Ca²⁺ entry at the plasma membrane and uptake by acidic stores is regulated by the activity of the V‐H⁺‐ATPase in <i>Toxoplasma gondii</i>

Abstract

Ca²⁺ is a universal intracellular signal that regulates many cellular functions. In Toxoplasma gondii, the controlled influx of extracellular and intracellular Ca²⁺ into the cytosol initiates a signaling cascade that promotes pathogenic processes like tissue destruction and dissemination. In this work, we studied the role of proton transport in cytosolic Ca²⁺ homeostasis and the initiation of Ca²⁺ signaling. We used a T. gondii mutant of the V-H⁺ -ATPase, a pump previously shown to transport protons to the extracellular medium, and to control intracellular pH and membrane potential and we show that proton gradients are important for maintaining resting cytosolic Ca²⁺ at physiological levels and for Ca²⁺ influx. Proton transport was also important for Ca²⁺ storage by acidic stores and, unexpectedly, the endoplasmic reticulum. Proton transport impacted the amount of polyphosphate (polyP), a phosphate polymer that binds Ca²⁺ and concentrates in acidocalcisomes. This was supported by the co-localization of the vacuolar transporter chaperone 4 (VTC4), the catalytic subunit of the VTC complex that synthesizes polyP, with the V-ATPase in acidocalcisomes. Our work shows that proton transport regulates plasma membrane Ca²⁺ transport and control acidocalcisome polyP and Ca²⁺ content, impacting Ca²⁺ signaling and downstream stimulation of motility and egress in T. gondii.