Δcsar1 parasites exhibit multiple defects in cell cycle. (A) The percentage of soluble tubulin (versus membrane-insoluble assembled cytoskeleton) was quantified in n = 4 biological replicates (averaged from n = 4 technical replicates) of parental ERK7^AID versus Δcsar1 parasites. Significance estimated by unpaired two-sided Student’s t test. Western blots for these data are in Fig. S4. (B) Representative images of Δcsar1 parasites dividing normally (two buds) or abnormally (three buds). (C) Quantification of non-binary division for ERK7^AID and Δcsar1 parasites. “Buds” and “mature,” respectively, indicate actively dividing parasites or newly divided, mature parasites. (D) Representative image of a recently divided Δcsar1 parasite showing the remains of a nonviable parasite (white arrow). (E and F) Representative images of (E) ERK7^AID parasites dividing synchronously within a normal rosette vacuole and (F) Δcsar1 parasites dividing asynchronously (white arrow: nonbudding; orange arrow: early buds; other parasites: late buds) in a non-rosette vacuole. (G and H) Quantification of (G) Rosette/non-rosette and (H) Asynchronous division phenotypes. Phenotypes quantified from three biological replicates of n ≥ 100 vacuoles. Error bars are 95% confidence interval of mean. P values are 1-way ANOVA followed by Tukey’s test (***, P <0.001). For both t tests and Tukey’s tests, data distribution was assumed to be normal, but this was not formally tested.