Rearrangements of Piezo1 blade domains correlate with pore opening

Mechanosensitive Piezo channels are anticipated to open their pore by flattening of large transmembrane domains called blades. Yet, direct experimental evidence for the coupling between blade motions and pore opening remains sparse. Here, we report unambiguous correlations between flow-induced Piezo1 opening and fluorimetric signals from conformation-sensitive probes genetically-inserted at two blade locations, one extracellular distal and the other intracellular proximal. Inhibition of gating motions near the pore with disulfide crosslinks reduced the amplitude of fluorescence signals from the most distal probe, consistent with long-range pore-blade conformational coupling. Interestingly, both probes remained fluorimetrically silent when Piezo1 opening was evoked with hypotonic shocks, cellular indentations, or application of the chemical activator Yoda1. This work provides direct experimental evidence that the Piezo1 blades act as mechanosensory domains and suggests that different mechanical stimuli activate Piezo1 through distinct mechanotransduction pathways. ### Competing Interest Statement The authors have declared no competing interest.