Figure 2. GRA15 enhances IFNγ‐induced PVM decoration with autophagy‐related proteins and endo‐lysosomal‐mediated vacuole destruction in HFFs.

• A–G
  HFFs were stimulated with for 24 h with 10 U/ml IFNγ or left unstimulated and subsequently infected with RH, Pru, or PruΔgra15 parasites for 3 h. The percentage of vacuoles that stained positive for (A) total ubiquitin (n = 3 for RH, n = 5 for Pru, and n = 3 for PruΔgra15), (B) K63‐linked ubiquitin (n = 3 for RH, n = 3 for Pru, and n = 3 for PruΔgra15), (C) p62 (n = 3 for RH, n = 10 for Pru, n = 7 for PruΔgra15, and n = 4 for Pru Δgra15 + GRA15), (D) NDP52 (n = 3 for RH, n = 3 for Pru, and n = 3 for PruΔgra15), (E) LC3B (n = 3 for RH, n = 3 for Pru, n = 3 for PruΔgra15, and n = 3 for Pru Δgra15 + GRA15), (F) GABARAP (n = 3 for RH, n = 3 for Pru, and n = 3 for PruΔgra15), and (G) LAMP1 (n = 3 for RH, n = 3 for Pru, and n = 3 for PruΔgra15) is shown in the left bar diagram. On the right‐hand side, a representative fluorescent image is shown for the Toxoplasma Pru strain, which expresses GFP. DNA was stained with Hoechst 33258. Scale bar is 10 μm. The yellow box inside each representative image is shown as inset pictures with magnification.
• H
  The number of PVs per 20× objective field was counted and compared between IFNγ‐stimulated and IFNγ + bafilomycin A1 (100 nM)‐treated HFFs 24 h p.i. with Pru strain. Images from at least six fields were taken for each condition (n = 3).

Data information: Each dot represents one experiment. Each time, at least 100 different vacuoles were scored and analyzed. Statistical analysis was done by two‐way ANOVA followed with Tukey's multiple comparison test (A–G) and one‐way ANOVA for (H). Data are represented as mean ± SEM.Source data are available online for this figure.