View metadata, citation and similar papers at core.ac.uk brought to you by CORE provided by Elsevier - Publisher Connector R620 Dispatch Synapse formation: If it looks like a duck and quacks like a duck… Isabel Cantallops and Hollis T. Cline Neuroligin and neurexin form an intercellular adhesion Scheiffele et al. [1] used a powerful experimental system complex sufficient to trigger formation of functional to demonstrate that expression of neuroligin is sufficient presynaptic elements in vitro. This single molecular to trigger the establishment of presynaptic specializations interaction appears to initiate clustering of synaptic in pontine axons. They co-cultured pontine explants with vesicles, assembly of vesicle-release machinery and non-neuronal heterologous cells, HEK cells, which do not morphological changes at the presynaptic membrane. express endogenous neuroligin. The pontine explants extend axons onto the HEK cells and will form presynap- Address: Cold Spring Harbor Laboratory, Cold Spring Harbor, New York 11724, USA. tic junctions if the transfected HEK cells produce neuroli- gin. The presynaptic specializations were recognized at Current Biology 2000, 10:R620–R623 the light microscope level as a clustering of synapsin 1 immunoreactivity, and at the electron microscope level as 0960-9822/00/$ – see front matter © 2000 Elsevier Science Ltd. All rights reserved. parallel plasma membranes with electron-dense material and clusters of small clear synaptic vesicles. Scheiffele Synapses are asymmetric junctions between neurons et al. [1] further showed that the presynaptic vesicles through which signals pass from the presynaptic neuron underwent cycles of exocytosis and endocytosis in the to the postsynaptic neuron. The presynaptic axon termi- presence of a depolarizing concentration of K+ ions. These nal is specialized to convert an electrical signal into an observations indicate that the axonal specializations intercellular chemical signal via calcium-dependent exo- induced by neuroligin look like presynaptic terminals and cytosis of neurotransmitter. The postsynaptic element is act like presynaptic terminals. specialized to detect transmitter and convert the chemical signal back into an electrical signal, which is then trans- A structure/function analysis of neuroligin revealed a close mitted through the neuron. In addition, complex signal correlation between the protein’s neurexin-binding and transduction events occur both presynaptically and post- vesicle-clustering activities [1]. The cytoplasmic tail of synaptically, which regulate the formation of synaptic neuroligin was found to be required for targeting to the connections and the strength of synaptic transmission. membrane, but not for either neurexin binding or synaptic The sequence of events that occurs during the establish- vesicle clustering; the extracellular acetylcholinesterase ment of synaptic contacts and the molecular interactions homology domain, however, was required for both that trigger these events in the central nervous system neurexin binding and vesicle clustering. Swapping the remain unclear. acetylcholinesterase homology domain with the catalyti- cally active esterase domain of acetylcholinesterase Recent work by Scheiffele et al. [1] has shed new light on blocked activity. This essential function of the esterase the potential role of the intercellular neuroligin–neurexin homology domain is interesting because the early complex in triggering presynaptic specializations during expression of esterases in the developing central nervous synaptogenesis. Neuroligins are heterophilic transmem- system (CNS) has long been a mystery. Recent studies brane cell adhesion molecules that bind β-neurexins [2]. [5,6] suggest that a non-catalytic function of acetyl- Both neuroligins and β-neurexins are enriched in synaptic cholinesterase may be required for normal CNS morpho- membranes in adult brain tissue, and in particular neuroligin genesis through an interaction with neurexin. Although localizes to the postsynaptic membrane of glutamatergic the data reported by Scheiffele et al. [1] do not give synapses [3]. β-neurexins are transmembrane proteins with detailed information about the type of interaction an extracellular laminin-like domain and a cytoplasmic tail between the esterase homology domain of neuroligin and with a PDZ-binding domain. Neuroligin is composed of an the binding sites in neurexin, comparison of the conserved extracellular non-catalytic acetylcholinesterase homology and divergent regions of the esterase domains in neuroli- domain, which is required for neurexin binding, and a gin and acetylcholinesterase will provide a starting point cytoplasmic tail with a PDZ-binding domain. Neuroligin for more detailed structure/function studies. has been implicated in mental retardation [4], suggesting a developmental function for neuroligin–neurexin. On The pièce de resistance in this series of experiments was the the basis of neurexin’s synaptic location, heterophilic demonstration that neuroligin–neurexin binding plays a binding properties and interesting binding partners (see role in synaptogenesis between pontine axons and their Figure 1), Scheiffele et al. [1] hypothesized that the inter- normal target, cerebellar granule cells. Using the pontine action between neuroligin and neurexin might be impor- explants co-cultured with HEK cells, Scheiffele et al. [1] tant in synaptogenesis. demonstrated that a soluble neurexin–immunoglobulin Dispatch R621 Figure 1 Schematic illustration of some components of Synaptic vesicles the glutamatergic synapse in adult brain. Domains Presynaptically, the PDZ-binding domain of Munc18-1 SH3 neurexin binds CASK, which in turn binds Mint CaM PTB kinase and Veli. These proteins affect presynaptic domain GK signal transduction events and indirectly affect PDZ Mint Veli CASK vesicle transport and neurotransmission (LIN-10) (LIN-7) (LIN-2) through an association between Mint-1 and N-cadherin Munc18-1. Postsynaptically, the cytoplasmic Presynaptic L1 membrane Ca2+ channel portion of neuroligin includes a PDZ-binding EphR Synaptic cleft Neuroligin domain through which it binds postsynaptic Neurexin scaffold proteins, including PSD-95 and NMDAR K+ channel AMPAR Postsynaptic S-SCAM. PSD-95 and S-SCAM bind NMDA membrane receptors (NMDAR) and K+ channels, and likely form extensive submembrane scaffolds nRapGEP SynGAP PSD-95 S-SCAM GRIP/ABP that cluster transmembrane proteins and regulate the physical proximity of regulatory Current Biology proteins — such as CaMKII, SynGAP and nRapGEP — with their effectors. Grip/ABP, spectrometric analysis to identify proteins evidence that the formation and maintenance a scaffold protein with seven PDZ domains, associated with the NMDA receptors in the of synaptic contacts includes a combination of binds AMPA receptors (AMPAR) and Ephrin PSD of adult brain, finding that cell adhesion cell adhesion and synaptic signaling receptors (EphR) in a complex that is distinct molecules L1 and N-cadherin are part of the machinery. Symbols for the domains of the from the NMDA receptor complex. Recently NMDA receptors complex, in addition to different proteins are indicated in the upper two groups [20,21] have used mass neuroligin–neurexin. This provides additional right corner. fusion protein blocked neuroligin-induced synaptic vesicle vesicle protein VAMP, tagged with green fluorescent clustering, presumably by interfering with the binding of protein (GFP), to monitor the establishment of the presy- endogenous neurexin on pontine axons with the exoge- naptic elements in hippocampal cultures. Together, these nous neuroligin in HEK cells. Scheiffele et al. [1] then observations suggest that a heterogeneous population of took advantage of this finding to test whether soluble vesicular bodies, including small clear synaptic vesicles, neurexin–immunoglobulin fusion protein affected cluster- travel within an axon and are ‘captured’ at particular sites ing of synapsin 1 in co-cultures of pontine explants and along the axon. The restricted mobility of the vesicles then granule cells: they found that the soluble fusion protein results in their coalescence at one location, which becomes decreased the number of synapsin 1 clusters per cell by recognizable as a presynaptic specialization. The data of half, indicating that the neuroligin–neurexin interaction Scheiffele et al. [1] now suggest that intercellular neuroli- likely plays a role in synaptogenesis in this system. gin–neurexin signaling is the event that initiates the capture of synaptic vesicles at a particular presynaptic location. Synapse formation requires intimate communication between the presynaptic and postsynaptic cells, so that The capture of synaptic vesicles at the designated presynaptic elements do not develop in the absence of presynaptic site likely involves signaling through a protein postsynaptic specializations and vice versa. Synaptogene- complex that associates with the PDZ-binding domain of sis is therefore likely to be a multistep process, in which the cytoplasmic tail of neurexin. The PDZ-binding domain progression from step to step requires feedback across the of neurexin binds CASK, which in turn associates with pro- synapse (Figure 2). One amazing feature of the observa- teins known as Mint and Veli (Figures 1 and 2). By inter- tions reported by Scheiffele et al. [1] is that neuroligin is acting with these proteins, neurexins can affect signal sufficient to trigger significant presynaptic specializa- transduction, synaptic vesicle