NFASC

Abstract: Basket axon collaterals synapse onto the Purkinje soma/axon initial segment (AIS) area to form specialized structures, the pinceau, which are critical for normal cerebellar function. Mechanistic details of how the pinceau become organized during cerebellar development are poorly understood. Loss of cytoskeletal adaptor protein Ankyrin G (AnkG) results in mislocalization of the cell adhesion molecule Neurofascin (Nfasc) at the Purkinje AIS and abnormal organization of the pinceau. Loss of Nfasc in adult Purkinje neurons leads to slow disorganization of the Purkinje AIS and pinceau morphology. Here, we used mouse conditional knock-out techniques to show that selective loss of Nfasc, specifically in Purkinje neurons during early development, prevented maturation of the AIS and resulted in loss of Purkinje neuron spontaneous activity and pinceau disorganization. Loss of Nfasc in both Purkinje and basket neurons caused abnormal basket axon collateral branching and targeting to Purkinje soma/AIS, leading to extensive pinceau disorganization, Purkinje neuron degeneration, and severe ataxia. Our studies reveal that the Purkinje Nfasc is required for AIS maturation and for maintaining stable contacts between basket axon terminals and the Purkinje AIS during pinceau organization, while the basket neuron Nfasc in combination with Purkinje Nfasc is required for proper basket axon collateral outgrowth and targeting to Purkinje soma/AIS. Thus, cerebellar pinceau organization requires coordinated mechanisms involving specific Nfasc functions in both Purkinje and basket neurons.

Abstract: The formation of paranodal axo-glial junctions is critical for the rapid and efficient propagation of nerve impulses. Genetic ablation of genes encoding the critical paranodal proteins Caspr, contactin (Cont), and the myelinating glia-specific isoform of Neurofascin (Nfasc(NF155)) results in the disruption of the paranodal axo-glial junctions, loss of ion channel segregation, and impaired nerve conduction, but the mechanisms regulating their interactions remain elusive. Here, we report that loss of immunoglobulin (Ig) domains 5 and 6 in Nfasc(NF155) in mice phenocopies complete ablation of Nfasc(NF155). The mutant mice lack paranodal septate junctions, resulting in the diffusion of Caspr and Cont from the paranodes, and redistribution of the juxtaparanodal potassium channels toward the nodes. Although critical for Nfasc(NF155) function, we find that Ig5-6 are dispensable for nodal Nfasc(NF186) function. Moreover, in vitro binding assays using Ig5-6 deletion constructs reveal their importance for the association of Nfasc(NF155) with Cont. These findings provide the first molecular evidence demonstrating domain-specific requirements controlling the association of the paranodal tripartite complex in vivo. Our studies further emphasize that in vivo structure/function analysis is necessary to define the unique protein-protein interactions that differentially regulate the functions of Neurofascins during axonal domain organization.