title: Assembly of juxtaparanodes in myelinating DRG culture: Differential clustering of the Kv1/Caspr2 complex and scaffolding protein 4.1B.
abstract: The precise distribution of ion channels at the nodes of Ranvier is essential for the efficient propagation of action potentials along myelinated axons. The voltage-gated potassium channels Kv1.1/1.2 are clustered at the juxtaparanodes in association with the cell adhesion molecules, Caspr2 and TAG-1 and the scaffolding protein 4.1B. In the present study, we set up myelinating cultures of DRG neurons and Schwann cells to look through the formation of juxtaparanodes in vitro. We showed that the Kv1.1/Kv1.2 channels were first enriched at paranodes before being restricted to distal paranodes and juxtaparanodes. In addition, the Kv1 channels displayed an asymmetric expression enriched at the distal juxtaparanodes. Caspr2 was strongly co-localized with Kv1.2 whereas the scaffolding protein 4.1B was preferentially recruited at paranodes while being present at juxtaparanodes too. Kv1.2/Caspr2 but not 4.1B, also transiently accumulated within the nodal region both in myelinated cultures and developing sciatic nerves. Studying cultures and sciatic nerves from 4.1B KO mice, we further showed that 4.1B is required for the proper targeting of Caspr2 early during myelination. Moreover, using adenoviral-mediated expression of Caspr-GFP and photobleaching experiments, we analyzed the stability of paranodal junctions and showed that the lateral stability of paranodal Caspr was not altered in 4.1B KO mice indicating that 4.1B is not required for the assembly and stability of the paranodal junctions. Thus, developing an adapted culture paradigm, we provide new insights into the dynamic and differential distribution of Kv1 channels and associated proteins during myelination.
authors: Aix-Marseille Université, CNRS, Centre de Recherche en Neurobiologie et Neurophysiologie de Marseille-UMR7286, Marseille, France.
authors: Bruno
authors: B
authors: Hivert
authors: Aix-Marseille Université, CNRS, Centre de Recherche en Neurobiologie et Neurophysiologie de Marseille-UMR7286, Marseille, France.
authors: Delphine
authors: D
authors: Pinatel
authors: Aix-Marseille Université, CNRS, Centre de Recherche en Neurobiologie et Neurophysiologie de Marseille-UMR7286, Marseille, France.
authors: Marilyne
authors: M
authors: Labasque
authors: INSERM U1051 Institut des Neurosciences de Montpellier, Montpellier, France.
authors: Nicolas
authors: N
authors: Tricaud
authors: INSERM UMRS 839 Institut du Fer à Moulin, Paris.
authors: Laurence
authors: L
authors: Goutebroze
authors: Aix-Marseille Université, CNRS, Centre de Recherche en Neurobiologie et Neurophysiologie de Marseille-UMR7286, Marseille, France.
authors: Catherine
authors: C
authors: Faivre-Sarrailh
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D058969:Contactin 2 / Q000378:metabolism; D004546:Elapid Venoms / Q000493:pharmacokinetics; D004622:Embryo, Mammalian; D036681:Fluorescence Recovery After Photobleaching; D005727:Ganglia, Spinal / Q000166:cytology*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D051662:Kv1.1 Potassium Channel / Q000235:genetics / Q000378:metabolism*; D051663:Kv1.2 Potassium Channel / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D008840:Microfilament Proteins / Q000235:genetics / Q000378:metabolism*; D008954:Models, Biological; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D011901:Ranvier's Nodes / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar; D012583:Schwann Cells / Q000378:metabolism*
chemical_list: C568163:CNTNAP2 protein, mouse; C548675:Cntn2 protein, mouse; D058969:Contactin 2; D004546:Elapid Venoms; C488402:Epb41l3 protein, mouse; D051663:Kv1.2 Potassium Channel; D008565:Membrane Proteins; D008840:Microfilament Proteins; D009419:Nerve Tissue Proteins; D051662:Kv1.1 Potassium Channel; D049452:Green Fluorescent Proteins; C020273:dendrotoxin
keywords: Caspr; FRAP; Schwann cells; cell adhesion; node of Ranvier; potassium channels