Sarcoendoplasmic reticulum calcium ATPase is an essential and druggable lipid-dependent ion pump in Toxoplasma gondii

Toxoplasma gondii is a common intracellular pathogenic protist causing acute and chronic infections in many warm-blooded organisms. Calcium homeostasis is pivotal for its asexual reproduction in mammalian host cells, and sarcoendoplasmic reticulum calcium-ATPase (SERCA) is considered vital for maintaining ion homeostasis within the parasite. This work studied the physiological relevance, structure-function relationship, mechanism, and therapeutic value of SERCA in the acutely-infectious tachyzoite stage of T. gondii. A conditional depletion of SERCA, located in the endoplasmic reticulum, by auxin-inducible degradation is lethal for the parasite due to severe defects in its replication, gliding motility, and invasion. The observed phenotypes are caused by dysregulated calcium ion homeostasis and microneme secretion in the absence of TgSERCA. Furthermore, ectopic expression of TgSERCA restored the lytic cycle of a phosphatidylthreonine-null and phosphatidylserine-enriched mutant with perturbed calcium homeostasis, motility and invasion. These lipids are expressed in the parasite ER, co-localizing with TgSERCA. Last but not least, the structure-function modeling and ligand docking of TgSERCA with a library comprising >5000 chemicals identified two compounds (RB-15, NR-301) that inhibited the lytic cycle by affecting the tachyzoite locomotion, invasion, microneme discharge, and calcium levels. In conclusion, we demonstrate TgSERCA as an indispensable lipid-assisted calcium pump in T. gondii and report small molecules with therapeutic potential against toxoplasmosis.