Abstract
The proton–activated chloride (PAC) channel plays critical roles in ischemic neuron death, but its activation mechanisms remain elusive. Here, we investigated the gating of PAC channels using its novel bifunctional modulator C77304. C77304 acted as a weak activator of the PAC channel, causing moderate activation by acting on its proton gating. However, at higher concentrations, C77304 acted as a weak inhibitor, suppressing channel activity. This dual function was achieved by interacting with 2 modulatory sites of the channel, each with different affinities and dependencies on the channel’s state. Moreover, we discovered a protonation–independent voltage activation of the PAC channel that appears to operate through an ion–flux gating mechanism. Through scanning–mutagenesis and molecular dynamics simulation, we confirmed that E181, E257, and E261 in the human PAC channel serve as primary proton sensors, as their alanine mutations eliminated the channel’s proton gating while sparing the voltage–dependent gating. This proton–sensing mechanism was conserved among orthologous PAC channels from different species. Collectively, our data unveils the polymodal gating and proton–sensing mechanisms in the PAC channel that may inspire potential drug development.
Funder
National Natural Science Foundation of China
the Science and Technology Innovation Program of Hunan Province
the Natural Science Foundation of Hunan Province
the Research Foundation of the Education Department of Hunan Province
the Fundamental Research Funds for the Central Universities
High-performance Computing Center (HPCC) of Nanjing University
Publisher
Public Library of Science (PLoS)
Subject
General Agricultural and Biological Sciences,General Immunology and Microbiology,General Biochemistry, Genetics and Molecular Biology,General Neuroscience