Munc18a does not alter fusion rates mediated by neuronal SNAREs, synaptotagmin, and complexin.
Bottom Line: Moreover, a phosphorylation mimic mutant of Munc18a with reduced affinity to syntaxin-1A results in less reduction of vesicle association.In summary, Munc18a does not directly affect fusion, although it has an effect on the t-SNARE complex, depending on the presence of other factors and experimental conditions.Our results suggest that Munc18a primarily acts at the prefusion stage.
Affiliation: From the Departments of Molecular and Cellular Physiology, Neurology and Neurological Sciences, Structural Biology, and Photon Science and.Show MeSH
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Mentions: So far, there is little evidence from reconstitution experiments that Munc18a plays a major role in fusion itself, although stimulatory effects by SM proteins on SNARE-dependent lipid mixing were observed using ensemble fluorescence assays (16–18). A stimulatory effect of Munc18a was also observed in conjunction with neuronal SNAREs using a single vesicle-vesicle lipid-mixing assay (19). However, all of these experiments only used neuronal SNAREs in addition to Munc18a. Subsequent ensemble experiments that reconstituted neuronal SNAREs in liposomes along with neuron-specific proteins synaptotagmin-1 and complexin-1 showed only a relatively small increase of spontaneous (i.e. at zero Ca2+ concentration) lipid mixing and content mixing upon Munc18a inclusion when complexin-1 was present (see Fig. 3 (D and F) in Ref. 20).
Affiliation: From the Departments of Molecular and Cellular Physiology, Neurology and Neurological Sciences, Structural Biology, and Photon Science and.