Cloning, expression, and localization of a rat brain high-affinity glycine transporter

Creators: Guastella, John; Brecha, Nicholas; Weigmann, Christine; Lester, Henry A.; Davidson, Norman

Abstract

A cDNA clone encoding a glycine transporter has been isolated from rat brain by a combined PCR and plaque-hybridization strategy. mRNA synthesized from this clone (designated GLYT1) directs the expression of sodium-and chloride-dependent, high-affinity uptake of [3H]glycine by Xenopus oocytes. [3H]Glycine transport mediated by clone GLYT1 is blocked by sarcosine but is not blocked by methylaminoisobutyric acid or L-alanine, a substrate specificity similar to that described for a previously identified glycine-uptake system called system Gly. In situ hybridization reveals that GLYT1 is prominently expressed in the cervical spinal cord and brainstem, two regions of the central nervous system where glycine is a putative neurotransmitter. GLYT1 is also strongly expressed in the cerebellum and olfactory bulb and is expressed at lower levels in other brain regions. The open reading frame of the GLYT1 cDNA predicts a protein containing 633 amino acids with a molecular mass of ≈70 kDa. The primary structure and hydropathicity profile of GLYT1 protein reveal that this protein is a member of the sodium- and chloride-dependent superfamily of transporters that utilize neurotransmitters and related substances as substrates.

Additional Information

© 1992 by the National Academy of Sciences. Contributed by Norman Davidson, May 6, 1992. We thank M. Quick for oocyte preparation, L. Czyzyk for assistance with nucleotide sequencing, and A. Figl for assistance with oligonucleotide synthesis. This work was supported by National Institutes of Health Grant EY04067, Neuroendocrine Anatomy Core Grant DK4130, and Veterans Administration Medical Research Funds to N.B.; National Institutes of Health Grant NS-11756 to H.A.L. and N.D.; and a National Institutes of Health Postdoctoral Fellowship to J.G. The sequence reported in this paper has been deposited in the GenBank data base (accession no. M95413). The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

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