Functional diversity of voltage-sensing phosphatases in two urodele amphibians

Authors

  • Joshua Mutua,

    Corresponding author
    1. Graduate School of Frontier Biosciences, Osaka University, Suita, Osaka, Japan
    • Correspondence

      Yasushi Okamura, Integrative Physiology, Department of Physiology, Graduate School of Medicine, Osaka University, Yamada-Oka 2-2, Suita, Osaka 565-0871, Japan.

      Tel: +81-6-68793311

      Fax: +81-6-68793319

      E-mail: yokamura@phys2.med.osaka-u.ac.jp

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  • Yuka Jinno,

    1. Laboratory of Integrative Physiology, Department of Physiology, Graduate School of Medicine, Osaka University, Suita, Osaka, Japan
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  • Souhei Sakata,

    1. Laboratory of Integrative Physiology, Department of Physiology, Graduate School of Medicine, Osaka University, Suita, Osaka, Japan
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  • Yoshifumi Okochi,

    1. Laboratory of Integrative Physiology, Department of Physiology, Graduate School of Medicine, Osaka University, Suita, Osaka, Japan
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  • Shuichi Ueno,

    1. Laboratory of Molecular Developmental Biology, Department of Applied Molecular Biosciences, Graduate School of Medicine, Yamaguchi University, Yamaguchi, Japan
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  • Hidekazu Tsutsui,

    1. Laboratory of Integrative Physiology, Department of Physiology, Graduate School of Medicine, Osaka University, Suita, Osaka, Japan
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  • Takafumi Kawai,

    1. Laboratory of Integrative Physiology, Department of Physiology, Graduate School of Medicine, Osaka University, Suita, Osaka, Japan
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  • Yasuhiro Iwao,

    1. Laboratory of Molecular Developmental Biology, Department of Applied Molecular Biosciences, Graduate School of Medicine, Yamaguchi University, Yamaguchi, Japan
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  • Yasushi Okamura

    Corresponding author
    1. Graduate School of Frontier Biosciences, Osaka University, Suita, Osaka, Japan
    2. Laboratory of Integrative Physiology, Department of Physiology, Graduate School of Medicine, Osaka University, Suita, Osaka, Japan
    • Correspondence

      Yasushi Okamura, Integrative Physiology, Department of Physiology, Graduate School of Medicine, Osaka University, Yamada-Oka 2-2, Suita, Osaka 565-0871, Japan.

      Tel: +81-6-68793311

      Fax: +81-6-68793319

      E-mail: yokamura@phys2.med.osaka-u.ac.jp

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  • Funding Information

    This study was supported by Ministry of Education, Culture, Sports, Science & Technology (MEXT) of Japan. Grants-in-Aid for basic research S (no. 25253016) and A (no. 21229003).

Abstract

Voltage-sensing phosphatases (VSPs) share the molecular architecture of the voltage sensor domain (VSD) with voltage-gated ion channels and the phosphoinositide phosphatase region with the phosphatase and tensin homolog (PTEN), respectively. VSPs enzymatic activities are regulated by the motions of VSD upon depolarization. The physiological role of these proteins has remained elusive, and insights may be gained by investigating biological variations in different animal species. Urodele amphibians are vertebrates with potent activities of regeneration and also show diverse mechanisms of polyspermy prevention. We cloned cDNAs of VSPs from the testes of two urodeles; Hynobius nebulosus and Cynops pyrrhogaster, and compared their expression and voltage-dependent activation. Their molecular architecture is highly conserved in both Hynobius VSP (Hn-VSP) and Cynops VSP (Cp-VSP), including the positively-charged arginine residues in the S4 segment of the VSD and the enzymatic active site for substrate binding, yet the C-terminal C2 domain of Hn-VSP is significantly shorter than that of Cp-VSP and other VSP orthologs. RT-PCR analysis showed that gene expression pattern was distinct between two VSPs. The voltage sensor motions and voltage-dependent phosphatase activities were investigated electrophysiologically by expression in Xenopus oocytes. Both VSPs showed “sensing” currents, indicating that their voltage sensor domains are functional. The phosphatase activity of Cp-VSP was found to be voltage dependent, as shown by its ability to regulate the conductance of coexpressed GIRK2 channels, but Hn-VSP lacked such phosphatase activity due to the truncation of its C2 domain.

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