Convergent activation of Ca²⁺ permeability in two-pore channel 2 through distinct molecular routes

TPC2 is a pathophysiologically relevant lysosomal ion channel that is activated directly by the phosphoinositide PI(3,5)P₂ and indirectly by the calcium ion (Ca²⁺)-mobilizing molecule NAADP through accessory proteins that associate with the channel. TPC2 toggles between PI(3,5)P₂-induced, sodium ion (Na⁺)-selective and NAADP-induced, Ca²⁺-permeable states in response to these cues. To address the molecular basis of polymodal gating and ion-selectivity switching, we investigated the mechanism by which NAADP and its synthetic functional agonist, TPC2-A1-N, induced Ca²⁺ release through TPC2 in human cells. Whereas NAADP required the NAADP-binding proteins JPT2 and LSM12 to evoke endogenous calcium ion signals, TPC2-A1-N did not. Residues in TPC2 that bind to PI(3,5)P₂ were required for channel activation by NAADP but not for activation by TPC2-A1-N. The cryptic voltage-sensing region of TPC2 was required for the actions of TPC2-A1-N and PI(3,5)P₂ but not for those of NAADP. These data mechanistically distinguish natural and synthetic agonist action at TPC2 despite convergent effects on Ca²⁺ permeability and delineate a route for pharmacologically correcting impaired NAADP-evoked Ca²⁺ signals.