Abstract
Protein serine/threonine phosphatases (PSPs), found in various plants and protozoa, are involved in the regulation of various biological processes. However, very little is known about the role of PSPs in the pathogenicity of the apicomplexan protozoan Toxoplasma gondii. Herein, the subcellular localization of 17 PSPs (PP5, PP7, EFPP, SLP, PPM3F, PPM4, PPM5A, PPM5B, PPM6, PPM8, PPM9, PPM12, PPM14, PPM18, CTD1, CTD2, and CTD3) was examined by 6× HA tagging of endogenous genes in C-terminal. The PSPs were detected in the cytoplasm (PP5, EFPP, PPM8, and CTD2), dense granules (SLP), nucleus (PPM4 and PPM9), inner membrane complex (PPM12), basal complex (CTD3), and apical pole (PP7). The remaining PSPs exhibited low or undetectable level of expression. To characterize the contribution of these genes to the infectivity of T. gondii, knock-out (KO) strains of type I RH strain deficient in the 17 psp genes and KO type II Pru strain deficient in pp7 and slp genes were constructed. The pathogenicity of individual RHΔpsp mutants was characterized in vitro using plaque, egress, and intracellular replication assays, and mouse infection, while pathogenicity of PruΔpp7 and PruΔslp mutant strains was evaluated by examining the parasite lytic cycle in vitro and assessment of brain cyst burden in mice. No significant differences were observed between 16 RHΔpsp strains and wild-type (WT) RH strain. However, RHΔpp7 exhibited significantly lower invasion efficiency and parasitophorous vacuole formation in vitro, and less virulence in mice compared with other RHΔpsp and WT strains. In addition, PruΔpp7 exhibited marked attenuation of virulence and significant reduction in the brain cyst burden in mice compared with PruΔslp and WT strains, suggesting the key role of PP7 in the virulence of T. gondii. Comparative transcriptomic profiling of the 17 psp genes showed that they may play different roles in the pathogenesis of different genotypes or life cycle stages of T. gondii. These findings provide new insight into the role of PSPs in the pathogenesis of T. gondii.
Highlights
The causative agent of toxoplasmosis, Toxoplasma gondii, is an obligate intracellular protozoan parasite (Robert-Gangneux and Dardé, 2012; Smith et al, 2021), which can infect many warmblooded animals and humans (Elsheikha et al, 2021)
immunofluorescence assay (IFA) results showed that PP5, EFPP, PPM8, and CTD2 were detected in the tachyzoite cytoplasm (Figure 1)
CTD3 accumulated in the tachyzoite basal complex in a punctate manner, which was similar to the partial localization of T. gondii Centrin2 (TgCEN2) (Figure 1) (Leung et al, 2019)
Summary
The causative agent of toxoplasmosis, Toxoplasma gondii, is an obligate intracellular protozoan parasite (Robert-Gangneux and Dardé, 2012; Smith et al, 2021), which can infect many warmblooded animals and humans (Elsheikha et al, 2021). Humans acquire infection via waterborne transmission, foodborne transmission, and congenital (transplacental) infection (Yarovinsky, 2014). T. gondii infection in immunocompetent individuals generally do not lead to illness; infection of immunocompromised individuals such as those with AIDS or malignancies, can lead to encephalitis and retinochoroiditis or even death (Wang et al, 2017). Congenital infection of the fetus can cause neonatal blindness and cognitive impairment (Wang et al, 2017; Elsheikha et al, 2021). There is a need for the identification of new pharmacological targets for the development of novel therapeutics for toxoplasmosis
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