 3833-3841Crossref (162) 10Gunawardana S.C. Liu Y.J. Macdonald M.J. Straub S.G. Sharp G.W. Am. Endocrinol. 2004; 287: E828-E833Crossref (12) 11Jensen M.V. Ronnebaum S.M. Burgess Sherry A.D. Newgard 2008; 295: E1287-E1297Crossref (189) Scholar).Important support channel-independent pathway (GSIS) comes studies showing can elicit increase conditions channels held open by application diazoxide high K+ (12Gembal Henquin 1992; 89: 1288-1295Crossref (421) 13Sato Y. Aizawa Komatsu Okada N. Yamada 41: 438-443Crossref (175) Scholar) rodents 14Nenquin Aguilar-Bryan L. Bryan Chem. 279: 32316-32324Abstract Full Text PDF (87) 15Szollosi Br. Pharmacol. 2010; 159: 669-677Crossref (33) 16Ravier Miki Seino S. Endocrinology. 150: 33-45Crossref (55) 17Quoix Cheng-Xue Mattart Zeinoun Z. Guiot Beauvois M.C. 58: 412-421Crossref (61) 18Henquin Diabetologia. 52: 739-751Crossref (350) These mitochondrial generates other signals besides stimulus-secretion coupling (11Jensen 19Hasan N.M. Longacre Stoker S.W. Boonsaen Jitrapakdee Kendrick Wallace MacDonald 283: 28048-28059Abstract (62) 20MacDonald Hasan Cell. Biochem. 313: 195-202Crossref (11) 21Lu D. Mulder H. Zhao Jensen Kamzolova 2002; 99: 2708-2713Crossref (219) 22MacDonald Fahien L.A. Brown L.J. Buss J.D. 288: E1-E15Crossref (198) 23Maechler Antinozzi P.A. IUBMB Life. 2000; 50: 27-31Crossref (28) Several molecules, including GTP, glutamate, malonyl-CoA/LC-CoA, α-ketoglutarate, NADPH, have proposed candidate factors GSIS 24Meredith Rabaglia M.E. Metz S.A. 1995; 96: 811-821Crossref (47) 25Corkey B.E. Glennon Chen K.S. Deeney J.T. Prentki 264: 21608-21612Abstract 26Prentki Vischer Regazzi Corkey 267: 5802-5810Abstract 27Newgard Lu Schissler Boucher 51: S389-S393Crossref 28Broca C. Brennan Petit Newsholme Maechler FEBS Lett. 545: 167-172Crossref (46) 29Ivarsson Quintens Dejonghe Tsukamoto K. 't Veld Renström E. Schuit F.C. 2132-2142Crossref (209) 30Joseph Ilkayeva O. Palmieri F. Alárcon Rhodes C.J. 2006; 281: 35624-35632Abstract (133) 31Kibbey R.G. Pongratz R.L. Romanelli A.J. Cline Shulman G.I. Cell 2007; 5: 253-264Abstract (124) Scholar).ARNT/HIF-1β member basic helix-loop-helix/PER/AhR/ARNT/Sim family considered obligate heterodimerization partner members family, such HIF-1α, HIF-2α, HIF-3α, AhR (32Furness Lees Whitelaw M.L. 581: 3616-3625Crossref (70) shown homodimerize regulate genes typically contain palindromic E-box (CACGTG) signature their promoter sequence (33Antonsson Arulampalam V. Pettersson Poellinger 270: 13968-13972Abstract (88) 34Arpiainen Lämsä Pelkonen Yim S.H. Gonzalez F.J. Hakkola 369: 640-652Crossref (16) Gene human its target markedly (35Gunton J.E. Kulkarni R.N. Hawthorne W.J. Tseng Y.H. Roberson R.S. Ricordi O'Connell P.J. Kahn C.R. 122: 337-349Abstract (393) importance was evidenced diminished β-cell-specific knock-out mice well Min6 cells effectively silenced siRNA technology Scholar).In aim obtain footprint low identify affected factor. impairment become eminent when INS-1-derived 832/13 cells. Our novel findings plays biphasic anaplerosis mechanism ARNT/HIF-1β-regulated independent ATP likely involves altered KATP-independent release.DISCUSSIONExpression showed 90% It also demonstrated control diabetes, HNF4α, receptor, IRS-2, Akt2, which (44Vaxillaire Froguel Endocr. 29: 254-264Crossref (117) 45Stoffel Duncan 94: 13209-13214Crossref (332) 46Withers D.J. Gutierrez J.S. Towery Burks Ren J.M. Previs Zhang Bernal Pons Bonner-Weir White M.F. Nature. 1998; 391: 900-904Crossref (1325) 47Kubota Tobe Terauchi Eto Yamauchi Suzuki Tsubamoto Komeda Nakano Satoh Sekihara Sciacchitano Lesniak Nagai Kimura Akanuma Taylor S.I. Kadowaki 49: 1880-1889Crossref (423) assessed profile provide insight into underlying mechanisms impair under these conditions.Based available literature, hypothesized would required GSIS. Although confirmed dramatically reduced, found utilization only decreased 31%, not altered. To gain employed metabolomics approach assess wide cellular metabolites. As expected, intermediates siARNT1-treated cells; however, most dramatic reductions seen TCA suggesting response anaplerosis.Anaplerosis suggested β-cells, abundant, entry cycle occurs via PDH PC almost equal proportions, giving rise two separate pools, one feeding acetyl-CoA (PDH pathway) leading net accumulation (PC (21Lu 48MacDonald Arch. 305: 205-214Crossref (79) 49MacDonald Metabolism. 42: 1229-1231Abstract 50Khan Ling Z.C. Landau B.R. 271: 2539-2542Abstract (73) 51Schuit De Vos Farfari Moens Pipeleers Brun 272: 18572-18579Abstract pathway, will transfer reducing equivalents respiratory chain, hyperpolarization increased production. cycling, LC-CoA hypothesis results idea anaplerotic related (PC, OGC) lower levels. at glucose, associated nonsignificant basal siARNT1- siARNT2-treated suggests secretion.Pyruvate cycling numerous groups 37Joseph Odegaard Muehlbauer 282: 31592-31600Abstract (60) 38Ronnebaum Stevens R.D. Becker T.C. 30593-30602Abstract (180) 40Jensen 22342-22351Abstract (107) 52Ronnebaum Hohmeier H.E. Zhou Y.P. Qian MacNeil Howard Thornberry 28909-28917Abstract (57) 53Brown 284: 35359-35367Abstract (26) 54Xu Han Long Y.S. Lock Weir G.C. Epstein P.N. Y.Q. 2281-2289Crossref 55Pongratz Kibbey 200-207Abstract (106) 56Guay Madiraju S.R. Aumais Joly 35657-35665Abstract (125) Increased input proper functioning pathways. Pyruvate generate potentially signaling molecules NADPH. NADPH produced three pathways, pyruvate/malate pyruvate/citrate pyruvate/isocitrate pathway. A NADPH-producing malate MEc isocitrate dehydrogenase (38Ronnebaum strong evidence pyruvate-isocitrate exists Scholar), others (54Xu Which pathway(s) unknown. down-regulated MEc).Gunton et al. suppression complete abnormalities ARNT/HIF-1β-silenced primary ARNT mice. Here, 60% compared perifused pseudoislets. cause responsiveness, initially measured oxidation. At mm did affect oxidation, agreement observation deviate conditions. exposed 16.7 clearly suppressed ∼31% cells, glycolysis accompanied drop earlier report several enzymes lowered upon effect oxidative surprising considering β-cells.Others established either does pacemaker, glucokinase, transporters GLUT1 GLUT2 Scholar); there GK. experiments GK expressions lower, overall activity 6-phosphate similar concentration. Therefore, seems steps downstream attained properties function impaired. possible explanation enzymes, glucose-6-phosphate isomerase, phosphofructokinase, aldolase, limiting substrate flow-through notion, observed dihydroxyacetone than siControl-treated glucose. reflected stimulation.The finding profound OGC metabolites, even though unaltered, unexpected result. collective metabolite preserved placed stimulatory regime occur expense cycle.Several arguments brought forward negatively First, disproportionate 65% detected sacrificed expenses oxidation.Second, end product malate, citrate, isocitrate, collectively flux-determining pyruvate-malate, pyruvate-citrate, shuttle, 50–70%. reduced. such, metabolites amount being concentrations, becomes interesting perspective act Scholar).A third argument affects follows FAS CPT1α, virtually absent observations export citrate/isocitrate carrier (CIC), provides essential source carbon lipogenesis (30Joseph severely abrogated relevant malonyl-CoA serve (25Corkey CIC pharmacologic inhibition protein isolated rat impairments show lost l-glutamate raised represent another facet signal controls (42Maechler 1999; 402: 685-689Crossref (436) Scholar).Our concept lowering affecting decreasing dependent called cycling. link between yet established.In summary, performed extensive survey proof causes flux, secretion, Considering knockdown, shift consistent marked anaplerosis. furthermore supported glutamate completely Taken together, absolutely keep glucose-responsive state ensuring sufficient flow through (Fig. 8). release.