Molecular determinants involved in the asymmetrical distribution of glycine transporters in polarized cells.
The asymmetrical distribution of neurotransmitter transporters on the cell surface of neurons seems to be a generalized feature of these proteins, and is thought to be important for an appropriate removal of neurotransmitters from the extracellular milieu. To study the subcellular distribution of the glycine transporter isoforms (GLYT1a, GLYT1b, GLYT2a and GLYT2b), these proteins were expressed in epithelial cells [Madin-Darby canine kidney (MDCK) cells] and in cultured hippocampal neurons, as models of polarized cells. The localization of the transporters was assessed by immunofluorescence assays. Our results indicated that the subcellular distribution of glycine transporters is dependent on both the protein isoform and the cell type. By using site-directed mutagenesis we have been able to identify signals for basolateral/somatodendritic localization in the alternative amino terminal region of GLYT1 and in two di-leucine motifs that are located in the carboxyl tail of this protein. Moreover, the N-glycosylation sites located in the large extracellular loop of GLYT2 are involved in apical localization of this protein in polarized MDCK cells. These results contribute to define the mechanisms of asymmetrical distribution of transporters on the cell surface of polarized cells.
Zafra
F
F
Centro de Biología Molecular Severo Ochoa, Universidad Autónoma de Madrid, 28049 Madrid, Spain. fzafra@cbm.uam.es
Giménez
C
C
D026921:Amino Acid Transport Systems, Neutral / Q000737:chemistry / Q000378:metabolism*; D000818:Animals; D004285:Dogs; D050487:Glycine Plasma Membrane Transport Proteins; D017403:In Situ Hybridization; D008954:Models, Biological; D020033:Protein Isoforms; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D051381:Rats
D026921:Amino Acid Transport Systems, Neutral; D050487:Glycine Plasma Membrane Transport Proteins; D020033:Protein Isoforms; C493269:Slc6a5 protein, rat; C493266:Slc6a9 protein, rat