Roles of Rho-family small GTPases in establishment and maintenance for hearing and balancing

• Project/Area Number: 26460340
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: General pharmacology
• Research Institution: Kobe University
• Principal Investigator: Ueyama Takehiko 神戸大学, バイオシグナル総合研究センター, 准教授 (80346254)
• Co-Investigator(Kenkyū-buntansha): 坂口 博史 京都府立医科大学, 医学(系)研究科(研究院), 准教授 (00515223)
• Co-Investigator(Renkei-kenkyūsha): SAITO Naoaki 神戸大学, バイオシグナル総合研究センター, 教授 (60178499)
• Project Period (FY): 2014-04-01 – 2017-03-31
• Project Status: Completed (Fiscal Year 2016)
• Budget Amount: ¥5,070,000 (Direct Cost: ¥3,900,000、Indirect Cost: ¥1,170,000); Fiscal Year 2016: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000); Fiscal Year 2015: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000); Fiscal Year 2014: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000)
• Keywords: 聴覚 / 平衡覚 / 低分子量G蛋白質 / 聴毛 / 耳石 / DIA1 / DFNA1 / 感音難聴 / 遺伝性難聴 / Cdc42 / RhoA / DIAPH1 / アクチン / Rho-family GTPase / Rac / 内耳有毛細胞 / 聴力 / 活性酸素種

Outline of Final Research Achievements

(1) We generated inner ear hair cell (HC)-specific KO mice to analyze the role of Cdc42 in HCs. HCs of Cdc42-KO mice developed normally but progressively degenerated after maturation, resulting in progressive hearing loss particularly at high frequencies. Adenovirus-encoded GFP-Cdc42 expression in HCs and fluorescence resonance energy transfer (FRET) imaging of HCs from transgenic mice expressing Cdc42-FRET biosensor indicated Cdc42 presence/activation at stereociliary membranes in cochlear HCs.
(2) We found that the amount of active RhoA (GTP-form) is increased in Cdc42-KD cells. DIA1 is a downstream molecule of RhoA signaling pathways, and nucleates and elongates unbranched/straight actin. We discovered a novel patient-derived DIAPH1 mutation (c.3610C>T) in two unrelated Japanese families. Mice expressing the DIA1(R1204X) mutant experienced progressive deafness beginning at high　frequencies and HC loss with various morphological abnormalities in stereocilia at the basal turn.

Research Products

• [Journal Article] A novel Rac1-GSPT1 signaling pathway controls astrogliosis following central nervous system injury (2017)
  • Author(s): Ishii T, Ueyama T, Shigyo M, Kohta M, Kondoh T, Kuboyama T, Uebi T, Hamada T, Gutmann DH, Aiba A, Kohmura E, Tohda C, Saito N
  • Journal Title: J Biol Chem Volume: 292 Issue: 4 Pages: 1240-1250
  • DOI: 10.1074/jbc.m116.748871
  • Peer Reviewed / Open Access / Int'l Joint Research / Acknowledgement Compliant
• [Journal Article] Constitutive activation of DIA1 (DIAPH1) via C-terminal truncation causes human sensorineural hearing loss. (2016)
  • Author(s): Ueyama T, Ninoyu Y, Nishio S, Miyoshi T, Torii H, Nishimura K, Sugahara K, Sakata H, Thumkeo D, Sakaguchi H, Watanabe N, Usami S, Saito N, Kitajiri S.
  • Journal Title: EMBO Molecular Medicine Volume: 8 Issue: 11 Pages: 1310-1324
  • DOI: 10.15252/emmm.201606609
  • NAID: 120005850206
  • Peer Reviewed / Open Access / Int'l Joint Research / Acknowledgement Compliant
• [Journal Article] The Extracellular A-loop of Dual Oxidases Affects the Specificity of Reactive Oxygen Species Release. (2015)
  • Author(s): Ueyama T, Sakuma M, Ninoyu Y, Hamada T, Dupuy C, Geiszt M, Leto TL, Saito N.
  • Journal Title: J Biol. Chem Volume: 290 Issue: 10 Pages: 6495-6506
  • DOI: 10.1074/jbc.m114.592717
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Journal Article] Identification and characterization of PKCγ, a kinase associated with SCA14, as an amyloidogenic protein. (2015)
  • Author(s): Takahashi H, Adachi N, Shirafuji T, Danno S, Ueyama T, Vendruscolo M, Shuvaev AN, Sugimoto T, Seki T, Hamada D, Irie K, Hirai H, Sakai N, *Saito N.
  • Journal Title: Hum Mol Genet. Volume: 24 Issue: 2 Pages: 525-539
  • DOI: 10.1093/hmg/ddu472
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Journal Article] Maintenance of stereocilia and apical junctional complexes by Cdc42 in cochlear hair cells (2014)
  • Author(s): Ueyama T, Sakaguchi H, Nakamura T, Goto A, Morioka S, Shimizu A, Nakao K, Hishikawa Y, Ninoyu Y, Kassai H, Suetsugu S, Koji T, Fritzsch B, Yonemura S, Hisa Y, Matsuda M, Aiba A, Saito N.
  • Journal Title: J Cell Sci. Volume: 127 Pages: 2040-2052
  • DOI: 10.1242/jcs.143602
  • NAID: 120006986582
  • Peer Reviewed / Open Access
• [Journal Article] The role of Pak-interacting exchange factor-β phosphorylation at serines 340 and 583 by PKCγ in dopamine release. (2014)
  • Author(s): Shirafuji T, Ueyama T, Yoshino K, Takahashi H, Adachi N, Ago Y, Koda K, Nashida T, Hiramatsu N, Matsuda T, Toda T, Sakai N, Saito N.
  • Journal Title: J Neurosci Volume: 34 Issue: 28 Pages: 9268-9280
  • DOI: 10.1523/jneurosci.4278-13.2014
  • NAID: 120005619641
  • Peer Reviewed / Open Access
• [Journal Article] RANTES/CCL5 mediated-biological effects depend on the s 1 yndecan-4/PKC signaling pathway (2014)
  • Author(s): Maillard L, Saito N, Hlawaty H, Friand V, Suffee N, Chmilewsky F, Haddad O, Laguillier C, Guyot E, Ueyama T, Oudar O, Sutton A, Charnaux N
  • Journal Title: Biology Open Volume: 3 Pages: 995-1004
  • Peer Reviewed / Open Access
• [Journal Article] Diacylglycerol kinase  regulates antigen-induced mast cell degranulation by mediating Ca2+ influxes (2014)
  • Author(s): Sakuma M, Shirai Y, Ueyama T, Saito N
  • Journal Title: Biochem Biophys Res Commun Volume: 445 Pages: 340-345
  • Peer Reviewed / Open Access
• [Journal Article] Hydrogen peroxide generated by DUOX1 regulates the expression levels of specific differentiation markers in normal human keratinocyte (2014)
  • Author(s): Choi H, Park J, Kim H, Noh M, Ueyama T, Bae Y, Lee TR, Shin DW
  • Journal Title: J Dermatol Sci Volume: 74 Pages: 56-63
  • Peer Reviewed
• [Presentation] The molecular mechanism of hearing loss caused by a novel DIA1 mutant (R1204X) (2017)
  • Author(s): Yuzuru Ninoyu, Shin-ichiro Kitajiri, Takushi Miyoshi, Dean Thumkeo, Hirofumi Sakaguchi, Naoki Watanabe, Shin-ichi Usami, Naoaki Saito, Takehiko Ueyama
  • Organizer: 第90回日本薬理学会年会
  • Place of Presentation: 長崎新聞文化ホール（長崎県・長崎市）
  • Year and Date: 2017-03-15
• [Presentation] Analysis of hearing loss causing from failed maintenance of auditory hair’s integrity (2016)
  • Author(s): Takehiko Ueyama
  • Organizer: 第８９回日本薬理学会年会、シンポジウム
  • Place of Presentation: パシフィコ横浜（神奈川県・横浜市）
  • Year and Date: 2016-03-09
  • Invited