Abstract
The secretory pathway in neurons requires efficient targeting of cargos and regulatory proteins to their release sites. Tomosyn contributes to synapse function by regulating synaptic vesicle (SV) and dense-core vesicle (DCV) secretion. While there is large support for the presynaptic accumulation of tomosyn in fixed preparations, alternative subcellular locations have been suggested. Here we studied the dynamic distribution of tomosyn-1 (Stxbp5) and tomosyn-2 (Stxbp5l) in mouse hippocampal neurons and observed a mixed diffuse and punctate localization pattern of both isoforms. Tomosyn-1 accumulations were present in axons and dendrites. As expected, tomosyn-1 was expressed in about 75% of the presynaptic terminals. Interestingly, also bidirectional moving tomosyn-1 and -2 puncta were observed. Despite the lack of a membrane anchor these puncta co-migrated with synapsin and neuropeptide Y, markers for respectively SVs and DCVs. Genetic blockade of two known tomosyn interactions with synaptotagmin-1 and its cognate SNAREs did not abolish its vesicular co-migration, suggesting an interplay of protein interactions mediated by the WD40 and SNARE domains. We hypothesize that the vesicle-binding properties of tomosyns may control the delivery, pan-synaptic sharing and secretion of neuronal signaling molecules, exceeding its canonical role at the plasma membrane.
Highlights
Neural communication is established by the controlled release of signaling molecules from synaptic vesicles (SVs) and large dense-core vesicles (DCVs)
Besides a diffuse distribution in neurites and accumulation at synapses, tomosyn co-localized with moving SVs and DCVs in living neurons
The presence of at least a third type of tomosyn-containing transport organelles was suggested by fast-moving tomosyn puncta that did not co-migrate with synapsin- or Neuropeptide Y (NPY)-mCherry (Fig 5E and 5F)
Summary
Neural communication is established by the controlled release of signaling molecules from synaptic vesicles (SVs) and large dense-core vesicles (DCVs). Coordinated transport is essential to deliver secretory vesicles and their cargos to sites of release. For synapse formation in young neurons, multiple active zone proteins are packaged and co-transported in piccolo-bassoon transport vesicles (PTVs) [1,2], while synaptic vesicle components are transported by synaptic vesicle precursor (SVP) organelles [3,4]. Vesicle targeting of tomosyn by redundant interactions project 91111009 to the VU/VUmc EM facility and 91113022 to AG)
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