The Dually Localized EF-Hand Domain-Containing Protein TgEFP1 Regulates the Lytic Cycle of .

The propagation of the obligate intracellular parasite is tightly regulated by calcium signaling. However, the mechanisms by which calcium homeostasis and fluxes are regulated in this human pathogen are not fully understood. To identify Toxoplasma's calcium homeostasis network, we have characterized a novel EF-hand domain-containing protein, which we have named TgEFP1. We have determined that TgEFP1 localizes to a previously described compartment known as the plant-like vacuole or the endosomal-like compartment (PLV/ELC), which harbors several proteins related to ionic regulation. Interestingly, partial permeabilization techniques showed that TgEFP1 is also secreted into the parasitophorous vacuole (PV), within which the parasite divides. Ultrastructure expansion microscopy confirmed the unusual dual localization of TgEFP1 at the PLV/ELC and the PV. Furthermore, we determined that the localization of TgEFP1 to the PV, but not to the PLV/ELC, is affected by disruption of Golgi-dependent transport with Brefeldin A. Knockout of TgEFP1 results in faster propagation in tissue culture, hypersensitivity to calcium ionophore-induced egress, and premature natural egress. Thus, our work has revealed an interplay between the PV and the PLV/ELC and a role for TgEFP1 in the regulation of calcium-dependent events.