Synapses are specialized contact sites that mediate information flow between neurons and their targets. Important physical interactions across the synapse are mediated by synaptic adhesion molecules. These adhesions regulate formation of synapses during development and play a role during mature synaptic function. Importantly, genes regulating synaptogenesis and axon regeneration are conserved across the animal phyla. Genetic screens in the nematode Caenorhabditis elegans have identified a number of molecules required for synapse patterning and assembly. C. elegans is able to survive even with its neuronal function severely compromised. This is in comparison with Drosophila and mice where increased complexity makes them less tolerant to impaired function. Although this fact may reflect differences in the function of the homologous proteins in the synapses between these organisms, the most likely interpretation is that many of these components are equally important, but not absolutely essential, for synaptic transmission to support the relatively undemanding life style of laboratory maintained C. elegans. Here, we review research on the major group of synaptic proteins, involved in the presynaptic machinery in C. elegans, showing a strong conservation between higher organisms and highlight how C. elegans can be used as an informa- tive tool for dissecting synaptic components, based on a simple nervous system organization. Keywords C. elegans · Synapse · Synaptic proteins · Synaptogenesis · Synaptic vesicles Background exocytosis (Katz 1971, 1979; Sudhof 2004). A precise neu- ronal reaction requires that SVs are clustered adjacent to Transmission of a signal within a neuron is carried by depo- the release site or presynaptic active zone. Here the SVs are larization of the resting membrane potential typically involv- docked and held in contact with the cell membrane by the ing a transient reversal of the resting membrane potential. docking complex, where they are primed for fusion. Then, a 2+ The depolarization leads to the opening of voltage-gated depolarization induces the opening of Ca channels, and the 2+ 2+ Ca channels in the presynaptic membrane. Then, the open-rising Ca concentration stimulates SV-plasma membrane 2+ ing of these channels causes a rapid influx of Ca into the fusion. For cells to respond rapidly and reliably to incom- presynaptic terminal, facilitating synaptic vesicles fusion ing depolarizing potentials, they must maintain a sufficient with the presynaptic plasma membrane of the neuron, and supply of vesicles containing neurotransmitter close to the causing neurotransmitters to be released into the synaptic active zone where the content is released from the presynap- cleft. Neurotransmitters in the synaptic cleft diffuse away tic neuron. However, there are neurons that use only graded from the release site and have ready access to binding sites voltage signals. These ‘non-spiking’ neurons that encode on synaptic receptors localized on both the post- and pre- information as graded potentials typically have higher infor- synaptic membrane. They bind to their cognate receptors mation rates compared to ‘spiking neurons’ (DiCaprio et al. and elicit a transduction cascade to bring about the cellular 2007). Graded potentials are a consequence of the passive response. In nerve terminals, neurotransmitters are packaged electrical property of the membrane and depolarizing poten- 2+ into synaptic vesicles (SVs) and released by Ca -induced tials the result of a coordinated response (van Steveninck and Laughlin 1996). Finally, neurotransmitter release can occur by action potential-independent spontaneous vesicle fusion. * Fernando Calahorro Although for decades it was thought that spontaneous trans- F.Calahorro@soton.ac.uk mission was a consequence of ‘leaky’ synapse, recent data show that this alternative mechanism underpins signalling Biological Sciences, University of Southampton, Life Sciences Building 85, Southampton SO17 1BJ, UK Vol.:(0123456789) 1 3 4 Page 2 of 13 Invertebrate Neuroscience (2018) 18:4 roles involve in synapse maturation and homoeostatic plas- pool (occupying most of the vesicle clusters and only recy- ticity (Ramirez and Kavalali 2011). cling upon C strong stimulation) (Sudhof 2013). Neurotransmitters are secreted from neurons by two types Key insights into the molecular mechanisms of synaptic of vesicles that are classified by their size and appearance in events have come from research using genetic model systems electron micrographs. Small clear synaptic vesicles (SCV) such as the nematode Caenorhabditis elegans (Richmond (40–60 nm diameter) contain small molecule, so-called clas- 2005). Of particular note are studies employing mutants with sical transmitters, such as glutamate, GABA and acetylcho- uncoordinated locomotion (unc genes) (Brenner 1974) to line, that activate postsynaptic ionotropic receptors mediat- define key synaptic determinants, optogenetics coupled with ing fast synaptic transmission, and metabotropic receptors high-pressure freezing to resolve the relationship between mediating a more slow and sustained transmission. Dense- docking and fusion (Watanabe et al. 2013) and genetic core vesicles (DCVs) (60–120 nm diameter) are character- manipulation of syntaxin to define priming events (McEwen ized by their electron dense appearance and larger diameter and Kaplan 2008). Here we provide a review of regulated relative to SCVs, containing neuropeptides and biogenic exocytosis in C. elegans by presynaptic elements, highlight- amine neuromodulators such as serotonin and dopamine. ing C. elegans synapse as a powerful tool to dissect synaptic DCVs dock at the plasma membrane but might be excluded components and understanding key synaptic processes. We from active zones (Hammarlund et al. 2008). There are comment on the opportunity for future research directions similarities in the fusion machinery for both vesicle types, deploying this model organism. but also there are differences in the kinetics of exocytosis, docking localization and physiological regulation of release (Martin 2003; Rettig and Neher 2002), suggesting that there Attachment of vesicles to the cytoskeleton are proteins and mechanisms that are distinct for SCV- and DCV-mediated exocytosis. The principal synaptic protein which functions as a cytoskel- Current models describing the molecular mechanism of eton anchor for vesicles in the reserve vesicle pool is syn- 2+ Ca -regulated synaptic vesicle exocytosis and endocytosis apsin (Fig. 2). Synapsins comprise a family of synaptic divide the process into multiple steps, leading ‘preferred’ vesicle proteins that have been identified in a variety of models: docking, priming, fusion, exocytosis (Fig. 1) (Jung invertebrate and vertebrate species (Stavoe et al. 2012; and Haucke 2007; Sudhof 1995, 2004). This process is facil- Cesca et al. 2010). In C. elegans, there is a homologue of itated by the formation of a complex between molecules on synapsin protein (SNN-1) which is most similar to verte- the synaptic vesicle and molecules attached to the plasma brate synapsin II. In vertebrates, synapsins present highly 2+ membrane. Ca binding to synaptotagmin triggers release conserved domains among the different isoforms. The best by stimulating synaptotagmin binding to a molecular com- characterized domains are: domain A containing a phos- plex composed of SNARE (‘soluble NSF attachment recep- phorylation site for PKA/CaMKI that regulates binding to tor’) and SM (‘Sec1/Munc18-like’) proteins that mediates synaptic vesicles, domain C containing ATP binding sites membrane fusion during exocytosis. Synaptic vesicles con- and domain E that regulates the reserve pool of synaptic taining synaptotagmin are positioned at the active zone, the vesicles. In C. elegans, snn-1 presents a conserved domain site of vesicle fusion, by a protein complex containing RIM organization with PKA/CaMKI site within domain A, sev- proteins. RIM proteins simultaneously activate docking and eral ATP binding sites in domain C and a highly conserved 2+ priming of synaptic vesicles and recruit Ca channels to domain E (Cesca et al. 2010). ssn-1 is expressed in neurons active zones, thereby connecting within a single complex exhibiting patterns consistent with localization to vesicles 2+ the primed synaptic vesicles to Ca channels. This archi- in presynaptic regions. Very little is known about the exact 2+ 2+ tecture allows direct flow of Ca ions from Ca channels role of SNN-1 in C. elegans, since snn-1 mutants display to synaptotagmin, mediating tight millisecond coupling of predominantly wild-type phenotypes. However, a detailed 2+ a depolarization to neurotransmitter release. Influx of Ca analysis of specific synapses in snn- 1 mutants reveals syn- then leads to the rapid completion of membrane fusion and aptic vesicle clustering defects in the sensory neuron AIY the release of the neurotransmitter (Rizo and Sudhof 2002). (Stavoe et al. 2012) and resistance to paralysis on aldicarb Finally, after fusion the vesicular components are recycled (Sieburth et al. 2005). This latter phenotype is indicative of through endocytosis to replenish the synaptic vesicle pools. reduced acetylcholine release at the body wall neuromuscu- Three functional and morphological classes of vesicle pools lar junction as the paralysis is induced by accumulation of have been assigned: the readily releasable pool (docked at acetylcholine in the presence of the cholinesterase inhibi- active zones and ‘ready to go’ upon stimulation), the recy- tor aldicarb. This assay has been extensively deployed to cling pool (scattered throughout the nerve terminals and resolve genetic determinants of cholinergic transmission in recycling upon moderate stimulation) and finally the reserve C. elegans (Mahoney et al. 2006). 1 3 Invertebrate Neuroscience (2018) 18:4 Page 3 of 13 4 Fig. 1 Molecular mechanisms of biogenesis and exocytosis of synap- based on ‘preferred’ models where docking is before priming. The tic vesicles. Under resting conditions, synaptic vesicles are stored in last interpretation is supported by evidence, among others, such as the cytoplasm of the nerve terminal. Vesicles are loaded with neuro- rab-3 and unc-18 knockouts present an alteration in vesicle docking transmitter through an active processes requiring a neurotransmitter although the docking is not completely disrupted (Nonet et al. 1997; transporter and a vacuolar-type proton pump ATPase that provides Weimer et al. 2003). Finally, synaptic vesicles are regenerated within a pH and electrochemical gradient. These transporters are selective the nerve terminal probably through one of the three proposed path- for different classes of transmitters. The identity of many of these ways (not shown in the diagram): a pathway in which vesicles endo- transporters was determined through the molecular characterization cytose by closure of the fusion pore and are refilled with neurotrans- of C. elegans mutants. Filled vesicles dock at the active zone (repre- mitters while remaining docked to the active zone (kiss-and-stay); sented by a thick grey line), where they undergo a priming reaction a local recycling pathway that is clathrin independent but results in 2+ that makes them competent for Ca -triggered fusion-pore opening. mixing vesicles with the reserve pool after endocytosis (kiss-and- Priming involves all steps required to acquire release preparation of run); and a pathway whereby vesicles undergo clathrin-mediated the exocytosis complex. In special situations—i.e., during sustained endocytosis and recycle either directly or via endosomes, ultrafast activity, the priming could precede docking, resulting in immediate endocytosis removes membrane added by vesicle fusion at the lat- fusion of vesicles. After exocytosis, the vesicle proteins remain clus- eral edge of the active zone. Large endocytic vesicles then fuse to tered in the plasma membrane to be recycled by endocytosis. The endosomes, and in this way, newly formed synaptic vesicles can be double arrow between docking and priming representations indi- recruited back to the active zone cates that priming can precede docking instead to the interpretations The synaptic vesicle clustering in mammals is regu- proteins that link synaptic vesicles to the presynaptic lated by F-actin protein through two different pathways, cytoskeletal matrix by interacting with actin (Bahler and one dependent on synapsins and the other independent Greengard 1987; De Camilli et al. 1983; Li et al. 1995). (Nelson et al. 2013). In mammals, the synapsin family They are expressed in neurons exhibiting patterns con- consists of at least 9 isoforms encoded by 3 distinct genes sistent with localization to vesicles in presynaptic regions which are characterized by a mosaic of conserved and var- (Sieburth et al. 2005), having multiple functions within iable domains (Fornasiero et al. 2010). Specifically, the presynaptic terminals, including anchoring of synaptic N-terminal portion of all synapsins is highly conserved, vesicles to the actin cytoskeleton, recruitment of them to whereas the C-terminal portion is variable because of het- a reserve pool and regulation of the fusion of SVs (De erogeneous combinations of two different domains (Kao Camilli et al. 1990; Pieribone et al. 1995; Hilfiker et al. et al. 1999; Porton et al. 2011). Synapsins are vesicle 1998). They are also implicated in neuronal development, 1 3 4 Page 4 of 13 Invertebrate Neuroscience (2018) 18:4 Fig. 2 Molecular protein complexes that organize the secretory channels, RIM proteins directly bind to the vesicle protein Rab3, to machinery at the presynaptic active zone. The vesicle clusters dock the priming factor Munc-13. Munc-13 directly activates the SNARE at the active zone through Rab proteins, CAPs protein (UNC-31), protein assembly. Both RIM and Munc-13 proteins are tightly regu- Munc-18 (UNC-18) and tomosyn. RIM (UNC-10) protein places the lated in a manner that determines presynaptic plasticity. The diagram 2+ priming factor Munc-13 and Ca channels into close proximity to is based on the Sudhof’s synaptic model (Sudhof 2013) and repre- synaptic vesicles and SNARE protein complex-dependent (synapto- sents a magnified view of vesicle docking shown in Fig. 1 2+ brevin, SNAP-25, syntaxin) fusion machinery. In addition to Ca synaptogenesis and maintenance of mature synapses (Fer- and tomosyn (Fig. 2). Rab proteins present a key role regu- reira et al. 2000). lating the recruitment of vesicles to the active zone in C. Studies in mice show that mutants lacking synapsin I appear elegans. The Rab proteins are a large family of monomeric to develop normally and do not have gross anatomical abnor- GTPases, conserved from yeast to humans, which through malities. However, in these mutants, the giant fibre terminals, specific scaffolding with distinct interactors specify presyn- in the CA3 area of the hippocampus, are significantly smaller, aptic function (Bock et al. 2001; Stenmark and Olkkonen the number of synaptic vesicles is reduced, and the presynaptic 2001). In C. elegans, there are 31 members of the Rab fam- structures altered (Takei et al. 1995). Furthermore, suppression ily, 29 of which are also found in humans as orthologues of synapsin II leads to an inhibition of developing and synapse (Gallegos et al. 2012). Probably, the most extensively stud- formation in hippocampal neurons. Similarly, a depletion of ied Rab proteins in C. elegans are RAB-27/AEX-6 and synapsin III affects the extension of processes and axon differ - RAB-3, homologues of human RAB-27 and RAB-3, respec- entiation in hippocampal neurons (Bloom et al. 2003; Ferreira tively. C. elegans rab-27 is expressed in neurons and in the et al. 1995, 2000; Takei et al. 1995). intestine. In the nervous system, rab-27 localizes to synapse- rich regions of the nervous system (nerve ring, dorsal and ventral cord) and partially co-localizes with synaptic vesicle- Docking associated rab-3, although RAB-27 immunostaining is nor- mal in rab-3 mutants, suggesting that RAB-27 localization The vesicle cluster that represents the reserve pool dock at is independent of RAB-3 function (Mahoney et al. 2006). the active zone through a subset of synaptic proteins include This expression profile is consistent with that of mammalian Rab proteins, CAPs protein (UNC-31), Munc-18 (UNC-18) Rab27B, which is also expressed in both brain and intestine 1 3 Invertebrate Neuroscience (2018) 18:4 Page 5 of 13 4 as well as other secretory cells. Both RAB-27 and RAB-3 syntaxin from the closed conformation into the open form present a key role in synaptic transmission process in C. giving the opportunity for syntaxin to form the SNARE elegans, regulating the recruitment of vesicles to the active core complex required for priming and docking (Basu et al. zone or sequestration of vesicles near release sites (Mahoney 2005; Betz et al. 1997; Hammarlund et al. 2008). Finally, the et al. 2006). In vertebrates, Rab molecules regulate vesicu- DCVBD domain mediates CAPS targeting to DCV (Gris- lar trafficking in many different transport pathways for both hanin et al. 2002). exocytosis and endocytosis in neural and non-neuronal tis- CAPS was first identified as an essential protein for sues, but in C. elegans RAB-3 in neurons specifically plays noradrenaline release from PC12 cells and recognized as a crucial role in regulating synaptic vesicle-mediated release being orthologous to C. elegans UNC-31 which had previ- (Nonet et al. 1997). As a consequence, C. elegans rab-3 ously been shown to be involved in neurosecretion (Walent mutants present slight behavioural abnormalities. They are et al. 1992). As the name of the protein suggests, mutations resistant to the paralytic action of the cholinesterase inhibitor in unc-31 result in uncoordinated motor behaviour and aldicarb suggesting that cholinergic transmission is gener- the worms are constitutively lethargic with slow and soft ally depressed (Mahoney et al. 2006; Nonet et al. 1997), as movements (Avery et al. 1993). In addition, unc-31 mutants well as exhibiting an altered morphology of neuromuscular feed constitutively and they have defects in egg laying and junctions (Nonet et al. 1997). There is a depletion of ≈ 40% failures in recovery from dauer, a metabolically quiescent of normal levels in vesicle population at synapses (identi- developmental larval stage of C. elegans (Avery et al. 1993; fied by electron microscopy) accompanied by an elevation Dalliere et al. 2016). Consistent with this, the expression of these populations in inter-synaptic regions of the axons pattern of unc-31 reveals a broad distribution in the nervous consistent with a deficit in SCV trafficking (Mahoney et al. system (Charlie et al. 2006). 2006; Nonet et al. 1997). In addition, extracellular electro- UNC-31/CAPS has been associated with exocytosis physiological recordings reveal an impairment of synaptic mediated by DCVs (Berwin et al. 1998), and in line with transmission in the pharyngeal nervous system (Nonet et al. this, in C. elegans it has been found that loss of the sin- 1997). There is only one isoform of rab-27 in C. elegans, gle isoform of UNC-31 decreases neuropeptide secretion while two isoforms are found in mammals. Like the rab- accompanied by an increase in neuropeptide abundance in 3 mutants, C. elegans rab-27 mutants are slightly aldicarb motor axons (Sieburth et al. 2007). Mammals express two resistant indicative of a reduction in cholinergic signalling, isoforms of CAPS, CAPS1 and CAPS2, with similar func- and exhibiting defecation defects consistent with neuromus- tions but which differ in their spatiotemporal expression cular transmission dysfunction in anterior body wall muscle pattern (Sadakata et al. 2007c; Speidel et al. 2003). CAPS1 contraction and expulsion steps of the defecation motor pro- is essential for the uptake or storage of catecholamines in gram (Mahoney et al. 2006). DCVs (Speidel et al. 2005), while CAPS2 appears to be In humans, the number of Rab genes reaches up to 60 required for DCV-mediated neurotrophin secretion in the where 33 of them have been identie fi d by proteomic analyses cerebellum (Sadakata et al. 2007a, c). In addition, in CAPS- in synaptic vesicle fractions (Takamori et al. 2006). Of these, 1/CAPS-2 double null mutants DCV secretion is severely Rab-3A, 3B, 3C, 3D and 27-B are involved in exocytosis, reduced (Farina et al. 2015). While early studies seemed to while Rab-4, 5, 10, 11B and 14 are intermediates of synap- indicate that CAPS is not required for exocytosis of gluta- tic vesicle recycling such as early endosomes (Binotti et al. mate-containing SCVs (Tandon et al. 1998), this has been 2016). Specifically, Rab-3A and Rab-27B are the best inves- revised with further investigation that provides evidence for 2+ tigated and play overlapping roles during Ca -triggered a more overlapping functional role with SCV-mediated exo- neurotransmitter release in mammals (Schluter et al. 2002). cytosis. CAPS1 and CAPS2 double knockout mice exhibit Another indispensable actor for synaptic vesicle-medi- specific priming defects in glutamatergic transmission (Jock - ated exocytosis is UNC-31. This is also known as CAPS usch et al. 2007). In Drosophila melanogaster, in which a 2+ (Ca -dependent activator protein for secretion) and is a single gene encodes dCAPS, there is a ≈ 50% loss in evoked multi-domain protein containing, from the N to the C ter- glutamatergic transmission at the neuromuscular junction, as minus, a dynactin 1 binding domain (DBD), a C2 domain, well as an accumulation of synaptic vesicles at active zones a PH domain, a (M)UNC-13 homology domain (MHD) (Renden et al. 2001). and finally a DCV binding domain (DCVBD) (Ann et al. Munc-18 proteins are the mammalian homologue of 1997). The DBD is required for CAPS sorting (Sadakata UNC-18 proteins in C. elegans and are a member of the 2+ et al. 2007b). The C2 domain, as a Ca sensor, mediates Sec1/Munc18-like (SM) protein family. Munc-18 is a 2+ Ca -dependent binding to phospholipids (Rizo and Sudhof key synaptic protein acting during multiple stages of 1998). The PH domain interacts with acidic phospholipids the exocytosis including vesicle priming, docking and and binds with plasma membranes (Lemmon 2008). The fusion. Although during these steps syntaxin interactions MHD domain directly interacts with syntaxin, transforming are required, Munc-18 also regulates vesicle fusion via 1 3 4 Page 6 of 13 Invertebrate Neuroscience (2018) 18:4 syntaxin-independent interactions. These syntaxin interac- a subset of neurons in head and tail ganglia (Dybbs et al. tions are possible through a Munc-18 closed conformation 2005). The synapses in C. elegans tom-1 mutants present of syntaxin binding. In C. elegans, UNC-18 is a protein no changes in neuronal outgrowth or in synaptogenesis but required in neurons for synaptic vesicle-mediated exocy- exhibit prolonged evoked postsynaptic responses. This lat- tosis. Characterization of unc-18 reveals a localization in ter phenotype is accompanied by an increase in the number ventral-cord motor neurons and some unidentified head neu- of plasma membrane-contacting vesicles (Gracheva et al. rons in the adult hermaphrodite (Gengyo-Ando et al. 1993). 2006). Thus, tomosyn-deficient mutants have increased syn- A similar expression is observed in males, but also a strong aptic transmission, an increased pool of primed vesicles and expression in the gonad (Schindelman et al. 2006). unc-18 increased abundance of UNC-13 (a synaptic protein involved 2+ mutants are deficient in synaptic transmission with a reduc-in Ca -triggered fusion-pore opening described in the next tion in neurotransmitter release and a consequent resistance sections) at synapses (McEwen et al. 2006). This indicates to aldicarb (Graham et al. 2011; Gracheva et al. 2010). The that priming is negatively regulated by TOM-1 and that there introduction of a gain of function mutation in a functionally is a fine balance between tomosyn and UNC-13, with the important domain (3b) within the UNC-18 protein confers availability of open syntaxin a possible mechanism for this a hypersensitivity to aldicarb (Graham et al. 2011). Munc- regulation (McEwen et al. 2006). 18 has a function in several exocytosis processes requiring Overall, these findings from C. elegans along with stud- syntaxin-dependent interactions; however, data based on C. ies using mouse models have shown tomosyn has a diffuse elegans studies reveal a key role of domain 3b of Munc-18 distribution in neurites and is accumulated at synapses co- in transducing regulation of vesicles fusion independent of localized with both moving SCVs and DCVs, regulating closed-conformation syntaxin binding (Graham et al. 2011). their mediated secretion. This suggests a function control- This fact highlights C. elegans as a powerful and key tool to ling the delivery, synaptic sharing and secretion of neuronal discover functional analysis of synaptic proteins, enhanced signalling molecules (Geerts et al. 2017). by the availability of CRISPR editing. Physiological data and electron micrographs of C. elegans neuromuscular junc- tion provide evidence that in the absence of UNC-18 the Priming size of the ready releasable pool of vesicles is drastically reduced (Weimer et al. 2003). Thus, unc-18 mutants present After the docking of vesicles at the active zone, they undergo a reduction in docked vesicles at the active zone, indicating a priming reaction regulated by a few key synaptic ele- that UNC-18 functions as a facilitator of vesicle docking ments, syntaxins (UNC-64), synaptobrevin (SNB-1) and (Weimer et al. 2003). Overall, the release defects in unc-18 SNAP-25 (RIC-4) (Fig. 2). Nevertheless, there might be a mutants are associated with the lack of two morphologi- regulatory-coupled process between docking and priming cally distinct vesicle pools: those tethered within 25 nm of of the synaptic vesicles (Fig. 1); namely, synapses lack- the plasma membrane and those docked with the plasma ing priming proteins, such as Munc-13 or SNARE, have a membrane (Gracheva et al. 2010). reduced or absence docking of vesicles (Imig et al. 2014). TOM-1 has also a role regulating the macromolecular Syntaxins are a family of transmembrane proteins that par- complex binding between the SNARE proteins syntaxin, ticipate in SNARE complexes to mediate membrane fusion SNAP-25 and synaptobrevin, three synaptic molecules events including exocytosis in different compartments of participating in the priming step. Tomosyn is a soluble the nervous system such as axons, the soma/dendrites or protein first isolated from rat brain as a syntaxin binding astrocytes. In C. elegans, the unc-64 gene encodes syntaxin, partner capable of disrupting Munc18–syntaxin-1a com- a plasma membrane receptor for intracellular vesicles that plexes (Fujita et al. 1998). Tomosyn has two recognizable is orthologous to vertebrate syntaxin 1A and Drosophila domains, an N-terminal domain rich in WD40 repeats and Syx1A. It is expressed in neural cells, especially in motor a C-terminal SNARE domain with high sequence homol- neurons and neurons constituting head ganglions (Ogawa ogy to the R-SNARE domain of synaptobrevin (Hatsuzawa et al. 1998; Yamashita et al. 2009). UNC-64 is required for et al. 2003; Masuda et al. 1998). C. elegans tomosyn (TOM- normal locomotion and possibly also for insulin secretion 1) is a cytosolic syntaxin binding protein implicated in the and is an essential component of the core synaptic vesicle modulation of both constitutive and regulated exocytosis that fusion machinery (McEwen and Kaplan 2008). UNC-64 negatively regulates synaptic vesicle priming in C. elegans interacts with UNC-13, UNC-18 and SNB-1/synaptobrevin (Gracheva et al. 2006). Thus, tomosyn inhibits synaptic (Sassa et al. 1999). Thus, it has been shown that loss of the vesicle priming through its synaptobrevin SNARE motif, N-terminal binding interaction between the syntaxin UNC- which forms an inhibitory SNARE complex with syntaxin 64 and the protein UNC-18 severely impairs neuromuscular and SNAP-25 (McEwen et al. 2006). The expression of tom- synaptic transmission in C. elegans, resulting in an uncoor- 1 is observed in ventral nerve cord motor neurons and in dinated phenotype (Munson and Bryant 2009). In addition, 1 3 Invertebrate Neuroscience (2018) 18:4 Page 7 of 13 4 unc-64–null mutants are unable to move and develop beyond useful tool dissecting the role of synaptobrevins in neuro- the first larval stage (Saifee et al. 1998). C. elegans unc-64 transmitter release. However, using high-density cultures of and mammalian syntaxin-1A are functional orthologues as hippocampal neurons from embryos, a drastic reduction in 2+ shown by the observation that unc-64–null mutant worms Ca -triggered vesicle fusion has been observed (Schoch expressing the mammalian syntaxin-1A wild type are able et al. 2001). to move, grow and reproduce (Park et al. 2016). Another The SNAP protein family consists of several homolo- example of the power of C. elegans to further analyse, in gous proteins of which SNAP-25 is essential for SV fusion this case, the structure/function relationship of syntaxin-1, (Delgado-Martinez et al. 2007). The C. elegans orthologue is the recent discovery that syntaxin-1 N-peptide is criti- of vertebrate SNAP-25, ric-4, appears to be expressed selec- cal when syntaxin adopts an ‘open’ conformation to bend tively in the nervous system including the nerve ring, com- towards Munc-18 (Park et al. 2016). missures, and ventral and dorsal nerve cords (Hwang and Similar to C. elegans, mammalian syntaxins present a Lee 2003). Little is known about the role of ric-4 at the C. typical domain organization where the N-terminal region elegans synapse; however, it is known that the loss of ric-4 contains two different motifs: a short N‐terminal peptide function via RNAi experiments leads to aldicarb resistance, (‘N‐peptide’) that binds to Munc18‐1 (Dulubova et al. 2007), indicating that ric-4 plays a role in synapse structure and and a larger H ‐domain that consists of an autonomously function (Sieburth et al. 2005). Studies using mouse models abc folded three‐helical bundle (Bracher and Weissenhorn show that the deletion of SNAP-25 leads to reduced neuronal 2004). Perhaps, the best-studied membrane–fusion complex survival and impaired arborisation, reduced spontaneous is that mediating synaptic vesicle fusion through syntaxin release, and arrest of evoked release in the surviving neurons 1A/1B (Teng et al. 2001). It has been suggested that STX1A (Delgado-Martinez et al. 2007). In addition, the neurons of and STX1B are functionally redundant. Thus, STX1A KO SNAP-25 null mutant mice (SNAP-25 KO) contain fewer mice show a normal lifespan, and hippocampal neurons with DCVs and have reduced DCV fusion probability in surviv- normal neurotransmission, indicating that STX1B function- ing neurons at DIV14 (days in vitro). Others SNAP family ally compensates the function of STX1A (Fujiwara et al. members such us SNAP-23, SNAP-29 and SNAP-47 are also 2006; Gerber et al. 2008). However, complete loss or partial present in neurons and in synaptic vesicle purifications (Holt loss of STX1B in mice caused a pre-weaning death, sug- et al. 2006). Overexpression of SNAP-29 inhibits synap- gesting that STX1A and STX1B have differential functions tic vesicle fusion possibly via inhibiting SNARE complex (Mishima et al. 2014). disassembly (Pan et al. 2005). Finally, SNAP-47 binds to Synaptobrevins are vesicle-associated proteins involved plasma membrane SNAREs in vitro, but is predominantly in neurotransmitter release (Nonet et al. 1998). In C. elegans, located on intracellular membranes (Holt et al. 2006). SNB-1 is broadly present in nervous system, in neurons in the head ganglia and motor neurons in ventral nerve cord. Particularly, the abundance of SNB-1 in GABAergic motor Ion channel regulation and fusion neurons is controlled by MEC-15, one of a small number of F-box proteins evolutionarily conserved from C. elegans to The priming reaction makes the vesicles competent for 2+ mammal (Sun et al. 2013). C. elegans null snb-1 mutants are Ca -triggered fusion-pore opening. The major elements not viable and die soon after hatching (Nonet et al. 1998). among the synaptic proteins involved in priming of SVs are In an attempt to generate viable C. elegans snb-1-deficient synaptotagmin (SNT-1), synaptogyrin (SNG-1), Munc-13 mutants, the I97D substitution in snb-1(e1563) changes a (UNC-13) and RIM (UNC-10) (Fig. 2). 2+ hydrophobic residue to a charged residue in the TMD of One key factor in Ca regulation and vesicle fusion is the 2+ synaptobrevin, leading to a synaptobrevin with reduction in Ca sensor synaptotagmin, which consists of a short N-ter- function. snb-1 mutants carrying this substitution are via- minal luminal segment, a single transmembrane α-helix, 2+ ble, with grossly normal locomotion (Sandoval et al. 2006). an unstructured linker, and two Ca binding C2 domains, These mutants are resistant to the acetylcholinesterase termed C2A and C2B, respectively. In C. elegans, the two inhibitor aldicarb, indicating that cholinergic transmission synaptotagmin isoforms, snt-1a and snt-1b, are expressed in is impaired, and present abnormal electropharyngeograms neurons, where snt-1a is typically expressed at higher levels which are extracellular recordings of the pharyngeal neuro- and in a larger subset of neurons. In addition, snt-1b is exclu- muscular network (Nonet et al. 1998). sively expressed in the excretory duct cell and a subset of Studies on synaptobrevins in mouse are difficult to carry tail neurons including DVB, a GABAergic neuron required out since synaptobrevin 1 and 2 mutants, the two isoforms for defecation (Nonet et al. 1993; Mathews et al. 2007). extensively expressed in the central nervous system (Schoch Behaviourally, snt-1 mutants present locomotory defects in et al. 2001), immediately die after birth (Nystuen et al. 2007; swimming behaviour, as well as in the defecation motor pro- Schoch et al. 2001). In this sense, C. elegans has been a gram (Mathews et al. 2007). Evoked synaptic transmission 1 3 4 Page 8 of 13 Invertebrate Neuroscience (2018) 18:4 is dependent on interactions between synaptotagmin and the across the nervous system (Abraham et al. 2011). In mouse SNARE complex, comprised of syntaxin, SNAP-25 and syn- and C. elegans, synaptogyrin is completely dispensable for aptobrevin. It has been shown that snt-1 C. elegans mutants nervous system development and performance of basic neu- present a significant reduction in this evoked transmission, ronal functions (Abraham et al. 2006; Eshkind and Leube indicating its key role in SNARE complex assembly (Yu 1995; McMahon et al. 1996). Thus, C. elegans mutants lack- et al. 2013). On the other hand, snt-1 mutants present large ing or overexpressing synaptogyrin present an increased irregular cisternae associated with abnormal endocytosis, sensitivity to the epileptogenic GABA antagonist pentyl- indicating a defect impacting at this level of the vesicle enetetrazole (PTZ), showing a reduced convulsive thresh- cycle (Yu et al. 2013). In addition, morphometric analyses old (Abraham et al. 2011). This suggests that modulation of NMJ (neuromuscular junction) in snt-1 mutants reveal of the synaptic vesicle cycle is fine-tuned by the specific a reduction in vesicle density, a phenotype associated with amount of synaptogyrin, since both decrease and increase an endocytosis defect (Yu et al. 2013). snt-1 mutants also in synaptogyrin result in an altered sensitivity to PTZ and show a reduction in absolute numbers of docked vesicle, aldicarb (Abraham et al. 2011). In addition, detailed analysis and this docking defect appeared to be a consequence of an also uncovers mildly altered motility and decreased recruit- overall reduced vesicle density, since the fraction of docked ment of synaptobrevin though not of RAB-3 to synapses, vesicles as a function of total vesicles in snt-1 mutants was suggesting that synaptogyrin presents a distinct modulatory not significantly reduced compared to wild type (Yu et al. and redundant neuronal function in C. elegans (Abraham 2013). Both reductions in absolute docked vesicles and in et al. 2011). evoked transmission suggest that SNT-1 has additional func- Another conserved core components of the presynaptic tion beyond exocytosis, consistent with the well-documented active zone are the UNC-13/Munc13 family. They are essen- 2+ role of snt-1 as a Ca sensor promoting vesicle fusion (Yu tial for both evoked and spontaneous SV release (Augustin et al. 2013). In addition, SNT-1 in C. elegans is crucial for et al. 1999; Richmond et al. 1999). These proteins con- the SVs (synaptic vesicles) association of RAB-3 protein. tain multiple protein interaction domains and involved in SNT-1 promotes the GTP-bound state of RAB-3 by inhibit- many aspect aspects of presynaptic release. All the UNC- ing RAB-3 GAP, and thus, the catalytic subunit of RAB-3 13/Munc13 isoforms contain a diacylglycerol binding C GAP (RBG-1) localizes on SVs and directly binds to SNT-1 domain followed by a MUN domain including the MHD 2+ (Cheng et al. 2015). Ca treatment disrupts the direct asso- (Munc13 homology domain) flanked by C B and C C 2 2 ciation between SNT-1 and RBG-1 (a Rab-3 GTPase). In domains. The MUN domain, structurally similar to the vesi- 2+ addition, Ca binding activity of SNT-1 is essential for the cle tethering factors of the CATCHR (Complex Associated dissociation of RAB-3 from SVs (Cheng et al. 2015). with Tethering Containing Helical rods) family (Li et al. Complementary studies in mouse models have shown that 2011), is necessary for vesicle priming (Basu et al. 2005; 2+ synaptotagmins (specifically Syt1 and Syt2) are Ca sen- Madison et al. 2005; Stevens et al. 2005) through binding to sors for both synchronous and fast neurotransmitter release SNARE and Munc18 (Betz et al. 1997; Ma et al. 2011). The (Sun et al. 2007; Xu et al. 2009). Overall, synaptotagmins N-terminal regions of UNC-13/Munc13 isoforms are diver- 2+ act as a cooperative Ca receptor in exocytosis, binding gent in amino acid sequences and have been hypothesized to 2+ Ca at physiological concentrations. This binding is specific contribute to the distinct properties of SV exocytosis in dif- 2+ for Ca and involves the cytoplasmic domain of synaptotag- ferent types of synapses (Augustin et al. 2001; Rosenmund min (Geppert et al. 1994; Pang et al. 2006). et al. 2002). Synaptogyrin and synaptophysin are tetraspan membrane In C. elegans, the unc-13 locus produces two main proteins, the major vesicle proteins, characterized by four isoforms that differ at the N-terminal region (Kohn et al. membrane-spanning domains that are tyrosine-phospho- 2000). The expression of unc-13 is in all neurons of rylated (Arthur and Stowell 2007; Evans and Cousin 2005; both head and tail ganglia, as well as ventral nerve cord Hubner et al. 2002). They are abundant and evolutionary (Maruyama et al. 2001). Using genetic mutations that conserved synaptic vesicle membrane proteins (Abraham eliminate function of all isoforms or only UNC-13L dem- et al. 2006) whose functions are poorly defined, and their onstrate an essential role of UNC-13L in neurotransmitter depletion does not interfere with proper neuronal develop- release (Richmond et al. 1999). The kinetic components of ment and basic neuronal function in both C. elegans and release are thought to be mediated by SVs in different spa- mammals (Abraham et al. 2006; Eshkind and Leube 1995; tial domains of the nerve terminal. Rapid (or synchronous) McMahon et al. 1996). In contrast to vertebrates, in C. ele- release occurs within a few milliseconds and is proposed 2+ gans the synaptogyrin but not the synaptophysin orthologue to consist of fusion of SVs that are close to Ca entry is predominant in neurons (Abraham et al. 2006; Hubner sites. However, slow release occurs over tens to hundreds et al. 2002; Nonet 1999; Ruvinsky et al. 2007), expressed in of milliseconds and is thought to be mediated by fusion 2+ all 26 GABAergic neurons, as well as in a subset of neurons of SVs that are farther from Ca channels (Neher and 1 3 Invertebrate Neuroscience (2018) 18:4 Page 9 of 13 4 Sakaba 2008). In C. elegans, the UNC-13L isoform is co- Concluding remarks localized at presynaptic terminals concentrated near dense projections. By contrast, the UNC-13S isoform presents a Through this review, we discussed the synaptic release diffuse distribution in axons. This suggests that UNC-13L machinery, and how the powerful genetic model C. elegans and UNC-13S may mediate different forms of release and contributes to elucidating core processes of synaptic trans- distinct effects on synaptic transmission (Hu et al. 2013). mission. This is facilitated by the ability to maintain viable In this sense, it has been shown that UNC-13L is involved mutants of C. elegans for synaptic proteins that are other- in both fast and slow release of SVs, while the short iso- wise essential in mice. It has broad relevance as C. elegans form UNC-13S is required for the slow release (Hu et al. harbours the same elaborate elements for neurotransmis- 2013). In more detail, other studies using a unique unc- sion as mammals, and thus, it has been instrumental in key 13 mutant that specifically deletes the C A domain of discoveries relating to the synaptic vesicle cycle. However, UNC-13L show that the precise position of UNC-13 in some key processes have yet to be elucidated; for exam- the active zone depended on the C A domain. In addition, ple, the precise physicochemical mechanisms underlying the C A domain regulates the release probability of SVs, fusion, the roles of key synaptic proteins with overlapping likely through positioning UNC-13L to the active zone, functions within complex neural networks and understand- and that this domain also has a significant influence in ing how synaptic vesicles recaptured by clathrin-mediated spontaneous release (Zhou et al. 2013). Importantly, early endocytosis are placed back into the vesicle pool; all these works using C. elegans have shown the role of PKC/DAG aspects demand further study which may be supported by in the modulation of UNC-13. Thus, the binding of DAG the C. elegans model. Advances in techniques to study the by UNC-13 drives its membrane association and regulates vesicle cycle in the intact living synapse in combination the exocytosis function of UNC-13 (Lackner et al. 1999). with genetic manipulation will accelerate progress in the Later, studies in Drosophila have also shown how the PLC field, shedding more light on these intricate processes. In and DAG modulation as well as G-proteins regulates the this sense, C. elegans is an excellent system to facilitate synaptic levels of DUNC-13, critical determinant of SV discovery in this field, thanks to a simple, genetically trac- fusion probability (Aravamudan and Broadie 2003). table nervous system that is evolutionarily conserved with RIM proteins are presynaptic scaffolding proteins spe- mammalian neurons, as well as providing new routes to cifically localized to the active zone and found to bind understand the dynamic processes underlying neurotrans- several presynaptic proteins, like Munc13-1, Rab3a and mitter exocytosis. 2+ voltage-gated Ca channels (Betz et al. 2001; Kaeser Acknowledgements We would like to apologize to the colleagues et al. 2011; Schoch et al. 2002; Wang et al. 1997). RIM whose work could not be cited due to space constraints. We are very proteins are encoded by four genes (Rims1–4); the Rim1 grateful to Lindy Holden-Dye and Vincent O’Connor for their critical and 2 genes give rise to five RIM isoforms, called RIM1α, discussion and comments. RIM1β, RIM2α, RIM2β and RIM2γ (Kaeser et al. 2008; Wang and Sudhof 2003). Studies using RIM1α constitu- Compliance with ethical standards tive knockout mice and RIM mutants in C. elegans found roles for RIM1 in transmitter release, presumably via Conflict of interest The authors declare that they have no competing interests. determining readily releasable vesicle pool size (Calakos et al. 2004; Koushika et al. 2001; Schoch et al. 2002). Open Access This article is distributed under the terms of the Crea- RIM mutants isolated in C. elegans (unc-10) are viable but tive Commons Attribution 4.0 International License (http://creat iveco exhibit behavioural and pharmacological defects, indica- mmons.or g/licenses/b y/4.0/), which permits unrestricted use, distribu- tive of synaptic dysfunction according to the localization tion, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the of RIM at the active zone. Although RIM was originally Creative Commons license, and indicate if changes were made. identified as a RAB-3 binding partner, the consequence of a loss of function mutation is more severe than rab-3 mutants, suggesting that it possesses additional functions (Gracheva et al. 2008). Electrophysiological analysis of References unc-10 worms revealed both reduced evoked release of SVs and spontaneous synaptic event frequency, thus impli- Abraham C, Hutter H, Palfreyman MT, Spatkowski G, Weimer RM, cating RIM in release (Koushika et al. 2001). UNC-10 is Windoffer R et al (2006) Synaptic tetraspan vesicle membrane 2+ co-localized with the Ca channel, UNC-2 at C. elegans proteins are conserved but not needed for synaptogenesis and neuronal function in Caenorhabditis elegans. Proc Natl presynaptic densities and synaptic release in unc-10 and 2+ Acad Sci U S A 103(21):8227–8232. https ://doi.org/10.1073/ rab-3 mutants exhibit reduced C a sensitivity (Gracheva pnas.05094 00103 et al. 2008a). 1 3 4 Page 10 of 13 Invertebrate Neuroscience (2018) 18:4 Abraham C, Bai L, Leube RE (2011) Synaptogyrin-dependent Cheng Y, Wang J, Wang Y, Ding M (2015) Synaptotagmin 1 directs modulation of synaptic neurotransmission in Caenorhabditis repetitive release by coupling vesicle exocytosis to the Rab3 elegans. Neuroscience 190:75–88. https ://doi.org/10.1016/j. cycle. Elife. https ://doi.org/10.7554/eLife .05118 neuro scien ce.2011.05.069 Dalliere N, Bhatla N, Luedtke Z, Ma DK, Woolman J, Walker RJ et al Ann K, Kowalchyk JA, Loyet KM, Martin TF (1997) Novel (2016) Multiple excitatory and inhibitory neural signals converge 2+ Ca -binding protein (CAPS) related to UNC-31 required for to fine-tune Caenorhabditis elegans feeding to food availability. 2+ Ca -activated exocytosis. J Biol Chem 272(32):19637–19640 FASEB J 30(2):836–848. https ://doi.org/10.1096/fj.15-27925 7 Aravamudan B, Broadie K (2003) Synaptic drosophila UNC-13 De Camilli P, Cameron R, Greengard P (1983) Synapsin I (protein I), a is regulated by antagonistic G-protein pathways via a pro- nerve terminal-specific phosphoprotein. I. Its general distribution teasome-dependent degradation mechanism. J Neurobiol in synapses of the central and peripheral nervous system demon- 54(3):417–438. https ://doi.org/10.1002/neu.10142 strated by immunofluorescence in frozen and plastic sections. J Arthur CP, Stowell MH (2007) Structure of synaptophysin: a hexa- Cell Biol 96(5):1337–1354 meric MARVEL-domain channel protein. Structure 15(6):707– De Camilli P, Benfenati F, Valtorta F, Greengard P (1990) The syn- 714. https ://doi.org/10.1016/j.str.2007.04.011 apsins. Annu Rev Cell Biol 6:433–460. https ://doi.org/10.1146/ Augustin I, Rosenmund C, Sudhof TC, Brose N (1999) Munc13-1 is annur ev.cb.06.11019 0.00224 5 essential for fusion competence of glutamatergic synaptic vesi- Delgado-Martinez I, Nehring RB, Sorensen JB (2007) Differential cles. Nature 400(6743):457–461. https ://doi.org/10.1038/22768 abilities of SNAP-25 homologs to support neuronal function. Augustin I, Korte S, Rickmann M, Kretzschmar HA, Sudhof TC, J Neurosci 27(35):9380–9391. https ://doi.or g/10.1523/Jneur Herms JW et al (2001) The cerebellum-specific Munc13 isoform osci.5092-06.2007 Munc13-3 regulates cerebellar synaptic transmission and motor DiCaprio RA, Billimoria CP, Ludwar BC (2007) Information rate learning in mice. J Neurosci 21(1):10–17 and spike-timing precision of proprioceptive afferents. J Neuro- Avery L, Bargmann CI, Horvitz HR (1993) The Caenorhabditis ele- physiol 98(3):1706–1717. https:// doi.org/10.1152/jn.00176.2 007 gans unc-31 gene affects multiple nervous system-controlled Dulubova I, Khvotchev M, Liu S, Huryeva I, Sudhof TC, Rizo J (2007) functions. Genetics 134(2):455–464 Munc18-1 binds directly to the neuronal SNARE complex. Proc Bahler M, Greengard P (1987) Synapsin I bundles F-actin in a phos- Natl Acad Sci U S A 104(8):2697–2702. https://doi.or g/10.1073/ phorylation-dependent manner. Nature 326(6114):704–707. https pnas.06113 18104 ://doi.org/10.1038/32670 4a0 Dybbs M, Ngai J, Kaplan JM (2005) Using microarrays to facilitate Basu J, Shen N, Dulubova I, Lu J, Guan R, Guryev O et al (2005) positional cloning: Identification of tomosyn as an inhibitor of A minimal domain responsible for Munc13 activity. Nat Struct neurosecretion. PLoS Genet 1(1):6–16. https ://doi.org/10.1371/ Mol Biol 12(11):1017–1018. https://doi.or g/10.1038/nsmb1001 journ al.pgen.00100 02 Berwin B, Floor E, Martin TF (1998) CAPS (mammalian UNC-31) Eshkind LG, Leube RE (1995) Mice lacking synaptophysin repro- protein localizes to membranes involved in dense-core vesicle duce and form typical synaptic vesicles. Cell Tissue Res exocytosis. Neuron 21(1):137–145 282(3):423–433 Betz A, Okamoto M, Benseler F, Brose N (1997) Direct interaction Evans GJ, Cousin MA (2005) Tyrosine phosphorylation of synapto- of the rat unc-13 homologue Munc13-1 with the N terminus of physin in synaptic vesicle recycling. Biochem Soc Trans 33(Pt syntaxin. J Biol Chem 272(4):2520–2526 6):1350–1353. https ://doi.org/10.1042/BST20 05135 0 Betz A, Thakur P, Junge HJ, Ashery U, Rhee JS, Scheuss V et al (2001) Farina M, van de Bospoort R, He E, Persoon CM, van Weering JR, Functional interaction of the active zone proteins Munc13-1 and Broeke JH et al (2015) CAPS-1 promotes fusion competence of RIM1 in synaptic vesicle priming. Neuron 30(1):183–196. https stationary dense-core vesicles in presynaptic terminals of mam- ://doi.org/10.1016/S0896 -6273(01)00272 -0 malian neurons. Elife. https ://doi.org/10.7554/eLife .05438 Binotti B, Jahn R, Chua JJ (2016) Functions of Rab proteins at presyn- Ferreira A, Han HQ, Greengard P, Kosik KS (1995) Suppres- aptic sites. Cells. https ://doi.org/10.3390/cells 50100 07 sion of synapsin II inhibits the formation and maintenance of Bloom O, Evergren E, Tomilin N, Kjaerulff O, Low P, Brodin L synapses in hippocampal culture. Proc Natl Acad Sci U S A et al (2003) Colocalization of synapsin and actin during syn- 92(20):9225–9229 aptic vesicle recycling. J Cell Biol 161(4):737–747. https ://doi. Ferreira A, Kao HT, Feng J, Rapoport M, Greengard P (2000) Synapsin org/10.1083/jcb.20021 2140 III: developmental expression, subcellular localization, and role Bock JB, Matern HT, Peden AA, Scheller RH (2001) A genomic in axon formation. J Neurosci 20(10):3736–3744 perspective on membrane compartment organization. Nature Fornasiero EF, Bonanomi D, Benfenati F, Valtorta F (2010) The 409(6822):839–841. https ://doi.org/10.1038/35057 024 role of synapsins in neuronal development. Cell Mol Life Sci Bracher A, Weissenhorn W (2004) Crystal structure of the Habc 67(9):1383–1396. https ://doi.org/10.1007/s0001 8-009-0227-8 domain of neuronal syntaxin from the squid Loligo pealei reveals Fujita Y, Shirataki H, Sakisaka T, Asakura T, Ohya T, Kotani H et al conformational plasticity at its C-terminus. BMC Struct Biol 4:6. (1998) Tomosyn: a syntaxin-1-binding protein that forms a https ://doi.org/10.1186/1472-6807-4-6 novel complex in the neurotransmitter release process. Neuron Brenner S (1974) The genetics of Caenorhabditis elegans. Genetics 20(5):905–915 77(1):71–94 Fujiwara T, Mishima T, Kofuji T, Chiba T, Tanaka K, Yamamoto Calakos N, Schoch S, Sudhof TC, Malenka RC (2004) Multiple A et al (2006) Analysis of knock-out mice to determine the roles for the active zone protein RIM1alpha in late stages of role of HPC-1/syntaxin 1A in expressing synaptic plasticity. J neurotransmitter release. Neuron 42(6):889–896. https ://doi. Neurosci 26(21):5767–5776. https ://doi.or g/10.1523/JNEUR org/10.1016/j.neuro n.2004.05.014 OSCI.0289-06.2006 Cesca F, Baldelli P, Valtorta F, Benfenati F (2010) The synapsins: Gallegos ME, Balakrishnan S, Chandramouli P, Arora S, Azameera key actors of synapse function and plasticity. Prog Neurobiol A, Babushekar A et al (2012) The C. elegans rab family: iden- 91(4):313–348. https://doi.or g/10.1016/j.pneurobio.2010.04.006 tification, classification and toolkit construction. PLoS ONE Charlie NK, Schade MA, Thomure AM, Miller KG (2006) Presynaptic 7(11):e49387. https ://doi.org/10.1371/journ al.pone.00493 87 UNC-31 (CAPS) is required to activate the G alpha(s) pathway of Geerts CJ, Mancini R, Chen N, Koopmans FTW, Li KW, Smit AB et al the Caenorhabditis elegans synaptic signaling network. Genetics (2017) Tomosyn associates with secretory vesicles in neurons 172(2):943–961. https ://doi.org/10.1534/genet ics.105.04957 7 1 3 Invertebrate Neuroscience (2018) 18:4 Page 11 of 13 4 through its N-and C-terminal domains. PLoS ONE. https ://doi. 416, 2014). Neuron 84(4):882. https://d oi.org/10.1016/j.neuro org/10.1371/journ al.pone.01809 12 n.2014.11.003 Gengyo-Ando K, Kamiya Y, Yamakawa A, Kodaira K, Nishiwaki K, Jockusch WJ, Speidel D, Sigler A, Sorensen JB, Varoqueaux F, Rhee JS Miwa J et al (1993) The C. elegans unc-18 gene encodes a pro- et al (2007) CAPS-1 and CAPS-2 are essential synaptic vesicle tein expressed in motor neurons. Neuron 11(4):703–711 priming proteins. Cell 131(4):796–808. https://doi.or g/10.1016/j. Geppert M, Goda Y, Hammer RE, Li C, Rosahl TW, Stevens CF et al cell.2007.11.002 2+ (1994) Synaptotagmin I: a major Ca sensor for transmitter Jung N, Haucke V (2007) Clathrin-mediated endocytosis at syn- release at a central synapse. Cell 79(4):717–727 apses. Traffic 8(9):1129–1136. https ://doi.or g/10.111 Gerber SH, Rah JC, Min SW, Liu XR, de Wit H, Dulubova I et al 1/j.1600-0854.2007.00595 .x (2008) Conformational switch of syntaxin-1 controls synap- Kaeser PS, Kwon HB, Chiu CQ, Deng L, Castillo PE, Sudhof TC tic vesicle fusion. Science 321(5895):1507–1510. https ://doi. (2008) RIM1alpha and RIM1beta are synthesized from distinct org/10.1126/scien ce.11631 74 promoters of the RIM1 gene to mediate differential but over - Gracheva EO, Burdina AO, Holgado AM, Berthelot-Grosjean M, lapping synaptic functions. J Neurosci 28(50):13435–13447. Ackley BD, Hadwiger G et al (2006) Tomosyn inhibits synaptic https ://doi.org/10.1523/JNEUR OSCI.3235-08.2008 vesicle priming in Caenorhabditis elegans. PLoS Biol 4(8):e261. Kaeser PS, Deng LB, Wang Y, Dulubova I, Liu XR, Rizo J et al 2+ https ://doi.org/10.1371/journ al.pbio.00402 61 (2011) RIM proteins tether Ca channels to presynaptic active Gracheva EO, Hadwiger G, Nonet ML, Richmond JE (2008) Direct zones via a direct PDZ-domain interaction. Cell 144(2):282– interactions between C. elegans RAB-3 and Rim provide a mech- 295. https ://doi.org/10.1016/j.cell.2010.12.029 anism to target vesicles to the presynaptic density. Neurosci Lett Kao HT, Porton B, Hilfiker S, Stefani G, Pieribone VA, DeSalle R 444(2):137–142. https ://doi.org/10.1016/j.neule t.2008.08.026 et al (1999) Molecular evolution of the synapsin gene family. Gracheva EO, Maryon EB, Berthelot-Grosjean M, Richmond JE (2010) J Exp Zool 285(4):360–377 Differential regulation of synaptic vesicle tethering and docking Katz B (1971) Quantal mechanism of neural transmitter release. Sci- by UNC-18 and TOM-1. Front Synaptic Neurosci 2:141. https: // ence 173(3992):123–126 doi.org/10.3389/fnsyn .2010.00141 Katz B (1979) Elementary components of synaptic transmission. Graham ME, Prescott GR, Johnson JR, Jones M, Walmesley A, Naturwissenschaften 66(12):606–610 Haynes LP et al (2011) Structure-function study of mammalian Kohn RE, Duerr JS, McManus JR, Duke A, Rakow TL, Maruy- Munc18-1 and C. elegans UNC-18 implicates domain 3b in the ama H et al (2000) Expression of multiple UNC-13 proteins regulation of exocytosis. PLoS ONE 6(3):e17999. https ://doi. in the Caenorhabditis elegans nervous system. Mol Biol Cell org/10.1371/journ al.pone.00179 99 11(10):3441–3452 Grishanin RN, Klenchin VA, Loyet KM, Kowalchyk JA, Ann K, Martin Koushika SP, Richmond JE, Hadwiger G, Weimer RM, Jorgensen 2+ TF (2002) Membrane association domains in Ca -dependent EM, Nonet ML (2001) A post-docking role for active zone activator protein for secretion mediate plasma membrane and protein Rim. Nat Neurosci 4(10):997–1005. https ://doi. 2+ dense-core vesicle binding required for Ca -dependent exocyto- org/10.1038/nn732 sis. J Biol Chem 277(24):22025–22034. https://doi.or g/10.1074/ Lackner MR, Nurrish SJ, Kaplan JM (1999) Facilitation of synap- jbc.M2016 14200 tic transmission by EGL-30 G(q)alpha and EGL-8 PLC beta: Hammarlund M, Watanabe S, Schuske K, Jorgensen EM (2008) CAPS DAG binding to UNC-13 is required to stimulate acetylcholine and syntaxin dock dense core vesicles to the plasma membrane release. Neuron 24(2):335–346. https://doi.or g/10.1016/S0896 in neurons. J Cell Biol 180(3):483–491. https://doi.or g/10.1083/-6273(00)80848 -X jcb.20070 8018 Lemmon MA (2008) Membrane recognition by phospholipid-bind- Hatsuzawa K, Lang T, Fasshauer D, Bruns D, Jahn R (2003) The ing domains. Nat Rev Mol Cell Biol 9(2):99–111. https ://doi. R-SNARE motif of tomosyn forms SNARE core complexes org/10.1038/nrm23 28 with syntaxin 1 and SNAP-25 and down-regulates exocytosis. J Li X, Rosahl TW, Sudhof TC, Francke U (1995) Mapping of syn- Biol Chem 278(33):31159–31166. https ://doi.org/10.1074/jbc. apsin II (SYN2) genes to human chromosome 3p and mouse M3055 00200 chromosome 6 band F. Cytogenet Cell Genet 71(3):301–305 Hilfiker S, Schweizer FE, Kao HT, Czernik AJ, Greengard P, Augustine Li W, Ma C, Guan R, Xu YB, Tomchick DR, Rizo J (2011) The GJ (1998) Two sites of action for synapsin domain E in regulat- crystal structure of a Munc13 C-terminal module exhib- ing neurotransmitter release. Nat Neurosci 1(1):29–35. https :// its a remarkable similarity to vesicle tethering factors. doi.org/10.1038/229 Str ucture 19(10):1443–1455. https ://doi.or g/ 10.1016/j. Holt M, Varoqueaux F, Wiederhold K, Takamori S, Urlaub H, str.2011.07.012 Fasshauer D et al (2006) Identification of SNAP-47, a novel Ma C, Li W, Xu YB, Rizo J (2011) Munc13 mediates the transition Qbc-SNARE with ubiquitous expression. J Biol Chem from the closed syntaxin-Munc18 complex to the SNARE 281(25):17076–17083. https://doi.or g/10.1074/jbc.M513838200 complex. Nat Struct Mol Biol 18(5):542-U206. https ://doi. Hu ZT, Tong XJ, Kaplan JM (2013) UNC-13L, UNC-13S, and org/10.1038/nsmb.2047 Tomosyn form a protein code for fast and slow neurotransmitter Madison JM, Nurrish S, Kaplan JM (2005) UNC-13 interaction release in Caenorhabditis elegans. Elife. https://doi.or g/10.7554/ with syntaxin is required for synaptic transmission. Curr Biol eLife .00967 15(24):2236–2242. https ://doi.org/10.1016/j.cub.2005.10.049 Hubner K, Windoffer R, Hutter H, Leube RE (2002) Tetraspan vesi- Mahoney TR, Liu Q, Itoh T, Luo S, Hadwiger G, Vincent R et al (2006) cle membrane proteins: synthesis, subcellular localization, and Regulation of synaptic transmission by RAB-3 and RAB-27 in functional properties. Int Rev Cytol 214:103–159 Caenorhabditis elegans. Mol Biol Cell 17(6):2617–2625. https Hwang SB, Lee J (2003) Neuron cell type-specific SNAP-25 expression ://doi.org/10.1091/mbc.E05-12-1170 driven by multiple regulatory elements in the nematode Caeno- Martin TF (2003) Tuning exocytosis for speed: fast and slow modes. rhabditis elegans. J Mol Biol 333(2):237–247 Biochim Biophys Acta 1641(2–3):157–165 Imig C, Min SW, Krinner S, Arancillo M, Rosenmund C, Sudhof TC Maruyama H, Rakow TL, Maruyama IN (2001) Synaptic exocytosis et al (2014) The morphological and molecular nature of syn- and nervous system development impaired in Caenorhabditis aptic vesicle priming at presynaptic active zones (vol 84, pg elegans unc-13 mutants. Neuroscience 104(2):287–297 1 3 4 Page 12 of 13 Invertebrate Neuroscience (2018) 18:4 Masuda ES, Huang BC, Fisher JM, Luo Y, Scheller RH (1998) Caenorhabditis elegans. Mol Biol Cell 27(4):669–685. https :// Tomosyn binds t-SNARE proteins via a VAMP-like coiled coil. doi.org/10.1091/mbc.E15-09-0638 Neuron 21(3):479–480 Pieribone VA, Shupliakov O, Brodin L, Hilfiker-Rothenfluh S, Czernik Mathews EA, Mullen GP, Crowell JA, Duerr JS, McManus JR, Duke A AJ, Greengard P (1995) Distinct pools of synaptic vesicles in et al (2007) Differential expression and function of synaptotag- neurotransmitter release. Nature 375(6531):493–497. https://doi. min 1 isoforms in Caenorhabditis elegans. Mol Cell Neurosci org/10.1038/37549 3a0 34(4):642–652. https ://doi.org/10.1016/j.mcn.2007.01.009 Porton B, Wetsel WC, Kao HT (2011) Synapsin III: role in neuronal McEwen JM, Kaplan JM (2008) UNC-18 promotes both the antero- plasticity and disease. Semin Cell Dev Biol 22(4):416–424. https grade trafficking and synaptic function of syntaxin. Mol Biol ://doi.org/10.1016/j.semcd b.2011.07.007 Cell 19(9):3836–3846. https://doi.or g/10.1091/mbc.E08-02-0160 Ramirez DMO, Kavalali ET (2011) Differential regulation of sponta- McEwen JM, Madison JM, Dybbs M, Kaplan JM (2006) Antago- neous and evoked neurotransmitter release at central synapses. nistic regulation of synaptic vesicle priming by Tomosyn and Curr Opin Neurobiol 21(2):275–282. https ://doi.org/10.1016/j. UNC-13. Neuron 51(3):303–315. https://doi.or g/10.1016/j.neuro conb.2011.01.007 n.2006.06.025 Renden R, Berwin B, Davis W, Ann K, Chin CT, Kreber R et al McMahon HT, Bolshakov VY, Janz R, Hammer RE, Siegelbaum SA, (2001) Drosophila CAPS is an essential gene that regulates Sudhof TC (1996) Synaptophysin, a major synaptic vesicle pro- dense-core vesicle release and synaptic vesicle fusion. Neuron tein, is not essential for neurotransmitter release. Proc Natl Acad 31(3):421–437 Sci U S A 93(10):4760–4764 Rettig J, Neher E (2002) Emerging roles of presynaptic proteins in ++ Mishima T, Fujiwara T, Sanada M, Kofuji T, Kanai-Azuma M, Aka- Ca -triggered exocytosis. Science 298(5594):781–785. https:// gawa K (2014) Syntaxin 1B, but not Syntaxin 1A, is necessary doi.org/10.1126/scien ce.10753 75 for the regulation of synaptic vesicle exocytosis and of the read- Richmond J (2005) Synaptic function. WormBook. https ://doi. ily releasable pool at central synapses. PLoS ONE. https ://doi.org/10.1895/wormb ook.1.69.1 org/10.1371/journ al.pone.00900 04 Richmond JE, Davis WS, Jorgensen EM (1999) UNC-13 is required Munson M, Bryant NJ (2009) A role for the syntaxin N-terminus. Bio- for synaptic vesicle fusion in C. elegans. Nat Neurosci chem J 418(1):e1–e3. https ://doi.org/10.1042/BJ200 82389 2(11):959–964 Neher E, Sakaba T (2008) Multiple roles of calcium ions in the regula- Rizo J, Sudhof TC (1998) C -domains, structure and function of a uni- 2+ tion of neurotransmitter release. Neuron 59(6):861–872. https ://versal Ca -binding domain. J Biol Chem 273(26):15879–15882 doi.org/10.1016/j.neuro n.2008.08.019 Rizo J, Sudhof TC (2002) Snares and Munc18 in synaptic vesicle Nelson JC, Stavoe AK, Colon-Ramos DA (2013) The actin cytoskel- fusion. Nat Rev Neurosci 3(8):641–653. https://doi.or g/10.1038/ eton in presynaptic assembly. Cell Adhes Migr 7(4):379–387. nrn89 8 https ://doi.org/10.4161/cam.24803 Rosenmund C, Sigler A, Augustin I, Reim K, Brose N, Rhee JS (2002) Nonet ML (1999) Visualization of synaptic specializations in live C. Differential control of vesicle priming and short-term plastic- elegans with synaptic vesicle protein-GFP fusions. J Neurosci ity by Munc13 isoforms. Neuron 33(3):411–424. https ://doi. Methods 89(1):33–40org/10.1016/S0896 -6273(02)00568 -8 Nonet ML, Grundahl K, Meyer BJ, Rand JB (1993) Synaptic function Ruvinsky I, Ohler U, Burge CB, Ruvkun G (2007) Detection of broadly is impaired but not eliminated in C. Elegans mutants lacking syn- expressed neuronal genes in C. elegans. Dev Biol 302(2):617– aptotagmin. Cell 73(7):1291–1305. https ://doi.or g/10.1016/0092- 626. https ://doi.org/10.1016/j.ydbio .2006.09.014 8674(93)90357 -V Sadakata T, Washida M, Furuichi T (2007a) Alternative splicing vari- Nonet ML, Staunton JE, Kilgard MP, Fergestad T, Hartwieg E, Horvitz ations in mouse CAPS2: differential expression and functional HR et al (1997) Caenorhabditis elegans rab-3 mutant synapses properties of splicing variants. BMC Neurosci 8:25. https ://doi. exhibit impaired function and are partially depleted of vesicles. org/10.1186/1471-2202-8-25 J Neurosci 17(21):8061–8073 Sadakata T, Washida M, Iwayama Y, Shoji S, Sato Y, Ohkura T et al Nonet ML, Saifee O, Zhao H, Rand JB, Wei L (1998) Synaptic (2007b) Autistic-like phenotypes in Cadps2-knockout mice and transmission deficits in Caenorhabditis elegans synaptobrevin aberrant CADPS2 splicing in autistic patients. J Clin Invest mutants. J Neurosci 18(1):70–80 117(4):931–943. https ://doi.org/10.1172/JCI29 031 Nystuen AM, Schwendinger JK, Sachs AJ, Yang AW, Haider NB Sadakata T, Washida M, Morita N, Furuichi T (2007c) Tissue distri- 2+ (2007) A null mutation in VAMP1/synaptobrevin is associated bution of Ca -dependent activator protein for secretion family with neurological defects and prewean mortality in the lethal- members CAPS1 and CAPS2 in mice. J Histochem Cytochem wasting mouse mutant. Neurogenetics 8(1):1–10. https ://doi. 55(3):301–311. https ://doi.org/10.1369/jhc.6A703 3.2006 org/10.1007/s1004 8-006-0068-7 Saifee O, Wei LP, Nonet ML (1998) The Caenorhabditis elegans unc- Ogawa H, Harada S, Sassa T, Yamamoto H, Hosono R (1998) Func- 64 locus encodes a syntaxin that interacts genetically with syn- tional properties of the unc-64 gene encoding a Caenorhabditis aptobrevin. Mol Biol Cell 9(6):1235–1252 elegans syntaxin. J Biol Chem 273(4):2192–2198 Sandoval GM, Duerr JS, Hodgkin J, Rand JB, Ruvkun G (2006) A Pan PY, Cai Q, Lin L, Lu PH, Duan SM, Sheng ZH (2005) SNAP- genetic interaction between the vesicular acetylcholine trans- 29-mediated modulation of synaptic transmission in cultured hip- porter VAChT/UNC-17 and synaptobrevin/SNB-1 in C. elegans. pocampal neurons. J Biol Chem 280(27):25769–25779. https :// Nat Neurosci 9(5):599–601. https ://doi.org/10.1038/nn168 5 doi.org/10.1074/jbc.M5023 56200 Sassa T, Harada S, Ogawa H, Rand JB, Maruyama IN, Hosono R Pang ZP, Melicoff E, Padgett D, Liu Y, Teich AF, Dickey BF et al (1999) Regulation of the UNC-18-Caenorhabditis elegans syn- (2006) Synaptotagmin-2 is essential for survival and contributes taxin complex by UNC-13. J Neurosci 19(12):4772–4777 2+ to Ca triggering of neurotransmitter release in central and neu- Schindelman G, Whittaker AJ, Thum JY, Gharib S, Sternberg PW romuscular synapses. J Neurosci 26(52):13493–13504. https :// (2006) Initiation of male sperm-transfer behavior in Caenorhab- doi.org/10.1523/JNEUR OSCI.3519-06.2006 ditis elegans requires input from the ventral nerve cord. BMC Park S, Bin NR, Michael Rajah M, Kim B, Chou TC, Kang SY et al Biol. https ://doi.org/10.1186/1741-7007-4-26 (2016) Conformational states of syntaxin-1 govern the neces- Schluter OM, Khvotchev M, Jahn R, Sudhof TC (2002) Locali- sity of N-peptide binding in exocytosis of PC12 cells and zation versus function of Rab3 proteins. Evidence for a 1 3 Invertebrate Neuroscience (2018) 18:4 Page 13 of 13 4 common regulatory role in controlling fusion. J Biol Chem neurons in C. elegans. PLoS ONE 8(3):e59132. https ://doi. 277(43):40919–40929. https://doi.or g/10.1074/jbc.M203704200 org/10.1371/journ al.pone.00591 32 Schoch S, Deak F, Konigstorfer A, Mozhayeva M, Sara Y, Sudhof Takamori S, Holt M, Stenius K, Lemke EA, Gronborg M, Riedel D TC et al (2001) SNARE function analyzed in synaptobrevin/ et al (2006) Molecular anatomy of a trafficking organelle. Cell VAMP knockout mice. Science 294(5544):1117–1122. https :// 127(4):831–846. https ://doi.org/10.1016/j.cell.2006.10.030 doi.org/10.1126/scien ce.10643 35 Takei Y, Harada A, Takeda S, Kobayashi K, Terada S, Noda T et al Schoch S, Castillo PE, Jo T, Mukherjee K, Geppert M, Wang Y et al (1995) Synapsin I deficiency results in the structural change in (2002) RIM1 alpha forms a protein scaffold for regulating neuro- the presynaptic terminals in the murine nervous system. J Cell transmitter release at the active zone. Nature 415(6869):321–326. Biol 131(6 Pt 2):1789–1800 https ://doi.org/10.1038/41532 1a Tandon A, Bannykh S, Kowalchyk JA, Banerjee A, Martin TF, Balch Sieburth D, Ch’ng Q, Dybbs M, Tavazoie M, Kennedy S, Wang D WE (1998) Differential regulation of exocytosis by calcium and et al (2005) Systematic analysis of genes required for synapse CAPS in semi-intact synaptosomes. Neuron 21(1):147–154 structure and function. Nature 436(7050):510–517. https ://doi. Teng FYH, Wang Y, Tang BL (2001) The syntaxins. Genome Biol org/10.1038/natur e0380 9 2(11):reviews3012.1–3012.7 Sieburth D, Madison JM, Kaplan JM (2007) PKC-1 regulates secre- van Steveninck RRD, Laughlin SB (1996) The rate of information tion of neuropeptides. Nat Neurosci 10(1):49–57. https ://doi. transfer at graded-potential synapses. Nature 379(6566):642– org/10.1038/nn181 0 645. https ://doi.org/10.1038/37964 2a0 Speidel D, Varoqueaux F, Enk C, Nojiri M, Grishanin RN, Martin TF Walent JH, Porter BW, Martin TF (1992) A novel 145 kd brain cyto- 2+ 2+ et al (2003) A family of Ca -dependent activator proteins for solic protein reconstitutes Ca -regulated secretion in permeable secretion: comparative analysis of structure, expression, localiza- neuroendocrine cells. Cell 70(5):765–775 tion, and function. J Biol Chem 278(52):52802–52809. https :// Wang Y, Sudhof TC (2003) Genomic definition of RIM proteins: evolu- doi.org/10.1074/jbc.M3047 27200 tionary amplification of a family of synaptic regulatory proteins. Speidel D, Bruederle CE, Enk C, Voets T, Varoqueaux F, Reim K et al Genomics 81(2):126–137 (2005) CAPS1 regulates catecholamine loading of large dense- Wang Y, Okamoto M, Schmitz F, Hofmann K, Sudhof TC (1997) Rim core vesicles. Neuron 46(1):75–88. https ://doi.org/10.1016/j. is a putative Rab3 effector in regulating synaptic-vesicle fusion. neuro n.2005.02.019 Nature 388(6642):593–598 Stavoe AK, Nelson JC, Martinez-Velazquez LA, Klein M, Samuel AD, Watanabe S, Liu Q, Davis MW, Hollopeter G, Thomas N, Jorgensen Colon-Ramos DA (2012) Synaptic vesicle clustering requires NB et al (2013) Ultrafast endocytosis at Caenorhabditis elegans a distinct MIG-10/Lamellipodin isoform and ABI-1 down- neuromuscular junctions. Elife. https ://doi.org/10.7554/eLif e stream from Netrin. Genes Dev 26(19):2206–2221. https ://doi. .00723 org/10.1101/gad.19340 9.112 Weimer RM, Richmond JE, Davis WS, Hadwiger G, Nonet ML, Jor- Stenmark H, Olkkonen VM (2001) The rab GTPase family. Genome gensen EM (2003) Defects in synaptic vesicle docking in unc-18 Biol 2(5):reviews3007.1–3007.7 mutants. Nat Neurosci 6(10):1023–1030. https://doi.or g/10.1038/ Stevens DR, Wu ZX, Matti U, Junge HJ, Schirra C, Becherer U et al nn111 8 (2005) Identification of the minimal protein domain required for Xu J, Pang ZP, Shin OH, Sudhof TC (2009) Synaptotagmin-1 func- 2+ priming activity of Munc13-1. Curr Biol 15(24):2243–2248. tions as a Ca sensor for spontaneous release. Nat Neurosci https ://doi.org/10.1016/j.cub.2005.10.055 12(6):759–766. https ://doi.org/10.1038/nn.2320 Sudhof TC (1995) The synaptic vesicle cycle: a cascade of protein- Yamashita M, Iwasaki K, Doi M (2009) The non-neuronal syntaxin protein interactions. Nature 375(6533):645–653. https ://doi. SYN-1 regulates defecation behavior and neural activity in C. org/10.1038/37564 5a0 elegans through interaction with the Munc13-like protein AEX- Sudhof TC (2004) The synaptic vesicle cycle. Annu Rev Neurosci 1. Biochem Biophys Res Commun 378(3):404–408. https ://doi. 27:509–547. https ://doi.or g/10.1146/annur ev .neur o .26.04100 org/10.1016/j.bbrc.2008.11.064 2.13141 2 Yu SC, Klosterman SM, Martin AA, Gracheva EO, Richmond JE Sudhof TC (2013) Neurotransmitter release: the last millisecond in (2013) Differential roles for snapin and synaptotagmin in the the life of a synaptic vesicle. Neuron 80(3):675–690. https://doi. synaptic vesicle cycle. PLoS ONE 8(2):e57842. https ://doi. org/10.1016/j.neuro n.2013.10.022org/10.1371/journ al.pone.00578 42 Sun J, Pang ZP, Qin D, Fahim AT, Adachi R, Sudhof TC (2007) A Zhou KM, Stawicki TM, Goncharov A, Jin YS (2013) Position of 2+ dual-Ca -sensor model for neurotransmitter release in a central UNC-13 in the active zone regulates synaptic vesicle release synapse. Nature 450(7170):676–682. http s ://doi.org/1 0.1038/ probability and release kinetics. Elife. https ://doi.org/10.7554/ natur e0630 8eLife .01180 Sun Y, Hu Z, Goeb Y, Dreier L (2013) The F-box protein MEC-15 (FBXW9) promotes synaptic transmission in GABAergic motor 1 3
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