Extended Data Figure 8. IgA-Low cultured bacteria can degrade IgA.

Primary intestinal epithelial cell monolayers were pre-treated with 10 μM DAPT + 1μg ml⁻¹ LPS on days 1 and 2 post-seeding to induce differentiation and pIgR expression. Some wells were left untreated as negative controls. On day 3 post-seeding, 3 μg of normal mouse IgA was added to the lower compartment of the Transwells. Different subsets of the DAPT+LPS-treated Transwells were also treated with combinations of the following in the apical compartment: live IgA-Low bacterial cultures (either the pelleted bacterial or supernatant fraction), freeze/thawed (F/T) IgALow bacterial cultures, and a 1X protease inhibitor (PI) cocktail. Apical Transwell supernatants were collected at 3 hours (a) and 6 hours (b), and the amount of IgA was measured by anti-mouse IgA ELISA. The dotted lines represent the limit of detection by ELISA. All values are indicated as mean±s.e.m. One-way ANOVA: (a) F=26.32, P<0.0001, n=8(Untreated), n=8(DAPT+LPS), n=6(IgA-Low culture, pellet), n=3(IgA-Low culture, sup), n=4(IgA-Low culture, F/T), and n=4(IgA-Low culture, +PI); (b) F=35.57, P<0.0001, n=8(Untreated), n=8(DAPT+LPS), n=6(IgA-Low culture, pellet), n=3(IgA-Low culture, sup), n=3(IgA-Low culture, F/T), and n=4(IgA-Low culture, +PI). Means with different letters are significantly different by Tukey's multiple comparison test; ND, not detected.