Figure 3.

Association between Autophagy- and Rac1-mediated C. jejuni invasion pathway. (A) HeLa cells were treated with 10 μM Cholchicine (CLC), 300 nM Cytochalasin D (CyD) with or without 1 μM Torin 1 for 3 h. The number of invaded C. jejuni was estimated by gentamicin protection assay at 1 h p. i. (B) HeLa cells were treated with 1 μM Torin 1 for 3 h, and 3 mM Methyl-beta-cyclodextrin (MβCD) was additionally treated for last 1 h. Intracellular bacterial number was measured by gentamicin protection assay at 1 h p. i. (C) Empty vector (EV) and, constitutive active form (CA) and dominant negative form (DN) of RhoA, Rac1, and Cdc42 were transfected in HeLa cells. The number of intracellular C. jejuni was estimated by gentamicin protection assay at 1 h p. i. (D) Intracellular active Rac1 level at 6 h p. i. was measured by Western blotting. (E) Band intensity of active Rac1 was quantified. (F) CA-Rac1 or DN-Rac1 expressed HeLa cells were treated with 200 nM Bafilomycin A1 (Baf) for 3 h. The number of intracellular C. jejuni was estimated by gentamicin protection assay at 1 h p. i. (G) HeLa cells were co-transfected EV, CA-Rac1, or DN-Rac1 and siNC or siLC3B. The number of intracellular C. jejuni was estimated by gentamicin protection assay at 1 h p. i. All data are expressed as means ± standard deviations from 3 independent experiments. Differences were evaluated with Two-tailed Student’s t-test. *p < 0.05, **p < 0.001.