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. 2020 Oct 10;9(10):2266. doi: 10.3390/cells9102266

Figure 2.

Figure 2

ROS production by gallic acid was required for the repression of rRNA transcription mediated by KDM2A in MCF-7 cells. (A) Gallic acid increases ROS production in MCF-7 cells. MCF-7 cells cultured with cell-permeable ROS probe DCFDA were cultured with or without 50 μM gallic acid (GA) in the presence or absence of 0.5 mM N-acetyl-l-cysteine (NAC) or l-glutathione (GSH). At the indicated times, the increases in the signal intensities of DCF by ROS in cells were measured. (B) Effects of anti-oxidants, NAC and GSH, on the reduction of rRNA transcription by gallic acid. MCF-7 cells treated with or without 50 μM gallic acid in the presence or absence of 0.5 mM NAC or 0.5 mM GSH for 4 h. Total RNAs were isolated and analyzed by qRT-PCR to detect pre-rRNA (left panel) and KDM2A mRNA (right panel). The ratios of the values for cells treated with various conditions to those for cells treated without gallic acid and anti-oxidants are shown. (C) Effects of anti-oxidants, NAC and GSH, on the reduction of H3K36me2 marks in the rDNA promoter by gallic acid. MCF-7 cells were cultured for 4 h with or without 50 μM gallic acid in the presence or absence of NAC or GSH. The levels of H3K36me2, H3K36me3, and KDM2A in the rDNA promoter were analyzed by ChIP assays. The results are expressed as fold changes to the values in various conditions to those without gallic acid and anti-oxidants. All experiments were performed more than three times, and the mean values with standard deviations are indicated. * p < 0.05.