Fig. 5.

The C. trachomatis effector CpoS blocks cell death in infected endometrial epithelia. (A) EMO cells remain viable after C. trachomatis infection. Live spinning-disk confocal images of an uninfected EMO and an EMO infected with GFP-expressing C. trachomatis (Ct-L2) for 24 h in medium containing propidium iodide (PI). Scale bars: 50 µm. (B–D) EMOs infected with a C. trachomatis cpoS mutant induce cell death. (B) EMOs were infected with a GFP-expressing cpoS mutant (M007), incubated with PI for 22 h and imaged by live 3D deconvolution microscopy. Dashed box indicates the region shown in the zoom image. Dashed line outlines the EMO. Scale bars: 50 µm; zoom, 5 µm. (C) Maximum projection images of PI staining from 3D spinning-disk confocal images of EMOs infected as in B with either CT-L2 or M007. Dashed lines outline the EMOs. Scale bars: 50 µm. (D) Boxplots showing the number of PI puncta per EMO and the percentage of PI-positive inclusions (n=10 EMOs per replicate). Boxplots show the median and interquartile range, with whiskers indicating the minimum and maximum values. For the L2 data in the right-hand plot, only the median line is shown; because most of the values are zero, this line overlaps with the x axis. Statistical significance was measured for each replicate using a Welch's t-test. (E) Live imaging of cell death in EMOs. Time-lapse spinning-disk confocal microscopy of an EMO infected with GFP-expressing cpoS mutants (M007) cultured in the presence of PI. Left panels: maximum projections at the beginning (top) and end (bottom) of image acquisition. Dashed boxes mark regions of interest. Scale bars: 50 µm. Right panels: enlarged regions of interest at the indicated time points showing (1) an inclusion lysing before the infected cell becomes PI-positive (white arrowheads), and (2) an inclusion gradually becoming positive for PI (white arrowheads). Time display (h:min). Scale bars: 20 µm.