The EGFR pathway is required for enterocyte delamination upon infection. (a) Immunostaining of gut epithelium with Armadillo (red) and DAPI (blue). In WT flies, the Armadillo signal was strong in progenitors and lower in enterocytes. Upon infection, Armadillo staining disappeared from enterocyte membranes, but remained intense in progenitor cells. At later stages, both progenitors and newly synthesized enterocytes displayed a strong Armadillo signal. Guts with enterocytes depleted of EGFR activity (UAS-EGFRDN) displayed low Armadillo staining in enterocytes that did not change upon infection. In the absence of infection, guts with activated EGFR (UAS-EGFRACT) in enterocytes exhibited a pattern of Armadillo staining similar to Ecc15-infected WT guts. (b) Histological sections of guts from flies with WT (top) and EGFR-depleted enterocytes (bottom) with and without infection. A strong multilayering of epithelial cells (t = 4 hours), followed by their blebbing, and delamination (t = 8 hours) are observed in WT guts following infection. Delaminating cells contain multiple large vacuoles. In contrast, the multilayering, blebbing and delamination of enterocytes was not observed in guts from flies depleted of EGFR in enterocytes. PM, peritrophic matrix; L, lumen; E, enterocyte; B, bacteria. (c) In contrast to WT (Figure 2e), apoptotic enterocytes (anticleaved caspase 3-positive) were detected within the epithelial layer in flies with enterocytes depleted of EGFR activity (UAS-EGFRDN). Neither delaminated cells nor enterocytes within the epithelium were apoptotic in guts with activated EGFR (UAS-EGFRACT) in enterocytes. (d) Quantification of delaminating and apoptotic cells in the guts of flies with WT enterocytes, enterocytes depleted of EGFR activity (UAS-EGFRDN) or enterocytes expressing an activated form of EGFR (UAS-EGFRACT). Cells within the epithelium layer and delaminating cells were counted from three histological sections (N = 16 guts) for each genotype in UC and Ecc15-infected flies. The proportions of living and dead cells per section were determined with anticleaved caspase 3 and DAPI staining.