Imaging myosin-driven stress fiber contraction with molecular resolution by high-speed AFM

2023 Fiscal Year Final Research Report

• Project/Area Number: 20H03218
• Research Category: Grant-in-Aid for Scientific Research (B)
• Allocation Type: Single-year Grants
• Section: 一般
• Review Section: Basic Section 43040:Biophysics-related
• Research Institution: Kanazawa University
• Principal Investigator: Franz Clemens 金沢大学, ナノ生命科学研究所, 准教授 (50837664)
• Project Period (FY): 2020-04-01 – 2023-03-31
• Keywords: AFM / SICM / myosin / actin / stress fiber / contractility

Outline of Final Research Achievements

By combining cell de-roofing and HS-AFM imaging, this project established a method to image contracting actin stress fibers (SFs) to near molecular resolution within native cellular environments. Achieving these goals required (1) the development of suitable de-roofing methods, (2) establishment of an ultrawide scanner system for large resolution HS-AFM imaging in combination with fluorescence microscopy, and (3) developing an optimized AFM tip design for imaging highly corrugated intracellular structures. Ultrawide HS-AFM scanning generated the first molecular-resolution maps of entire native SFs, providing novel insight into SF ultrastructure. Furthermore, arrangement and conformational changes of individual non-muscle myosin II motors during SF contraction could be visualized for the first time. In parallel, SICM imaging was also adapted for intracellular imaging in de-roofed cells for the first time, providing additional structural and mechanical insight into SF contraction.

Free Research Field

Bioimaging

Academic Significance and Societal Importance of the Research Achievements

アクチンストレスファイバー（SF）の収縮は、マトリックス接着、遊走、メカノセンシングなど、多くの細胞プロセスに影響を与える一方、SF機能の誤制御は、アテローム性動脈硬化症、骨粗鬆症、がんなどの疾患の原因となりうる。このプロジェクトで得られたSFの超微細構造とSF収縮を駆動する分子メカニズムに関する新たな知見は、SFの正常および異常機能についての理解を広げるものである。