Calreticulin (CALR) promotes ionophore-induced microneme secretion in Toxoplasma gondii

Abstract

The flow of calcium ions (Ca²⁺) is involved in numerous vital activities of Toxoplasma gondii. Calreticulin is a type of Ca²⁺-binding protein in the endoplasmic reticulum (ER) that is involved in Ca²⁺ signaling pathway regulation, Ca²⁺ storage, and protein folding. In this work, the calreticulin (CALR), a protein predicted to possess a conserved domain of calreticulin in T. gondii, was characterized. The CALR localized in the ER. Using reverse genetics, we discovered that CALR is not necessary for the lytic cycle, including invasion and replication. However, depletion of CALR affected microneme secretion triggered by A23187, which is a Ca²⁺ ionophore used to increase cytoplasmic Ca²⁺ concentration. Furthermore, we discovered that CALR influences Ca²⁺ release. Transcriptomic comparison between Δcalr and Δku80 parasites showed that 226 genes in the Δcalr parasites were significantly downregulated (p < 0.05). The cellular biological functions of the downregulated genes were mainly involved in calmodulin-dependent protein kinase pathways. Furthermore, in the absence of CALR, tachyzoites were still able to cause acute infection in mice. These results imply that by influencing ER Ca²⁺ release content, CALR may further impair the ionophore-induced secretion of the parasite. However, this protein is not required for the completion of the parasite's lytic cycle or for the acute virulence of the parasite.

Keywords: Calreticulin; Ionophore-induced secretion; Microneme proteins; Toxoplasma gondii.