Li, S.; Wijerathne, T. D.; Bhatt, A.; Jiang, W.; Lacroix, J.; Han, W.; Luo, Y. L.

Mechanosensitive PIEZO channels are thought to open their pore through tension-induced flattening of large transmembrane arm domains. Yet, the structural basis of this activation remains unclear. Here, we uncover the conformational coupling between arm flattening and pore opening in PIEZO2 by capturing protein motions across length scales using hybrid-resolution molecular dynamics simulations. Sampling multiple microsecond-long trajectories under physiological activation tension show that arm flattening correlates with anticlockwise rotation of the pore domain and with clockwise twisting of inner pore helices, enabling dilation and hydration of a transmembrane pore gate. These clockwork motions enable PIEZO2 to populate two open states with distinct conductance depending on applied membrane tension, in agreement with single-channel electrophysiology. Pore opening is accompanied by the separation of pore helices, creating interhelical gaps which become filled with lipids, resulting in the fully conducting pore being walled by both lipids and protein. The fully open PIEZO2 state recapitulates minimal pore size, conductance, ion selectivity, and outward rectification of chloride currents observed electrophysiologically. This work reveals how tension-induced large-scale rearrangements of the PIEZO2 arms funnel into subtle and dynamic gating motions, providing invaluable structural insights for future structure-function and drug discovery studies.