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[Preprint]. 2026 May 24:2026.05.21.726867. [Version 1] doi: 10.64898/2026.05.21.726867

Figure 3: TGIF disrupts parasite replication.

Figure 3:

(A) Immunofluorescence showing the localization of trans-Golgi markers GalNac-GFP (magenta, top), SORTLR (visualized with an anti-SORTLR antibody, magenta, middle) and Rab6 (EmGFP-Rab6, magenta, bottom) in TGIF-Ty parasites treated with ethanol (control) or IAA (knockdown) for 15 and 48 hours. Anti-MLC1 or anti-IMC1 antibodies (yellow) used to visualize the periphery of parasites. 48 hours after TGIF knockdown, parasites exhibit both normal and abnormal morphologies (yellow arrow). Dashed white boxes denote area used to make insets to highlight mislocalization of GalNac in parasites with normal morphology. Magenta arrow indicates SORTLR and GalNac missing from parasites with abnormal morphology. Scale bars are 5μm and 1μm. Representative images from at least two experimental replicates (N > 20 vacuoles per condition). (B) Immunofluorescence showing parasite periphery labeled with IMC3 (anti-IMC3, grey) in TGIF-Ty parasites treated with ethanol (control) or IAA (knockdown) for 12, 24, 36, and 48 hours. Images are representative of the average number of parasites per vacuole at each time point. Scale bars are 5μm. Representative images from three experimental replicates (N > 100 vacuoles per time point per condition). (C) Graph shows parasite growth rate in control and TGIF knockdowns. After 60 hours of growth control parasites were extracellular. (D) Anti-IMC3 immunofluorescence showing abnormal parasite morphology in TGIF-Ty parasites treated with ethanol (control) or IAA (knockdown) for 48 and 60 hours in interphase (top) and replicating (bottom). In the bottom panels, the number of daughters per parasite are indicated. Scale bars are 5μm. Representative images from three experimental replicates (N > 100 vacuoles per time point per condition). (E-F) Quantification of the percent of parasites with abnormal morphologies, and percent of parasites replicating asynchronously after 48 hours of growth. Colored circles indicate averages from each independent replicates. P-values from paired Student’s T-tests are indicated.