FIG 1.

CPB does not play a major role in tachyzoite growth and protein turnover. (A) Immunoblot of tachyzoite lysates probed for T. gondii CPL and CPB along with MIC2 as a loading control. Protein bands are labeled as proform (pro), intermediate form (int), or mature form (mat) based on their molecular weights. (B) Recently invaded intracellular tachyzoites were stained with anti-CPB (violet) and anti-CPL (blue). Arrowheads indicate labeling of the VAC. Scale bars, 2 μm. (C) T. gondii tachyzoites were cultured in HFF monolayers. Samples were collected at 24 and 48 h postinfection, fixed, and stained with anti-SAG1 antibody, and the number of parasites per vacuole was quantified. Percentages of various replication stages for each strain are quantified. Results represent means from 2 or 3 biological replicates. The total numbers of parasitophorous vacuoles counted across 3 biological replicates were the following: Pru, 147 and 152; PΔcpb, 144 and 145; PΔcpl, 153 and 136 (values are for 24 and 48 h postinfection, respectively). Only statistically significant differences are represented. Statistical significance was determined by unpaired two-tailed Student’s t test performed on the mean number of parasites per vacuole for each strain across 3 biological replicates. (D) Assay quantification of tachyzoite ingestion of mCherry showing means ± standard deviations from 3 biological replicates. The data generated were analyzed using the following number of tachyzoites per replicate: Pru, n = 469, 464, and 231; PΔcpb, n = 374, 613, and 213; and PΔcpl, n = 427, 299, and 216. All strains were compared, and only significant differences are shown. Unpaired two-tailed t test with Welch’s corrections was performed on the means from 3 biological replicates.