Cloning and expression of a spinal cord- and brain-specific glycine transporter with novel structural features.

Abstract

A novel glycine transporter (GLYT2) was cloned from a rat brain cDNA library. GLYT2 is about 48 and 50% homologous to the previously cloned mouse glycine transporter (GLYT1) and rat proline transporter (PROT), respectively. GLYT2 differs from GLYT1 in molecular structure, tissue specificity, and pharmacological properties. The cDNA of GLYT2 encodes for 799 amino acid residues with an extended amino-terminal peptide containing 200 amino acids before the first transmembrane domain. Potential phosphorylation sites for protein kinase C, cAMP-dependent kinase, and calmodulin-dependent kinase were identified in the amino-terminal region. GLYT2 mRNA was shown to be specifically localized in spinal cord, brain stem, and to a lesser extent in the cerebellum. In contrast, GLYT1 mRNA distribution in the brain has been found previously to be more ubiquitous. Xenopus oocytes injected with GLYT2 cRNA transport glycine with a Km of 17 microM, and the uptake of glycine is resistant to inhibition by sarcosine. The experimental data suggests GLYT2 might play a major role in the termination of the inhibitory effect of glycine in the brain stem and spinal cord of vertebrates. On the other hand, the main function of GLYT1 may be in the modulation of excitatory nerve terminals. Two types of GLYT1 cDNA, GLYT1a and GLYT1b, were cloned from the mouse brain library. They differ only at their amino-terminal sequences, and GLYT1b contains two additional potential phosphorylation sites for proline-dependent kinase. Cloning of the gene encoding the GLYT1 revealed that the two variants resulted from a differential splicing.