A conserved pressure-driven mechanism for regulating cytosolic osmolarity

Controlling intracellular osmolarity is essential to all cellular life. Cells that live in hypo-osmotic environments, such as freshwater, must constantly battle water influx to avoid swelling until they burst. Many eukaryotic cells use contractile vacuoles to collect excess water from the cytosol and pump it out of the cell. Although contractile vacuoles are essential to many species, including important pathogens, the mechanisms that control their dynamics remain unclear. To identify the basic principles governing contractile vacuole function, we investigate here the molecular mechanisms of two species with distinct vacuolar morphologies from different eukaryotic lineages: the discoban Naegleria gruberi and the amoebozoan slime mold Dictyostelium discoideum. Using quantitative cell biology, we find that although these species respond differently to osmotic challenges, they both use vacuolar-type proton pumps for filling contractile vacuoles and actin for osmoregulation, but not to power water expulsion. We also use analytical modeling to show that cytoplasmic pressure is sufficient to drive water out of contractile vacuoles in these species, similar to findings from the alveolate Paramecium multimicronucleatum. These analyses show that cytoplasmic pressure is sufficient to drive contractile vacuole emptying for a wide range of cellular pressures and vacuolar geometries. Because vacuolar-type proton-pump-dependent contractile vacuole filling and pressure-dependent emptying have now been validated in three eukaryotic lineages that diverged well over a billion years ago, we propose that this represents an ancient eukaryotic mechanism of osmoregulation. [Display omitted] •Vacuolar-type proton pumps contribute to contractile vacuole filling in Naegleria•Actin networks are dispensable for vacuole emptying in Naegleria and Dictyostelium•Cytoplasmic pressure is sufficient to drive emptying in Naegleria and Dictyostelium•Modeling shows pressure can drive emptying in a wide variety of cells Contractile vacuoles—organelles that collect water and pump it out of cells—are essential for osmoregulation in species from across the eukaryotic tree. Despite their variable morphology, Velle et al. show that mechanisms used by Paramecium also drive contractile vacuole activity in Naegleria and Dictyostelium.