Fig. 9

Localization of TgDCX and its orthologues in Toxoplasma. a-b Deconvolved wide-field images of dividing TgDCX knockout (a, “ΔTgDCX”, two examples) and RHΔku80Δhx (b, “WT”) parasites transiently expressing TgDCX-eGFP driven by the T. gondii α-tubulin promoter. TgDCX-eGFP is highly enriched in the mother conoid (green arrowhead) and daughter conoids (green arrows) and is absent from the cortical microtubules of mother parasites. However, in contrast to expression regulated by the endogenous promoter, when expression is driven by this nearly constitutive (see d) α1-tubulin promoter, in some cases TgDCX-eGFP signal is also detected on the daughter cortical microtubules, centrosomes (cyan arrowheads), and basal complexes (cyan arrows). Dashed cyan lines in a outline two of four parasites in the same parasitophorous vacuole. Insets: 1.5x. The lower panels show merged DIC and fluorescence (in red) images. c Deconvolved wide-field images of RHΔku80Δhx (WT) parasites expressing FP tagged DCX orthologues. Two examples are shown for CvDCX1. In the left example, dashed blue lines outline 4 of the 8 parasites in the vacuole. In the right example, the dashed blue oval outlines two nearly mature daughters, shown enlarged 1.5x in the oval inset with white outline. Note that among the eight orthologues, only CvDCX1 closely mimics the pattern of localization shown by TgDCX (when expressed under this T. gondii α1-tubulin promoter). Green arrows: daughter conoids. Green arrowheads: mother conoids. Cyan arrowhead: centrosome. d Time course of RNA expression levels [21] in Toxoplasma gondii for α1-tubulin (green) and TgDCX (red). Tubulin expression is nearly constitutive, whereas TgDCX varies by more than 30-fold across the cell-cycle