Fig. 4

H. pylori infection induces demethylation modification of the promoter region of GNB4. A Pyrophosphate sequencing of the promoter region of GNB4 (5 CpG sites, chr3:179451774–179451745) in MKN45 cells uninfected or infected with H. pylori strains (26695 and SS1) for 6 h. The upper part of the panel shows the primer sequence regions for methylation-specific PCR (MSP). The bottom part of the panel shows a representative image of the results of pyrophosphate sequencing. B Methylation levels of the CpG#5 site (chr3:179451746–179451745) in three independent experiments via pyrophosphate sequencing. C MSP analysis showed GNB4 promoter region undergoes decreased methylation levels after H. pylori infection (26695 and SS1; 6 h) in MKN45 and GES1 cells. M, methylation-specific PCR product; U, unmethylation-specific PCR product. D Western blot analysis of GNB4 in MKN45 cells treated with different concentrations of decitabine (DNA methylation inhibitor) for 24 h. E Compared to H. pylori-negative (HP−) GC samples, the promoter methylation level of GNB4 in H. pylori-positive (HP+) GC samples was significantly decreased (verified by MSP). F Mass spectrometry-based analysis of the difference in methylation levels of the indicated CpG island region (53 CpG sites; − 235 to 130 bp; location of the GNB4 promoter) between HP+ and HP− patients with GC