PI(3,5)P₂ controls vacuole potassium transport to support cellular osmoregulation

Lysosomes are dynamic organelles with critical roles in cellular physiology. The lysosomal signaling lipid phosphatidylinositol 3,5-bisphosphate (PI(3,5)P₂) is a key regulator that has been implicated to control lysosome ion homeostasis, but the scope of ion transporters targeted by PI(3,5)P₂ and the purpose of this regulation is not well understood. Through an unbiased screen in Saccharomyces cerevisiae, we identified loss-of-function mutations in the vacuolar H⁺-ATPase (V-ATPase) and in Vnx1, a vacuolar monovalent cation/proton antiporter, as suppressor mutations that relieve the growth defects and osmotic swelling of vacuoles (lysosomes) in yeast lacking PI(3,5)P₂. We observed that depletion of PI(3,5)P₂ synthesis in yeast causes a robust accumulation of multiple cations, most notably an ∼85 mM increase in the cellular concentration of potassium, a critical ion used by cells to regulate osmolarity. The accumulation of potassium and other cations in PI(3,5)P₂-deficient yeast is relieved by mutations that inactivate Vnx1 or inactivate the V-ATPase and by mutations that increase the activity of a vacuolar cation export channel, Yvc1. Collectively, our data demonstrate that PI(3,5)P₂ signaling orchestrates vacuole/lysosome cation transport to aid cellular osmoregulation.