Lysosome Ca2+ Store Refilling Mechanisms.

Abstract

Lysosomes are acidic intracellular vesicles containing hydrolases that degrade intracellular and extracellular debris delivered through endocytic trafficking and autophagy. Lysosome function requires the establishment of luminal ionic homeostasis for ions including H+ and Ca2+, which are 1,000-5,000 times more concentrated in the lysosome lumen than in the cytosol. Lysosomal H+ homeostasis is required to activate hydrolases and Ca2+ efflux through lysosomal ion channels serves as signals required for precise delivery of hydrolases and cargo and the timely removal of catabolites. Impaired lysosomal Ca2+ homeostasis results in lysosomal dysfunction, lysosomal storage diseases (LSDs), and has been implicated more broadly in neurodegenerative phenotypes. The molecular mechanisms by which lysosomes acquire and refill Ca2+ are unknown. We developed a physiological assay to monitor lysosomal Ca2+ store refilling using specific activators of lysosomal Ca2+ channel TRPML1 to repeatedly induce lysosomal Ca2+ release. In contrast to the prevailing view that lysosomal acidification drives Ca2+ into the lysosome, inhibiting the V-ATPase H+ pump did not prevent Ca2+ refilling. Instead, pharmacological and genetic depletion or chelation of endoplasmic reticulum (ER) Ca2+ prevented lysosomal Ca2+ stores from refilling. More specifically, antagonists of ER IP3 receptors rapidly and completely blocked Ca2+ refilling to lysosomes. Reducing ER Ca2+ or blocking IP3 receptors resulted in a dramatic lysosome storage phenotype. By closely apposing each other, the ER may serve as a direct and primary source of Ca2+ to the lysosome. These findings may clarify seemingly overlapping ER and lysosome Ca2+ stores in some studies and shed light on why ER Ca2+ homeostasis is often involved in LSDs and neurodegenerative diseases.