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Open AccessArticle

Mapping the Proteome of the Synaptic Cleft through Proximity Labeling Reveals New Cleft Proteins

1
Department of Neuroscience, Tufts University School of Medicine, Boston, MA 02111, USA
2
Department of Physiology and Program in Neuroscience, University of Maryland School of Medicine, Baltimore, MD 21201, USA
3
Yale/NIDA Neuroproteomics Center, New Haven, CT 06511, USA
4
W.M. Keck Biotechnology Resource Laboratory, Yale University School of Medicine, 300 George Street, New Haven, CT 06511, USA
5
Department of Molecular Biophysics and Biochemistry, Yale University School of Medicine, New Haven, CT 06520, USA
*
Authors to whom correspondence should be addressed.
Proteomes 2018, 6(4), 48; https://doi.org/10.3390/proteomes6040048
Received: 16 October 2018 / Revised: 15 November 2018 / Accepted: 18 November 2018 / Published: 28 November 2018
(This article belongs to the Special Issue Neuroproteomics)
Synapses are specialized neuronal cell-cell contacts that underlie network communication in the mammalian brain. Across neuronal populations and circuits, a diverse set of synapses is utilized, and they differ in their molecular composition to enable heterogenous connectivity patterns and functions. In addition to pre- and post-synaptic specializations, the synaptic cleft is now understood to be an integral compartment of synapses that contributes to their structural and functional organization. Aiming to map the cleft proteome, this study applied a peroxidase-mediated proximity labeling approach and used the excitatory synaptic cell adhesion protein SynCAM 1 fused to horseradish peroxidase (HRP) as a reporter in cultured cortical neurons. This reporter marked excitatory synapses as measured by confocal microcopy and was targeted to the edge zone of the synaptic cleft as determined using 3D dSTORM super-resolution imaging. Proximity labeling with a membrane-impermeant biotin-phenol compound restricted labeling to the cell surface, and Label-Free Quantitation (LFQ) mass spectrometry combined with ratiometric HRP tagging of membrane vs. synaptic surface proteins was used to identify the proteomic content of excitatory clefts. Novel cleft candidates were identified, and Receptor-type tyrosine-protein phosphatase zeta was selected and successfully validated. This study supports the robust applicability of peroxidase-mediated proximity labeling for synaptic cleft proteomics and its potential for understanding synapse heterogeneity in health and changes in diseases such as psychiatric disorders and addiction. View Full-Text
Keywords: synapse; synaptic cleft; trans-synaptic adhesion; proximity labeling; SynCAM; Cadm; Receptor-type tyrosine-protein phosphatase zeta; R-PTP-zeta; Ptprz1 synapse; synaptic cleft; trans-synaptic adhesion; proximity labeling; SynCAM; Cadm; Receptor-type tyrosine-protein phosphatase zeta; R-PTP-zeta; Ptprz1
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MDPI and ACS Style

Cijsouw, T.; Ramsey, A.M.; Lam, T.T.; Carbone, B.E.; Blanpied, T.A.; Biederer, T. Mapping the Proteome of the Synaptic Cleft through Proximity Labeling Reveals New Cleft Proteins. Proteomes 2018, 6, 48.

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