A Tyrosine-based Motif Localizes a Drosophila Vesicular Transporter to Synaptic Vesicles in Vivo

Department of Psychiatry and Biobehavioral Sciences, Semel Institute for Neuroscience and Human Behavior, Hatos Center for Neuropharmacology, David Geffen School of Medicine, UCLA, Los Angeles, California 90095-1761, USA.
Journal of Biological Chemistry (Impact Factor: 4.57). 03/2010; 285(10):6867-78. DOI: 10.1074/jbc.M109.073064
Source: PubMed


Vesicular neurotransmitter transporters must localize to synaptic vesicles (SVs) to allow regulated neurotransmitter release at the synapse. However, the signals required to localize vesicular proteins to SVs in vivo remain unclear. To address this question we have tested the effects of mutating proposed trafficking domains in Drosophila orthologs of the vesicular monoamine and glutamate transporters, DVMAT-A and DVGLUT. We show that a tyrosine-based motif (YXXY) is important both for DVMAT-A internalization from the cell surface in vitro, and localization to SVs in vivo. In contrast, DVGLUT deletion mutants that lack a putative C-terminal trafficking domain show more modest defects in both internalization in vitro and trafficking to SVs in vivo. Our data show for the first time that mutation of a specific trafficking motif can disrupt localization to SVs in vivo and suggest possible differences in the sorting of VMATs versus VGLUTs to SVs at the synapse.

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Available from: Richard W Daniels, Jun 20, 2014
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    • "VAchT dileucine motif (LL), and possibly a tyrosine-based motif YNYY, allow internalization and localization to SLMVs (Kim and Hersh, 2004; Tan et al., 1998). Recently, it was reported that deletion of the Drosophila VGLUT C-terminal domain has a consistent effect in reducing internalization and localization to SVs of the transporter (Grygoruk et al., 2010). VGLUT2 also has three putative glycosylation sites at amino acid residues Asn100, Asn101 and Asn470 (UniProtKB Database), which were also correlated with a decrease in VMAT2 location to SVs in vivo (Cruz-Muros et al., 2008). "
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    • "If DVGLUT function is required for lethality, then overexpression of non-functional DVGLUT transgenes should suppress this lethality. Since there is little known about residues important for VGLUT function, we performed an in vivo chemical mutagenesis screen to find DVGLUT mutants that are viable following expression from the strong BG380-Gal4 driver (Grygoruk et al., 2010). From this screen we isolated approximately 50 independent mutants that change amino acids in a GFP-tagged DVGLUT transgene. "
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