NAADP mobilizes calcium from acidic organelles through two-pore channels

Ca(2+) mobilization from intracellular stores represents an important cell signaling process 1 which is regulated, in mammalian cells, by inositol 1,4,5-trisphosphate (InsP(3)), cyclic ADP ribose (cADPR) and nicotinic acid adenine dinucleotide phosphate (NAADP). InsP(3) and cADPR release Ca(2+) from sarco / endoplasmic reticulum (S/ER) stores through activation of InsP(3) and ryanodine receptors (InsP(3)Rs and RyRs). By contrast, the nature of the intracellular stores targeted by NAADP and molecular identity of the NAADP receptors remain controversial 1,2, although evidence indicates that NAADP mobilizes Ca(2+) from lysosome-related acidic compartments 3,4. Here we show that two-pore channels (TPCs) comprise a family of NAADP receptors, with TPC1 and TPC3 being expressed on endosomal and TPC2 on lysosomal membranes. Membranes enriched with TPC2 exhibit high affinity NAADP binding and TPC2 underpins NAADP-induced Ca(2+) release from lysosome-related stores that is subsequently amplified by Ca(2+)-induced Ca(2+) release via InsP(3)Rs. Responses to NAADP were abolished by disrupting the lysosomal proton gradient and by ablating TPC2 expression, but only attenuated by depleting ER Ca(2+) stores or blocking InsP(3)Rs. Thus, TPCs form NAADP receptors that release Ca(2+) from acidic organelles, which can trigger additional Ca(2+) signals via S/ER. TPCs therefore provide new insights into the regulation and organization of Ca(2+) signals in animal cells and will advance our understanding of the physiological role of NAADP.