Regulation of glycine transporters

Abstract

The regulation of neurotransmitter transporters is a central aspect of their physiology. Recent studies that focused on syntaxin-1 transporter interactions led to the postulation that syntaxin-1 is somehow implicated in protein trafficking. Because syntaxin-1 is involved in the exocytosis of neurotransmitters and it interacts with glycine transporter 2 (GLYT2), we stimulated exocytosis in synaptosomes and examined its effect on GLYT2 surface-expression and transport activity. We found that GLYT2 is rapidly trafficked first towards the plasma membrane and then internalized under conditions that stimulate vesicular glycine release. However, when syntaxin-1 was inactivated by pre-treatment of synaptosomes with the botulinum neurotoxin C, GLYT2 was unable to reach the plasma membrane but still was able to leave it. These results indicate the existence of a SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor)-mediated regulatory mechanism that controls the surface expression of GLYT2. Syntaxin-1 is involved in the transport of GLYT2 to, but not its retrieval from, the plasma membrane. Immunogold-labelling on purified vesicular preparations from synaptosomes showed that GLYT2 is present in small synaptic-like vesicles. This may represent neurotransmitter transporter that is being trafficked. The subcellular distribution of the glycine transporters was further examined in PC12 cells that were stably transfected with the fusions of GLYT1 and GLYT2 with green fluorescent protein. There was a clear difference in their intracellular distribution, GLYT1 being present mainly on the plasma membrane and GLYT2 being localized mainly on large, dense-core vesicles. We are trying to find signal sequences responsible for this differential localization.