Figure 4.

Interaction of autophagy signaling and Rac1 signaling in C. jejuni infection. (A) HeLa cells were transfected with EV, CA-Rac1, and DN-Rac1. Cells were lysed and subjected to immunoprecipitation with Sepharose beads, and analyzed by Western blotting by using anti-Flag antibody. (B) HeLa cells were transfected with vectors as shown, and subjected to immunoprecipitation. Cell lysates were analyzed by Western blotting by using anti-LC3B antibody. (C) Representative immunofluorescence images of CA-Rac1 expressed HeLa cells infected with C. jejuni which stained with 10 ng/ml 5(6)-Carboxyfluorescein diacetate (CFDA) solution. Cells were immunostained by anti-LC3 antibody and DAPI, and observed by confocal microscope. (D) The rate of colocalization of C. jejuni and LC3 was quantified in EV, CA-Rac1, DN-Rac1, CA-RhoA, and CA-Cdc42 expressed HeLa cells. CA-Rac1 transfected cells were treated with 200 nM Bafilomycin A1 (Baf) for 3 h or 300 nM Cytochalasin D (CyD) for 1 h before infection. The differences of signnificance were tested in comparison with CA-Rac1. EV: n = 48, CA-Rac1: n = 73, DN-Rac1: n = 59, CA-RhoA: n = 65, CA-Cdc42: n = 83, CA-Rac1 + Baf: n = 55, CA-Rac1 + CyD: n = 51 of bacterial cells. The data of confocal microscope were pooled from 4 independent experiments. Differences were evaluated with Two-tailed Student’s t-test. *p < 0.05, **p < 0.001.