Phosphorylation by extracellular signal-regulated kinase of a multidomain adaptor protein, vinexin, at synapses

• Published in: Journal of neurochemistry Vol. 100; no. 2; pp. 545 - 554
• Main Authors: Ito, Hidenori, Usuda, Nobuteru, Atsuzawa, Kimie, Iwamoto, Ikuko, Sudo, Kaori, Katoh-Semba, Ritsuko, Mizutani, Kosuke, Morishita, Rika, Deguchi, Takashi, Nozawa, Yoshinori, Asano, Tomiko, Nagata, Koh-ichi
• Format: Journal Article
• Language: English
• Published: Oxford, UK Oxford, UK : Blackwell Publishing Ltd 01.01.2007; Blackwell Publishing Ltd; Blackwell
• Subjects: Actins - metabolism; Adaptor Proteins, Signal Transducing - genetics; Adaptor Proteins, Signal Transducing - immunology; Adaptor Proteins, Signal Transducing - metabolism; Analytical, structural and metabolic biochemistry; Animals; Antibodies, Monoclonal - metabolism; Biological and medical sciences; Brain - cytology; Cell physiology; Cells; Cells, Cultured; Cercopithecus aethiops; Enzyme Inhibitors - pharmacology; Enzymes and enzyme inhibitors; Extracellular Signal-Regulated MAP Kinases - metabolism; extracellular signal‐regulated kinase; Female; Flavonoids - pharmacology; Fundamental and applied biological sciences. Psychology; Kinases; Male; Microscopy, Immunoelectron - methods; Molecular and cellular biology; Mutagenesis - physiology; neuron; Neurons; Neurons - metabolism; Neurons - ultrastructure; phosphorylation; Phosphorylation - drug effects; Protein Structure, Tertiary; Proteins; Rats; Rats, Sprague-Dawley; Rodents; Serine - metabolism; Signal transduction; synapse; Synapses - metabolism; Synapses - ultrastructure; Synaptophysin - metabolism; Transfection - methods; Transferases; vinexin; Alternative splicing; Phosphorylation; extracellular signal-regulated kinase; Cell function; Organization; Extracellular signal-regulated protein kinase; Rat; Enzyme; Rodentia; Central nervous system; Telencephalon; neuron; Adhesion; Protein; Encephalon; Signal transduction; Vertebrata; Synapse; Mammalia; Developmental stage; vinexin
• ISSN: 0022-3042; 1471-4159
• DOI: 10.1111/j.1471-4159.2006.04222.x

Vinexin is an adaptor protein that is supposed to play pivotal roles in cell adhesion, cytoskeletal organization and signaling. At least three splice variants, vinexinα, β and γ, 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 vinexinα is relatively abundant in the telencephalon regions of the adult rat brain. An immunohistochemical study showed the localization of vinexinα 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. Vinexinβ is reported to be phosphorylated by extracellular signal-regulated kinase (ERK) at Ser189, which is equivalent to Ser593 of vinexinα. 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.