Abstract
The proteome of the presynaptic active zone controls neurotransmitter release and the short- and long-term structural and functional dynamics of the nerve terminal. The proteinaceous inventory of the presynaptic active zone has recently been reported. This review will evaluate the subcellular fractionation protocols and the proteomic approaches employed. A breakthrough for the identification of the proteome of the presynaptic active zone was the successful employment of antibodies directed against a cytosolic epitope of membrane integral synaptic vesicle proteins for the immunopurification of synaptic vesicles docked to the presynaptic plasma membrane. Combining immunopurification and subsequent analytical mass spectrometry, hundreds of proteins, including synaptic vesicle proteins, components of the presynaptic fusion and retrieval machinery, proteins involved in intracellular and extracellular signaling and a large variety of adhesion molecules, were identified. Numerous proteins regulating the rearrangement of the cytoskeleton are indicative of the functional and structural dynamics of the presynapse. This review will critically discuss both the experimental approaches and prominent protein candidates identified. Many proteins have not previously been assigned to the presynaptic release sites and may be directly involved in the short- and long-term structural modulation of the presynaptic compartment. The identification of proteinaceous constituents of the presynaptic active zone provides the basis for further analyzing the interaction of presynaptic proteins with their targets and opens novel insights into the functional role of these proteins in neuronal communication.
Highlights
Half a century of subcellular fractionation of brain tissue and protein identification culminated in the identification of the proteome of synaptic vesicles and the presynaptic active zone from murine brain
The purification of the presynaptic active zone was preceded by subcellular fractionation of metabolically intact nerve endings, named synaptosomes [5], which had already been reported in the early sixties of the last century [6,7]
Refinement of subcellular fractionation, immunopurification and mass spectrometry culminated in the isolation and identification of the proteinaceous inventory of the subcellular compartment containing synaptic vesicles docked to the presynaptic plasma membrane, comprising the presynaptic active zone
Summary
Half a century of subcellular fractionation of brain tissue and protein identification culminated in the identification of the proteome of synaptic vesicles and the presynaptic active zone from murine brain. The purification of the presynaptic active zone was preceded by subcellular fractionation of metabolically intact nerve endings, named synaptosomes [5], which had already been reported in the early sixties of the last century [6,7] It is beyond the scope of this review to introduce all individual experimental steps eventually leading to highly purified fractions of the presynaptic compartment comprising the active zone. During exo- and endo-cytosis, synaptic vesicles are tightly bound via a quadruple helical SNARE complex to the presynaptic plasma membrane [19] This allows the immunopurification of the active zone employing antibodies directed against a cytosolic epitope of membrane integral vesicle proteins. We discuss recent information concerning the proteome of the presynaptic active zone derived from mouse brain focusing on those proteins that are potentially involved in the short- and long-term structural regulation of the mature presynaptic compartment
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