Association of H3K4me3 and CBX7 Levels to the Down-Regulation of <i>GKN</i>1 Gene Expression in Gastric Cancer Cells

Gastrokine 1 (GKN1) is a highly secreted gastric mucosal protein in normal individuals but strongly down-regulated or totally absent in gastric cancer subjects. An epigenetic mechanism might be responsible for GKN1 gene silencing probably through the activity of a transcription factor in association with the enzymes SUV39H1 and HDACs on the GKN1 promoter. In fact, compared to non-tumor tissues, a high increase of H3K9me3 level was observed in the corresponding tumor ones. Because H3K4me3 seems to be a possible epigenetic mark for active euchromatin, we try to verify the H3K4me3 level on the GKN1 promoter in gastric cancer tumor specimens. In addition, we also attempt to highlight if CBX7 could be the possible regulatory transcription factor correlated to GKN1 gene promoter. Therefore, we evaluated if the CBX7 expression levels could be associated with GKN1 down-regulation in gastric cancer. To this purpose, 2 pairs of non-tumor and tumor surgical specimens from patients with gastric cancer were analyzed for H3K4me3 by chromatin immunoprecipitation (ChiP) assays, and 9 pairs were instead analyzed by Western blotting for GKN1, and CBX7 expression levels, respectively. The results suggested that the observed increase of H3K4me3 in tumor samples was not in agreement with its proposed function whereas the expression of CBX7 was not associated with the down-regulation of GKN1. In particular, the expression levels of CBX7 in tumor samples might suggest a survival role in gastric cancer.

Down-Regulation of GKN1 in Gastric Cancer Is Not Associated with the RUNX3 Expression

Gastrokine 1 (GKN1) is a gastric mucosal protein highly expressed and secreted in normal individuals but during Helicobacter pylori infection or in gastric carcinogenesis it is strongly down-regulated or totally absent. In gastric cancer, the GKN1 gene is silenced through an epigenetic mechanism most likely mediated by a transcription factor that promotes on GKN1 promoter the activity of the enzymes SUV39H1 and HDACs. Because RUNX3 is a potential candidate in the regulation of molecular carcinogenesis process of stomach cancers, we tried to assess if RUNX3 could be involved in GKN1 down-regulation in GC. 17 paired of non-tumoral and tumoral surgical specimens from patients with gastric cancer were analyzed for GKN1 and RUNX3 by Western blotting and chromatin immunoprecipitation (Chip) assays. The overall results indicated that RUNX3 expression was not associated with the down-regulation of GKN1. The expression levels of RUNX3 in non-tumoral and tumoral samples suggest that RUNX3 does not act as a tumor suppressor but that it might play a complex oncogenic role in gastric cancer cells.