The voltage-gated sodium channel TPC1 confers endolysosomal excitability

Abstract

The physiological function and molecular regulation of plasma membrane potential have been extensively studied, but how intracellular organelles sense and control membrane potential is not well understood. Using whole-organelle patch clamp recording, we show that endosomes and lysosomes are electrically excitable organelles. In a subpopulation of endolysosomes, a brief electrical stimulus elicits a prolonged membrane potential depolarization spike. The organelles have a previously uncharacterized, depolarization-activated, noninactivating Na(+) channel (lysoNaV). The channel is formed by a two-repeat six-transmembrane-spanning (2×6TM) protein, TPC1, which represents the evolutionary transition between 6TM and 4×6TM voltage-gated channels. Luminal alkalization also opens lysoNaV by markedly shifting the channel's voltage dependence of activation toward hyperpolarization. Thus, TPC1 is a member of a new family of voltage-gated Na(+) channels that senses pH changes and confers electrical excitability to organelles.

The lysosomal Ca2+ release channel TRPML1 regulates lysosome size by activating calmodulin

“…The percentage of P2X4-expressing cells with enlarged lysosomes (induced by MA) was decreased from 79.00 Ϯ 2.64 to 30.00 Ϯ 2.66% by TRPML1 co-expression. In contract, co-expression of two-pore channel 2 (TPC2), a Na ϩ release channel (37)(38)(39), and TRPML1-DDKK, a non-conducting TRPML1 mutant (21), did not increase the percentage of cells with enlarged lysosomes induced by MA in P2X4-expressing cells (Fig. 3B), suggesting that lysosome size is specifically controlled by TRPML1.…”

Two-pore channels (TPCs): Novel voltage-gated ion channels with pleiotropic functions

“…15 A decrease in cytoplasmic free Mg 2C is suggested to depolarize the lysosomal membrane potential to affect the transport of all electrogenic coupled and uncoupled transporters, including facilitation of Ca 2C release from the lysosomes. 15 In this line, TPC1 and TPC3 have been probed to be regulated by membrane potential, 11,14,16,63 whereas TPC2 seems to be independent of membrane potential changes, 12,14,72 in spite of the presence of the positively charged voltagesensing motifs in the S4 transmembrane domain. And some studies have reported that acidic luminal pH activates TPCs, 11,14,72 however, in others it was found that acidic pH inhibits TPCs opening.…”

“…54 The three types of TPC have positively charged voltage-sensing motifs in the S4 transmembrane domain characteristic of the VGICs superfamily, however, it was suggested that only TPC1 and TPC3 are regulated by voltage, 14,63 whereas TPC2 is not. 8,16 TPC1 mRNA transcripts have a molecular weight around 5 kb and those for TPC2 around 3 kb in murine (Table 1). 1,60 However, northern blot and RT-PCR analysis of TPC1 expression in rats and mice confirmed the presence of 2 TPC1 transcripts with different size, being now referred as TPC1A (the first characterized isoform, » 5 kb) and TPC1B (a smaller isoform, » 4 kb).…”

“…And some studies have reported that acidic luminal pH activates TPCs, 11,14,72 however, in others it was found that acidic pH inhibits TPCs opening. 12,16 TPCs have been also suggested to function as nutrient sensors linked to mTOR action: nutrient replete cells have high adenosine triphosphate (ATP), which presumably enables mTOR to phosphorylate TPCs and/or its associated proteins and maintain the channel in a closed state. During cell starvation, ATP levels fall, mTOR delocalizes from TPCs, and the channel opens.…”

“…4 Still, on the other hand, in 2012 and 2013 2 independent studies refuted that mammalian TPCs were Ca 2C release channels activated by NAADP, probing that they are not Ca 2C but Na C release channels that are not activated by NAADP but by phosphatidylinositol 3,5-bisphosphate (PtdIns(3,5)P2) and inhibited by the mammalian target of rapamycin (mTOR), 5,6 which caused a great commotion regarding TPCs regulation and function among the scientific community. [7][8][9][10] Nowadays, it is accepted that mammalian TPCs not only function as Ca 2C or Na C release channels, but also as H C and K C channels, 11,12 and it has been demonstrated that TPCs can be activated by other signals apart from NAADP and PI (3,5)P2, such as the leucine-rich repeat kinase 2 (LRRK2) 13 or action potentials, 14 and inhibited by Mg 2C concentrations, 15 Ca 2C and Na C ion channels inhibitors, 16,17 or c-Jun N-terminal kinase (JNK) and p38 kinase, 15 apart from mTOR. So that, it seems reasonable that, according to the cellular context, TPCs could be differentially regulated and exert different functions.…”