Semaphorin3A signalling is mediated via sequential Cdk5 and GSK3β phosphorylation of CRMP2: implication of common phosphorylating mechanism underlying axon guidance and Alzheimer's disease

Authors

  • Yutaka Uchida,

    1. Department of Molecular Pharmacology and Neurobiology, Yokohama City University Graduate School of Medicine, Yokohama 236-0004, Japan
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  • Toshio Ohshima,

    Corresponding author
    1. Laboratory for Developmental Neurobiology, Brain Science Institute, The Institute of Physical and Chemical Research (RIKEN), Wako 351-0198, Japan
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  • Yukio Sasaki,

    1. Department of Molecular Pharmacology and Neurobiology, Yokohama City University Graduate School of Medicine, Yokohama 236-0004, Japan
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    • a

      Present address: Department of Neuroscience, Rose Kennedy Center for Mental Retardation, Albert Einstein College of Medicine, Bronx, NY 10461, USA

  • Hiromi Suzuki,

    1. Laboratory for Developmental Neurobiology, Brain Science Institute, The Institute of Physical and Chemical Research (RIKEN), Wako 351-0198, Japan
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  • Shigeki Yanai,

    1. Department of Molecular Pharmacology and Neurobiology, Yokohama City University Graduate School of Medicine, Yokohama 236-0004, Japan
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  • Naoya Yamashita,

    1. Department of Molecular Pharmacology and Neurobiology, Yokohama City University Graduate School of Medicine, Yokohama 236-0004, Japan
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  • Fumio Nakamura,

    1. Department of Molecular Pharmacology and Neurobiology, Yokohama City University Graduate School of Medicine, Yokohama 236-0004, Japan
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  • Kohtaro Takei,

    1. Department of Molecular Pharmacology and Neurobiology, Yokohama City University Graduate School of Medicine, Yokohama 236-0004, Japan
    2. CREST, Japan Science and Technology Corporation, Kawaguchi 332-0012, Japan
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  • Yasuo Ihara,

    1. Department of Neuropathology, Faculty of Medicine, University of Tokyo, 113-0033, Japan
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  • Katsuhiko Mikoshiba,

    1. Laboratory for Developmental Neurobiology, Brain Science Institute, The Institute of Physical and Chemical Research (RIKEN), Wako 351-0198, Japan
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  • Papachan Kolattukudy,

    1. Biomolecular Science Center, University of Central Florida, Biomolecular Science, Orlando, Florida 32816, USA
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  • Jerome Honnorat,

    1. Institut National de la Sante et de la Recherche Medicale U 433, Institut Federatif des Neurosciences de Lyon, Hôpital Neurologique, 69003 Lyon, France
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  • Yoshio Goshima

    Corresponding author
    1. Department of Molecular Pharmacology and Neurobiology, Yokohama City University Graduate School of Medicine, Yokohama 236-0004, Japan
    2. CREST, Japan Science and Technology Corporation, Kawaguchi 332-0012, Japan
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  • Communicated by: Kozo Kaibuchi

Correspondence: E-mail: goshima@med.yokohama-cu.ac.jp and ohshima@brain.riken.go.jp

Abstract

Collapsin response mediating protein-2 (CRMP2) has been identified as an intracellular protein mediating Semaphorin3A (Sema3A), a repulsive guidance molecule. In this study, we demonstrate that cyclin-dependent kinase 5 (Cdk5) and glycogen synthase kinase 3β (GSK3β) plays a critical role in Sema3A signalling. In In vitro kinase assay, Cdk5 phosphorylated CRMP2 at Ser522, while GSK3β did not induce any phosphorylation of CRMP2. Phosphorylation by GSK3β was exclusively observed in Cdk5-phosphorylated CRMP2, but barely in CRMP2T509A. These results indicate that Cdk5 primarily phosphorylates CRMP2 at Ser522 and GSK3β secondarily phosphorylates at Thr509. The dual-phosphorylated CRMP2, but not non-phosphorylated or single-phosphorylated CRMP2, is recognized with the antibody 3F4, which is highly reactive with the neurofibrillary tangles of Alzheimer's disease. 3F4 recognized the CRMP2 in the wild-type but not cdk5−/− mouse embryonic brain lysates. The phosphorylation of CRMP2 at Ser522 caused reduction of its affinity to tubulin. In dorsal root ganglion neurones, Sema3A stimulation enhanced the levels of the phosphorylated form of CRMP2 detected by 3F4. Over-expression of CRMP2 mutant substituting either Ser522 or Thr509 to Ala attenuates Sema3A-induced growth cone collapse response. These results suggest that the sequential phosphorylation of CRMP is an important process of Sema3A signalling and the same mechanism may have some relevance to the pathological aggregation of the microtubule-associated proteins.

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