The vacuole-like structures are not associated with the autophagic clearance of R. parkeri. (A) Immunofluorescence microscopy of infected A549 cells at 24 hpi showing endogenous p62 localization and indicated bacterial strains. Scale bar, 1 µm. (B) Percentage of indicated bacteria in A showing p62 localization. Data are mean ± SD of three independent experiments (≥4 FOV with ≥200 bacteria quantified per condition per independent experiment). (C) Western blot analysis of ATG5 expression in HeLa cell lysates from D. GAPDH, loading control. (D) Percentage of WT or ATG5^−/− HeLa cells displaying BERCs with sca2::Tn mutant bacteria (48 hpi). The mean ± SEM of three independent experiments (large outlined circles) are superimposed over the raw data the percent of cells with BERCs per FOV (small unlined circles) (>75 cells quantified per condition per independent experiment) (one-way ANOVA with Dunnett’s post hoc test). (E) Live-cell imaging snapshots at 28 hpi of ompB::Tn mutant (magenta) infecting A549-mCherry-Sec61β cells (cyan). Scale bar, 5 µm; inset scale bar, 1 µm. (F) Percentage of RpBFP or ompB::Tn mutant bacteria forming vacuole-like ER structures. RpBFP data are independent from that shown in Figs. 1 and 2. Each dot represents the mean percentage of bacteria interacting with the ER for three independent experiments (>500 bacteria per independent experiment). Error bars represent SEM. Source data are available for this figure: SourceData FS2.