Reconstitution of Rab2 targeting to ER-Golgi intermediate compartment in semi-intact cells

2020 Fiscal Year Final Research Report

• Project/Area Number: 17K07379
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: Cell biology
• Research Institution: Tokyo Institute of Technology
• Principal Investigator: Kano Fumi 東京工業大学, 科学技術創成研究院, 准教授 (10361594)
• Project Period (FY): 2017-04-01 – 2021-03-31
• Keywords: 再構成 / オルガネラ / タンパク質局在 / Rab2

Outline of Final Research Achievements

Rab2, a member of small GTPase protein family, specifically targeted to ER-Golgi intermediate compartment (ERGIC). To reveal its regulatory mechanism, we reconstituted the targeting of GFP-tagged Rab2 recombinant protein to ERGIC in semi-intact HeLa cells, the plasma membrane of which was permeabilized with streptococcal pore forming toxin, streptolysin O. The targeting assay using the inhibitory antibodies revealed the involvement of two proteins, X and Y (the names of the proteins can not be open to the public since the paper relating to the results is now under submission), in the Rab2 targeting among the Rab2-interacting proteins. X was found interacting with Rab2 in cytoplasm and functioned for retaining the GTP-bound form of Rab2, while Y was bound to Rab2 near ERGIC membrane. These results suggested that Rab2 targeting to ERGIC was sequentially regulated by interacting with two distinct proteins.

Free Research Field

細胞生物学

Academic Significance and Societal Importance of the Research Achievements

セミインタクト細胞アッセイは細胞内構造の維持が必要不可欠なタンパク質の局在化研究に適した細胞側ツールであると言える。本研究で構築するアッセイは汎用性が高く、蛋白質の局在化と機能相関が重要である細胞生物学分野において学術的に意義がある。また、本申請研究で注目するRab2は、小胞輸送だけでなく、シグナル伝達、インスリン分泌・分解制御、オートファジーなどの細胞機能にも関与する。他にも自閉症スペクトラム障害や精神分裂病でRab2A点変異が報告されるなどRab2の生理機能の重要性はますます注目されており、その局在化機構解明はこれらの生命現象にも深く関わる重要なテーマである。