Spatially Resolved Mapping of Voltage‐Gated Proton Channel Activity Reveals Delayed Proton Transport in Local Microenvironments

Voltage-gated proton channel Hv1 is a proton-selective channel that lacks the pore domain typically found in other ion channels, and it is thought to participate in signal transduction by inducing transient, localized changes in proton concentration. However, the contribution of Hv1 channel distribution and activity to the overall pH homeostatic process is still not well understood. Here a single-molecule photobleaching-based quantitative imaging method is developed with an Hv1-fused ratiometric pHluorin probe to study the distribution and dynamics of the proton channel in living bacteria. It is shown that the Hv1 channel does not close sharply when voltage is removed and can directly cause local fluctuations of intracellular pH gradient. Especially, the proton transport through Hv1 is significantly delayed in hypertonic environments, offering novel perspectives at cellular level for neuropathological research. This spatially localized pH microenvironment created by the Hv1 channel may play a crucial role in signal transduction in neuron cells.