Figure 2.

Yptb binds to M cells using invasin. (a-b) Differentiated uninfected HIE25 ileal monolayers (a) RT- or (b) RT+ were stained for apical β1 integrin (red) and DAPI (nuclei-blue). (c) XZ plane of RT+ monolayer shows β1 integrin⁺ M cell and neighboring non-M cells stained with phalloidin (F-actin-cyan). (d) Fluorescence intensity of F-actin was measured for each M cell and corresponding adjacent non-M cells and divided by cell area. Each point represents an M cell or the average of the corresponding neighboring non-M cells. Bars indicate mean and SD. Data were pooled from 3 images from 3 independent experiments. (e-i) Differentiated HIE25 RT+ monolayers were infected for 5 hours with 5 × 10⁶ CFU (e) WT, (f) ΔyadA, (g) Δinv, or (h) Δinv/yadA YPIII Yptb expressing GFP (green) and stained with anti-β1 integrin antibody (red) and DAPI (nuclei-blue). Magnified insets of an M cell are shown in upper right corner. XY planes are maximum intensity projections. (i) The percentage of Yptb-associated cells per field was plotted according to the number of Yptb (1, 2–5, or 6+) bound to a cell. (j) The number of Yptb-associated cells per field was plotted according to the number of Yptb (1, 2–5, or 6+) bound to a cell. (i-j) Data were pooled from 3+ independent experiments with 2–4 fields analyzed per Transwell and averaged. Each symbol represents the average from one Transwell. Bars indicate mean and SEM. Statistical significance was determined using a two-way ANOVA with Tukey’s post hoc multiple comparison test, with comparisons between columns within each row