Identification of a mammalian H+‐myo‐inositol symporter expressed predominantly in the brain

Inositol and its phosphorylated derivatives play a major role in brain function, either as osmolytes, second messengers or regulators of vesicle endo‐ and exocytosis. Here we describe the identification and functional characterization of a novel H+‐myo‐ inositol co‐transporter, HMIT, expressed predominantly in the brain. HMIT cDNA encodes a 618 amino acid polypeptide with 12 predicted transmembrane domains. Functional expression of HMIT in Xenopus oocytes showed that transport activity was specific for myo‐inositol and related stereoisomers with a Michaelis–Menten constant of ∼100 μM, and that transport activity was strongly stimulated by decreasing pH. Electrophysiological measurements revealed that transport was electrogenic with a maximal transport activity reached at pH 5.0. In rat brain membrane preparations, HMIT appeared as a 75–90 kDa protein that could be converted to a 67 kDa band upon enzymatic deglycosylation. Immunofluorescence microscopy analysis showed HMIT expression in glial cells and some neurons. These data provide the first characterization of a mammalian H+‐coupled myo‐ inositol transporter. Predominant central expression of HMIT suggests that it has a key role in the control of myo‐inositol brain metabolism.

[1]  W. Sherman,et al.  The measurement of myo-inositol, myo-inosose-2 and scyllo-inositol in mammalian tissues. , 1968, Biochimica et biophysica acta.

[2]  R. Spector.  THE SPECIFICITY AND SULFHYDRYL SENSITIVITY OF THE INOSITOL TRANSPORT SYSTEM OF THE CENTRAL NERVOUS SYSTEM , 1976, Journal of neurochemistry.

[3]  D. Kaplan,et al.  The TrkB-Shc Site Signals Neuronal Survival and Local Axon Growth via MEK and PI3-Kinase , 2000, Neuron.

[4]  J. Rothman,et al.  A possible docking and fusion particle for synaptic transmission , 1995, Nature.

[5]  P. Séguéla,et al.  Mammalian ASIC2a and ASIC3 Subunits Co-assemble into Heteromeric Proton-gated Channels Sensitive to Gd3+ * , 2000, The Journal of Biological Chemistry.

[6]  B. Thorens,et al.  Agonist-induced internalization and recycling of the glucagon-like peptide-1 receptor in transfected fibroblasts and in insulinomas. , 1995, The Biochemical journal.

[7]  S. K. Woo,et al.  Tonicity-responsive enhancer binding protein, a rel-like protein that stimulates transcription in response to hypertonicity. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[8]  G. Prestwich,et al.  Binding kinetics and ligand specificity for the interactions of the C2B domain of synaptogmin II with inositol polyphosphates and phosphoinositides. , 2000, Biochemistry.

[9]  P. Good,et al.  Oligomerization and maturation of Na,K-ATPase: functional interaction of the cytoplasmic NH2 terminus of the beta subunit with the alpha subunit , 1996, The Journal of cell biology.

[10]  R. Zimmerman,et al.  In vivo brain myo-inositol levels in children with Down syndrome. , 1999, The Journal of pediatrics.

[11]  C. Kahn,et al.  Insulin action and the insulin signaling network. , 1995, Endocrine reviews.

[12]  P. Chomczyński,et al.  Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. , 1987, Analytical biochemistry.

[13]  J. Falck,et al.  A Functional Phosphatidylinositol 3,4,5-Trisphosphate/Phosphoinositide Binding Domain in the Clathrin Adaptor AP-2 α Subunit. IMPLICATIONS FOR THE ENDOCYTIC PATHWAY* , 1996, The Journal of Biological Chemistry.

[14]  R. Thomas.  Review Lecture: Experimental displacement of intracellular pH and the mechanism of its subsequent recovery. , 1984, The Journal of physiology.

[15]  A. Harwood,et al.  Lithium therapy and signal transduction. , 2000, Trends in pharmacological sciences.

[16]  K. Tokuyasu.  A study of positive staining of ultrathin frozen sections. , 1978, Journal of ultrastructure research.

[17]  M. Lazdunski,et al.  The Acid-sensitive Ionic Channel Subunit ASIC and the Mammalian Degenerin MDEG Form a Heteromultimeric H+-gated Na+ Channel with Novel Properties* , 1997, The Journal of Biological Chemistry.

[18]  K. Mikoshiba,et al.  Distinct roles of C2A and C2B domains of synaptotagmin in the regulation of exocytosis in adrenal chromaffin cells. , 1997, Proceedings of the National Academy of Sciences of the United States of America.

[19]  David R Kaplan,et al.  Neurotrophin signal transduction in the nervous system , 2000, Current Opinion in Neurobiology.

[20]  Michael J. Berridge,et al.  Neural and developmental actions of lithium: A unifying hypothesis , 1989, Cell.

[21]  S. Schmid,et al.  Phosphatidylinositol-4,5-bisphosphate is required for endocytic coated vesicle formation , 1998, Current Biology.

[22]  S. Rapoport,et al.  Polyol profiles in Down syndrome. myo-Inositol, specifically, is elevated in the cerebrospinal fluid. , 1995, The Journal of clinical investigation.

[23]  I. Zagon,et al.  Spectrin subtypes in mammalian brain: an immunoelectron microscopic study , 1986, The Journal of neuroscience : the official journal of the Society for Neuroscience.

[24]  R. Irvine,et al.  Inositol phospholipids: Translocation, translocation, translocation … , 1998, Current Biology.

[25]  A. Yamauchi,et al.  Cloning of the cDNa for a Na+/myo-inositol cotransporter, a hypertonicity stress protein. , 1992, The Journal of biological chemistry.

[26]  M. Lazdunski,et al.  A proton-gated cation channel involved in acid-sensing , 1997, Nature.

[27]  T. Risler,et al.  Effect of extracellular pH on the myo-inositol transporter SMIT expressed in Xenopus oocytes , 1998, Pflügers Archiv.

[28]  Marino Zerial,et al.  EEA1 links PI(3)K function to Rab5 regulation of endosome fusion , 1998, Nature.

[29]  J. Hartwig,et al.  Structure of the novel membrane-coating material in proton-secreting epithelial cells and identification as an H+ATPase , 1987, The Journal of cell biology.

[30]  M. Waterfield,et al.  Signalling through phosphoinositide 3-kinases: the lipids take centre stage. , 1999, Current opinion in cell biology.

[31]  D. McCormick,et al.  Essential Role of Phosphoinositide Metabolism in Synaptic Vesicle Recycling , 1999, Cell.

[32]  P. Marquet,et al.  A quantitative analysis of l‐glutamate‐regulated Na+ dynamics in mouse cortical astrocytes: implications for cellular bioenergetics , 2000, The European journal of neuroscience.

[33]  M. Hediger,et al.  Molecular physiology of sodium-glucose cotransporters. , 1994, Physiological reviews.

[34]  W. Sherman,et al.  A sensitive and specific method for the measurement of monophosphoinositide at a microregional level in brain. , 1973, Analytical biochemistry.

[35]  K. Kaila,et al.  Modulation of pH by neuronal activity , 1992, Trends in Neurosciences.

[36]  L. Cantley,et al.  Regulatory interactions in the recognition of endocytic sorting signals by AP‐2 complexes , 1997, The EMBO journal.

[37]  M. Uldry,et al.  GLUTX1, a Novel Mammalian Glucose Transporter Expressed in the Central Nervous System and Insulin-sensitive Tissues* , 2000, The Journal of Biological Chemistry.

[38]  Y. Jan,et al.  A New ER Trafficking Signal Regulates the Subunit Stoichiometry of Plasma Membrane KATP Channels , 1999, Neuron.