title: Removal of the mechanoprotective influence of the cytoskeleton reveals PIEZO1 is gated by bilayer tension.
abstract: Mechanosensitive ion channels are force-transducing enzymes that couple mechanical stimuli to ion flux. Understanding the gating mechanism of mechanosensitive channels is challenging because the stimulus seen by the channel reflects forces shared between the membrane, cytoskeleton and extracellular matrix. Here we examine whether the mechanosensitive channel PIEZO1 is activated by force-transmission through the bilayer. To achieve this, we generate HEK293 cell membrane blebs largely free of cytoskeleton. Using the bacterial channel MscL, we calibrate the bilayer tension demonstrating that activation of MscL in blebs is identical to that in reconstituted bilayers. Utilizing a novel PIEZO1-GFP fusion, we then show PIEZO1 is activated by bilayer tension in bleb membranes, gating at lower pressures indicative of removal of the cortical cytoskeleton and the mechanoprotection it provides. Thus, PIEZO1 channels must sense force directly transmitted through the bilayer.
authors.affiliation: Victor Chang Cardiac Research Institute, Darlinghurst, New South Wales 2010, Australia.
authors.forename: Charles D
authors.initials: CD
authors.lastname: Cox
authors.affiliation: Department of Physiology and Biophysics, State University of New York at Buffalo, Buffalo, New York 14214, USA.
authors.forename: Chilman
authors.initials: C
authors.lastname: Bae
authors.affiliation: Department of Physiology and Biophysics, State University of New York at Buffalo, Buffalo, New York 14214, USA.
authors.forename: Lynn
authors.initials: L
authors.lastname: Ziegler
authors.affiliation: Department of Physiology and Biophysics, State University of New York at Buffalo, Buffalo, New York 14214, USA.
authors.forename: Silas
authors.initials: S
authors.lastname: Hartley
authors.affiliation: Victor Chang Cardiac Research Institute, Darlinghurst, New South Wales 2010, Australia.
authors.forename: Vesna
authors.initials: V
authors.lastname: Nikolova-Krstevski
authors.affiliation: Victor Chang Cardiac Research Institute, Darlinghurst, New South Wales 2010, Australia.
authors.forename: Paul R
authors.initials: PR
authors.lastname: Rohde
authors.affiliation: Victor Chang Cardiac Research Institute, Darlinghurst, New South Wales 2010, Australia.|St Vincent's Clinical School, University of New South Wales, Darlinghurst, New South Wales 2010, Australia.
authors.forename: Chai-Ann
authors.initials: CA
authors.lastname: Ng
authors.affiliation: Department of Physiology and Biophysics, State University of New York at Buffalo, Buffalo, New York 14214, USA.|The Centre for Single Molecule Biophysics, State University of New York at Buffalo, Buffalo, New York 14214, USA.
authors.forename: Frederick
authors.identifier: http://orcid.org/0000-0002-4324-2240
authors.initials: F
authors.lastname: Sachs
authors.affiliation: Department of Physiology and Biophysics, State University of New York at Buffalo, Buffalo, New York 14214, USA.|The Centre for Single Molecule Biophysics, State University of New York at Buffalo, Buffalo, New York 14214, USA.
authors.forename: Philip A
authors.initials: PA
authors.lastname: Gottlieb
authors.affiliation: Victor Chang Cardiac Research Institute, Darlinghurst, New South Wales 2010, Australia.|St Vincent's Clinical School, University of New South Wales, Darlinghurst, New South Wales 2010, Australia.
authors.forename: Boris
authors.identifier: http://orcid.org/0000-0001-8422-7082
authors.initials: B
authors.lastname: Martinac
mesh_terms: D002470:Cell Survival / Q000502:physiology; D003599:Cytoskeleton / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D007473:Ion Channels / Q000378:metabolism*; D008051:Lipid Bilayers / Q000378:metabolism
chemical_list: D007473:Ion Channels; D008051:Lipid Bilayers; C515204:PIEZO1 protein, human