1. Overexpression of AP2IV-3 enhances tissue cyst formation.
Overexpression of AP2IV-3 enhances tissue cyst formation. The 72-h capacity of the type III CTG parent strain to form tissue cysts compared to that of a CTG transgenic clone expressing DDAP2IV-3 was evaluated after a shift to mild-to-strong alkaline medium plus or minus Shield 1 at 100 nM, as indicated in the color key. The fraction of DBA-positive parasites in 100 randomly selected vacuoles was determined in triplicate. The increases in cyst formation by the DDAP2IV-3 parasites compared to that by the CTG parent control caused by alkaline stress alone or in combination with Shield 1 was statistically significant (*, P < 0.05; **, P < 0.01; ***, P < 0.001; pairwise data comparisons diagramed for the pH 7.4 data were repeated for the pH 7.8 and 8.2 data). Note also that type III CTG parental tachyzoites have a lower propensity to form tissue cysts spontaneously and are more resistant to milder alkaline pH medium conditions than Pru parental strain tachyzoites (Fig. 2B, blue bars). (Inset) Diagram of the DDAP2IV-3 overexpression construct and Western blot assay of DDAP2IV-3 expression with or without Shield 1. Note that DDAP2IV-3 was induced to detectable levels by pH 8.2 medium alone and to higher levels when Shield 1 was added to transgenic parasites grown in pH 7.0 and 8.2 media. The level of nuclear T. gondii PCNA1, as revealed by rabbit polyclonal antibody staining, was included as a loading control.
Dong-Pyo Hong et al. mSphere 2017; doi: /mSphere