Protonation of Human PIEZO1 Ion Channel Stabilizes Inactivation [Molecular Biophysics]

January 5th, 2015 by Bae, C., Sachs, F., Gottlieb, P. A.

PIEZO1 is a recently cloned eukaryotic cation selective channel that opens with mechanical force. We found that extracellular protonation inhibits channel activation by ≈ 90% by increased occupancy in the closed or inactivated states. Titration between pH 6.3 and 8.3 exhibited a pK of ≈ 6.9. The steepness of the titration data suggests positive cooperativity, implying the involvement of at least two protonation sites. Whole-cell recordings yielded results similar to patches and pH 6.5 reduced whole-cell currents by > 80%. The effects were reversible. To assess whether pH acted on the open or inactivated state, we tested a doubly mutated PIEZO1 that does not inactivate. Cell-attached patches and whole-cell currents from this mutant channel were pH insensitive. Thus, protonation appears to be associated with domain(s) of the channel involved with inactivation. pH also did not affect mutant channels with point mutations at position 2456 that are known to exhibit slow inactivation. To determine if the physical properties of the membrane were altered by pH and thereby affected channel gating, we measured patch capacitance during mechanical stimuli at pH 6.5 and 7.3. The rate constants for changes in patch capacitance were independent of pH suggesting that bilayer mechanics were not involved. In summary, low pH stabilizes the inactivated state. This effect may be important when channels are activated in pathologic conditions where the pH is reduced such as during ischemia.