Abstract
Helicobacter pylori, a microaerophilic spiral gram-negative bacterium, can cause diseases such as gastritis, duodenal ulcer, gastric ulcer, and gastric cancer. Among the virulence factors of Helicobacter pylori, cytotoxin associated gene A (CagA) is found to associate with gastric cancer. CagA is a highly antigenic membrane protein, and translocates into animal cells by type-IV secretion system. CagA is then phosphorylated on tyrosine residues by host cell kinase and triggers intracellular signaling pathways. Cells may subsequently develope into humming-bird phenotype. The EPIYA (Glu-Pro-Iso-Tyr-Ala) motifs located at CagA C-terminals were considered to associate with CagA phosphorylation. The aim of this investigation is to identify the relationship among CagA phosphorylation, EPIYA motif and clinical diseases. The results showed that CagA phosphorylation is significantly greater in gastric carcinoma strains than that in duodenal ulcer strains (p=0.017). Of 156 strains, 153 strains of Helicobacter pylori had 0.6-kb PCR amplified fragment at the C-terminal region by use of Yamaoka’s PCR analysis. Further sequence analysis revealed 3 EPIYA motifs in the C-terminal region. The other 3 strains had 0.67-kb fragment, which was found to contain 4 EPIYA motifs by sequence analysis. CagA phosphorylation analysis showed that two strains with four EPIYA motifs had significantly higher level of phosphorylation in CagA than other strains, suggesting the positive correlation between the number of EPIYA motifs and the degree of phosphorylation in CagA. The infection of AGS cells using Helicobacter pylori with different numbers of EPIYA motifs (three or four) showed similar humming bird phenotype. We conclude that the level of CagA phosphorylation of Helicobacter pylori in infected cells can be considered as an important indicator to evaluate the severity of clinical diseases.