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

Acid-sensing ion channels (ASICs) are neuronal H ⁺ -gated, Na ⁺ -permeable channels involved in learning, fear sensing, pain sensation and neurodegeneration. An increase in the extracellular Ca ²⁺ concentration shifts the pH dependence of ASIC1a to more acidic values. Here, we predicted candidate residues for Ca ²⁺ binding on ASIC1a, based on available structural information and molecular dynamics simulations; the function of channels carrying mutations of these residues was then measured. We identify several residues in cavities previously associated with pH-dependent gating, whose mutation decreased the Ca ²⁺ -induced shift in ASIC1a pH dependence, likely due to a disruption of Ca ²⁺ binding. We show also that Mg ²⁺ shares some of the binding sites with Ca ²⁺ , and that some of the 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.