RT Journal Article
SR Electronic
T1 Cotrafficking of SV2 and Synaptotagmin at the Synapse
JF The Journal of Neuroscience
JO J. Neurosci.
FD Society for Neuroscience
SP 5569
OP 5578
DO 10.1523/JNEUROSCI.4781-09.2010
VO 30
IS 16
A1 Jia Yao
A1 Amy Nowack
A1 Patricia Kensel-Hammes
A1 Richard G. Gardner
A1 Sandra M. Bajjalieh
YR 2010
UL http://www.jneurosci.org/content/30/16/5569.abstract
AB Synaptic vesicles are specialized cycling endosomes that contain a unique constellation of membrane proteins. Proteins are sorted to vesicles by short amino acid sequences that serve as binding sites for clathrin adaptor proteins. Here we show that a tyrosine-based endocytosis motif in the vesicle protein SV2 is required for trafficking to synaptic vesicles of both SV2 and the calcium sensor protein synaptotagmin. Aberrant neurotransmission in cultured hippocampal neurons lacking SV2 was rescued by expression of wild-type SV2A, but not by SV2A-Y46A, a mutant containing a disrupted endocytosis motif in SV2A's cytoplasmic N terminus. Neurons expressing SV2A-Y46A had significantly more SV2 on the plasma membrane, indicating reduced internalization. A screen for proteins that preferentially bound wild-type SV2A identified multiple endocytosis-related proteins, and in vitro binding studies confirmed binding to the clathrin adaptors AP2, EPS15, and amphiphysin 2/Bin1. Neurons lacking SV2 contained less synaptotagmin and had a higher proportion of synaptotagmin on the plasma membrane. Expression of either wild-type SV2A or SV2A-Y46A restored synaptotagmin expression levels; however, only wild-type SV2A restored a normal proportion of synaptotagmin on the plasma membrane. These findings indicate that SV2 influences the expression and trafficking of synaptotagmin via separate mechanisms. Synaptic vesicles immunoisolated from SV2A/B double knock-out mice had significantly less synaptotagmin than vesicles isolated from wild-type mice. Our results indicate that SV2 plays a major role in regulating the amount of synaptotagmin in synaptic vesicles and provide an explanation for the observation that synapses lacking SV2 have fewer vesicles competent for calcium-induced fusion.