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. 2021 Jan 1;11(2):700–714. doi: 10.7150/thno.47800

Figure 6.

Figure 6

PLAGL2 modulates Snail1 stability by activating USP37 transcription. (A) A significant positive correlation between PLAGL2 mRNA and USP37 mRNA in GC tissues could be observed both in GEPIA database(left) and our data(right). (B) WB and qRT-PCR analyses of Snail1expression in PLAGL2 knockdown SGC7901 cell and PLAGL2 overexpression AGS cell. (C) The ChIP DNA was amplified by qRT-PCR, and then the products of qRT-PCR were electrophoresed on a 2% agarose gel. (D) The schematic representation of USP37 promoter. The sequences of wild-type and mutant PLAGL2 binding sites were indicated. (E) Luciferase activity assays were performed in PLAGL2 knockdown SGC7901 cell and PLAGL2 overexpression AGS cell, which were then transfected with wild type (USP37-WT) or mutant-type (USP37-mut) USP37 promoter-reporter plasmids. (F) WB of Snail1 protein level in SGC7901 cell cotransfected with Lenti-shPLAGL2 and USP37 plasmid and AGS cell cotransfected with Lenti-PLAGL2 and USP37 siRNA. (G-H) The stable PLAGL2 knockdown (SGC7901-shRNA) cell was transfected with USP37 plasmid, and stable overexpression (AGS-PLAGL2) cell was transfected with the USP37 siRNA. The role of USP37 in PLAGL2-induced proliferation was examined by CCK8 (G). Transwell assays detected the effect of USP37 on PLAGL2-induced migration. Scale bars, 200 µm (H). (I) WB analysis of the expression level of EMT-related proteins in cotransfected SGC7901 and AGS cells. (J) The qRT-PCR analysis of the expression level of EMT-related genes in cotransfected SGC7901 and AGS cells.