The NAADP/ TPC2/Ca2+ Signaling Antagonizes Autophagosome Maturation

Abstract

Autophagy is an evolutionarily conserved lysosomal degradation pathway, yet the underlying mechanisms remains poorly understood. Nicotinic acid adenine dinucleotide phosphate (NAADP) elicits Ca2+ release from lysosomes via two pore channel 2 (TPC2) in many cell types. Here we found that overexpression of TPC2 in Hela cells, which lack the expression of endogenous TPC2, inhibited autophagosomal‐lysosomal fusion by raising lysosomal pH. Treatment of TPC2 overexpressing cells with a cell permeant NAADP agonist, NAADP‐AM, further inhibited the fusion, whereas Ned‐19, a NAADP antagonist, promoted the fusion. Moreover, intracellular acidification mitigated the fusion blockage in TPC2 overexpressing cells. Interestingly, autophagy is gradually induced during in vitro neural differentiation of mouse embryonic stem (ES) cells initiated by the monolayer culture, accompanying with the marked downregulation of endogenous TPC2. Consistently, TPC2 knockdown or overexpression in mouse ES cells promoted or inhibited the fusion between autophagosome and lysosome during early neural differentiation, respectively. Correspondingly, TPC2 knockdown accelerated mouse ES cells entry into neural lineages, whereas TPC2 overexpression in ES cells markedly inhibited it. The development of novel agonists or antagonists of NAADP should be a good approach to control the maturation processes of the autophagosome.