Spring-like mechanics enable rapid inactivation and stochastic single-channel gating of the mechanically activated...

PIEZO ion channels generate force-induced macroscopic currents that rapidly inactivate but display repetitive single-channel opening and closing under steady membrane tension. The mechanism underlying these gating characteristics remains unresolved. In the force-induced flattened state of PIEZO1, its top cap domain swings between up and down states via the spring-like extension and compression of the connecting linkers, driving the opening and closing of the pore. We identify that the linkers and key residues contribute to the activation and inactivation of the macroscopic current, as well as to the open dwell time and conductance of the single-channel current. Steered molecular dynamics simulations reveal that the linkers behave like entropic springs, whose elastic energy matches the steady-state single-channel gating energy. Taken together, we propose that the linkers function as compressive entropic springs to store and release energy upon their compression and extension, which drives activation, inactivation, as well as stochastic single-channel gating of PIEZO channels.