Steven Tait

Neurofascins Are Required to Establish Axonal Domains for Saltatory Conduction

Voltage-gated sodium channels are concentrated in myelinated nerves at the nodes of Ranvier flanked by paranodal axoglial junctions. Establishment of these essential nodal and paranodal domains is determined by myelin-forming glia, but the mechanisms are not clear. Here, we show that two isoforms of Neurofascin, Nfasc155 in glia and Nfasc186 in neurons, are required for the assembly of these specialized domains. In Neurofascin-null mice, neither paranodal adhesion junctions nor nodal complexes are formed. Transgenic expression of Nfasc155 in the myelinating glia of Nfasc-/- nerves rescues the axoglial adhesion complex by recruiting the axonal proteins Caspr and Contactin to the paranodes. However, in the absence of Nfasc186, sodium channels remain diffusely distributed along the axon. Our study shows that the two major Neurofascins play essential roles in assembling the nodal and paranodal domains of myelinated axons; therefore, they are essential for the transition to saltatory conduction in developing vertebrate nerves.

Neurofascin Is a Glial Receptor for the Paranodin/Caspr-Contactin Axonal Complex at the Axoglial Junction

In myelinated fibers of the vertebrate nervous system, glial-ensheathing cells interact with axons at specialized adhesive junctions, the paranodal septate-like junctions. The axonal proteins paranodin/Caspr and contactin form a cis complex in the axolemma at the axoglial adhesion zone, and both are required to stabilize the junction. There has been intense speculation that an oligodendroglial isoform of the cell adhesion molecule neurofascin, NF155, expressed at the paranodal loop might be the glial receptor for the paranodin/Caspr-contactin complex, particularly since paranodin/Caspr and NF155 colocalize to ectopic sites in the CNS of the dysmyelinated mouse Shiverer mutant. We report that the extracellular domain of NF155 binds specifically to transfected cells expressing the paranodin/Caspr-contactin complex at the cell surface. This region of NF155 also binds the paranodin/Caspr-contactin complex from brain lysates in vitro. In support of the functional significance of this interaction, NF155 antibodies and the extracellular domain of NF155 inhibit myelination in myelinating cocultures, presumably by blocking the adhesive relationship between the axon and glial cell. These results demonstrate that the paranodin/Caspr-contactin complex interacts biochemically with NF155 and that this interaction is likely to be biologically relevant at the axoglial junction.

Peripheral demyelination and neuropathic pain behavior in periaxin-deficient mice

The Prx gene in Schwann cells encodes L- and S-periaxin, two abundant PDZ domain proteins thought to have a role in the stabilization of myelin in the peripheral nervous system (PNS). Mice lacking a functional Prx gene assemble compact PNS myelin. However, the sheath is unstable, leading to demyelination and reflex behaviors that are associated with the painful conditions caused by peripheral nerve damage. Older Prx-/- animals display extensive peripheral demyelination and a severe clinical phenotype with mechanical allodynia and thermal hyperalgesia, which can be reversed by intrathecal administration of a selective NMDA receptor antagonist We conclude that the periaxins play an essential role in stabilizing the Schwann cell-axon unit and that the periaxin-deficient mouse will be an important model for studying neuropathic pain in late onset demyelinating disease.

Glial and neuronal isoforms of Neurofascin have distinct roles in the assembly of nodes of Ranvier in the central nervous system

Rapid nerve impulse conduction in myelinated axons requires the concentration of voltage-gated sodium channels at nodes of Ranvier. Myelin-forming oligodendrocytes in the central nervous system (CNS) induce the clustering of sodium channels into nodal complexes flanked by paranodal axoglial junctions. However, the molecular mechanisms for nodal complex assembly in the CNS are unknown. Two isoforms of Neurofascin, neuronal Nfasc186 and glial Nfasc155, are components of the nodal and paranodal complexes, respectively. Neurofascin-null mice have disrupted nodal and paranodal complexes. We show that transgenic Nfasc186 can rescue the nodal complex when expressed in Nfasc−/− mice in the absence of the Nfasc155–Caspr–Contactin adhesion complex. Reconstitution of the axoglial adhesion complex by expressing transgenic Nfasc155 in oligodendrocytes also rescues the nodal complex independently of Nfasc186. Furthermore, the Nfasc155 adhesion complex has an additional function in promoting the migration of myelinating processes along CNS axons. We propose that glial and neuronal Neurofascins have distinct functions in the assembly of the CNS node of Ranvier.

A Critical Role for Neurofascin in Regulating Action Potential Initiation through Maintenance of the Axon Initial Segment

Summary The axon initial segment (AIS) is critical for the initiation and propagation of action potentials. Assembly of the AIS requires interactions between scaffolding molecules and voltage-gated sodium channels, but the molecular mechanisms that stabilize the AIS are poorly understood. The neuronal isoform of Neurofascin, Nfasc186, clusters voltage-gated sodium channels at nodes of Ranvier in myelinated nerves: here, we investigate its role in AIS assembly and stabilization. Inactivation of the Nfasc gene in cerebellar Purkinje cells of adult mice causes rapid loss of Nfasc186 from the AIS but not from nodes of Ranvier. This causes AIS disintegration, impairment of motor learning and the abolition of the spontaneous tonic discharge typical of Purkinje cells. Nevertheless, action potentials with a modified waveform can still be evoked and basic motor abilities remain intact. We propose that Nfasc186 optimizes communication between mature neurons by anchoring the key elements of the adult AIS complex.

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.

A novel rat tetraspan protein in cells of the oligodendrocyte lineage

Abstract : The tetraspanin/transmembrane 4 superfamily gene superfamily encodes proteins that span the plasma membrane four times. Tetraspan proteins are implicated in proliferation, motility, and differentiation in various cell types, and in some cells they may link plasma membrane proteins into signalling complexes. Using a subtractive cDNA library prepared from oligodendrocytes and their progenitor cells, we have identified Tspan‐2 as a member of this superfamily. In situ hybridization analysis revealed robust expression in cells of the oligodendrocyte lineage in comparison with the Plp gene, a well‐characterized marker for myelin‐forming glia in the CNS. Rat Tspan‐2 mRNA is restricted to the nervous system and is detectable by northern blot shortly after birth in the CNS. Subsequently the gene is up‐regulated strongly between postnatal day 3 and 10, and expression levels continue to rise up to postnatal day 22. These data indicate that Tspan‐2 is likely to play a role in signalling in oligodendrocytes in the early stages of their terminal differentiation into myelin‐forming glia and may also function in stabilizing the mature sheath.

A functional FERM domain binding motif in neurofascin

The L1 family of transmembrane cell adhesion receptors are involved in the development of the nervous system and consist of L1, neuron-glial-related cell adhesion molecule and neurofascin. All three receptors have a short cytoplasmic tail which is known to bind to the cytoskeletal associated protein ankyrin. Ezrin is a cytoplasmic binding protein known to link plasma membrane proteins to the cytoskeleton and has been shown to be a binding partner for L1. Here we show that neurofascin can also interact directly with ezrin. However, the mechanism of interaction of L1 and neurofascin with ezrin is by different mechanisms. We also show that the neurofascin isoform, Nfasc155, co-localizes with ezrin in transfected HEK293 cells but also in interdigitating Schwann cells at the node of Ranvier.

Etr-r3/mNapor, encoding an ELAV-type RNA binding protein, is expressed in differentiating cells in the developing rodent forebrain

The gene encoding human neuroblastoma apoptosis-related RNA binding protein (NAPOR) has rat and mouse homologues, Etr-r3 and mNapor, cloned recently by ourselves and others. Etr-r3/mNapor is transcribed in embryonic brain but its detailed expression patterns are unknown. It has been hypothesised that its expression coincides with the occurrence of programmed cell death, but existing expression data are insufficient to evaluate this possibility. We used northern blots and in situ hybridisation to show that Etr-r3/mNapor is expressed by most postmitotic differentiating cells throughout the forebrain during embryogenesis and the first weeks of postnatal life. Expression is particularly high in the developing cerebral cortex, in nuclei of the ventral telencephalon, in dorsal thalamus and in hypothalamus. In contrast, Etr-r3/mNapor is expressed by very few cells in proliferative regions, where cell death rates are highest.