Molecular mechanisms of protein sorting in the synaptic vesicle life cycle.

Synaptic vesicle (SV) proteins are synthesized at the level of the cell body and transported along the axons in precursor vesicles which undergo cycles of exo-endocytosis in transit to either the distal growth cone or the nerve terminal. I sought to investigate the mechanisms underlying the sorting of SV components and to determine whether and how SV exocytosis is regulated prior to synaptogenesis. SV fusion is underlain by the interaction of vesicle-associated membrane protein (VAMP) 2 with plasma membrane partners. Fluorescence resonance energy transfer analysis reveals that VAMP2 interacts with Synaptophysin I (SypI), a major resident of SVs, on the SV membrane and that exocytotic stimuli cause dissociation of this complex at a stage preceding SV fusion. This observation is consistent with the idea that SypI limits VAMP2 availability for fusogenic complexes. The interaction between SypI and VAMP2 occurs early along the exocytotic pathway and is required in order for SypI to govern the sorting of VAMP2 to SVs. The control of VAMP2 sorting by its negative regulator SypI establishes a mechanism which prevents the fusion at inappropriate sites of SVs directed to the nerve terminal. The study of SV dynamics in developing hippocampal neurons reveals that cAMP-dependent pathways affect SV distribution and recycling in the axonal growth cone and that these effects are mediated by the SV-associated phosphoprotein synapsin I. Synapsin I and its phosphorylation by cAMP-dependent protein kinase A play a pivotal role in regulating SV organization and dynamics in neuronal growth cones and in determining the formation of synaptic contacts. These results provide new clues as to the bases of the well known activity of synapsin I in synapse maturation and indicate that molecular mechanisms similar to those operating at mature nerve terminals are active in developing neurons to regulate the SV life cycle prior to synaptogenesis