Abstract
Vesicular trafficking of presynaptic and postsynaptic components is emerging as a general cellular mechanism for the delivery of scaffold proteins, ion channels, and receptors to nascent and mature synapses. However, the molecular mechanisms leading to the selection of cargos and their differential transport to subneuronal compartments are not well understood, in part because of the mixing of cargos at the plasma membrane and/or within endosomal compartments. In the present study, we have explored the cellular mechanisms of active zone precursor vesicle assembly at the Golgi in dissociated hippocampal neurons of Rattus norvegicus. Our studies show that Piccolo, Bassoon, and ELKS2/CAST exit the trans-Golgi network on a common vesicle that requires Piccolo and Bassoon for its proper assembly. In contrast, Munc13 and synaptic vesicle proteins use distinct sets of Golgi-derived transport vesicles, while RIM1α associates with vesicular membranes in a post-Golgi compartment. Furthermore, Piccolo and Bassoon are necessary for ELKS2/CAST to leave the Golgi in association with vesicles, and a core domain of Bassoon is sufficient to facilitate formation of these vesicles. While these findings support emerging principles regarding active zone differentiation, the cellular and molecular analyses reported here also indicate that the Piccolo-Bassoon transport vesicles leaving the Golgi may undergo further changes in protein composition before arriving at synaptic sites.
Publication types
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Research Support, N.I.H., Extramural
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Research Support, Non-U.S. Gov't
MeSH terms
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Age Factors
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Animals
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Antibodies / pharmacology
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Autoantigens / metabolism
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Axons / metabolism
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Carrier Proteins / genetics
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Carrier Proteins / immunology
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Carrier Proteins / metabolism
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Cell Membrane / drug effects
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Cell Membrane / metabolism
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Cells, Cultured
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Chlorocebus aethiops
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Cytoskeletal Proteins / metabolism
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Embryo, Mammalian
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Gene Expression Regulation / physiology
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Golgi Apparatus / metabolism*
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Green Fluorescent Proteins / genetics
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Green Fluorescent Proteins / metabolism
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Hippocampus / cytology
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Membrane Glycoproteins / metabolism
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Membrane Proteins / metabolism
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Microscopy, Confocal
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Nerve Tissue Proteins / genetics
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Nerve Tissue Proteins / immunology
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Nerve Tissue Proteins / metabolism
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Neurons / ultrastructure*
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Neuropeptides / metabolism
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Presynaptic Terminals / metabolism*
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Protein Binding / drug effects
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Protein Transport / genetics
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Protein Transport / physiology
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RNA, Small Interfering / metabolism
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Rats
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Rats, Sprague-Dawley
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Synaptophysin / metabolism
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Time Factors
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Transfection
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Transport Vesicles / metabolism*
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Tubulin / metabolism
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rab GTP-Binding Proteins
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trans-Golgi Network / metabolism
Substances
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Antibodies
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Autoantigens
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Bsn protein, mouse
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Carrier Proteins
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Cytoskeletal Proteins
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Erc1 protein, mouse
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Golgin subfamily A member 2
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Membrane Glycoproteins
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Membrane Proteins
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Nerve Tissue Proteins
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Neuropeptides
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Pclo protein, mouse
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RNA, Small Interfering
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Sv2a protein, rat
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Synaptophysin
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Tgoln2 protein, rat
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Tubulin
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Unc13a protein, rat
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enhanced green fluorescent protein
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Green Fluorescent Proteins
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rab GTP-Binding Proteins