SnapShot: - Complexes Stéphane Baudouin and Peter Scheiffele Biozentrum of the University of Basel, 4056 Basel, Switzerland

908 Cell 141, May 28, 2010 ©2010 Elsevier Inc. DOI 10.1016/j.cell.2010.05.024 See online version for legend and references. SnapShot: Neuroligin-Neurexin Complexes Stéphane Baudouin and Peter Scheiffele Biozentrum of the University of Basel, 4056 Basel, Switzerland

The neuroligin-neurexin complex is a heterophilic adhesion system at neuronal synapses. Both and have synaptogenic activities that organize pre- and postsynaptic compartments in a bidirectional manner. Multiple , alternative promoters, and extensive generate neuroligin and neurexin variants that may underlie synapse-specific functions. In human patients, mutations in neuroligin and neurexin genes are associated with neurodevelopmental disorders, highlighting the importance of these for cognitive function.

Generation of Isoform Diversity In rodents and humans, neuroligins are encoded by four and five genes, respectively (NLGN1, 2, 3, 4, and NLGN4Y, which is only in humans). Alternative splicing at one (in NLGN2, 3, and 4) or two sites (in NLGN1) designated A and B generates twelve variants with insertions in the extracellular domain, such as NL1(−) lacking any insertions and NL1B contain- ing an insertion at site B; constitutive exons are depicted in blue, alternative exons in red. Vertebrate neurexins are encoded by three genes (NRXN1, 2, and 3), each of which is controlled by two alternative promoters (α and β), resulting in six primary transcripts (NRX1α, NRX1β, NRX2α, etc.). Complex alternative splicing (usage of alternative exons, splice donor, and acceptor sites) at five sites (1, 2, 3, 4, and 5) generates more than 2000 alternative splice variants (such as NRX1α4(+) or NRX1α4(−) that include or lack insertion at alternative splice site 4, respectively). In the fruit fly Drosophila melanogaster, NLGN and NRXN genes have critical roles in the assembly of glutamatergic neuromuscular junctions but are less complex and apparently encode fewer variants.

Biochemical Interactions of Neuroligins and Neurexins Neuroligin (NL) and neurexin (NRX) proteins each contain a single transmembrane domain, short cytoplasmic domains with juxtamembrane Band4.1 interaction sites, and C-terminal PDZ-binding motifs. The extracellular sequences of NLs contain a large cholinesterase-homology (ChE) domain. NRXα extracellular sequences consist of repeats of G-like (LG) and epidermal growth factor (EGF) domains, whereas NRXβs carry a single LG domain. NL-NRX complexes form through interactions via the ChE and LG domains, differ between NRXα and NRXβ variants, and are further regulated by alternative splicing. Notably, NLs and NRXs also interact with multiple additional binding partners (see below). Three examples of isoform-dependent interactions are illustrated: NRXβs interact with some ligands independently of splice insertions at site 4 (common ligands), whereas interaction with other ligands is regulated by an insertion in site 4 (NRXβ4(−)-selective ligands). In NRXαs, NRXα4(−)-selective ligands interact only with NRXαs lacking an insertion at site 4 but not containing an insertion at this site. Finally, α-dystroglycan (α-DG) interacts in splice variant-dependent manner with NRXβ4(−) and NRXα2(−) variants, that is, interactions with LG domains 2 and 4 are inhibited by inclusion of splice insertions. Alterations in ligand binding due to alternative splice insertions range from subtle to large shifts in affinity, which modulate or abolish interactions, respectively.

Synaptic Complexes Nucleated by the Neuroligin-Neurexin Complex NL1 and NL2 isoforms exhibit synapse-specific localization and functions. NL1B is concentrated at glutamatergic synapses and central cholinergic synapses, whereas NL2A is at GABAergic and glycinergic synapses. NL3 has been detected at both glutamatergic and GABAergic synapses in vitro, but localization of NL3 and NL4 in vivo is currently unknown. At glutamatergic synapses, NL1 couples to the scaffolding protein PSD95 that may in turn interact with NMDA-type glutamate receptors. NL1 may interact with the extracellular matrix

protein thrombospondin (TSP) that is secreted from astroglia. At GABAergic synapses, NL2 couples via the cytoplasmic tail to collybistin, , and GABAA-receptors. NRXs are concentrated in the presynaptic terminal, but dendritic pools are also detected in vivo. Presynaptically, NRXs bind through PDZ-domain interactions to the scaffold- ing proteins Mint/X11alpha and Lin2/Cask. Cask may exhibit a kinase activity that can phosphorylate NRX. Through the Cask/Mint/Lin7 complex, NRX may couple to presynaptic voltage-gated calcium channels (Cav2.1 and Cav2.2). NRXs also engage in extracellular interactions with ligands other than the NLs: with the black widow spider toxin α-latrotoxin (LTX), at glutamatergic synapses with leucine-rich repeat transmembrane proteins (LRRTM1 and 2), and at GABAergic synapses with α-DG. In vitro, specific neurexin splice variants exhibit differential ability to promote assembly of postsynaptic glutamatergic and GABAergic structures, but cell type-specific NRX isoform repertoires are currently unknown.

Disorder-Associated Mutations in NRXN and NLGN Genes Multiple structural variants in the NLGN and NRXN genes, including copy number variations (CNVs), point mutations, and missense mutations (denoted by a “*”), are associated with -spectrum disorders and . Further disease-associated mutations have been identified in SHANK3, a cytoplasmic component of the NL-NRX complex. The NL3 R451C and NL4 R87W mutations result in degradation of the mutant protein in the endoplasmic reticulum. However, 10% of NL3 R451C are transported to the cell surface and retain some functionality. Genetically modified mouse mutants (NLGN1, NLGN3 R451C, NLGN4, and NRXN1) exhibit behavioral abnormalities that resemble endo- phenotypes of the disorders.

Acknowledgments

Work in the authors’ laboratory is supported by grants from the Swiss National Fund, the National Institute on Drug Abuse, and the Biozentrum of the University of Basel. This work was supported in part by a grant from F. Hoffmann-La Roche.

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908.e1 Cell 141, May 28, 2010 ©2010 Elsevier Inc. DOI 10.1016/j.cell.2010.05.024