Identification of the modulatory Ca ²⁺ -binding sites of acid-sensing ion channel 1a

Abstract

Acid-sensing ion channels (ASICs) are neuronal Na + -permeable ion channels activated by extracellular acidification. ASICs are involved in learning, fear sensing, pain sensation and neurodegeneration. Increasing the extracellular Ca 2+ concentration decreases the H + sensitivity of ASIC1a, suggesting a competition for binding sites between H + and Ca 2+ ions. Here, we predicted candidate residues for Ca 2+ binding on ASIC1a, based on available structural information and our molecular dynamics simulations. With functional measurements, we identified several residues in cavities previously associated with pH-dependent gating, whose mutation reduced the modulation by extracellular Ca 2+ of the ASIC1a pH dependence of activation and desensitization. This occurred probably owing to a disruption of Ca 2+ binding. Our results link one of the two predicted Ca 2+ -binding sites in each ASIC1a acidic pocket to the modulation of channel activation. Mg 2+ regulates ASICs in a similar way as does Ca 2+ . We show that Mg 2+ shares some of the binding sites with Ca 2+ . Finally, we provide evidence that some of the ASIC1a Ca 2+ -binding sites are functionally conserved in the splice variant ASIC1b. Our identification of divalent cation-binding sites in ASIC1a shows how Ca 2+ affects ASIC1a gating, elucidating a regulatory mechanism present in many ion channels.