mTORC1 controls lysosomal Ca ²⁺ release through the two-pore channel TPC2

Abstract

Two-pore segment channel 2 (TPC2) is a ubiquitously expressed, lysosomally targeted ion channel that aids in terminating autophagy and is inhibited upon its association with mechanistic target of rapamycin (mTOR). It is controversial whether TPC2 mediates lysosomal Ca2+ release or selectively conducts Na+ and whether the binding of nicotinic acid adenine dinucleotide phosphate (NAADP) or phosphatidylinositol 3,5-bisphosphate [PI(3,5)P2] is required for the activity of this ion channel. We show that TPC2 is required for intracellular Ca2+ signaling in response to NAADP or to mTOR inhibition by rapamycin. In pulmonary arterial myocytes, rapamycin and NAADP evoked global Ca2+ transients that were blocked by depletion of lysosomal Ca2+ stores. Preincubation of cells with high concentrations of rapamycin resulted in desensitization and blocked NAADP-evoked Ca2+ signals. Moreover, rapamycin and NAADP did not evoke discernable Ca2+ transients in myocytes derived from Tpcn2 knockout mice, which showed normal responses to other Ca2+-mobilizing signals. In HEK293 cells stably overexpressing human TPC2, shRNA-mediated knockdown of mTOR blocked rapamycin- and NAADP-evoked Ca2+ signals. Confocal imaging of a genetically encoded Ca2+ indicator fused to TPC2 demonstrated that rapamycin-evoked Ca2+ signals localized to lysosomes and were in close proximity to TPC2. Therefore, inactivation of mTOR may activate TPC2 and consequently lysosomal Ca2+ release.

Energetic adaptations: Metabolic control of endocytic membrane traffic

“…In addition to control of lysosomal expansion through regulated transcription and translation, mTORC1 may also control various aspects of lysosomal membrane dynamics. mTORC1 controls lysosomal Ca 2+ dynamics, either by control of Ca 2+ channels TPC2 or mucolipin 1 (MCOLN1), and upregulation of MCOLN1 activity during starvation is essential for lysosomal adaptation to enhance degradative capacity . As localized Ca 2+ signals control lysosomal fusion (Figure ), this suggests that mTORC1 may modulate lysosomal fusion to coordinate nutrient supply, although this phenomenon remains to be fully elucidated.…”

The protein interaction networks of mucolipins and two-pore channels

“…Through recording primary peritoneal macrophages, cardiomyocytes, fibroblasts, hepatocytes, and TPC1/TPC2-overexpressing HEK293 cells, the authors demonstrated endolysosomal ATP-sensitive Na + currents to be attributable to activity of TPC1 and TPC2, but not TRPML1 (TRPML1 and mTOR are discussed in Section 1.3) [33]. Ogunbayo et al [21] recently demonstrated mTORC1 to not only regulate lysosomal Na + release, but also TPC2/NAADP-mediated Ca 2+ release both in pulmonary arterial smooth muscle cells (PASMCs) and stably expressing HEK293 cells [21]. Indeed, rapamycin elicited similar Ca 2+ signals as NAADP in wild-type PASMCs through lysosomal Ca 2+ release, while neither NAADP nor rapamycin evoked similar signals in Tpc2 −/− PAMSCs [21].…”

“…Ogunbayo et al [21] recently demonstrated mTORC1 to not only regulate lysosomal Na + release, but also TPC2/NAADP-mediated Ca 2+ release both in pulmonary arterial smooth muscle cells (PASMCs) and stably expressing HEK293 cells [21]. Indeed, rapamycin elicited similar Ca 2+ signals as NAADP in wild-type PASMCs through lysosomal Ca 2+ release, while neither NAADP nor rapamycin evoked similar signals in Tpc2 −/− PAMSCs [21]. While TPC2 sensitivity to ATP requires mTORC1 kinase activity, the mTOR target site on TPC2 remains uncharacterised.…”

“…For example, both channel families are gated by the endolysosomal phosphoinositide PI(3,5)P 2 , conferring Na + currents upon activation. TRPML channels are widely accepted to be Ca 2+ permeable, yet the recent discovery of NAADP-activated TPC-mediated Ca 2+ currents remains heavily debated, as at least two groups claim that TPCs are predominantly Na + permeable channels activated by PI(3,5)P 2 , while several other groups show that TPCs are also Ca 2+ permeable and can be activated by both PI(3,5)P 2 and NAADP [15][16][17][18][19][20][21]. Resolution of these ongoing debates is of particular importance, as the channels are proposed therapeutic targets to treat disorders such as cancer [22,23], neurodegeneration [24][25][26][27], metabolic/cardiovascular disorders [12,28], and infectious diseases [29][30][31][32].…”

Membrane transport proteins in melanosomes: Regulation of ions for pigmentation

“…Both NAADP and PI(3,5)P 2 are intracellular second messengers in response to various stimuli. NAADP-sensing TPC2 mediates Ca 2+ release in lysosome and endolysosome [58,59,[110][111][112][113][114]. PI(3,5)P 2 -dependent TPC activation induces Na + release [105,[115][116][117][118].…”