Fig. 5
p-TGIF2 recruits HDAC1 to promote EMT and metastasis in LUAD. (A) ChIP-Seq data from the Cistrome Data Browser database showing the TGIF2, HDAC1 and HDAC3 binding sites on the human CDH1 genomic locus. (B) ChIP-qPCR of HDAC1 on the CDH1 promoter at the indicated regions in H1299 cells. Mouse IgG served as a negative control. (C) Western blot assays of H1299 cell samples immunoprecipitated with anti-HDAC1 and anti-TGIF2 antibody. (D) Western blot assays of TGIF2AA-overexpressing H1299 cell samples immunoprecipitated with anti-HDAC1 and anti-Flag antibody. (E) Colocalization of TGIF2 and HDAC1 was examined in control and TGIF2AA-overexpressing H1299 cells by immunofluorescence. Scale bars, 5 μm. (F) Western blot for E-cadherin, TGIF2 and p-TGIF2 in H1299 cells with the indicated dose of HDAC1 inhibitor (MGCD0103). (G) Schematic of the phosphorylation-mimicking mutations (TGIF2DD) of the two MAPK sites in the TGIF2 coding sequence. (H) Western blots for E-cadherin, Vimentin, HDAC1, TGIF2 and p-TGIF2 in H1299 cells. (I) Immunofluorescence analysis of E-cadherin and Vimentin in the indicated H1299 cells. Scale bars, 20 μm. (J) Transwell assay of H1299 cell migration according to the ectopic expression of TGIF2DD or HDAC1 knockdown. Scale bars, 100 μm. (K) Wound healing assay of the distance migrated by H1299 cells. Scale bars, 100 μm. **, p < 0.01; ***, p < 0.001; ns, not significant. All data are representative of three repeated experiments