FIG 6.

Relevance of the afadin-binding sequence in the nectin-4 cytoplasmic tail for MV epithelial infection. (A to C) Localization of human nectin-4 in PAE either transduced from the basolateral surface with an adenoviral vector expressing human nectin-4 (A) or human N4ΔGHLV (B) or mock transduced (C). Two days later, cells were fixed with 2% paraformaldehyde, permeabilized with 0.2% Triton X-100, and incubated overnight with rabbit polyclonal antibodies against human nectin-4. Nectin-4 was visualized with a secondary antibody conjugated to Alexa Fluor 546 (red). Cell nuclei were visualized using DAPI (blue). For each panel, both an xy en face view (top) and an xz vertical view (bottom) are shown. Images were acquired with a confocal laser scanning microscope. White arrows indicate basolateral localization; yellow arrows indicate junctional expression. (D) Analysis of Ad-nectin-4 and Ad-N4ΔGHLV expression levels in CHO cells. β-tub., β-tubulin. (E) PAE transduced with an adenoviral vector expressing either human nectin-4 or N4ΔGHLV were infected from either the apical or the basolateral surface with MV-GFP, and the infectious centers were counted. An adenoviral vector expressing mCherry served as a negative control. Results are means for 3 experiments, each with samples from 3 donors; *, P < 0.01.