Figure 6.

Lack of HIF1A favors LC3-associated phagocytosis. (A) Control and HIF1A-silenced T84 cells were infected with AIEC LF82 at a MOI of 10 for the indicated times. Cells were processed for immunoblotting using anti phospho- or total ULK1 antibodies. The time course indicated that bacteria induced ULK1 (Ser555) phosphorylation only in T84-ShCTR cells. Results from 3 independent experiments were quantified as described in Materials and Methods; signal corresponding to p-ULK/ULK normalized to ACTB was calculated for each condition. The values of T84-ShCTR infected 4 h with bacteria cell samples were set as 1 and the fold change was calculated. *P < 0.05 as compared to uninfected T84-ShCTR cells. (B) Prior to infection cells were incubated with both blocking TLR5 (Pab-hTLR5) and CEACAM6 (clone 9A6) antibodies or with anti CEACAM6 and anti TLR5 separately and further processed as described in (A). (C) Control, HIF1A-silenced T84 cells or control cells preincubated with both anti TLR5 and anti CEACAM6 blocking antibodies were infected with AIEC LF82 at a MOI of 10 for 2 h. Cells were then processed for immunofluorescence analysis in order to analyze the colocalization of LF82-GFP with ATG16L1 and ULK1. The data are representative of 2 independent experiments. *P < 0.05 as compared to the number of noncolocalized LF82-GFP bacteria.