Lysosomes, the cell's degradation center, are filled with acidic hydrolases. Lysosomes generate nutrient-sensitive signals to regulate the import of H + , hydrolases, and endocytic and autophagic cargos, and the export of the degradation products (catabolites). In response to environmental and cellular signals, lysosomes change their positioning, number, morphology, size, composition, and activity within minutes to hours to meet the changing cellular needs. Ion channels in the lysosome are essential transducers that mediate signal-initiated Ca 2+ /Fe 2+ /Zn 2+ release and H + /Na + /K +dependent changes of membrane potential across the perimeter membrane. Dysregulation of lysosomal ion flux impairs lysosome movement, membrane trafficking, nutrient-sensing, membrane repair, organelle membrane contact, and lysosome biogenesis and adaptation. Hence, activation and inhibition of lysosomal channels by synthetic modulators may tune lysosome function to maintain cellular health and promote cellular clearance in lysosome storage disorders.

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LRRC8 family proteins on the plasma membrane play a critical role in cellular osmoregulation by forming volume-regulated anion channels (VRACs) necessary to prevent necrotic cell death. We demonstrate that intracellular LRRC8 proteins acting within lysosomes also play an essential role in cellular osmoregulation. LRRC8 proteins on lysosome membranes generate large lysosomal volume-regulated anion channel (Lyso-VRAC) currents in response to low cytoplasmic ionic strength conditions. When a double-leucine L706L707 motif at the C terminus of LRRC8A was mutated to alanines, normal plasma membrane VRAC currents were still observed, but Lyso-VRAC currents were absent. We used this targeting mutant, as well as pharmacological tools, to demonstrate that Lyso-VRAC currents are necessary for the formation of large lysosome-derived vacuoles, which store and then expel excess water to maintain cytosolic water homeostasis. Thus, Lyso-VRACs allow lysosomes of mammalian cells to act as the cell`s “bladder.” When Lyso-VRAC current was selectively eliminated, the extent of necrotic cell death to sustained stress was greatly increased, not only in response to hypoosmotic stress, but also to hypoxic and hypothermic stresses. Thus Lyso-VRACs play an essential role in enabling cells to mount successful homeostatic responses to multiple stressors.