Figure 3.

KMT5A downregulates CDH1 expression by inhibiting its transcription. (A) Western blot images demonstrating that knockdown of KMT5A expression level in 786-O cells led to the upregulation CDH1 protein levels. (B) Knockdown of KMT5A expression in 786-O cells increased the relative CDH1 mRNA levels, detected using a reverse transcription-qPCR assay. The means ± SD of relative mRNA levels from three independent experiments are represented. (C) KMT5A knockdown led to an increase in the CDH1 promoter activity in 786-O cells. The means ± SD of normalized luciferase activity from three independent experiments are represented. (D) Endogenous KMT5A was associated with the CDH1 promoter. Chromatin immunoprecipitation-qPCR was performed on sonicated chromatin from 786-O cells immunoprecipitated with anti-KMT5A antibody or isotype IgG. The CDH1 promoter segments were measured by qPCR against a 5% input. The means ± SD from three independent experiments are represented. Significant differences were confirmed by Student's t-test for comparing 2 groups or one-way analysis of variance for >2 groups. **P<0.01 and ***P<0.001, each group compared vs. the control group. KMT5A, N-lysine methyltransferase KMT5A; SCR, scrambled control; shKMT5A, short hairpin RNA targeting KMT5A; CDH1, cadherin-1; qPCR, quantitative polymerase chain reaction; SD, standard deviation.