TPC1 is a Proton Permeable Channel that can be Independently Activated by Cytosolic Calcium or NAADP

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Nicotinic acid adenine dinucleotide phosphate (NAADP) potently releases Ca2+ from acidic intracellular lysosomal Ca2+-stores. There is evidence that two-pore channels (TPCs), a family of ion-channels localised to the acidic stores, are involved in NAADP-mediated Ca2+-release. We have shown that human TPC type-2 (TPC2) possesses certain key biophysical properties that suggest it may function as a lysosomal NAADP-sensitive Ca2+-release channel [1]. We now investigate whether two-pore channel type-1 (TPC1) exhibits similar functional properties by reconstituting purified human TPC1 into artificial membranes under voltage-clamp conditions. Similar to TPC2, we find that TPC1 is impermeable to Cl- but is poorly selective among cations with conductance decreasing in the order K+>Na+>Ca2+. We also find that, unlike TPC2, TPC1 is highly permeable to protons; under bi-ionic conditions, relative permeability decreases in the order H+>>K+>Na+>Ca2+. Significantly, we show that the regulation of TPC1 gating is very different to that of TPC2. We previously demonstrated that TPC2 activity is not regulated by cytosolic Ca2+ [2] but here find that TPC1 can be activated by nanomolar concentrations of NAADP alone or by cytosolic Ca2+ alone. This fundamental difference in channel regulation will enable TPC1 to be activated independently from TPC2 allowing the endo-lysosomal system to respond to physiological needs by integrating a variety of cellular signals. We suggest that in the endolysosomal and lysosomal systems, TPC2 can effectively function as a NAADP-activated Ca2+-release channel. In contrast, TPC1 can leak protons from these acidic Ca2+-stores and although TPC1 is capable of providing some degree of Ca2+-release, this is heavily dependent on K+ and Na+ gradients across the lysosomal membrane, on the lysosomal membrane potential, the luminal [Ca2+] and the luminal pH.[1].Pitt et al.,(2010)JBC;285:35039-46.[2].Pitt et al.,(2011)Biophys.J;100:433a.BHF and Royal Society Edinburgh funded.