Identification of regulatory factors for sperm flagellar motility by using proteomic analysis and molecular imaging techniques

• Project/Area Number: 16K07337
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: Cell biology
• Research Institution: University of Tsukuba
• Principal Investigator: Shiba Kogiku 筑波大学, 生命環境系, 助教 (70533561)
• Research Collaborator: INABA KAZUO
• Project Period (FY): 2016-04-01 – 2019-03-31
• Project Status: Completed (Fiscal Year 2018)
• Budget Amount: ¥4,940,000 (Direct Cost: ¥3,800,000、Indirect Cost: ¥1,140,000); Fiscal Year 2018: ¥1,820,000 (Direct Cost: ¥1,400,000、Indirect Cost: ¥420,000); Fiscal Year 2017: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000); Fiscal Year 2016: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000)
• Keywords: 精子鞭毛運動 / カルシウム / cAMP / 鞭毛運動 / 走化性 / 波形調節 / 精子運動 / 精子 / プロテオミクス / 細胞構造・機能 / 鞭毛

Outline of Final Research Achievements

This research was carried out to identify regulatory factors for sperm flagellar motility by using proteomic analysis and molecular imaging techniques. Marine invertebrates were mainly used for this research. The following results were obtained. (1) Several axonemal proteins in ascidian sperm related to Ca2+-dependent changes in flagellar waveforms were identified. (2) The experimental system for functional analysis of regulatory factors were constructed and improved. (3) Experiments using caged compound revealed that cAMP and phosphorylation of axonemal proteins by cAMP-dependent kinase are involved in Ca2+-dependent regulation of flagellar waveform.

Academic Significance and Societal Importance of the Research Achievements

真核生物の鞭毛繊毛は、精子運動・体内の水流制御など細胞運動やシグナル受容において非常に重要な器官である。その構造・機能は進化を通して高度に保存されており、鞭毛繊毛運動制御機構の欠失はヒトをはじめ多くの生物で生命活動の維持に致命的な影響を与える。本研究の成果により精子運動制御における調節因子の役割および分子機構の一端が解明された。将来的には生殖治療や繁殖技術への応用が期待される。また高速度運動の分子イメージング技術を活かした本研究の進展は、さまざまな細胞運動における解析システムの開発にもつながると考えられる。

Research Products

• [Int'l Joint Research] UMR7009 LBDV, CNRS Sorbonne University(フランス)
• [Journal Article] A brown algal sex pheromone reverses the sign of phototaxis by cAMP/Ca2+ dependent signaling in brown algal male gametes of Mutimo cylindricus (Cutleriaceae) (2019)
  • Author(s): Nana Kinoshita-Terauchi, Kogiku Shiba, Taiki Umezawa, Fuyuhiko Matsuda, Taizo Motomura, Kazuo Inaba
  • Journal Title: Journal of Photochemistry and Photobiology B Volume: 192 Pages: 113-123
  • DOI: 10.1016/j.jphotobiol.2019.01.010
  • Peer Reviewed
• [Journal Article] Microtubule stabilizer reveals requirement of Ca2+-dependent conformational changes of microtubules for rapid coiling of haptonema in haptophyte algae (2019)
  • Author(s): Nomura Mami、Atsuji Kohei、Hirose Keiko、Shiba Kogiku、Yanase Ryuji、Nakayama Takeshi、Ishida Ken-ichiro、Inaba Kazuo
  • Journal Title: Biology Open Volume: 8 Pages: 036590-036590
  • DOI: 10.1242/bio.036590
  • NAID: 120007128081
  • Peer Reviewed / Open Access
• [Journal Article] Ca2+ efflux via plasma membrane Ca2+-ATPase mediates chemotaxis in ascidian sperm (2018)
  • Author(s): Yoshida Kaoru, Shiba Kogiku, Nakamoto Ayako, Ikenaga Jumpei, Matsunaga Shigeru, Inaba Kazuo, Yoshida Manabu
  • Journal Title: Scientific Reports Volume: 8 Issue: 1 Pages: 16622-16622
  • DOI: 10.1038/s41598-018-35013-2
  • Peer Reviewed / Open Access
• [Journal Article] 14-3-3εa directs the pulsatile transport of basal factors towards the apical domain for lumen growth in tubulogenesis. (2018)
  • Author(s): Yuji Mizotani, Mayu Suzuki, Kohji Hotta, Hidenori Watanabe, Kogiku Shiba, Kazuo Inaba, Etsu Tashiro, Kotaro Oka, Masaya Imoto
  • Journal Title: Proc. Natl. Acad. Sci. USA. Volume: 115 Issue: 38
  • DOI: 10.1073/pnas.1808756115
  • NAID: 120007133686
  • Peer Reviewed / Open Access
• [Journal Article] High doses of CRISPR/Cas9 ribonucleoprotein efficiently induce gene knockout with low mosaicism in the hydrozoan Clytia hemisphaerica through microhomology-mediated deletion (2018)
  • Author(s): Momose Tsuyoshi、De Cian Anne、Shiba Kogiku、Inaba Kazuo、Giovannangeli Carine、Concordet Jean-Paul
  • Journal Title: Scientific Reports Volume: 8 Issue: 1 Pages: 11734-11734
  • DOI: 10.1038/s41598-018-30188-0
  • NAID: 120007133847
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Release of Sticky Glycoproteins from Chlamydomonas Flagella During Microsphere Translocation on the Surface Membrane (2018)
  • Author(s): Kamiya Ritsu, Shiba Kogiku, Inaba Kazuo, Kato-Minoura Takako
  • Journal Title: Zoological Science Volume: 35 Issue: 4 Pages: 299-305
  • DOI: 10.2108/zs180025
  • NAID: 210000169673
  • Peer Reviewed
• [Journal Article] Microscopic analysis of sperm movement: links to mechanisms and protein components (2018)
  • Author(s): Inaba Kazuo, Shiba Kogiku
  • Journal Title: Microscopy Volume: 67 Issue: 3 Pages: 144-155
  • DOI: 10.1093/jmicro/dfy021
  • Peer Reviewed
• [Journal Article] A unique seminal plasma protein, zona pellucida 3-like protein, has Ca2+-dependent sperm agglutination activity. (2018)
  • Author(s): Ugwu, S. I., Shiba, K., Inaba, K., Morita, M.
  • Journal Title: Zool. Sci. Volume: 35 Issue: 2 Pages: 161-171
  • DOI: 10.2108/zs170150
  • NAID: 210000179747
  • Peer Reviewed / Int'l Joint Research
• [Journal Article] Inverse relationship of Ca2+-dependent flagellar response between animal sperm and prasinophyte algae (2017)
  • Author(s): Shiba Kogiku、Inaba Kazuo
  • Journal Title: J Plant Res. Volume: 130 Issue: 3 Pages: 465-473
  • DOI: 10.1007/s10265-017-0931-7
  • Peer Reviewed
• [Journal Article] Calaxin establishes basal body orientation and coordinates movement of monocilia in sea urchin embryos (2017)
  • Author(s): Mizuno Katsutoshi、Shiba Kogiku、Yaguchi Junko、Shibata Daisuke、Yaguchi Shunsuke、Pruliere Gerard、Chenevert Janet、Inaba Kazuo
  • Journal Title: Sci Rep. Volume: 7 Issue: 1 Pages: 10751-10751
  • DOI: 10.1038/s41598-017-10822-z
  • NAID: 120007134832
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Doublet 7 shortening, doublet 5-preferential poly-Glu reduction, and beating stall of sperm flagella in Ttll9-/- mice. (2016)
  • Author(s): Konno A, Ikegami K, Konishi Y, Yang HJ, Abe M, Yamazaki M, Sakimura K, Yao I, Shiba K, Inaba K, Setou M.
  • Journal Title: J Cell Sci. Volume: 129 Issue: 14 Pages: 2757-2766
  • DOI: 10.1242/jcs.185983
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Flagellar waveforms of gametes in the brown alga Ectocarpus siliculosus. (2016)
  • Author(s): Kinoshita N, Shiba K, Inaba K, Fu G, Nagasato C and Motomura T.
  • Journal Title: European Journal of Phycology Volume: 51 Issue: 2 Pages: 139-148
  • DOI: 10.1080/09670262.2015.1109144
  • NAID: 120006220013
  • Peer Reviewed / Int'l Joint Research
• [Presentation] cAMP modulates Ca2+-mediated regulation of sperm flagellar waveform (2018)
  • Author(s): Kogiku Shiba, Taichi Sakano, Toshiaki Furuta, Eiji Fujiwara, Shoji A Baba, Kazuo Inaba
  • Organizer: XIII International Symposium on Spermatology (ISS)
  • Int'l Joint Research
• [Presentation] cAMP modulates Ca2+-mediated regulation of sperm flagellar waveform (2018)
  • Author(s): 柴 小菊, 坂野 太一, 古田 寿昭, 藤原 英史, 馬場 昭次, 稲葉 一男
  • Organizer: 第56回日本生物物理学会年会
• [Presentation] カルシウムイオンを介した精子鞭毛波形制御におけるcAMP の調整作用 (2017)
  • Author(s): 柴 小菊，坂野 太一，古田 寿昭，藤原 英史，馬場 昭次，稲葉 一男
  • Organizer: 日本動物学会第88回大会
• [Presentation] cAMP modulates Ca2+-mediated regulation of sperm flagellar waveform (2017)
  • Author(s): Kogiku Shiba, Taichi Sakano, Toshiaki Furuta, Eiji Fujiwara, Shoji A Baba, Kazuo Inaba
  • Organizer: International Workshop Dynein 2017
  • Int'l Joint Research
• [Presentation] The role of calaxin in the propagation of asymmetric waves in sperm flagella (2016)
  • Author(s): Kogiku Shiba, Shoji A Baba, Eiji Fujiwara, Kazuo Inaba
  • Organizer: the 28th CDB Meeting "Cilia and Centrosomes: Current Advances and Future Directions"
  • Place of Presentation: RIKEN Center for Developmental Biology, Kobe, Hyogo, Japan
  • Year and Date: 2016-11-27
  • Int'l Joint Research
• [Presentation] Roles of calcium in the regulation of sperm flagellar movement. (2016)
  • Author(s): Kogiku Shiba
  • Organizer: 第54回日本生物物理学会年会
  • Place of Presentation: つくば国際会議場, 茨城県つくば市
  • Year and Date: 2016-11-25
  • Invited
• [Presentation] Regulation of asymmetry in sperm flagellar waveforms and intracellular calcium. (2016)
  • Author(s): Kogiku Shiba
  • Organizer: The joint meeting of the 22nd International Congress of Zoology and the 87th Meeting of the Zoological Society of Japan
  • Place of Presentation: Okinawa Institute of Science and Technology Graduate University and Okinawa Convention Center, Onna-son, Okinawa, Japan
  • Year and Date: 2016-11-14
  • Int'l Joint Research / Invited
• [Remarks] 筑波大学下田臨海実験センターHP
  • URL: http://www.shimoda.tsukuba.ac.jp/ronbun.html
• [Remarks] 筑波大学下田臨海実験センター稲葉研究室HP
  • URL: http://www.shimoda.tsukuba.ac.jp/~inaba/publish.html
• [Remarks] 筑波大学下田臨海実験センター稲葉研究室柴小菊HP
  • URL: http://www.shimoda.tsukuba.ac.jp/~inaba/kogiku.html
• [Remarks] 研究室ホームページ
  • URL: http://www.shimoda.tsukuba.ac.jp/~inaba/kogiku.html
• [Remarks] 筑波大学下田臨海実験センターホームページ
  • URL: http://www.shimoda.tsukuba.ac.jp/index.html
• [Remarks] 筑波大学下田臨海実験センター発表論文
  • URL: http://www.shimoda.tsukuba.ac.jp/ronbun.html
• [Remarks] 筑波大学下田臨海実験センター稲葉研究室発表論文
  • URL: http://www.shimoda.tsukuba.ac.jp/~inaba/publish.html