Dissecting cell membrane tension dynamics and its effect on Piezo1-mediated cellular mechanosensitivity using force-controlled nanopipettes

Year: 2024

Authors: Luechtefeld I., Pivkin IV., Gardini L., Zare-Eelanjegh E., Gdbelein C., Ihle SJ., Reichmuth AM., Capitanio M., Martinac B., Zambelli T., Vassalli M.

Autors Affiliation: Swiss Fed Inst Technol, Lab Biosensors & Bioelect, Zurich, Switzerland; Univ Svizzera Italiana, Inst Comp, Lugano, Switzerland; Swiss Inst Bioinformat, Lausanne, Switzerland; CNR, Natl Inst Opt, Florence, Italy; Univ Florence, European Lab Nonlinear Spect, Florence, Italy; Swiss Fed Inst Technol, Inst Microbiol, Zurich, Switzerland; Univ Florence, Phys & Astron Dept, Florence, Italy; Victor Chang Cardiac Res Inst, Darlinghurst, NSW, Australia; Univ Glasgow, James Watt Sch Engn, Glasgow, Scotland.

Abstract: The dynamics of cellular membrane tension and its role in mechanosensing, which is the ability of cells to respond to physical stimuli, remain incompletely understood, mainly due to the lack of appropriate tools. Here, we report a force-controlled nanopipette-based method that combines fluidic force microscopy with fluorescence imaging for precise manipulation of the cellular membrane tension while monitoring the impact on single-cell mechanosensitivity. The force-controlled nanopipette enables control of the indentation force imposed on the cell cortex as well as of the aspiration pressure applied to the plasma membrane. We show that this setup can be used to concurrently monitor the activation of Piezo1 mechanosensitive ion channels via calcium imaging. Moreover, the spatiotemporal behavior of the tension propagation is assessed with the fluorescent membrane tension probe Flipper-TR, and further dissected using molecular dynamics modeling. Finally, we demonstrate that aspiration and indentation act independently on the cellular mechanobiological machinery, that indentation induces a local pre-tension in the membrane, and that membrane tension stays confined by links to the cytoskeleton. FluidFM-based force-controlled nanopipettes enable control of mechanical stimuli for the investigation of Piezo1-induced mechanosensation in cell membranes.

Journal/Review: NATURE METHODS

Volume: 21 (6)      Pages from:   to:

More Information: The authors thank J. Voeroes and the whole Laboratory of Biosensors and Bioelectronics for useful discussions and continued support to the project. The authors also thank E. Sarajlic and SmartTip B.V. for their support in cantilever fabrication, S. Wheeler for his support in the workshop, and J. Kusch and the ScopeM facility at ETH Zurich for their microscopy support. I.L. was funded by the EUREKA Network (Eurostars Project E!11644 SOUL to T.Z.). E.Z.-E. was funded by the Swiss National Science Foundation (Sinergia CRSII5_202301/1 to T.Z.). I.V.P. acknowledges the support from the Swiss National Science Foundation (Grant 205321_173020). Simulations were carried out at the Swiss National Supercomputer Center (CSCS Projects s747 and u4). C.G. was funded by the Volkswagen foundation (Initiative ’Life’). M.C. and L.G. were supported by the European Union’s Horizon 2020 research and innovation program under grant agreement no 871124 Laserlab-Europe. B.M. acknowledges the National Health and Medical Research Council of Australia for a Principal Research Fellowship (APP1135974).Open access funding provided by Swiss Federal Institute of Technology Zurich.
KeyWords: Ion Channels; Atomic-force; Patch-clamp; Piezo1; Mechanobiology; Organization; Calibration; Domain
DOI: 10.1038/s41592-024-02277-8

Citations: 2
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