Entry failure is drastically enhanced in Δ toxofilin parasites. (A) Time lapses showing the initial engagement of the Δ toxofilin RON2mC parasite into the forming PV followed by their withdrawal. The two top panels show the DIC and fluorescent merged frames while the two bottom panels show the overlay between the parasite RON2mC (green) and HeLa cell Myr-Palm- GFP (red) signals; the pink arrowhead points to the start of entry and the blue line shows the initial forward trajectory of the tachyzoite. (B) Zoom from (A) on the time the tachyzoite rotates and starts to disengage, the white line follows the trajectory of the rhoptry-associated signal and the curved white arrow indicates the counter clockwise rotation. (C) Time lapse showing Δ toxofilin RON2mC parasite that releases RON2 into the PM and starts to penetrate (blue trajectory) but eventually withdraws (white trajectory) while RON2 moves backwards along the parasite surface. (D) Graphs corresponding to the time lapse (C) showing: the xy coordinates as a function of time (blue dots) and the fitting polynomial curve (red crosses), the path of the tachyzoite is labeled with arrows (blue = forward, grey = disengagement); and the radius curvature (RC) as a function of time. (E) Time lapse showing failure to complete entry while a faint signal of RON2mC is seen capped backwards at the posterior end of the tachyzoite, the pink arrowheads indicate the junction while parasite forward (blue) and backward (white) trajectories are drawn. (F) Graphs corresponding to the time lapse (E) showing: the xy coordinates as a function of time (blue dots) and the fitting polynomial curve (red crosses), and the radius curvature (RC) as a function of time; the RC shift is marked with a blue arrow.