Figure 4.

Helicobacter pylori-mediated suppression of claudin-7 is dependent on β-catenin. (A) MKN28 cells were transfected with a β-catenin reporter containing three tandem lymphoid enhancer factor/T-cell factor (LEF/TCF)-binding motifs upstream of the luciferase gene (Topflash) or a control construct containing mutant LEF/TCF sites (Fopflash), and then treated with 10 mM LiCl. Luciferase activity, reflecting β–catenin activation, was quantified. (B and C) MKN28 cells were incubated with LiCl for 24 h, and expression levels of claudin-7 or glyceraldehyde 3-phosphate dehydrogenase (GAPDH) were quantified by western blotting. (B) Representative western blot for claudin-7 and GAPDH in MKN28 cells in the presence or absence of LiCl. (C) Densitometric analysis demonstrating decreased expression of claudin-7 in LiCl-treated cells. (D and E) MKN28 cells were transfected with control or β-catenin-specific siRNA and levels of claudin-7 or GAPDH were quantified by western blotting. (D) Representative western blot for claudin-7 and GAPDH in MKN28 cells in the presence of non-targeting (NT) or β-catenin-specific siRNA, with or without H. pylori strain 60910. (E) Densitometric analysis demonstrating that decreased expression of claudin-7 in H. pylori-infected cells is abolished in the presence of β-catenin-specific siRNA. Data are expressed as means±SEM, n=3 independent replicates.