Journal
JOURNAL OF NEUROSCIENCE
Volume 34, Issue 45, Pages 15083-15096Publisher
SOC NEUROSCIENCE
DOI: 10.1523/JNEUROSCI.0348-14.2014
Keywords
microscopy; neurexin; neuroligin; split-GFP; synapses; SynView
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Funding
- National Institute of Mental Health-National Institutes of Health [MH052804]
- National Institute of Neurological Disorders and Stroke-National Institutes of Health [NS077906]
- Human Frontiers in Science Program Organization (HFSPO) [LT-000135/2009-L]
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Neurexins and neuroligins are synaptic cell-adhesion molecules that are essential for normal synapse specification and function and are thought to bind to each other trans-synaptically, but such interactions have not been demonstrated directly. Here, we generated neurexin-1 beta and neuroligin-1 and neuroligin-2 fusion proteins containing complementary split GFP fragments positioned such that binding of neurexin-1 beta to neuroligin-1 or neuroligin-2 allowed GFP reconstitution without dramatically changing their binding affinities. GFP fluorescence was only reconstituted from split-GFP-modified neurexin-1 beta and neuroligin-1 if and after neurexin-1 beta bound to its neuroligin partner; reassociation of the split-GFP components with each other did not mediate binding. Using trans-cellular reconstitution of GFP fluorescence from split-GFP-modified neurexin-1 beta and neuroligins as an assay, we demonstrate that trans-synaptic neurexin/neuroligin binding indeed occurred when mouse hippocampal neurons formed synapses onto non-neuronal COS-7 cells expressing neuroligins or when mouse hippocampal neurons formed synapses with each other. This visualization of synapses by neurexin/neuroligin binding prompted us to refer to this approach as SynView. Our data demonstrate that neurexin-1 beta forms a trans-synaptic complex with neuroligin-1 and neuroligin-2 and that this interaction can be used to label synapses in a specific fashion in vivo.
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