Figure 5: The consequences of GSK-3 downregulation on Snail and other oncogenic pathways. | Nature Communications

Figure 5: The consequences of GSK-3 downregulation on Snail and other oncogenic pathways.

From: Helicobacter pylori CagA promotes Snail-mediated epithelial–mesenchymal transition by reducing GSK-3 activity

Figure 5

(a) Flag-tagged wt Snail and PR mutant (S104,107A) were co-transfected with full-length CagA in 293 cells. The Snail protein abundance was detected with immunoblot analysis against anti-Flag antibody. Relative protein abundance of each loading control is denoted below the blots. (b) Flag-His-tagged wt Snail was co-transfected with control or CagA expression vector for 48 h. GSK-3-mediated phosphorylated Snail was determined following immunoprecipitation (1 × , 1/3 × , 1/3 × volume of the respective cell lysate for Snail semi-quantitation) with minimum amount of Ni-Ti beads to saturate protein binding and, subsequent immunoblot analysis with anti-phospho-β-catenin antibody (pSnail). (c) The abundance of GSK-3β regulated proteins, total β-catenin, phospho-β-catenin (S33/37/T41), MCL-1, T172-phospho-AMPK and total AMPK were detected after infection with control or 60190 or isogenic CagA deletion mutant strain (ΔCagA) in AGS and MKN28 cells (left and middle panels). The 293 cells transfected with control or full-length of wt CagA or PR mutant CagA were subjected to the same panel of antibodies (right panels). As a control of GSK-3 inhibition, the 40 mM of LiCl were treated for 3 h before protein harvest.

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