Enteropathogenic E. coli relies on collaboration between the formin mDia1 and the Arp2/3 complex for actin pedestal biogenesis and maintenance
Fig 1
Chemical inhibition of formins decreases actin pedestal assembly by EPEC, but not by KC12+EspFU.
(A) HeLa cells were pretreated with DMSO, CK666+CK869 (CKCK), Wiskostatin (WISKO), or SMIFH2, and infected with either EPEC or KC12+EspFU for 3.5 h. Cells were fixed and treated with antibodies to detect HA-Tir (red), phalloidin to visualize F-actin (green), and DAPI to stain DNA (blue). Scale bar, 25 µm. (B-C) The % of adherent EPEC or KC12+EspFU (defined by Tir staining) that associated with pedestals was quantified from experiments performed as in A. Each point represents a single infected cell (n = 25–30) harboring 10–50 bacteria, and bars display the mean % +/- SEM. (D-E) The F-actin pixel intensity in the pedestal-forming region (indicated by Tir staining) was quantified and normalized to adjacent pedestal-free areas of the cell, which were set to 1. Each point represents a single EPEC or KC12+EspFU pedestal-forming region, and lines show the mean intensity +/- 95% CI (n = 150 pedestal-forming regions, 15 cells). ** p<0.01, *** p<0.001, ns = not significant (ANOVA, Tukey post-hoc tests). Significance asterisks in B and C are in reference to the DMSO treated conditions.