A common molecular basis for membrane docking and functional priming of synaptic vesicles

Abstract

Soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complexes execute synaptic vesicle (SV) fusion. Vesicle fusion is preceded by an obligatory Munc13-dependent priming process that conveys fusion competence to SVs by facilitating SNARE complex assembly. Ultrastructural studies after chemical fixation indicated that vesicle docking to the plasma membrane is independent of Munc13s but these results may be misleading because aldehyde fixatives modify the localization of SVs with respect to the plasma membrane. To reinvestigate the role of Munc13s in vesicle docking, cultured hippocampal slices were immobilized using high-pressure freezing, which circumvents aldehyde artifacts. High-pressure freezing was combined with electron tomography to reach a resolution that allows the characterization of details of SV docking in a close-to-native state. In control slices, docked vesicles are not hemifused with the plasma membrane but linked to it and to dense material at the active zone by small strands. In slice cultures from Munc13-deficient mice, vesicles are not docked to the active zone plasma membrane. These results indicate that SV docking at the plasma membrane and functional priming are respective morphological and physiological manifestations of the same molecular process mediated by SNARE complexes and Munc13s.

Variable priming of a docked synaptic vesicle

“…However, at all synapses where AZM relationships have been examined by electron tomography, AZM macromolecules link the VM to the PM (2,11,12,14,27,37,(56)(57)(58)(59). This commonality, together with the high degree of homology of proteins involved in docking and priming at all synapses (60), suggests that our AZM-mediated variable force hypothesis (Fig.…”

“…At typical synapses, docking and fusion take place at structurally specialized regions along the PM called active zones (9, 10). Several lines of evidence suggest that the formation of the SNARE core complex occurs in the macromolecules composing the common active zone organelle, active zone material (AZM) (2,(11)(12)(13)(14), which is positioned near Ca 2+ channels concentrated in the PM at active zones (15-17). Influx of Ca 2+ through the channels after the arrival of a nerve impulse triggers fusion of the VM of docked SVs with the PM.…”

“…However, as described for frog neuromuscular junctions (20-23), the number of SVs that can fuse with the PM after arrival of an impulse varies greatly with differing concentrations of cytosolic Ca 2+ , indicating that priming is more complex than a simple binary transition. Biochemical and electrophysiological approaches have provided evidence that priming is mediated by interactions between the SNARE proteins and their regulators (7,(12)(13)(14)24) and can involve differences in positioning of docked SVs relative to Ca 2+ channels (25). Biochemistry has also led to the suggestion that primed SVs may become deprimed (26).…”

Protein scaffolds in the coupling of synaptic exocytosis and endocytosis

“…Small filaments that tether docked synaptic vesicles to the plasma membrane and to CAZ filaments have been observed by electron microscopy 105 . Cryoelectron tomography revealed that docked synaptic vesicles did not contact the active zone membrane directly but were linked to it by tethers of different lengths 106 .…”

<b>Inhibitory role of Munc13-1 in antigen-induced mast cell </b><b>degranulation </b>

“…Ca 2+ -dependent regulated exocytosis is generally subdivided into three processes, vesicle docking, priming, and fusion, in chronological order (19,31). Docking and priming are defined as a process that involves the physical contact between vesicles and the PM, and the subsequent formation of the trans-SNARE complex, which is ready for membrane fusion, respectively (19); however, they are thought to be closely linked processes, because a common molecular basis for their regulation has been discovered (28,34). One of the molecules that possibly regulates both docking and priming is a protein of the Munc13 family, which contains conserved domains for the assembly of the SNARE complex, and possesses membrane-binding activity (18).…”