Elucidation of a complete picture of the activation mechanism of Toll-like receptors in lipid bilayers by multilevel analysis

• Project/Area Number: 19H03164
• Research Category: Grant-in-Aid for Scientific Research (B)
• Allocation Type: Single-year Grants
• Section: 一般
• Review Section: Basic Section 43020:Structural biochemistry-related
• Research Institution: The University of Tokyo
• Principal Investigator: Ohto Umeharu 東京大学, 大学院薬学系研究科(薬学部), 准教授 (90451856)
• Project Period (FY): 2019-04-01 – 2022-03-31
• Project Status: Completed (Fiscal Year 2021)
• Budget Amount: ¥17,420,000 (Direct Cost: ¥13,400,000、Indirect Cost: ¥4,020,000); Fiscal Year 2021: ¥4,810,000 (Direct Cost: ¥3,700,000、Indirect Cost: ¥1,110,000); Fiscal Year 2020: ¥4,810,000 (Direct Cost: ¥3,700,000、Indirect Cost: ¥1,110,000); Fiscal Year 2019: ¥7,800,000 (Direct Cost: ¥6,000,000、Indirect Cost: ¥1,800,000)
• Keywords: 自然免疫 / TLR受容体 / クライオ電子顕微鏡 / タンパク質 / 構造解析 / シグナル伝達 / 膜受容体

Outline of Research at the Start

本課題では自然免疫系の一回膜貫通型のToll様受容体（TLR）を研究対象とする。TLRは様々な疾患に関与し、医学・薬学的な重要に創薬ターゲットであり、そのリガンド認識・活性化・シグナル伝達機構の正しい理解が必要とされている。本申請課題では、TLR全長の脂質二重膜上での構造および挙動を多階層で解析する。その結果はTLRの活性化機構のの正しい理解、さらにTLRを標的とした薬剤の開発に資するものである。

Outline of Final Research Achievements

Structural biology of the full-length of innate immune receptor TLR was investigated. Cryo-EM structural analysis of full-length TLR3 reconstituted into nanodisc was successfully performed. The density of extracellular domain of TLR3 was clearly observed, but transmembrane regions and intracellular TIR domains were poorly resolved. The structures of TLR3-UNC93B1 and TLR7-UNC93B1 were successfully determined, revealing the structural basis of the regulation mechanism of nucleic acid-sensing TLR localization by UNC93B1.

Academic Significance and Societal Importance of the Research Achievements

種々の病気の治療薬開発のターゲットであるTLR受容体に関して構造生物学的な研究を進めた。TLR全長の活性化機構の全貌はいまだ明らかになっておらず、本研究は今後の研究の基盤となることが期待される。一部のTLRの細胞内局在を制御するUNC93B1とTLRの複合体の構造解析に成功し、TLRの細胞内局在を制御する薬剤開発の構造基盤を明らかにした。

Research Products

• [Journal Article] Structural basis for the oligomerization-mediated regulation of NLRP3 inflammasome activation (2022)
  • Author(s): Ohto Umeharu、Kamitsukasa Yukie、Ishida Hanako、Zhang Zhikuan、Murakami Karin、Hirama Chie、Maekawa Sakiko、Shimizu Toshiyuki
  • Journal Title: Proceedings of the National Academy of Sciences Volume: 119 Issue: 11
  • DOI: 10.1073/pnas.2121353119
  • Peer Reviewed
• [Journal Article] The structure of NLRP9 reveals a unique C‐terminal region with putative regulatory function (2022)
  • Author(s): Kamitsukasa Yukie、Nakano Kenji、Murakami Karin、Hirata Kunio、Yamamoto Masaki、Shimizu Toshiyuki、Ohto Umeharu
  • Journal Title: FEBS Letters Volume: 596 Issue: 7 Pages: 876-885
  • DOI: 10.1002/1873-3468.14302
  • Peer Reviewed
• [Journal Article] Tetrasubstituted imidazoles as incognito Toll-like receptor 8 a(nta)gonists (2021)
  • Author(s): Yang Yi、Csakai Adam、Jiang Shuangshuang、Smith Christina、Tanji Hiromi、Huang Jian、Jones Torey、Sakaniwa Kentaro、Broadwell Lindsey、Shi Chengrui、Soti Subada、Ohto Umeharu、Fang Yaohui、Shen Shu、Deng Fei、Shimizu Toshiyuki、Yin Hang
  • Journal Title: Nature Communications Volume: 12 Issue: 1 Pages: 4351-4351
  • DOI: 10.1038/s41467-021-24536-4
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Improving particle quality in cryo-EM analysis using a PEGylation method (2021)
  • Author(s): Zhang Zhikuan、Shigematsu Hideki、Shimizu Toshiyuki、Ohto Umeharu
  • Journal Title: Structure Volume: 29 Issue: 10 Pages: 1192-1199.e4
  • DOI: 10.1016/j.str.2021.05.004
  • Peer Reviewed
• [Journal Article] Cryo-EM structures of Toll-like receptors in complex with UNC93B1 (2021)
  • Author(s): Ishida Hanako、Asami Jinta、Zhang Zhikuan、Nishizawa Tomohiro、Shigematsu Hideki、Ohto Umeharu、Shimizu Toshiyuki
  • Journal Title: Nature Structural & Molecular Biology Volume: 28 Issue: 2 Pages: 173-180
  • DOI: 10.1038/s41594-020-00542-w
  • Peer Reviewed / Open Access
• [Journal Article] Structural analysis reveals TLR7 dynamics underlying antagonism (2020)
  • Author(s): Tojo Shingo、Zhang Zhikuan、Matsui Hiroyuki、Tahara Masahiro、Ikeguchi Mitsunori、Kochi Mami、Kamada Mami、Shigematsu Hideki、Tsutsumi Akihisa、Adachi Naruhiko、Shibata Takuma、Yamamoto Masaki、Kikkawa Masahide、Senda Toshiya、Isobe Yoshiaki、Ohto Umeharu、Shimizu Toshiyuki
  • Journal Title: Nature Communications Volume: 11 Issue: 1 Pages: 1-11
  • DOI: 10.1038/s41467-020-19025-z
  • Peer Reviewed / Open Access
• [Journal Article] The effect of anesthetics on toll like receptor 9 (2020)
  • Author(s): Koutsogiannaki Sophia、Bu Weiming、Hou Lifei、Shibamura‐Fujiogi Miho、Ishida Hanako、Ohto Umeharu、Eckenhoff Roderic G.、Yuki Koichi
  • Journal Title: The FASEB Journal Volume: 34 Issue: 11 Pages: 14645-14654
  • DOI: 10.1096/fj.202000791rr
  • Peer Reviewed / Int'l Joint Research
• [Journal Article] Crystal structure of the FYCO1 RUN domain suggests possible interfaces with small GTPases (2020)
  • Author(s): Sakurai Shunya、Shimizu Toshiyuki、Ohto Umeharu
  • Journal Title: Acta Crystallographica Section F Structural Biology Communications Volume: 76 Issue: 8 Pages: 326-333
  • DOI: 10.1107/s2053230x20009012
  • Peer Reviewed
• [Journal Article] Crystal structure of TEX101, a glycoprotein essential for male fertility, reveals the presence of tandemly arranged Ly6/uPAR domains (2020)
  • Author(s): Masutani Mamiko、Sakurai Shunya、Shimizu Toshiyuki、Ohto Umeharu
  • Journal Title: FEBS Letters Volume: 594 Issue: 18 Pages: 3020-3031
  • DOI: 10.1002/1873-3468.13875
  • Peer Reviewed
• [Journal Article] Homeostatic and pathogenic roles of GM3 ganglioside molecular species in TLR4 signaling in obesity (2020)
  • Author(s): Kanoh Hirotaka、Nitta Takahiro、Go Shinji、Inamori Kei‐ichiro、Veillon Lucas、Nihei Wataru、Fujii Mayu、Kabayama Kazuya、Shimoyama Atsushi、Fukase Koichi、Zonca Asia、Cattaneo Anna、Letizia Marilena、Ciampa Maria、Mauri Laura、Prinetti Alessandro、Sonnino Sandro、Suzuki Akemi、Inokuchi Jin‐ichi et. al.
  • Journal Title: The EMBO Journal Volume: － Issue: 12 Pages: 1-20
  • DOI: 10.15252/embj.2019101732
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Rationally Designed Small-Molecule Inhibitors Targeting an Unconventional Pocket on the TLR8 Protein-Protein Interface (2020)
  • Author(s): Jiang Shuangshuang、Tanji Hiromi、Yin Kejun、Zhang Shuting、Sakaniwa Kentaro、Huang Jian、Yang Yi、Li Jing、Ohto Umeharu、Shimizu Toshiyuki、Yin Hang
  • Journal Title: Journal of Medicinal Chemistry Volume: 63 Issue: 8 Pages: 4117-4132
  • DOI: 10.1021/acs.jmedchem.9b02128
  • Peer Reviewed / Int'l Joint Research
• [Presentation] UNC93B1による核酸認識TLRの輸送機構の構造基盤 (2021)
  • Author(s): 石田英子、浅見仁太、大戸梅治、清水敏之
  • Organizer: 日本薬学会第141年会（広島）オンライン
• [Presentation] 一回膜貫通型Toll様受容体のリガンド認識とシグナル伝達機構の構造基盤 (2019)
  • Author(s): 大戸梅治
  • Organizer: 第19回日本蛋白質科学会 第71回日本細胞生物学会 合同年次大会
  • Invited
• [Presentation] ZOO systemを用いた自動回折データ収集の実際 (2019)
  • Author(s): 大戸梅治
  • Organizer: 第11回SPring-8ユーザー共同体(SPRUC)放射光構造生物学研究会
  • Invited
• [Remarks] 研究室ホームページ
  • URL: https://kouzou.f.u-tokyo.ac.jp/
• [Remarks] 研究室ホームページ
  • URL: http://www.f.u-tokyo.ac.jp/~kouzou/index.html