Variation in the frequencies of stable versus capped junctions during cell entry by RON2mC-expressing tachyzoites. (A) Time lapse of a GFP-tachyzoite using a capped junction to invade a NRK cell and further multiplying inside a competent PV, the parasite apex (blue line) and the junction (pink line) trajectories are shown, the pink arrowhead points to the junction site, white arrows mark the two daughter cells formed in the PV at about seven hours post-invasion; graph showing the tachyzoite (blue) and the junction (pink) speeds over time during the entry event with a capped junction. (B) Pie graphs showing the distribution of stable (blue), capped (orange) and end-capped (red) junctions in epithelial and fibroblastic cells. The absolute numbers are indicated; note the significantly higher frequency of all capping events in fibroblasts. (C) Scatter graph showing the average speed as a function of time for each stable (blue dots) or capped (pink dots) junction-associated event; note the highly significant differences in speed (v) and time (t) between the two types of junctions. (D) Time lapses showing tachyzoites that hit an already internalized parasite, which is marked with a white star during penetration into a HFF cell. The blue and pink lines reconstitute the trajectories of the parasite apex and the RON2-labeled junction (green) respectively; pink arrowheads define the first and last signs of the junction; graphs on the right show the tachyzoite (blue) and the junction (pink) speeds over time; note that the junction capping coincides with the time when parasites were forced to stop; all scale bars: 5 μm. HFF, human foreskin fibroblasts; NRK, normal rat kidney fibroblasts; PV, parasitophorous vacuole; RON2, RhOptry Neck.