Fig. 5

SMYD3 transactivates Slug expression through ANKHD1. a ChIP assays were performed in huh7-control and huh7-SMYD3 stable cell lines using antibodies against ANKHD1; immunoprecipitated DNA was measured by real-time PCR using primers for amplifying the SMYD3-binding regions in the SLUG gene promoter. b ChIP assays were performed in MHCC97H with and without SMYD3 inhibitor (BCI-121) treatment using antibodies against ANKHD1; immunoprecipitated DNA was measured by real-time PCR using primers for amplifying the SMYD3-binding regions in the SLUG gene promoter. c and d ChIP assays were performed in huh7-control and huh7-SMYD3 stable cell lines with or without ANKHD1 knockdown using antibodies against SMYD3 and H3K4me3; immunoprecipitated DNA was measured by real-time PCR using primers for amplifying the SMYD3-binding regions in the SLUG gene promoter. (E) Luciferase reporter assay of huh7 cells cotransfected with the indicated luciferase reporter (wide type SLUG gene promoter-luciferase construct or its SMYD3 binding site mutants), the empty vector or SMYD3 expression vector, and control siRNA or ANKHD1 siRNA. Data are shown as mean ± SD of three separate experiments. *P < 0.05