Elucidation of extracellular vesicle release mechanism stimulated by LFA-1 activatory signals

• Project/Area Number: 17K18253
• Research Category: Grant-in-Aid for Young Scientists (B)
• Allocation Type: Multi-year Fund
• Research Field: Functional biochemistry; Immunology
• Research Institution: Kansai Medical University
• Principal Investigator: KONDO Naoyuki 関西医科大学, 医学部, 助教 (30570840)
• Project Period (FY): 2017-04-01 – 2019-03-31
• Project Status: Completed (Fiscal Year 2018)
• Budget Amount: ¥4,420,000 (Direct Cost: ¥3,400,000、Indirect Cost: ¥1,020,000); Fiscal Year 2018: ¥2,210,000 (Direct Cost: ¥1,700,000、Indirect Cost: ¥510,000); Fiscal Year 2017: ¥2,210,000 (Direct Cost: ¥1,700,000、Indirect Cost: ¥510,000)
• Keywords: LFA-1 / インテグリン / Rap1 / NDR1 / Kindlin-3 / 一分子解析 / 免疫シナプス / 細胞外小胞 / CD63 / 生体分子 / シグナル伝達 / 免疫学 / 細胞・組織 / 蛋白質

Outline of Final Research Achievements

At the early stage of immune response, a supra molecular structure, immunological synapse (IS), is formed between immune cells for the efficient signal transduction. It has been reported that extracellular vesicles (EVs), which contain genetic information, are accumulated at contact plane of IS; however precise regulatory mechanism of EVs accumulation/release is still elusive. In this work, several mice deficient for activatory molecules of LFA-1, a leukocyte integrin, were constructed. Using T-cells isolated from these mice, it was revealed that release of EVs occurred at specific compartment at IS contact plane and that LFA-1 activatory signal cascade via Rap1/RAPL/Mst1/NDR1/Kindlin-3 was important for EVs accumulation/release.

Academic Significance and Societal Importance of the Research Achievements

本研究は,現在まで不明な点の多かったT細胞-抗原提示細胞間ISの細胞間情報伝達過程において, EVsが接着依存的に集積することで情報伝達に関与する可能性を示唆し, またその引き金の一つとしてインテグリンLFA-1の活性化シグナルが重要な働きを担うことを初めて解明した．この結果は，ISの破綻や過剰な活性化を伴う免疫不全やアレルギー疾患の新規標的として, LFA-1活性化シグナル経路が一つの有用な候補である可能性を示唆する．また本研究で確立した実験手法は, 細胞内でのLFA-1活性やシグナル強度の分子レベルでの評価にも用いることが可能であり,新規薬剤の評価系としても応用されることが期待される．

Research Products

• [Journal Article] 自己免疫疾患のイメージング：自己寛容の成立と維持における細胞間相互作用の可視化 (2018)
  • Author(s): 植田祥啓、近藤直幸、木梨達雄
  • Journal Title: 臨床免疫・アレルギー科 Volume: 69/4 Pages: 318-325
• [Journal Article] NDR1-Dependent Regulation of Kindlin-3 Controls High-Affinity LFA-1 Binding and Immune Synapse Organization. (2017)
  • Author(s): Kondo, N., Ueda, Y., Kita, T., Ozawa, M., Tomiyama, T., Yasuda, K., Lim, D.S. and Kinashi, T.
  • Journal Title: Mol. Cell Biol. Volume: 37 Issue: 8 Pages: 00424-16
  • DOI: 10.1128/mcb.00424-16
  • Peer Reviewed / Open Access / Int'l Joint Research / Acknowledgement Compliant
• [Presentation] LFA-1を介したリンパ球細胞接着の一分子解析 (2018)
  • Author(s): Naoyuki Kondo, Tatsuo Kinashi
  • Organizer: 日本分子生物学会
• [Presentation] Rap1 signaling to NDR1 kinase regulate immune synapse formation and cell polarity (2018)
  • Author(s): Tatsuo Kinashi, Naoyuki Kondo, Yoshihiro Ueda, Yuji Kamioka
  • Organizer: The 9th Xiamen winter symposium
  • Int'l Joint Research / Invited
• [Presentation] W747 talin1 binding site in cytoplasmic domain of the integrin beta2 subunit is crucial for T-cell migration andactivation (2018)
  • Author(s): Yoshihiro Ueda, Naoyuki Kondo, Yuji Kamioka, Tatsuo Kinashi
  • Organizer: The 47th annual meeting of the Japanese Society for Immunology
• [Presentation] Roles of Rap1, Talin-1 and Kindlin-3 in lymphocyte homing and mucosal lymph nodes (2018)
  • Author(s): Yuji Kamioka, Yoshihiro Ueda, Naoyuki Kondo, Tatsuo Kinashi
  • Organizer: The 47th annual meeting of the Japanese Society for Immunology
• [Presentation] Thermus protein as versatile tools for detailed analysis of functionally unknown protein and efficient protein production (2018)
  • Author(s): Naoyuki Kondo, Zene Matsuda, Ryoji Masui, Seiki Kuramitsu
  • Organizer: International workshop on 50th anniversary of Thermus thermophilus discovery
  • Int'l Joint Research
• [Presentation] NDR1 acts as a molecular hub for the organization of immunological synapse (2017)
  • Author(s): Kondo N, Ueda Y, Kinashi T
  • Organizer: The 46th Annual Meeting of the Japanese Society for Immunology
• [Presentation] Regulation of cell polarization by Rap1 via NDR/Rab8 axis upon chemokine stimulation (2017)
  • Author(s): Ueda Y, Kondo N, Kamioka Y, Kinashi T
  • Organizer: The 46th Annual Meeting of the Japanese Society for Immunology
• [Presentation] Roles of Rap1 and Kindlin-3 in lymphocyte homing to peripheral lymph nodes (2017)
  • Author(s): Kamioka Y, Ueda Y, Kondo N, Kinashi T
  • Organizer: The 46th Annual Meeting of the Japanese Society for Immunology