TPC Proteins Are Phosphoinositide- Activated Sodium-Selective Ion Channels in Endosomes and Lysosomes

Abstract

Summary Mammalian Two-Pore Channels (TPC1, 2; TPCN1, TPCN2) encode ion channels in intracellular endosomes and lysosomes and were proposed to mediate endolysosomal calcium release triggered by the second messenger, nicotinic acid adenine dinucleotide phosphate (NAADP). By directly recording TPCs in endolysosomes from wild-type and TPC double knockout mice, here we show that, in contrast to previous conclusions, TPCs are in fact sodium-selective channels activated by PI(3,5)P2, and are not activated by NAADP. Moreover, the primary endolysosomal ion is Na+, not K+, as had been previously assumed. These findings suggest that the organellar membrane potential may undergo large regulatory changes, and may explain the specificity of PI(3,5)P2 in regulating the fusogenic potential of intracellular organelles.

High susceptibility to fatty liver disease in two-pore channel 2-deficient mice

“…Given the proposed close structural assembly of TPCs and the fusion machinery ( Supplementary Fig. 5g), depolarization by Na þ efflux through TPC2 may also electrostatically facilitate fusion between two vesicular membranes as suggested by the model of Wang et al 17 Similar events may be relevant for TPC1 within the proximal endosomal pathway. Defects in the endolysosomal trafficking and cellular processing of LDL-cholesterol have been implicated in human diseases such as Niemann Pick disease and Wolman's disease (CE storage disease) where CEs accumulate in late endosomes and lysosomes due to a severely diminished activity of cholesterol export proteins NPC1 and NPC2 or lysosomal acid lipase 54 .…”

“…In this scenario, cations other than Ca 2 þ flowing through TPC2 would act on a downstream channel, which in turn provides Ca 2 þ . Na þ and K þ have also been shown to permeate through TPC2 channels 17,20 . Given the proposed close structural assembly of TPCs and the fusion machinery ( Supplementary Fig.…”

“…1a). This targeting strategy results in non-functional ion-channel proteins and is different from the strategies used for two mouse lines published before 17,18 . The deletion of the TPC2 gene was confirmed by genomic DNA analysis using Southern blot (not shown), PCR strategies, immunocytochemistry and western blotting ( Supplementary Fig.…”

“…There is evidence that TPC1 is mainly present in the proximal endosomal system, and TPC2 is predominantly expressed on late endosomes and lysosomes 16,18,19 . The activation mechanism of TPCs is complex and has been suggested to involve the second messenger nicotinic acid adenine dinucleotide phosphate (NAADP) and the endolysosomal membrane lipidphosphatidylinositol (3,5)bisphosphate (PI(3,5)P 2 ) 17,18,[20][21][22][23][24] . Independent of what the nature of the endogenous ligand of these channels might be, there is substantial evidence that on activation TPCs mediate the release of Ca 2 þ from lysosomal stores.…”

“…Interesting candidate proteins, which might regulate endolysosomal trafficking, and fusion processes are cation channels present in the membrane of endolysosomal vesicles. Recently identified cation channels that may generate Ca 2 þ signals during endolysosomal fusion events and thereby vesicle trafficking are the two-pore channels (TPCs) 16,17 . Two different TPC subtypes exist in mouse and man, TPC1 and TPC2.…”