The Role Of JMJD2B In The Process Of Helicobacter Pylori-induced Gastric Epithelial Cell Malignant Transformation

BackgroundGastric cancer is one of the most common malignancies worldwide and the third leading cause of death among all cancers in the world. China is the second high incidence area of gastric cancer in the world, second only to Japan. There are many known factors contributing to the development of gastric cancer. Chronic infection with H. pylori is the strongest identified risk factor of gastric cancer. Although numerous studies have been conducted in illuminating the pathogenesis of H. pylori infection, the detailed mechanisms remain undefined. Recent studies have reported that H. pylori infection induces epigenetic alterations, such as DNA methylation and histone modification, which play critical roles in oncogenic transformation.Histone demethylase JMJD2B, also known as KDM4B, is newly identified and characterized as a member of the histone demethylase JMJD2 family. JMJD2B specifically targets the trimethylated lysine 9 of histone H3 (H3K9) for demethylation to regulate chromatin structure or gene expression, which has been reported to play an epigenetic role in inflammation, and tumorigenesis. Our previous studies demonstrated that JMJD2B promoted cell proliferation, survival, invasion and metastasis in gastric cancer. However, the precise mechanism how JMJD2B is overexpressed in gastric cancer has not been elucidated. Because H. pylori is an important factor for the initiation of gastric epithelial cell malignant transformation, we investigate whether H. pylori infection regulates JMJD2B expression.Recent studies suggested that COX2 was upregulated in H. pylori-infeced gastritis and gastric cancer, which was critical in inflammatory response and tumorigenesis. Although the role of COX2 in H.pylori-induced gastric carcinogenesis has been illuminated, the precise molecular mechanism of H.pylori-induced COX2 activation is not well defined. It has been suggested that COX2 expression may be regulated by epigenetic mechanisms, such as DNA methylation and histone modifications, which is associated with the development and clinical outcome of gastric cancer. Therefore, we explore whether H. pylori-induced COX2 upregulation might be controlled by JMJD2B-mediated epigenetic mechanisms.PurposeThis study determined the role of histone demethylase JMJD2B in the transformation from H. pylori-induced chronic inflammation to gastric cancer and the underlying molecular mechanism of H. pylori associated carcinogenesis.Methods1. The regulatory mechanism of H. pylori infection on JMJD2B expression. QRT-PCR、western blot and luciferase reporter assay were used to analyze the regulation of H. pylori on JMJD2B expression. Colony formation assay was used to examine the biological function of JMJD2B on H.pylori-induced cell proliferation. QRT-PCR and western blot were applied to detect JMJD2B expression after β-catenin siRNA transfection. Moreover, luciferase reporter assay was used to explore the direct regulation of p-catenin on JMJD2B promoter.2. The role of JMJD2B in H. pylori-induced COX2 activation. QRT-PCR、western blot and luciferase reporter assay were separately used to determine the regulation of JMJD2B on COX2 expression after JMJD2B siRNA transfection. Co-IP and ChIP assay were performed to determine the molecular mechanism that JMJD2B induced COX2 expression.3. H. pylori regulates JMJD2B and COX2 expression in vivo. C57BL/6J mice were infected with H. pylori or PBS using intragastric administration and killed after inoculation. QRT-PCR was used to detect JMJD2B and COX2 mRNA levels.4. JMJD2B and COX2 expression in H. pylori-infected human gastric tissues. JMJD2B and COX2 expression was assessed by IHC in human gastric tissues, including normal tissues, superficial gastritis, atrophic gastritis and gastric cancer tissues. Then we analyzed the association between JMJD2B expression and H. pylori infection and COX2 expression.Results1. JMJD2B was induced by H. pylori infection in a CagA-independent manner.2. Beta-catenin directly bound to JMJD2B promoter and stimulated JMJD2B expression following H. pylori infection.3. JMJD2B was required for H. pylori-induced COX2 expression. JMJD2B, together with NF-κB, bound to COX2 promoter to enhance its transcription by demethylating H3K9me3 locally.4. JMJD2B and COX2 expression were both upregulated in H. pylori infected mice tissues in vivo.5. Immunohistochemistry staining revealed that JMJD2B and COX2 expression was gradually increased in human gastric tissues from gastritis to gastric cancer. Furthermore, JMJD2B expression was positively correlated with H. pylori infection and COX2 expression respectively.ConclusionsH. pylori infection induced P-catenin nuclear translocation and lymphocyte enhancer factor/T cell factor (LEF/TCF) transactivation. Beta-catenin, together with LEF/TCF bound to JMJD2B promoter and activated its expression. The upregulated JMJD2B, cooperating with NF-κB, bound to COX2 promoter to stimulate its transcription by demethylating H3K9me3 locally, which eventually promoted H. pylori-induced carcinogenic process. In the present work, we explored the underlying molecular mechanisms of JMJD2B expression induced by H. pylori in the initiation and progression of gastric cancer, suggesting that JMJD2B may serve as a novel target for the prevention and early detection of gastric cancer.