Transcriptional activation of the human claudin-18 gene promoter through two AP-1 motifs in PMA-stimulated MKN45 gastric cancer cells

Claudin-18 (CLDN18), a member of the claudin family of proteins that are structural components of tight junctions, has two alternatively spliced variants, claudin-18a1 and claudin-18a2, which are highly expressed in lung and stomach, respectively. Downregulation of claudin-18a2 is associated with gastric cancers of an intestinal phenotype; however, the mechanisms regulating its expression have not been defined. Here, we found that phorbol 12-myristate 13-acetate (PMA) treatment of MKN45 human gastric cancer cell line increased claudin-18a2 expression. In addition, this study aimed to characterize the human CLDN18a2 promoter. Using reporter gene assays and deletion analysis, we mapped the critical promoter region of the PMA-stimulated claudin-18a2 expression to the -923/-286 region. Electrophoretic mobility shift assays and mutational analyses revealed that two activator protein (AP)-1 binding sites played an important role in the expression of claudin-18a2 in PMA-stimulated MKN45 cells. Protein kinase C (PKC) and mitogen-activated protein kinase (MAPK) inhibitors suppressed the upregulation of claudin-18a2. These results indicate that the PKC/MAPK/AP-1 dependent pathway regulates claudin-18a2 expression in gastric cells.