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The CADM family of proteins consists of four neuronal specific adhesion molecules (CADM1, CADM2, CADM3 and CADM4) that mediate the direct contact and interaction between axons and glia. In the peripheral nerve, axon-Schwann cell interaction is essential for the structural organization of myelinated fibres and is primarily mediated by the binding of CADM3, expressed in axons, to CADM4, expressed by myelinating Schwann cells. We have identified-by whole exome sequencing-three unrelated families, including one de novo patient, with axonal Charcot-Marie-Tooth disease (CMT2) sharing the same private variant in CADM3, Tyr172Cys. This variant is absent in 230 000 control chromosomes from gnomAD and predicted to be pathogenic. Most CADM3 patients share a similar phenotype consisting of autosomal dominant CMT2 with marked upper limb involvement. High resolution mass spectrometry analysis detected a newly created disulphide bond in the mutant CADM3 potentially modifying the native protein conformation. Our data support a retention of the mutant protein in the endoplasmic reticulum and reduced cell surface expression in vitro. Stochastic optical reconstruction microscopy imaging revealed decreased co-localization of the mutant with CADM4 at intercellular contact sites. Mice carrying the corresponding human mutation (Cadm3Y170C) showed reduced expression of the mutant protein in axons. Cadm3Y170C mice showed normal nerve conduction and myelin morphology, but exhibited abnormal axonal organization, including abnormal distribution of Kv1.2 channels and Caspr along myelinated axons. Our findings indicate the involvement of abnormal axon-glia interaction as a disease-causing mechanism in CMT patients with CADM3 mutations., (© The Author(s) (2021). Published by Oxford University Press on behalf of the Guarantors of Brain. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Cell adhesion proteins of the Cadm (SynCAM/Necl) family regulate myelination and the organization of myelinated axons. In the peripheral nervous system (PNS), intercellular contact between Schwann cells and their underlying axons is believed to be mediated by binding of glial Cadm4 to axonal Cadm3 or Cadm2. Nevertheless, given that distinct neurons express different combinations of the Cadm proteins, the identity of the functional axonal ligand for Cadm4 remains to be determined. Here, we took a genetic approach to compare the phenotype of Cadm4 null mice, which exhibit abnormal distribution of Caspr and Kv1 potassium channels, with mice lacking different combinations of Cadm1 - Cadm3 genes. We show that in contrast to mice lacking the single Cadm1 , Cadm2 , or Cadm3 genes, genetic ablation of all three phenocopies the abnormalities detected in the absence of Cadm4. Similar defects were observed in double mutant mice lacking Cadm3 and Cadm2 (i.e., Cadm3 -/- /Cadm2 -/- ) or Cadm3 and Cadm1 (i.e., Cadm3 Myelination by Schwann cells enables fast conduction of action potentials along motor and sensory axons. In these nerves, Schwann cell-axon contact is mediated by cell adhesion molecules of the Cadm family. Cadm4 in Schwann cells regulates axonal ensheathment and myelin wrapping, as well as the organization of the axonal membrane, but the identity of its axonal ligands is not clear. Here, we reveal that Cadm mediated axon-glia interactions depend on a hierarchical adhesion code that involves multiple family members. Our results provide important insights into the molecular mechanisms of axon-glia communication, and the function of Cadm proteins in PNS myelin.-/- /Cadm1 -/- ), but not in mice lacking Cadm1 and Cadm2 (i.e., Cadm1 -/- /Cadm2 -/- ). Furthermore, axonal organization abnormalities were also detected in Cadm3 null mice that were heterozygous for the two other axonal Cadms. Our results identify Cadm3 as the main axonal ligand for glial Cadm4, and reveal that its absence could be compensated by the combined action of Cadm2 and Cadm1. SIGNIFICANCE STATEMENT Myelination by Schwann cells enables fast conduction of action potentials along motor and sensory axons. In these nerves, Schwann cell-axon contact is mediated by cell adhesion molecules of the Cadm family. Cadm4 in Schwann cells regulates axonal ensheathment and myelin wrapping, as well as the organization of the axonal membrane, but the identity of its axonal ligands is not clear. Here, we reveal that Cadm mediated axon-glia interactions depend on a hierarchical adhesion code that involves multiple family members. Our results provide important insights into the molecular mechanisms of axon-glia communication, and the function of Cadm proteins in PNS myelin., (Copyright © 2021 the authors.)

The nodes of Ranvier have clustered Na + and K + channels necessary for rapid and efficient axonal action potential conduction. However, detailed mechanisms of channel clustering have only recently been identified: they include two independent axon-glia interactions that converge on distinct axonal cytoskeletons. Here, we discuss how glial cell adhesion molecules and the extracellular matrix molecules that bind them assemble combinations of ankyrins, spectrins and other cytoskeletal scaffolding proteins, which cluster ion channels. We present a detailed molecular model, incorporating these overlapping mechanisms, to explain how the nodes of Ranvier are assembled in both the peripheral and central nervous systems.

The paranodal junctions flank mature nodes of Ranvier and provide a barrier between ion channels at the nodes and juxtaparanodes. These junctions also promote node assembly and maintenance by mechanisms that are poorly understood. Here, we examine their role in the accumulation of NF186, a key adhesion molecule of PNS and CNS nodes. We previously showed that NF186 is initially targeted/accumulates via its ectodomain to forming PNS (hemi)nodes by diffusion trapping, whereas it is later targeted to mature nodes by a transport-dependent mechanism mediated by its cytoplasmic segment. To address the role of the paranodes in this switch, we compared accumulation of NF186 ectodomain and cytoplasmic domain constructs in WT versus paranode defective (i.e., Caspr-null) mice. Both pathways are affected in the paranodal mutants. In the PNS of Caspr-null mice, diffusion trapping mediated by the NF186 ectodomain aberrantly persists into adulthood, whereas the cytoplasmic domain/transport-dependent targeting is impaired. In contrast, accumulation of NF186 at CNS nodes does not undergo a switch; it is predominantly targeted to both forming and mature CNS nodes via its cytoplasmic domain and requires intact paranodes. Fluorescence recovery after photobleaching analysis indicates that the paranodes provide a membrane diffusion barrier that normally precludes diffusion of NF186 to nodes. Linkage of paranodal proteins to the underlying cytoskeleton likely contributes to this diffusion barrier based on 4.1B and βII spectrin expression in Caspr-null mice. Together, these results implicate the paranodes as membrane diffusion barriers that regulate targeting to nodes and highlight differences in the assembly of PNS and CNS nodes. SIGNIFICANCE STATEMENT Nodes of Ranvier are essential for effective saltatory conduction along myelinated axons. A major question is how the various axonal proteins that comprise the multimeric nodal complex accumulate at this site. Here we examine how targeting of NF186, a key nodal adhesion molecule, is regulated by the flanking paranodal junctions. We show that the transition from diffusion-trapping to transport-dependent accumulation of NF186 requires the paranodal junctions. We also demonstrate that these junctions are a barrier to diffusion of axonal proteins into the node and highlight differences in PNS and CNS node assembly. These results provide new insights into the mechanism of node assembly and the pathophysiology of neurologic disorders in which impaired paranodal function contributes to clinical disability., (Copyright © 2020 the authors.)

The initiation of axoglial contact is considered a prerequisite for myelination, yet the role cell adhesion molecules (CAMs) play in mediating such interactions remains unclear. To examine the function of axoglial CAMs, we tested whether enhanced CAM-mediated adhesion between OLs and neurons could affect myelination. Here we show that increased expression of a membrane-bound extracellular domain of Cadm4 (Cadm4dCT) in cultured oligodendrocytes results in the production of numerous axoglial contact sites that fail to elongate and generate mature myelin. Transgenic mice expressing Cadm4dCT were hypomyelinated and exhibit multiple myelin abnormalities, including myelination of neuronal somata. These abnormalities depend on specific neuron-glial interaction as they were not observed when these OLs were cultured alone, on nanofibers, or on neurons isolated from mice lacking the axonal receptors of Cadm4. Our results demonstrate that tightly regulated axon-glia adhesion is essential for proper myelin targeting and subsequent membrane wrapping and lateral extension., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Postnatal synapse elimination plays a critical role in sculpting and refining neural connectivity throughout the central and peripheral nervous systems, including the removal of supernumerary axonal inputs from neuromuscular junctions (NMJs). Here, we reveal a novel and important role for myelinating glia in regulating synapse elimination at the mouse NMJ, where loss of a single glial cell protein, the glial isoform of neurofascin (Nfasc155), was sufficient to disrupt postnatal remodeling of synaptic circuitry. Neuromuscular synapses were formed normally in mice lacking Nfasc155, including the establishment of robust neuromuscular synaptic transmission. However, loss of Nfasc155 was sufficient to cause a robust delay in postnatal synapse elimination at the NMJ across all muscle groups examined. Nfasc155 regulated neuronal remodeling independently of its canonical role in forming paranodal axo-glial junctions, as synapse elimination occurred normally in mice lacking the axonal paranodal protein Caspr. Rather, high-resolution proteomic screens revealed that loss of Nfasc155 from glial cells was sufficient to disrupt neuronal cytoskeletal organization and trafficking pathways, resulting in reduced levels of neurofilament light (NF-L) protein in distal axons and motor nerve terminals. Mice lacking NF-L recapitulated the delayed synapse elimination phenotype observed in mice lacking Nfasc155, suggesting that glial cells regulate synapse elimination, at least in part, through modulation of the axonal cytoskeleton. Together, our study reveals a glial cell-dependent pathway regulating the sculpting of neuronal connectivity and synaptic circuitry in the peripheral nervous system., (Copyright © 2014 the authors 0270-6474/14/3412904-15$15.00/0.)

Clustering of Na(+) channels at the nodes of Ranvier is coordinated by myelinating glia. In the peripheral nervous system, axoglial contact at the nodes is mediated by the binding of gliomedin and glial NrCAM to axonal neurofascin 186 (NF186). This interaction is crucial for the initial clustering of Na(+) channels at heminodes. As a result, it is not clear whether continued axon-glial contact at nodes of Ranvier is required to maintain these channels at the nodal axolemma. Here, we report that, in contrast to mice that lack either gliomedin or NrCAM, absence of both molecules (and hence the glial clustering signal) resulted in a gradual loss of Na(+) channels and other axonal components from the nodes, the formation of binary nodes, and dysregulation of nodal gap length. Therefore, these mice exhibit neurological abnormalities and slower nerve conduction. Disintegration of the nodes occurred in an orderly manner, starting with the disappearance of neurofascin 186, followed by the loss of Na(+) channels and ankyrin G, and then βIV spectrin, a sequence that reflects the assembly of nodes during development. Finally, the absence of gliomedin and NrCAM led to the invasion of the outermost layer of the Schwann cell membrane beyond the nodal area and the formation of paranodal-like junctions at the nodal gap. Our results reveal that axon-glial contact mediated by gliomedin, NrCAM, and NF186 not only plays a role in Na(+) channel clustering during development, but also contributes to the long-term maintenance of Na(+) channels at nodes of Ranvier.

The interaction between myelinating Schwann cells and the axons they ensheath is mediated by cell adhesion molecules of the Cadm/Necl/SynCAM family. This family consists of four members: Cadm4/Necl4 and Cadm1/Necl2 are found in both glia and axons, whereas Cadm2/Necl3 and Cadm3/Necl1 are expressed by sensory and motor neurons. By generating mice lacking each of the Cadm genes, we now demonstrate that Cadm4 plays a role in the establishment of the myelin unit in the peripheral nervous system. Mice lacking Cadm4 (PGK-Cre/Cadm4(fl/fl)), but not Cadm1, Cadm2, or Cadm3, develop focal hypermyelination characterized by tomacula and myelin outfoldings, which are the hallmark of several Charcot-Marie-Tooth neuropathies. The absence of Cadm4 also resulted in abnormal axon-glial contact and redistribution of ion channels along the axon. These neuropathological features were also found in transgenic mice expressing a dominant-negative mutant of Cadm4 lacking its cytoplasmic domain in myelinating glia Tg(mbp-Cadm4dCT), as well as in mice lacking Cadm4 specifically in Schwann cells (DHH-Cre/Cadm4(fl/fl)). Consistent with these abnormalities, both PGK-Cre/Cadm4(fl/fl) and Tg(mbp-Cadm4dCT) mice exhibit impaired motor function and slower nerve conduction velocity. These findings indicate that Cadm4 regulates the growth of the myelin unit and the organization of the underlying axonal membrane.

Objectives: We asked whether autoantibodies against neurofascin (NF)186 or NF155, both localized at the nodes of Ranvier, are present in serum of patients with inflammatory neuropathy, and whether NF-specific monoclonal antibodies are pathogenic in vivo., Methods: We cloned human NF155 and NF186, and developed an ELISA and cell-based assay to screen for antibodies to human NF in a total of 434 donors including 294 patients with Guillain-Barré syndrome variants acute inflammatory demyelinating polyneuropathy (AIDP), acute motor axonal neuropathy, and chronic inflammatory demyelinating polyneuropathy (CIDP). We characterized reactive samples by isotyping, tissue section staining, and epitope mapping. We also injected NF-specific monoclonal antibodies IV into rats with experimental autoimmune neuritis., Results: We detected autoantibodies to NF by ELISA in 4% of patients with AIDP and CIDP, but not in controls. Most positive samples contained immunoglobulin G (IgG)1, IgG3, or IgG4 antibodies directed to only one isoform of NF. Two patients with CIDP showed particularly high (1:10,000 dilution) NF155-specific reactivity in both assays and stained paranodes. Two other patients with CIDP who benefited from plasma exchange exhibited antibodies to NF155 by ELISA, and upon affinity purification, antibodies to both isoforms were observed by both assays. Anti-NF monoclonal antibodies enhanced and prolonged induced neuritis in rats., Conclusions: Autoantibodies to NF are detected in a very small proportion of patients with AIDP and patients with CIDP, but may nevertheless be pathogenic in these cases.

Saltatory conduction requires high-density accumulation of Na(+) channels at the nodes of Ranvier. Nodal Na(+) channel clustering in the peripheral nervous system is regulated by myelinating Schwann cells through unknown mechanisms. During development, Na(+) channels are first clustered at heminodes that border each myelin segment, and later in the mature nodes that are formed by the fusion of two heminodes. Here, we show that initial clustering of Na(+) channels at heminodes requires glial NrCAM and gliomedin, as well as their axonal receptor neurofascin 186 (NF186). We further demonstrate that heminodal clustering coincides with a second, paranodal junction (PNJ)-dependent mechanism that allows Na(+) channels to accumulate at mature nodes by restricting their distribution between two growing myelin internodes. We propose that Schwann cells assemble the nodes of Ranvier by capturing Na(+) channels at heminodes and by constraining their distribution to the nodal gap. Together, these two cooperating mechanisms ensure fast and efficient conduction in myelinated nerves.

Accumulation of Na(+) channels at the nodes of Ranvier is a prerequisite for saltatory conduction. In peripheral nerves, clustering of these channels along the axolemma is regulated by myelinating Schwann cells through a yet unknown mechanism. We report the identification of gliomedin, a glial ligand for neurofascin and NrCAM, two axonal immunoglobulin cell adhesion molecules that are associated with Na+ channels at the nodes of Ranvier. Gliomedin is expressed by myelinating Schwann cells and accumulates at the edges of each myelin segment during development, where it aligns with the forming nodes. Eliminating the expression of gliomedin by RNAi, or the addition of a soluble extracellular domain of neurofascin to myelinating cultures, which caused the redistribution of gliomedin along the internodes, abolished node formation. Furthermore, a soluble gliomedin induced nodal-like clusters of Na+ channels in the absence of Schwann cells. We propose that gliomedin provides a glial cue for the formation of peripheral nodes of Ranvier.

Three cell adhesion molecules are present at the axoglial junctions that form between the axon and myelinating glia on either side of nodes of Ranvier. These include an axonal complex of contacin-associated protein (Caspr) and contactin, which was proposed to bind NF155, an isoform of neurofascin located on the glial paranodal loops. Here, we show that NF155 binds directly to contactin and that surprisingly, coexpression of Caspr inhibits this interaction. This inhibition reflects the association of Caspr with contactin during biosynthesis and the resulting expression of a low molecular weight (LMw), endoglycosidase H-sensitive isoform of contactin at the cell membrane, which remains associated with Caspr but is unable to bind NF155. Accordingly, deletion of Caspr in mice by gene targeting results in a shift from the LMw- to a HMw-contactin glycoform. These results demonstrate that Caspr regulates the intracellular processing and transport of contactin to the cell surface, thereby affecting its ability to interact with other cell adhesion molecules.

An axonal complex of cell adhesion molecules consisting of Caspr and contactin has been found to be essential for the generation of the paranodal axo-glial junctions flanking the nodes of Ranvier. Here we report that although the extracellular region of Caspr was sufficient for directing it to the paranodes in transgenic mice, retention of the Caspr-contactin complex at the junction depended on the presence of an intact cytoplasmic domain of Caspr. Using immunoelectron microscopy, we found that a Caspr mutant lacking its intracellular domain was often found within the axon instead of the junctional axolemma. We further show that a short sequence in the cytoplasmic domain of Caspr mediated its binding to the cytoskeleton-associated protein 4.1B. Clustering of contactin on the cell surface induced coclustering of Caspr and immobilized protein 4.1B at the plasma membrane. Furthermore, deletion of the protein 4.1B binding site accelerated the internalization of a Caspr-contactin chimera from the cell surface. These results suggest that Caspr serves as a "transmembrane scaffold" that stabilizes the Caspr/contactin adhesion complex at the paranodal junction by connecting it to cytoskeletal components within the axon.

Kallmann syndrome is a neurological disorder characterized by various behavioral and neuroanatomical defects. The X-linked form of this disease is caused by mutations in the KAL-1 gene, which codes for a secreted molecule that is expressed in restricted regions of the brain. Its molecular mechanism of action has thus far remained largely elusive. We show here that expression of the Caenorhabditis elegans homolog of KAL-1 in selected sensory and interneuron classes causes a highly penetrant, dosage-dependent, and cell autonomous axon-branching phenotype. In a different cellular context, heterologous C. elegans kal-1 expression causes a highly penetrant axon-misrouting phenotype. The axon-branching and -misrouting activities require different domains of the KAL-1 protein. In a genetic modifier screen we isolated several loci that either suppress or enhance the kal-1-induced axonal defects, one of which codes for an enzyme that modifies specific residues in heparan sulfate proteoglycans, namely heparan-6O-sulfotransferase. We hypothesize that KAL-1 binds by means of a heparan sulfate proteoglycan to its cognate receptor or other extracellular cues to induce axonal branching and axon misrouting.

Rapid action potential propagation in myelinated axons requires Na⁺ channel clustering at nodes of Ranvier. However, the mechanism of clustering at CNS nodes remains poorly understood. Here, we show that the assembly of nodes of Ranvier in the CNS involves three mechanisms: a glia-derived extracellular matrix (ECM) complex containing proteoglycans and adhesion molecules that cluster NF186, paranodal axoglial junctions that function as barriers to restrict the position of nodal proteins, and axonal cytoskeletal scaffolds (CSs) that stabilize nodal Na⁺ channels. We show that while mice with a single disrupted mechanism had mostly normal nodes, disruptions of the ECM and paranodal barrier, the ECM and CS, or the paranodal barrier and CS all lead to juvenile lethality, profound motor dysfunction, and significantly reduced Na⁺ channel clustering. Our results demonstrate that ECM, paranodal, and axonal cytoskeletal mechanisms ensure robust CNS nodal Na⁺ channel clustering.

Axon initial segment (AIS) cell surface proteins mediate key biological processes in neurons including action potential initiation and axo-axonic synapse formation. However, few AIS cell surface proteins have been identified. Here, we use antibody-directed proximity biotinylation to define the cell surface proteins in close proximity to the AIS cell adhesion molecule Neurofascin. To determine the distributions of the identified proteins, we use CRISPR-mediated genome editing for insertion of epitope tags in the endogenous proteins. We identify Contactin-1 (Cntn1) as an AIS cell surface protein. Cntn1 is enriched at the AIS through interactions with Neurofascin and NrCAM. We further show that Cntn1 contributes to assembly of the AIS extracellular matrix, and regulates AIS axo-axonic innervation by inhibitory basket cells in the cerebellum and inhibitory chandelier cells in the cortex. Few resident cell surface proteins have been identified at the axon initial segment. Here, Ogawa and colleagues use proximity labeling and proteomics to identify Contactin-1 as a transmembrane axon initial segment protein that regulates brain wiring. [ABSTRACT FROM AUTHOR]

Action potential propagation along myelinated axons depends on the geometry of the myelin unit and the division of the underlying axon to specialized domains. The latter include the nodes of Ranvier (NOR), the paranodal junction (PNJ) flanking the nodes, and the adjacent juxtaparanodal region that is located below the compact myelin of the internode. Each of these domains contains a unique composition of axoglial adhesion molecules (CAMs) and cytoskeletal scaffolding proteins, which together direct the placement of specific ion channels at the nodal and juxtaparanodal axolemma. In the last decade it has become increasingly clear that antibodies to some of these axoglial CAMs cause immune‐mediated neuropathies. In the current review we detail the molecular composition of the NOR and adjacent membrane domains, describe the function of different CAM complexes that mediate axon‐glia interactions along the myelin unit, and discuss their involvement and the underlying mechanisms taking place in peripheral nerve pathologies. This growing group of pathologies represent a new type of neuropathies termed "nodopathies" or "paranodopathies" that are characterized by unique clinical and molecular features which together reflect the mechanisms underlying the molecular assembly and maintenance of this specialized membrane domain. [ABSTRACT FROM AUTHOR]

The CADM family of proteins consists of four neuronal specific adhesion molecules (CADM1, CADM2, CADM3 and CADM4) that mediate the direct contact and interaction between axons and glia. In the peripheral nerve, axon-Schwann cell interaction is essential for the structural organization of myelinated fibres and is primarily mediated by the binding of CADM3, expressed in axons, to CADM4, expressed by myelinating Schwann cells. We have identified-by whole exome sequencing-three unrelated families, including one de novo patient, with axonal Charcot-Marie-Tooth disease (CMT2) sharing the same private variant in CADM3, Tyr172Cys. This variant is absent in 230 000 control chromosomes from gnomAD and predicted to be pathogenic. Most CADM3 patients share a similar phenotype consisting of autosomal dominant CMT2 with marked upper limb involvement. High resolution mass spectrometry analysis detected a newly created disulphide bond in the mutant CADM3 potentially modifying the native protein conformation. Our data support a retention of the mutant protein in the endoplasmic reticulum and reduced cell surface expression in vitro. Stochastic optical reconstruction microscopy imaging revealed decreased co-localization of the mutant with CADM4 at intercellular contact sites. Mice carrying the corresponding human mutation (Cadm3Y170C) showed reduced expression of the mutant protein in axons. Cadm3Y170C mice showed normal nerve conduction and myelin morphology, but exhibited abnormal axonal organization, including abnormal distribution of Kv1.2 channels and Caspr along myelinated axons. Our findings indicate the involvement of abnormal axon-glia interaction as a disease-causing mechanism in CMT patients with CADM3 mutations. [ABSTRACT FROM AUTHOR]

In myelinated peripheral axons, Kv1 potassium channels are clustered at the juxtaparanodal region and at an internodal line located along the mesaxon and below the Schmidt-Lanterman incisures. This polarized distribution is controlled by Schwann cells and requires specific cell adhesion molecules (CAMs). The accumulation of Kv1 channels at the juxtaparanodal region depends on the presence of Caspr2 at this site, as well as on the presence of Caspr at the adjacent paranodal junction. However, the localization of these channels along the mesaxonal internodal line still persists in the absence of each one of these CAMs. By generating mice lacking both Caspr and Caspr2 (caspr-/-/caspr2-/-), we now reveal compensatory functions of the two proteins in the organization of the axolemma. Although Kv1 channels are clustered along the inner mesaxon and in a circumferential ring below the incisures in the single mutants, in sciatic nerves of caspr-/-/caspr2-/- mice, these channels formed large aggregates that were dispersed along the axolemma, demonstrating that internodal localization of Kv1 channels requires either Caspr or Caspr2. Furthermore, deletion of both Caspr and Caspr2 also resulted in widening of the nodes of Ranvier, suggesting that Caspr2 (which is present at paranodes in the absence of Caspr) can partially compensate for the barrier function of Caspr at this site even without the formation of a distinct paranodal junction. Our results indicate that Caspr and Caspr2 are required for the organization of the axolemma both radially, manifested as the mesaxonal line, and longitudinally, demarcated by the nodal domains. [ABSTRACT FROM AUTHOR]

Clustered Kv1 K+ channels regulate neuronal excitability at juxtaparanodes of myelinated axons, axon initial segments, and cerebellar basket cell terminals (BCTs). These channels are part of a larger protein complex that includes cell adhesion molecules and scaffolding proteins. To identify proteins that regulate assembly, clustering, and/or maintenance of axonal Kv1 channel protein complexes, we immunoprecipitated Kv1.2α subunits, and then used mass spectrometry to identify interacting proteins.Wefound that a disintegrin and metalloproteinase 22 (ADAM22) is a component of the Kv1 channel complex and that ADAM22 coimmunoprecipitates Kv1.2 and the membrane-associated guanylate kinases (MAGUKs) PSD-93 and PSD-95. When coexpressed with MAGUKs in heterologous cells, ADAM22 and Kv1 channels are recruited into membrane surface clusters. However, coexpression of Kv1.2 with ADAM22 and MAGUKs does not alter channel properties. Among all the known Kv1 channel-interacting proteins, only ADAM22 is found at every site where Kv1 channels are clustered. Analysis of Caspr-null mice showed that, like other previously described juxtaparanodal proteins, disruption of the paranodal junction resulted in redistribution of ADAM22 into paranodal zones. Analysis of Caspr2-, PSD-93-, PSD-95-, and double PSD-93/PSD-95-null mice showed ADAM22 clustering at BCTs requires PSD-95, but ADAM22 clustering at juxtaparanodes requires neither PSD-93 nor PSD-95. In direct contrast, analysis of ADAM22-null mice demonstrated juxtaparanodal clustering of PSD-93 and PSD-95 requires ADAM22, whereas Kv1.2 and Caspr2 clustering is normal inADAM22-null mice. Thus,ADAM22is an axonal component of the Kv1 K+ channel complex that recruits MAGUKs to juxtaparanodes. [ABSTRACT FROM AUTHOR]

Clustering of Kv1 channels at the juxtaparanodal region (JXP) in myelinated axons depends on their association with the Caspr2/TAG-1 adhesion complex. The interaction between these channels and Caspr2 was suggested to depend on PDZ (PSD-95/Discs large/zona occludens-1) scaffolding proteins. Here, we show that at a subset of the JXP, PSD-93 colocalizes with Caspr2, K+ channels and its related protein postsynaptic density protein-95 (PSD-95). The localization of PSD-93 and PSD-95 depends on the presence of Caspr2, as both scaffolding proteins failed to accumulate at the JXP in mice lacking either Caspr2 or TAG-1. In contrast, Caspr2 and K+ channels still colocalized and associated in PSD-93, PSD-95 or double PSD-93/PSD-95 null mice. To directly evaluate the role of PDZ domain proteins in the function of Caspr2, we examined the ability of transgenic Caspr2 molecules lacking either their cytoplasmic domain (Caspr2dCT), or their PDZ-binding sequence (Caspr2dPDZ), to restore Kv1 channel clustering in Caspr2 null mice.Wefound that while Kv1 channels were distributed throughout internodes in nerves expressing Caspr2dCT, they were clustered at the JXP in axons expressing a full-length Caspr2 (Caspr2FL) or the Caspr2dPDZ transgene. Further proteomic analysis revealed that Caspr2 interacts with a distinct set of scaffolding proteins through its PDZ- and protein 4.1-binding sequences. These results demonstrate that while the molecular assembly of the JXP requires the cytoplasmic domain of Caspr2, its carboxy-terminal PDZ-binding motif is dispensable for Kv1 channel clustering. This mechanism is clearly distinct from the one operating at the axon initial segment, which requires PSD-93 for Kv1 channel clustering. [ABSTRACT FROM AUTHOR]

Myelination in the peripheral nervous system requires close contact between Schwann cells and the axon, but the underlying molecular basis remains largely unknown. Here we show that cell adhesion molecules (CAMs) of the nectin-like (Necl, also known as SynCAM or Cadm) family mediate Schwann cell–axon interaction during myelination. Necl4 is the main Necl expressed by myelinating Schwann cells and is located along the internodes in direct apposition to Necl1, which is localized on axons. Necl4 serves as the glial binding partner for axonal Necl1, and the interaction between these two CAMs mediates Schwann cell adhesion. The disruption of the interaction between Necl1 and Necl4 by their soluble extracellular domains, or the expression of a dominant-negative Necl4 in Schwann cells, inhibits myelination. These results suggest that Necl proteins are important for mediating axon-glia contact during myelination in peripheral nerves. [ABSTRACT FROM AUTHOR]

Synaptic scaffolding molecule (S-SCAM) is a synaptic protein, which harbors five or six PSD-95/Discs large/ZO-1 (PDZ), a guanylate kinase and two WW domains. It interacts with NMDA receptor subunits, neuroligin and β-catenin, and is involved in the accumulation of neuroligin at excitatory synapses. In this study, we have demonstrated S-SCAM is localized at inhibitory synapses in rat primary cultured hippocampal neurons. We have identified β-dystroglycan (β-DG) as a binding partner for S-SCAM at inhibitory synapses. WW domains of S-SCAM bind to three sequences of β-DG. We have also revealed that S-SCAM can interact with neuroligin 2, which is known to be exclusively localized at inhibitory synapses. The WW domains and the second PDZ domain of S-SCAM are involved in the interaction with neuroligin 2. β-DG, neuroligin 2 and S-SCAM form a tripartite complex in vitro. Neuroligin 2 is detected in the immunoprecipitates by anti-β-DG antibody from rat brain. S-SCAM, β-DG and neuroligin 2 are partially co-localized in rat hippocampal neurons. These data suggest that S-SCAM is associated with β-DG and neuroligin 2 at inhibitory synapses, and functions as a linker between the dystrophin glycoprotein complex and the neurexin–neuroligin complex. [ABSTRACT FROM AUTHOR]

We have examined the localization of contactin-associated protein (Caspr), the Shaker-type potassium channels, Kv1.1 and Kv1.2, their associated β subunit, Kvβ2, and Caspr2 in the myelinated fibers of the CNS. Caspr is localized to the paranodal axonal membrane, and Kv1.1, Kv1.2, Kvβ2 and Caspr2 to the juxtaparanodal membrane. In addition to the paranodal staining, an internodal strand of Caspr staining apposes the inner mesaxon of the myelin sheath. Unlike myelinated axons in the peripheral nervous system, there was no internodal strand of Kv1.1, Kv1.2, Kvβ2, or Caspr2. Thus, the organization of the nodal, paranodal, and juxtaparanodal axonal membrane is similar in the central and peripheral nervous systems, but the lack of Kv1.1/Kv1.2/Kvβ2/Caspr2 internodal strands indicates that the oligodendrocyte myelin sheaths lack a trans molecular interaction with axons, an interaction that is present in Schwann cell myelin sheaths. [ABSTRACT FROM AUTHOR]

Discusses the implications of the structural and functional similarity of receptor-like protein tyrosine phosphatase beta (RPTPbeta) to cell adhesion molecules (CAM). Facts about CAM; Similarities of RPTPbeta to CAM; Importance of RPTPbeta during development of the nervous system; Domains which made up the RPTPbeta molecule.

A membrane barrier important for assembly of the nodes of Ranvier is found at the paranodal junction. This junction is comprised of axonal and glial adhesion molecules linked to the axonal actin-spectrin membrane cytoskeleton through specific adaptors. In this issue, Zhang et al. (2013. J. Cell Biol. http://dx.doi.org/10.1083/jcb.201308116) show that axonal (311 spectrin maintains the diffusion barrier at the paranodal junction. Thus, 311 spectrin serves to compartmentalize the membrane of myelinated axons at specific locations that are determined either intrinsically (i.e., at the axonal initial segment), or by axoglial contacts (i.e., at the paranodal junction). [ABSTRACT FROM AUTHOR]

TDP-43 is extensively studied in neurons in physiological and pathological contexts. However, emerging evidence indicates that glial cells are also reliant on TDP-43 function. We demonstrate that deletion of TDP-43 in Schwann cells results in a dramatic delay in peripheral nerve conduction causing significant motor deficits in mice, which is directly attributed to the absence of paranodal axoglial junctions. By contrast, paranodes in the central nervous system are unaltered in oligodendrocytes lacking TDP-43. Mechanistically, TDP-43 binds directly to Neurofascin mRNA, encoding the cell adhesion molecule essential for paranode assembly and maintenance. Loss of TDP-43 triggers the retention of a previously unidentified cryptic exon, which targets Neurofascin mRNA for nonsense-mediated decay. Thus, TDP-43 is required for neurofascin expression, proper assembly and maintenance of paranodes, and rapid saltatory conduction. Our findings provide a framework and mechanism for how Schwann cell-autonomous dysfunction in nerve conduction is directly caused by TDP-43 loss-of-function. [ABSTRACT FROM AUTHOR]

Oligodendrocyte–axon contact is mediated by several cell adhesion molecules (CAMs) that are positioned at distinct sites along the myelin unit, yet their role during myelination remains unclear. Cadm4 and its axonal receptors, Cadm2 and Cadm3, as well as myelin-associated glycoprotein (MAG), are enriched at the internodes below the compact myelin, whereas NF155, which binds the axonal Caspr/contactin complex, is located at the paranodal junction that is formed between the axon and the terminal loops of the myelin sheath. Here we report that Cadm4-, MAG-, and Caspr-mediated adhesion cooperate during myelin membrane ensheathment. Genetic deletion of either Cadm4 and MAG or Cadm4 and Caspr resulted in the formation of multimyelinated axons due to overgrowth of the myelin away from the axon and the forming paranodal junction. Consequently, these mice displayed paranodal loops either above or underneath compact myelin. Our results demonstrate that accurate placement of the myelin sheath by oligodendrocytes requires the coordinated action of internodal and paranodal CAMs. [ABSTRACT FROM AUTHOR]

A high density of Na+ channels at nodes of Ranvier is necessary for rapid and efficient action potential propagation in myelinated axons. Na+ channel clustering is thought to depend on two axonal cell adhesion molecules that mediate interactions between the axon and myelinating glia at the nodal gap (i.e., NF186) and the paranodal junction (i.e., Caspr). Here we show that while Na+ channels cluster at nodes in the absence of NF186, they fail to do so in double conditional knockout mice lacking both NF186 and the paranodal cell adhesion molecule Caspr, demonstrating that a paranodal junction-dependent mechanism can cluster Na+ channels at nodes. Furthermore, we show that paranode-dependent clustering of nodal Na+ channels requires axonal bII spectrin which is concentrated at paranodes. Our results reveal that the paranodal junction-dependent mechanism of Na+channel clustering is mediated by the spectrin-based paranodal axonal cytoskeleton. [ABSTRACT FROM AUTHOR]

Summary Myelination occurs selectively around neuronal axons to increase the efficiency and velocity of action potentials. While oligodendrocytes are capable of myelinating permissive structures in the absence of molecular cues, structurally permissive neuronal somata and dendrites remain unmyelinated. Utilizing a purified spinal cord neuron-oligodendrocyte myelinating co-culture system, we demonstrate that disruption of dynamic neuron-oligodendrocyte signaling by chemical cross-linking results in aberrant myelination of the somatodendritic compartment of neurons. We hypothesize that an inhibitory somatodendritic cue is necessary to prevent non-axonal myelination. Using next-generation sequencing and candidate profiling, we identify neuronal junction adhesion molecule 2 (JAM2) as an inhibitory myelin-guidance molecule. Taken together, our results demonstrate that the somatodendritic compartment directly inhibits myelination and suggest a model in which broadly indiscriminate myelination is tailored by inhibitory signaling to meet local myelination requirements. [ABSTRACT FROM AUTHOR]

Fast neural conduction requires accumulation of Na+ channels at nodes of Ranvier. Dedicated adhesion molecules on myelinating cells and axons govern node organization. Among those, specific laminins and dystroglycan complexes contribute to Na+ channel clustering at peripheral nodes by unknown mechanisms. We show that in addition to facing the basal lamina, dystroglycan is found near the nodal matrix around axons, binds matrix components, and participates in initial events of nodogenesis. We identify the dystroglycan-ligand perlecan as a novel nodal component and show that dystroglycan is required for the selective accumulation of perlecan at nodes. Perlecan binds the clustering molecule gliomedin and enhances clustering of node of Ranvier components. These data show that proteoglycans have specific roles in peripheral nodes and indicate that peripheral and central axons use similar strategies but different molecules to form nodes of Ranvier. Further, our data indicate that dystroglycan binds free matrix that is not organized in a basal lamina. [ABSTRACT FROM AUTHOR]

Abstract: The neurexin superfamily is a group of transmembrane molecules mediating cell–cell contacts and generating specialized membranous domains in polarized epithelial and nerves cells. We describe here the domain organization and expression of the entire, core neurexin superfamily in the nematode Caenorhabditis elegans, which is composed of three family members. One of the superfamily members, nrx-1, is an ortholog of vertebrate neurexin, the other two, itx-1 and nlr-1, are orthologs of the Caspr subfamily of neurexin-like genes. Based on reporter gene analysis, we find that nrx-1 is exclusively expressed in most if not all cells of the nervous system and localizes to presynaptic specializations. itx-1 and nrx-1 reporter genes are expressed in non-overlapping patterns within and outside the nervous system. ITX-1 protein co-localizes with β-G-spectrin to a subapical domain within intestinal cells. These studies provide a starting point for further functional analysis of this family of proteins. [Copyright &y& Elsevier]

The interaction between gliomedin and the axonodal cell adhesion molecules (CAMs) neurofascin and NrCAM induces the clustering of Na+ channels at the nodes of Ranvier. We define new interactions of gliomedin that are essential for its clustering activity. We show that gliomedin exists as both transmembrane and secreted forms that are generated by proteolytic cleavage of the protein, and that only the latter is detected at the nodes of Ranvier. The secreted extracellular domain of gliomedin binds to Schwann cells and is incorporated into the extracellular matrix (ECM) in a heparin-dependent manner, suggesting the involvement of heparan sulfate proteoglycans (HSPGs). Furthermore, we show that the N-terminal region of gliomedin serves as an oligomerization domain that mediates self-association of the molecule, which is required for its binding to neurofascin and NrCAM. Our results indicate that the deposition of gliomedin multimers at the nodal gap by binding to HSPGs facilitates the clustering of the axonodal CAMs and Na+ channels. [ABSTRACT FROM AUTHOR]