Force-induced motions of the PIEZO1 blade probed with fluorimetry

• Published in: Cell reports (Cambridge) 2023-08, Vol.42 (8), p.112837-112837, Article 112837
• Main Authors: Ozkan, Alper D., Wijerathne, Tharaka D., Gettas, Tina, Lacroix, Jérôme J.
• Format: Article
• Language: English
• Subjects: conformational changes; CP: Molecular biology; cpGFP; cyclic permuted GFP; fluid shear stress; mechanotransduction; PIEZO1; site-specific fluorimetry
• ISSN: 2211-1247; 2211-1247
• DOI: 10.1016/j.celrep.2023.112837

Mechanical forces are thought to activate mechanosensitive PIEZO channels by changing the conformation of a large transmembrane blade domain. Yet, whether different stimuli induce identical conformational changes in this domain remains unclear. Here, we repurpose a cyclic permuted green fluorescent protein as a conformation-sensitive probe to track local rearrangements along the PIEZO1 blade. Two independent probes, one inserted in an extracellular site distal to the pore and the other in a distant intracellular proximal position, elicit sizable fluorescence signals when the tagged channels activate in response to fluid shear stress of low intensity. Neither cellular indentations nor osmotic swelling of the cell elicit detectable fluorescence signals from either probe, despite the ability of these stimuli to activate the tagged channels. High-intensity flow stimuli are ineffective at eliciting fluorescence signals from either probe. Together, these findings suggest that low-intensity fluid shear stress causes a distinct form of mechanical stress to the cell. [Display omitted] •Optical probes track PIEZO1 conformational changes at two distant blade positions•Two independent probes respond to flow, not indentations or osmotic swelling•Fluorescence signals from both probes correlate with flow-induced channel opening•Both probes respond to fluid shear stress stimuli of low, but not high, intensity Ozkan et al. use conformation-sensitive fluorescent probes to track force-induced rearrangements of PIEZO1’s mechanosensory blade domains. Two probes inserted far apart in the blade independently “light up” as channels activate in response to low-intensity fluid shear stress but not to other tested mechanical stimuli.