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. 2014 Jun 14;5(14):5369–5380. doi: 10.18632/oncotarget.2105

Figure 6. GRP78 induces APC downregulation via promotion of APC secretion.

Figure 6

(A) Analysis of GRP78 and APC interactions. Immunoprecipitation experiments were carried out on HT-29 (top) and DLD1 (bottom) lysates using anti-GRP78 and anti-β-catenin antibodies followed by Western blotting for APC. (B) APC was stained in DLD1 cells stably expressing GFP and GFP-GRP78 fusion protein. (C) Cell lysates from HT-29 cells were incubated with GST or GST-GRP78 glutathione beads. The precipitates were separated by SDS-PAGE and immunoblotted with anti-APC antibody. (D) GST-GRP78 pulldown of lysates from DLD1 cells. (E) GST-GRP78 pulldown of lysates from MDCK cells stably expressing the truncated N1-APC (aa 1-448), N2-APC (aa 449-781) and N3-APC (782-1082) respectively. The precipitates were separated by SDS-PAGE and immunoblotted with anti-GFP antibody. (F) Western botting analysis of APC and GRP78 secretion in conditioned medium from DLD1 cells grown with/without glucose. (G) Analysis of GRP78 and APC interactions in conditioned medium. Immunoprecipitation experiments were performed on glucose free conditioned medium from DLD1 cells using GRP78 antibody followed by Western blotting for APC. (H) Western botting analysis of APC in conditioned medium from DLD1 cells stably expressing GFP and GFP-GRP78 fusion protein.