Skip to main content
. 2021 Jan 1;11(5):2318–2333. doi: 10.7150/thno.48739

Figure 3.

Figure 3

Hypermethylation of GNA14 leads to downregulated expression in HCC. (A) Relative methylation level of three GNA14 probes (ch.9.991104F, cg12687527, cg17301902) in HCC patients in TCGA. (B) Pearson's correlation between methylation level and mRNA level of GNA14 in TCGA. (C) Kaplan-Meier survival analysis based on high or low GNA14 methylation. (D) The heatmap of the methyl target sequencing results indicated the methylation levels of 15 methylation sites in the target CpG islands (Genomic position 1: 80262823-80262968) of the GNA14 promoter region in 12 pairs of matched tumor tissues and normal tissues in our hospital. “T” refers to tumor tissue and “N” refers to normal tissue. The horizontal axis was the probe sites, and the number represents the location of this site on the target CpG island. For example, ”22” represents the 22th base site on CpG island. Each point represents the degree of DNA methylation of the tissue. The larger the size of the dot, the redder the color, indicating the higher the degree of methylation of the tissue. (E) Pearson's correlation between methylation level and mRNA level of GNA14 in different HCC tissues from our hospital. (F) Methyl-target sequencing results showing GNA14 promoter methylation level in HCC cell lines, L02 and HepLi5 cells. (G) ChIP-qPCR analysis showing the binding of DNMT1 and DNMT3A to the GNA14 promoter region in SK-Hep-1 and Hep3B. Histone 3 was positive control group. IgG was negative control group. (H) DNA methylation sequencing of SK-Hep-1 knocking down DNMT1 or DNMT3A. (I) Western blot analysis of GNA14 protein level after DNMTs (DNMT1 or DNMT3A) knockdown in SK-Hep-1. (J) qRT-PCR analysis of GNA14 mRNA level in HCC cell lines after 5-Azacytidine treatment. (K) DNA methylation sequencing of SK-Hep-1 and Hep3B cells treated with 5-Azacytidine. *p < 0.05. **p < 0.01. ***p < 0.001. ****p < 0.0001. Statistical significance was determined by unpaired t-test. Kaplan-Meier survival analysis was performed to analyze the survival percentage. Pearson analysis was used to analyze the correlation between two groups.