Phosphorylation-regulated axonal dependent transport of syntaxin 1 is mediated by a Kinesin-1 adapter

Abstract

Presynaptic nerve terminals are formed from preassembled vesicles that are delivered to the prospective synapse by kinesin-mediated axonal transport. However, precisely how the various cargoes are linked to the motor proteins remains unclear. Here, we report a transport complex linking syntaxin 1a (Stx) and Munc18, two proteins functioning in synaptic vesicle exocytosis at the presynaptic plasma membrane, to the motor protein Kinesin-1 via the kinesin adaptor FEZ1. Mutation of the FEZ1 ortholog UNC-76 in Caenorhabditis elegans causes defects in the axonal transport of Stx. We also show that binding of FEZ1 to Kinesin-1 and Munc18 is regulated by phosphorylation, with a conserved site (serine 58) being essential for binding. When expressed in C. elegans, wild-type but not phosphorylationdeficient FEZ1 (S58A) restored axonal transport of Stx. We conclude that FEZ1 operates as a kinesin adaptor for the transport of Stx, with cargo loading and unloading being regulated by protein kinases. T he formation and maintenance of presynaptic boutons are intricate but highly efficient processes during which the machinery for exocytosis and recycling of synaptic vesicles is assembled from preformed units. These units are delivered to the nascent synapse via molecular motor proteins of the kinesin superfamily (reviewed in Ref. 1).Although Kinesin-3 appears to be the main motor transporting synaptic vesicle precursors, recent evidence suggests that Kinesin-1 (KIF5) is also involved. In Drosophila, deletion of UNC-76/ fasciculation and elongation protein zeta 1 (FEZ1), a specific adaptor for Kinesin-1, left synaptic vesicles stranded in the axon (2, 3), showing that Kinesin-1 is needed at least during later phases of axonal transport. Transport of the synaptic vesicle protein synaptotagmin by the UNC-76/Kinesin-1 complex requires phosphorylation of UNC-76 by the UNC-51/ATG1 kinase, a prerequisite for UNC-76 to bind synaptotagmin (3). Deletion of this kinase phenocopies deletion of UNC-76. Indeed, phosphorylation-regulated interactions between cargo, adaptors, and kinesins have also been observed for other transport complexes such as the kinesin light chain/JIP1 (c-Jun N-terminal kinase-interacting protein 1) complex (4). This suggests that phosphorylation is a common mechanism for the regulation of kinesin-based transport complexes (5).Less is known about the involvement of Kinesin-1 in the transport of other classes of synaptic precursor vesicles. Transport of syntaxin 1a (Stx), an essential component of the exocytotic release apparatus residing in the presynaptic plasma membrane, is clearly distinct from synaptic vesicle precursors and appears to involve a complex between Kinesin-1 and the Stx-binding protein syntabulin (6, 7). Down-regulation or expression of dominant-negative syntabulin reduces but does not abolish membrane delivery of Stx, indicating the existence of other transport mechanisms (6). Moreover, proper intracellular trafficking of Stx and its function in exocytosis depends on Munc18 coexpression (8-14). Stx tra...

Munc18-1 redistributes in nerve terminals in an activity- and PKC-dependent manner

“…3 E). In developing neurons Munc18-1 is bound to transport vesicles (Chua et al, 2012). We observed Munc18-1-Venus puncta entering the bleached area in only 16% of Munc18-1-Venus axons (Fig.…”

“…During neuronal development, Munc18-1/syntaxin-1 and syntaxin-1 are transported by FEZ1-KIF5C (Chua et al, 2012) and syntabulin-KIF5B (Su et al, 2004;Cai et al, 2007) transport complexes. Our FRAP experiments in axons (Fig.…”

“…In developing neurons, syntaxin-1 is transported on vesicles via pathways dependent on the kinesin adaptor proteins syntabulin (Su et al, 2004) and FEZ1 (Chua et al, 2012). Munc18-1 is present in syntaxin-FEZ1-kinesin transport complexes (Chua et al, 2012).…”

“…Munc18-1 is present in syntaxin-FEZ1-kinesin transport complexes (Chua et al, 2012). At this stage, the syntaxin-1/Munc18-1 dimer may M unc18-1 is a soluble protein essential for synaptic transmission.…”

Schwannomin-interacting Protein 1 Isoform IQCJ-SCHIP1 Is a Multipartner Ankyrin- and Spectrin-binding Protein Involved in the Organization of Nodes of Ranvier

“…The functional relevance of this phenotype is not known because to our knowledge it was not described before. Interestingly, the coiled-coil region of SCHIP1 presents sequence homologies with a coiled-coil region within the C-terminal domain of the protein FEZ1 (8), which plays a role in kinesin-mediated anterograde transport of vesicles and mitochondria in axons (37)(38)(39). In addition, although we showed that the SD domain of ␤IV-spectrin is the primary binding site for SCHIP1, our results do not strictly exclude the possibility that SCHIP1 could interact with other ␤-spectrins such as ␤III-spectrin, which has been shown to interact with dynactin (40) and is implicated in dynein-mediated vesicular transport (41,42).…”