title: Phosphorylation by extracellular signal-regulated kinase of a multidomain adaptor protein, vinexin, at synapses.
abstract: Vinexin is an adaptor protein that is supposed to play pivotal roles in cell adhesion, cytoskeletal organization and signaling. At least three splice variants, vinexinalpha, beta and gamma, have so far been reported. In spite of the possible importance of vinexin, the properties and functions of vinexin in neuronal cells are almost unknown. Here we show that vinexin isoforms are expressed in rat brain in a developmental stage-dependent manner, and that vinexinalpha is relatively abundant in the telencephalon regions of the adult rat brain. An immunohistochemical study showed the localization of vinexinalpha in neurons and glia in the rat brain. In primary cultured rat hippocampal neurons, vinexin was found to be present at synapses and filopodia in growth cones by immunofluorescent analyses. Biochemical fractionation revealed the distribution of vinexin in synaptosomes. Nerve terminal localization of vinexin was confirmed by electron microscopy. Vinexinbeta is reported to be phosphorylated by extracellular signal-regulated kinase (ERK) at Ser189, which is equivalent to Ser593 of vinexinalpha. We thus constructed a site- and phosphorylation state-specific antibody to monitor the ERK-mediated phosphorylation of vinexin. In immunofluorescent analyses, the phosphorylation was observed at synapses formed among cultured rat hippocampal neurons and it was reduced by treatment of the cells with PD98059. In an immunoelectron microscopic examination, the phosphorylation signal was mainly detected on the postsynaptic side of synapses in the rat hippocampal neurons. As active ERK was co-localized with vinexin in synapses, the ERK signal is likely to be involved in the regulation of vinexin-dependent cellular processes in synapses. On the other hand, the phosphorylation was hardly detected in neurons cultured for 3 days, suggesting the presence of a yet unidentified regulatory mechanism of vinexin at the growth cone.
authors.affiliation: Department of Molecular Neurobiology, Institute for Developmental Research, Aichi Human Service Center, Kasugai, japan.
authors.forename: Hidenori
authors.initials: H
authors.lastname: Ito
authors.forename: Nobuteru
authors.initials: N
authors.lastname: Usuda
authors.forename: Kimie
authors.initials: K
authors.lastname: Atsuzawa
authors.forename: Ikuko
authors.initials: I
authors.lastname: Iwamoto
authors.forename: Kaori
authors.initials: K
authors.lastname: Sudo
authors.forename: Ritsuko
authors.initials: R
authors.lastname: Katoh-Semba
authors.forename: Kosuke
authors.initials: K
authors.lastname: Mizutani
authors.forename: Rika
authors.initials: R
authors.lastname: Morishita
authors.forename: Takashi
authors.initials: T
authors.lastname: Deguchi
authors.forename: Yoshinori
authors.initials: Y
authors.lastname: Nozawa
authors.forename: Tomiko
authors.initials: T
authors.lastname: Asano
authors.forename: Koh-Ichi
authors.initials: K
authors.lastname: Nagata
mesh_terms: D000199:Actins / Q000378:metabolism; D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D000818:Animals; D000911:Antibodies, Monoclonal / Q000378:metabolism; D001921:Brain / Q000166:cytology; D002478:Cells, Cultured; D002522:Chlorocebus aethiops; D004791:Enzyme Inhibitors / Q000494:pharmacology; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism*; D005260:Female; D005419:Flavonoids / Q000494:pharmacology; D008297:Male; D016253:Microscopy, Immunoelectron / Q000379:methods; D016296:Mutagenesis / Q000502:physiology; D009474:Neurons / Q000378:metabolism / Q000648:ultrastructure; D010766:Phosphorylation / Q000187:drug effects; D017434:Protein Structure, Tertiary; D051381:Rats; D017207:Rats, Sprague-Dawley; D012694:Serine / Q000378:metabolism; D013569:Synapses / Q000378:metabolism* / Q000648:ultrastructure; D016708:Synaptophysin / Q000378:metabolism; D014162:Transfection / Q000379:methods
chemical_list: D000199:Actins; D048868:Adaptor Proteins, Signal Transducing; D000911:Antibodies, Monoclonal; D004791:Enzyme Inhibitors; D005419:Flavonoids; C512725:Sorbs3 protein, rat; D016708:Synaptophysin; C085348:tau-1 monoclonal antibody; D012694:Serine; D048049:Extracellular Signal-Regulated MAP Kinases; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one