Cell Adhesion Molecule NrCAM Research Articles

Loss of Function of the Neural Cell Adhesion Molecule NrCAM Regulates Differentiation, Proliferation and Neurogenesis in Early Postnatal Hypothalamic Tanycytes.

Hypothalamic tanycytes are neural stem and progenitor cells, but little is known of how they are regulated. Here we provide evidence that the cell adhesion molecule, NrCAM, regulates tanycytes in the adult niche. NrCAM is strongly expressed in adult mouse tanycytes. Immunohistochemical and in situ hybridization analysis revealed that NrCAM loss of function leads to both a reduced number of tanycytes and reduced expression of tanycyte-specific cell markers, along with a small reduction in tyrosine hydroxylase-positive arcuate neurons. Similar analyses of NrCAM mutants at E16 identify few changes in gene expression or cell composition, indicating that NrCAM regulates tanycytes, rather than early embryonic hypothalamic development. Neurosphere and organotypic assays support the idea that NrCAM governs cellular homeostasis. Single-cell RNA sequencing (scRNA-Seq) shows that tanycyte-specific genes, including a number that are implicated in thyroid hormone metabolism, show reduced expression in the mutant mouse. However, the mild tanycyte depletion and loss of markers observed in NrCAM-deficient mice were associated with only a subtle metabolic phenotype.

Bi-allelic variants in neuronal cell adhesion molecule cause a neurodevelopmental disorder characterized by developmental delay, hypotonia, neuropathy/spasticity

Cell adhesion molecules are membrane-bound proteins predominantly expressed in the central nervous system along principal axonal pathways with key roles in nervous system development, neural cell differentiation and migration, axonal growth and guidance, myelination, and synapse formation. Here, we describe ten affected individuals with bi-allelic variants in the neuronal cell adhesion molecule NRCAM that lead to a neurodevelopmental syndrome of varying severity; the individuals are from eight families. This syndrome is characterized by developmental delay/intellectual disability, hypotonia, peripheral neuropathy, and/or spasticity. Computational analyses of NRCAM variants, many of which cluster in the third fibronectin type III (Fn-III) domain, strongly suggest a deleterious effect on NRCAM structure and function, including possible disruption of its interactions with other proteins. These findings are corroborated by previous invitro studies of murine Nrcam-deficient cells, revealing abnormal neurite outgrowth, synaptogenesis, and formation of nodes of Ranvier on myelinated axons. Our studies on zebrafish nrcamaΔ mutants lacking the third Fn-III domain revealed that mutant larvae displayed significantly altered swimming behavior compared to wild-type larvae (p < 0.03). Moreover, nrcamaΔ mutants displayed a trend toward increased amounts of α-tubulin fibers in the dorsal telencephalon, demonstrating an alteration in white matter tracts and projections. Taken together, our study provides evidence that NRCAM disruption causes a variable form of a neurodevelopmental disorder and broadens the knowledge on the growing role of the cell adhesion molecule family in the nervous system.

Alternative RNA splicing associated with axon regeneration after rat peripheral nerve injury

The intrinsic axon regeneration capacity is crucial for peripheral nerve regeneration after injury. Identifying key molecules involved in this process makes great contribution to the investigation of peripheral nerve injury repair. Alternative splicing (AS) is an important regulation mode of eukaryotic gene expression, which has been widely studied both in physiological and pathological processes. However, less is known about the role of AS in peripheral nerve regeneration. In this work, to identify the AS events associated with axon regeneration capacity, we analyzed the AS events during sciatic nerve injury repair by RNA sequencing (RNA-Seq) and replicate multivariate analysis of transcript splicing (rMATS). The differential AS events were underwent gene ontology enrichment and pathway analyses. Moreover, we identified a significantly increased AS event of neuronal cell adhesion molecule Nrcam (Nrcam-S), and demonstrated down-regulation of Nrcam-S by specific siRNAs inhibited axon regeneration of Dorsal Root Ganglion (DRG) neurons after sciatic nerve injury in vitro and in vivo. Additionally, we found expression levels of RNA binding proteins (RBPs) CUGBP Elav-like family member 3 (CELF3) and RNA binding protein fox-1 homolog 2 (Rbfox2) were markedly increased after sciatic nerve injury. Our data may serve as a resource useful for further understanding how AS contributes to molecular regulations in DRG during sciatic nerve regeneration.

Temporal Regulation of Dendritic Spines Through NrCAM-Semaphorin3F Receptor Signaling in Developing Cortical Pyramidal Neurons.

Neuron-glial related cell adhesion molecule NrCAM is a newly identified negative regulator of spine density that genetically interacts with Semaphorin3F (Sema3F), and is implicated in autism spectrum disorders (ASD). To investigate a role for NrCAM in spine pruning during the critical adolescent period when networks are established, we generated novel conditional, inducible NrCAM mutant mice (Nex1Cre-ERT2: NrCAMflox/flox). We demonstrate that NrCAM functions cell autonomously during adolescence in pyramidal neurons to restrict spine density in the visual (V1) and medial frontal cortex (MFC). Guided by molecular modeling, we found that NrCAM promoted clustering of the Sema3F holoreceptor complex by interfacing with Neuropilin-2 (Npn2) and PDZ scaffold protein SAP102. NrCAM-induced receptor clustering stimulated the Rap-GAP activity of PlexinA3 (PlexA3) within the holoreceptor complex, which in turn, inhibited Rap1-GTPase and inactivated adhesive β1 integrins, essential for Sema3F-induced spine pruning. These results define a developmental function for NrCAM in Sema3F receptor signaling that limits dendritic spine density on cortical pyramidal neurons during adolescence.

Severity of Demyelinating and Axonal Neuropathy Mouse Models Is Modified by Genes Affecting Structure and Function of Peripheral Nodes

Charcot-Marie-Tooth (CMT) disease is a clinically and genetically heterogeneous group of inherited polyneuropathies. Mutations in 80 genetic loci can cause forms of CMT, resulting in demyelination andaxonal dysfunction. The clinical presentation, including sensory deficits, distal muscle weakness, and atrophy, can vary greatly in severity and progression. Here, we used mouse models of CMT to demonstrate genetic interactions that result in a more severe neuropathy phenotype. The cell adhesion molecule Nrcam and the Na+ channel Scn8a (NaV1.6) are important components of nodes. Homozygous Nrcam and heterozygous Scn8a mutations synergized with both an Sh3tc2 mutation, modeling recessive demyelinating Charcot-Marie-Tooth type 4C, and mutations in Gars, modeling dominant axonal Charcot-Marie-Tooth type 2D. We conclude that genetic variants perturbing the structure and function of nodes interact with mutations affecting the cable properties of axons by thinning myelin or reducing axon diameter. Therefore, genes integral to peripheral nodes are candidate modifiers of peripheral neuropathy.

ConcurrentLpin1andNrcamMouse Mutations Result in Severe Peripheral Neuropathy with Transitory Hindlimb Paralysis

Peripheral neuropathy is a broad category of disorders with a diverse etiology, grouped together by their common pathogenic effect on the peripheral nervous system (PNS). Because of the heterogeneity observed to be responsible for these disorders, a forward genetics method of gene discovery was used to identify additional affected pathways. In this report, we describe the mutant mouse line 20884, generated by N-ethyl-N-nitrosourea mutagenesis, which is characterized by adult-onset transitory hindlimb paralysis. Linkage mapping revealed that two point mutations are responsible for the phenotype: a partial loss-of-function mutation in the gene for phosphatidate phosphatase Lpin1 and a truncation mutation in the gene that encodes the neuronal cell adhesion molecule NrCAM. To investigate how the 20884 Lpin1 and Nrcam mutations interact to produce the paralysis phenotype, the double mutant and both single mutants were analyzed by quantitative behavioral, histological, and electrophysiological means. The Lpin1(20884) mutant and the double mutant are characterized by similar levels of demyelination and aberrant myelin structures. Nevertheless, the double mutant exhibits more severe electrophysiological abnormalities than the Lpin1(20884) mutant. The Nrcam(20884) mutant is characterized by normal sciatic nerve morphology and a mild electrophysiological defect. Comparison of the double mutant phenotype with the two single mutants does not point to an additive relationship between the two defects; rather, the Lpin1(20884) and Nrcam(20884) defects appear to act synergistically to produce the 20884 phenotype. It is proposed that the absence of NrCAM in a demyelinating environment has a deleterious effect, possibly by impairing the process of remyelination.

Association of PTPRB gene polymorphism with drug addiction

Several lines of evidence support the involvement of protein tyrosine phosphatase receptor type beta (PTPRB) in addiction. Generally, PTPs interact with both neuronal receptors and cell adhesion molecules, and appear to play roles in neurite growth and neuronal differentiation. We previously identified a role of the cell adhesion molecule NrCAM in polysubstance abuse vulnerability in humans, as well as in the rewarding effects of abused drugs in animals. Furthermore, we have identified genomic regions containing several cell adhesion molecules as polysubstance abuse vulnerability loci by whole-genome association study. The present study of human chromosome 12 loci revealed that the Ser127Gly polymorphism in PTPRB is associated with substance abuse vulnerability in three independent case-control samples (European-American from COGA families, USA, n = 177, P = 0.047; European-American from Maryland, USA, n = 650, P = 0.018; and African-American from Maryland, USA, n = 331, P = 0.009). However, this polymorphism was not associated with alcoholism in Japanese subjects (n = 1,599, P = 0.37). To confirm the importance of PTPRB in responses to drugs of abuse the expression of Ptprb in mouse brain was examined after chronic morphine treatment and found to be up-regulated in some brain regions. Thus, PTPRB is an addiction-associated and drug-regulated gene whose variants may affect substance abuse vulnerability.

Under the Eye of Nr-CAM

Binocular vision relies upon the existence of contralateral and ispilateral projections from retinal ganglion cells. Contacts between visual axons and optic chiasm cells are critical for the sorting of crossed and uncrossed projections during development. In this issue of Neuron, a study by Williams et al. shows that the cell adhesion molecule Nr-CAM facilitates/promotes the decussation of contralateral axons across the chiasm.

A Role for Nr-CAM in the Patterning of Binocular Visual Pathways

Retinal ganglion cell (RGC) axons diverge within the optic chiasm to project to opposite sides of the brain. In mouse, contralateral RGCs are distributed throughout the retina, whereas ipsilateral RGCs are restricted to the ventrotemporal crescent (VTC). While repulsive guidance mechanisms play a major role in the formation of the ipsilateral projection, little is known about the contribution of growth-promoting interactions to the formation of binocular visual projections. Here, we show that the cell adhesion molecule Nr-CAM is expressed by RGCs that project contralaterally and is critical for the guidance of late-born RGCs within the VTC. Blocking Nr-CAM function causes an increase in the size of the ipsilateral projection and reduces neurite outgrowth on chiasm cells in an age- and region-specific manner. Finally, we demonstrate that EphB1/ephrin-B2-mediated repulsion and Nr-CAM-mediated attraction comprise distinct molecular programs that each contributes to the proper formation of binocular visual pathways.

NrCAM in Addiction Vulnerability: Positional Cloning, Drug-Regulation, Haplotype-Specific Expression, and Altered Drug Reward in Knockout Mice

Several lines of evidence support roles for the cell adhesion molecule NrCAM in addictions. Fine mapping within a chromosome 7 region that contains previously linked and associated genomic markers identifies NrCAM haplotypes that are associated with substance abuse vulnerabilities in four samples of abusers and controls. Differential display identifies NrCAM as a drug regulated gene. NrCAM is expressed in neurons linked to reward and memory. NrCAM displays haplotype-specific gene expression in human post-mortem brain samples. Knockout mice display reduced opiate- and stimulant-conditioned place preferences. These observations support NrCAM as a positionally cloned and drug-regulated gene whose variants are likely to change expression and alter substance abuse vulnerabilities in human addictions and animal models of drug reward.

The cell adhesion molecule NrCAM is crucial for growth cone behaviour and pathfinding of retinal ganglion cell axons

We investigated the role of the cell adhesion molecule NrCAM for axonal growth and pathfinding in the developing retina. Analysis of the distribution pattern of NrCAM in chick embryo retina sections and flat-mounts shows its presence during extension of retinal ganglion cell (RGC) axons; NrCAM is selectively present on RGC axons and is absent from the soma. Single cell cultures show an enrichment of NrCAM in the distal axon and growth cone. When offered as a substrate in addition to Laminin, NrCAM promotes RGC axon extension and the formation of growth cone protrusions. In substrate stripe assays, mimicking the NrCAM-displaying optic fibre layer and the Laminin-rich basal lamina, RGC axons preferentially grow on NrCAM lanes. The three-dimensional analysis of RGC growth cones in retina flat-mounts reveals that they are enlarged and form more protrusions extending away from the correct pathway under conditions of NrCAM-inhibition. Time-lapse analyses show that these growth cones pause longer to explore their environment, proceed for shorter time spans, and retract more often than under control conditions; in addition, they often deviate from the correct pathway towards the optic fissure. Inhibition of NrCAM in organ-cultured intact eyes causes RGC axons to misroute at the optic fissure; instead of diving into the optic nerve head, these axons cross onto the opposite side of the retina. Our results demonstrate a crucial role for NrCAM in the navigation of RGC axons in the developing retina towards the optic fissure, and also for pathfinding into the optic nerve.

Correction: Overlapping functions of the cell adhesion molecules Nr-CAM and L1 in cerebellar granule cell development

Correction: Overlapping functions of the cell adhesion molecules Nr-CAM and L1 in cerebellar granule cell development

Overlapping functions of the cell adhesion molecules Nr-CAM and L1 in cerebellar granule cell development.

The structurally related cell adhesion molecules L1 and Nr-CAM have overlapping expression patterns in cerebellar granule cells. Here we analyzed their involvement in granule cell development using mutant mice. Nr-CAM–deficient cerebellar granule cells failed to extend neurites in vitro on contactin, a known ligand for Nr-CAM expressed in the cerebellum, confirming that these mice are functionally null for Nr-CAM. In vivo, Nr-CAM–null cerebella did not exhibit obvious histological defects, although a mild size reduction of several lobes was observed, most notably lobes IV and V in the vermis. Mice deficient for both L1 and Nr-CAM exhibited severe cerebellar folial defects and a reduction in the thickness of the inner granule cell layer. Additionally, anti-L1 antibodies specifically disrupted survival and maintenance of Nr-CAM–deficient granule cells in cerebellar cultures treated with antibodies. The combined results indicate that Nr-CAM and L1 play a role in cerebellar granule cell development, and suggest that closely related molecules in the L1 family have overlapping functions.

Differential expression of the cell-adhesion molecule Nr-CAM in hyperplastic and neoplastic human pancreatic tissue

Nr-CAM is a member of the immunoglobulin superfamily of neural cell-adhesion molecules initially thought to be expressed mainly in the brain. Here we show the presence of Nr-CAM protein in normal human pancreas and characterize its expression in hyperplastic and neoplastic human pancreatic tissue. Nr-CAM is expressed on the cell surface in normal pancreatic acini with enhanced staining at cell—cell junctions, and weak or no surface staining is seen on normal ductal cells. Nr-CAM expression is markedly up-regulated in intraductal hyperplasia. Expression was well maintained in well or moderately differentiated carcinoma but was reduced or absent from most poorly differentiated tumors. In addition, 4 of 4 human pancreatic adenocarcinoma cell lines tested demonstrated little or no Nr-CAM expression. This differential regulation of Nr-CAM expression suggests that it may be involved in the pathogenesis and invasive/metastatic behavior of pancreatic cancers. HUM PATHOL 32:396-400. Copyright © 2001 by W.B. Saunders Company

Alternative Splicing of HumanNrCAMin Neural and Nonneural Tissues

Neural cell adhesion molecule NrCAM exists in a variety of isoforms as a result of alternative splicing of individual exons during RNA processing. In this report we demonstrate that many of the alternative splicing events described for chick are conserved in man and describe a novel variant of NrCAM cDNA. Furthermore, we show that NrCAM is expressed at significant levels outside the nervous system; in particular in pancreas, adrenal glands, and placenta and that expression in both brain and other tissues is accompanied by a very variable pattern of exon utilization in fetal and adult cells.