TRPML1-Induced Lysosomal Ca²⁺ Signals Activate AQP2 Translocation and Water Flux in Renal Collecting Duct Cells

Lysosomes are acidic Ca²⁺ storage organelles that actively generate local Ca²⁺ signaling events to regulate a plethora of cell functions. Here, we characterized lysosomal Ca²⁺ signals in mouse renal collecting duct (CD) cells and we assessed their putative role in aquaporin 2 (AQP2)-dependent water reabsorption. Bafilomycin A1 and ML-SA1 triggered similar Ca²⁺ oscillations, in the absence of extracellular Ca²⁺, by alkalizing the acidic lysosomal pH or activating the lysosomal cation channel mucolipin 1 (TRPML1), respectively. TRPML1-dependent Ca²⁺ signals were blocked either pharmacologically or by lysosomes' osmotic permeabilization, thus indicating these organelles as primary sources of Ca²⁺ release. Lysosome-induced Ca²⁺ oscillations were sustained by endoplasmic reticulum (ER) Ca²⁺ content, while bafilomycin A1 and ML-SA1 did not directly interfere with ER Ca²⁺ homeostasis per se. TRPML1 activation strongly increased AQP2 apical expression and depolymerized the actin cytoskeleton, thereby boosting water flux in response to an hypoosmotic stimulus. These effects were strictly dependent on the activation of the Ca²⁺/calcineurin pathway. Conversely, bafilomycin A1 led to perinuclear accumulation of AQP2 vesicles without affecting water permeability. Overall, lysosomal Ca²⁺ signaling events can be differently decoded to modulate Ca²⁺-dependent cellular functions related to the dock/fusion of AQP2-transporting vesicles in principal cells of the CD.