Bidirectional sensitivity of CALHM1 channel to protons from both sides of plasma membrane

Calcium homeostasis modulator 1 (CALHM1), a newly discovered voltage-dependent nonselective ion channel, has drawn attention for its role in neuronal activity and taste sensation. Its sluggish voltage-dependent activation is facilitated by lowering extracellular Ca²⁺ concentration ([Ca²⁺]_e). Here, we investigated the effects of extracellular and intracellular pH (pH_e and pH_i) on human CALHM1. When normalized to the amplitude of the CALHM1 current (I_CALHM1) under whole cell patch clamp at symmetrical pH 7.4, I_CALHM1 decreased at acidic pH_e or pH_i, whereas it sharply increased at alkaline pH_e or pH_i. The effects of pH were preserved in the inside-out configuration. The voltage dependence of I_CALHM1 showed leftward and rightward shifts at alkaline and acidic pH_e and pH_i, respectively. Site-directed mutagenesis of the water-accessible charged residues of the pore and nearby domains revealed that E17, K229, E233, D257, and E259 are nonadditively responsible for facilitation at alkaline pH_i. Identification of the pH_e-sensing residue was not possible because mutation of putative residues impaired membrane expression, resulting in undetectable I_CALHM1. Alkaline pH_e-dependent facilitation appeared gradually with depolarization, suggesting that the sensitivity to pH_e might be due to H⁺ diffusion through the open-state CALHM1. At pH_e 6.2, decreased [Ca²⁺]_e could not recover the inhibited I_CALHM1 but further augmented the increased I_CALHM1 at pH_e 8.6, suggesting that unidentified common residues might contribute to the [Ca²⁺]_e and acidic pH_e. This study is the first, to our knowledge, to demonstrate the remarkable pH sensitivity of CALHM1, which might contribute to the pH-dependent modulation of neuronal excitability or taste sensation.