Establishment of method for identification of ubiquitinated protein

• Project/Area Number: 17K19506
• Research Category: Grant-in-Aid for Challenging Research (Exploratory)
• Allocation Type: Multi-year Fund
• Research Field: Biomedical structure and function and related fields
• Research Institution: Hokkaido University
• Principal Investigator: Hatakeyama Shigetsugu 北海道大学, 医学研究院, 教授 (70294973)
• Research Collaborator: TAKAHASHI Hidehisa 横浜市立大学, 医学研究科, 教授 (20399819) ; WATANABE Masashi 北海道大学, 医学研究院, 講師 (10632424)
• Project Period (FY): 2017-06-30 – 2019-03-31
• Project Status: Completed (Fiscal Year 2018)
• Budget Amount: ¥6,500,000 (Direct Cost: ¥5,000,000、Indirect Cost: ¥1,500,000); Fiscal Year 2018: ¥3,250,000 (Direct Cost: ¥2,500,000、Indirect Cost: ¥750,000); Fiscal Year 2017: ¥3,250,000 (Direct Cost: ¥2,500,000、Indirect Cost: ¥750,000)
• Keywords: ユビキチン / TRIM型E3 / TUBE / Parkin / 蛋白質 / シグナル伝達 / プロテオーム / 生体分子

Outline of Final Research Achievements

Ubiquitination is a chemical modification that controls protein degradation and is one of the most important post-translational modifications that support many biological phenomena. The reaction system is mediated by ubiquitin ligases (E3) selectively recognizing substrate proteins. Therefore, it is important to identify specific substrates of each E3 and to determine the ubiquitination site of substrate proteins in understanding the biological phenomenon that they are associated with. However, due to various problems, it is still far from standardizing ubiquitinated substrate identification methods. In this study, we challenged to establish a novel technology using TUBE (artificial protein exhibiting high affinity binding to polyubiquitin chain) and ubiquitin remnant antibody allows comprehensive application to various types of ubiquitin ligase (E3) and the E3-substrates relationship was analyzed.

Academic Significance and Societal Importance of the Research Achievements

本申請で開発した基質同定法は、これまでの問題点を原理的に解消した全く新しい手法であり、すべてのE3に対して適用可能であるため、E3の機能解析が加速することが期待できると思われる。本手法の応用により、正常及び異常（疾患）に関与しているユビキチン化修飾の全貌解明が期待できる。したがって、将来的には疾患特異的ユビキチン化部位を標的とした創薬のシーズを発掘できる可能性がある。

Research Products

• [Journal Article] Loss of TRIM29 Alters Keratin Distribution to Promote Cell Invasion in Squamous Cell Carcinoma. (2018)
  • Author(s): Yanagi T, Watanabe M, Hata H, Kitamura S, Imafuku K, Yanagi H, Homma A, Wang L, Takahashi H, Shimizu H, Hatakeyama S.
  • Journal Title: Cancer Res Volume: 78 Issue: 24 Pages: 6795-6806
  • DOI: 10.1158/0008-5472.can-18-1495
  • Peer Reviewed
• [Journal Article] Brain-Derived Neurotrophic Factor Improves Limited Exercise Capacity in Mice With Heart Failure (2018)
  • Author(s): Matsumoto Junichi、et al. Hatakeyama Shigetsugu、Matsumoto Masaki、Nakayama Keiichi I.、Otsuka Yutaro、Sabe Hisataka、Tsutsui Hiroyuki、Anzai Toshihisa
  • Journal Title: Circulation Volume: 138 Issue: 18 Pages: 2064-2066
  • DOI: 10.1161/circulationaha.118.035212
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Anti-Sez6l2 antibody, detected in a patient with immune-mediated cerebellar ataxia, inhibits complex formation of GluR1 and Sez6l2 (2018)
  • Author(s): Yaguchi, H., Yabe, I., Takahashi, H., Watanabe, M., Nomura, T., Kano, T., Watanabe, M. and Hatakeyama, S.
  • Journal Title: J. Neurol. Volume: 265 Issue: 4 Pages: 962-965
  • DOI: 10.1007/s00415-018-8785-z
  • Peer Reviewed
• [Journal Article] TRIM59 promotes gliomagenesis by inhibiting TC45 dephosphorylation of STAT3 (2018)
  • Author(s): Sang, Y., Li, Y., Song, L., Alvarez, A.A., Zhang, W., Lv, D., Tang, J., Liu, F., Chang, Z., Hatakeyama, S., Hu, B., Cheng, S. and Feng, H.
  • Journal Title: Cancer Res. Volume: 78 Issue: 7 Pages: 1792-1804
  • DOI: 10.1158/0008-5472.can-17-2774
  • Peer Reviewed / Int'l Joint Research
• [Journal Article] Mutations in bassoon in individuals with familial and sporadic progressive supranuclear palsy-like syndrome (2018)
  • Author(s): Yabe I, Yaguchi H, Kato Y, Miki Y, Takahashi H, Tanikawa S, Shirai S, Takahashi I, Kimura M, Hama Y, Matsushima M, Fujioka S, Kano T, Watanabe M, Nakagawa S, Kunieda Y, Ikeda Y, Hasegawa M, Nishihara H, Ohtsuka T, Tanaka S, Tsuboi Y, Hatakeyama S, Wakabayashi K, Sasaki H
  • Journal Title: Sci Rep Volume: 8 Issue: 1 Pages: 819-819
  • DOI: 10.1038/s41598-018-19198-0
  • NAID: 120006377499
  • Peer Reviewed / Open Access
• [Journal Article] TRIM Family Proteins: Roles in Autophagy, Immunity and Carcinogenesis. (2017)
  • Author(s): Hatakeyama, S
  • Journal Title: Trends Biochem. Sci. Volume: 42 Issue: 4 Pages: 297-311
  • DOI: 10.1016/j.tibs.2017.01.002
  • Peer Reviewed / Acknowledgement Compliant
• [Journal Article] Fine-tuning of thymocyte development by ubiquitination-mediated stability control of the ESCRT protein CHMP5 (2017)
  • Author(s): Watanabe, M. and Hatakeyama, S.
  • Journal Title: Cell. Moll. Immnol. Volume: 14 Issue: 12 Pages: 957-959
  • DOI: 10.1038/cmi.2017.91
  • Peer Reviewed
• [Journal Article] Sez6l2 regulates phosphorylation of ADD and neuritogenesis (2017)
  • Author(s): Yaguchi, H., Yabe, I., Takahashi, H., Watanabe, M., Nomura, T., Kano, T., Matsumoto, M., Nakayama, K.I., Watanabe, M. and Hatakeyama, S.
  • Journal Title: Biochem. Biophys. Res. Commun. Volume: 494 Issue: 1-2 Pages: 234-241
  • DOI: 10.1016/j.bbrc.2017.10.047
  • Peer Reviewed
• [Presentation] TRIMファミリーの機能解析 (2019)
  • Author(s): 畠山鎮次
  • Organizer: 第２回ユビキチン研究会
• [Presentation] Comprehensive identification of E3 ubiquitin ligase substrates by fusion of TUBE and ligase trapping methods (2018)
  • Author(s): Masashi watanabe, Shigetsugu Hatakeyama
  • Organizer: The 2nd GI-CoRE GSQ, GSB & IGM Joint Symposium
  • Int'l Joint Research / Invited
• [Presentation] TRIMタンパク質の機能と病理学的解析 (2018)
  • Author(s): 畠山鎮次
  • Organizer: 第１回ユビキチン研究会
• [Presentation] ＴＲＩＭファミリータンパク質が関与する疾患 (2017)
  • Author(s): 畠山鎮次
  • Organizer: 第24回分子皮膚科学フォーラム
  • Invited
• [Presentation] ＴＲＩＭファミリータンパク質による細胞機能制御 (2017)
  • Author(s): 畠山鎮次
  • Organizer: GU Cancer Forum 2017
  • Invited
• [Book] 人体の構造と機能[2] 生化学 第14版 (2019)
  • Author(s): 畠山 鎮次
  • Total Pages: 304
  • Publisher: 医学書院
  • ISBN: 9784260035569