Department of Molecular Oncology, Graduate School of Medicine, Chiba University, Chiba, Japan.
Department of Pathology, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan.
Cancer Sci. 2021 Aug;112(8):3349-3362. doi: 10.1111/cas.14978. Epub 2021 Jun 13.
Epstein-Barr virus (EBV) is associated with approximately 10% of gastric cancers (GCs). We previously showed that EBV infection of gastric epithelial cells induces aberrant DNA methylation in promoter regions, which causes silencing of critical tumor suppressor genes. Here, we analyzed gene expressions and active histone modifications (H3K4me3, H3K4me1, and H3K27ac) genome-widely in EBV-positive GC cell lines and in vitro EBV-infected GC cell lines to elucidate the transcription factors contributing to tumorigenesis through enhancer activation. Genes associated with "signaling of WNT in cancer" were significantly enriched in EBV-positive GC, showing increased active β-catenin staining. Genes neighboring activated enhancers were significantly upregulated, and EHF motif was significantly enriched in these active enhancers. Higher expression of EHF in clinical EBV-positive GC compared with normal tissue and EBV-negative GC was confirmed by RNA-seq using The Cancer Genome Atlas cohort, and by immunostaining using our cohort. EHF knockdown markedly inhibited cell proliferation. Moreover, there was significant enrichment of critical cancer pathway-related genes (eg, FZD5) in the downstream of EHF. EBV protein LMP2A caused upregulation of EHF via phosphorylation of STAT3. STAT3 knockdown was shown to inhibit cellular growth of EBV-positive GC cells, and the inhibition was rescued by EHF overexpression. Our data highlighted the important role of EBV infection in gastric tumorigenesis via enhancer activation.
EB 病毒（EBV）与大约 10%的胃癌（GC）有关。我们之前表明，EBV 感染胃上皮细胞会导致启动子区域的异常 DNA 甲基化，从而导致关键肿瘤抑制基因沉默。在这里，我们在 EBV 阳性 GC 细胞系和体外 EBV 感染的 GC 细胞系中广泛分析了基因表达和活性组蛋白修饰（H3K4me3、H3K4me1 和 H3K27ac），以阐明通过增强子激活导致肿瘤发生的转录因子。与“癌症中 WNT 信号传导”相关的基因在 EBV 阳性 GC 中显著富集，显示出增加的活性 β-连环蛋白染色。邻近激活增强子的基因明显上调，并且在这些活性增强子中 EHF 基序显著富集。使用癌症基因组图谱队列的 RNA-seq 和我们队列的免疫染色证实，与正常组织和 EBV 阴性 GC 相比，临床 EBV 阳性 GC 中的 EHF 表达更高。EHF 敲低显着抑制细胞增殖。此外，在 EHF 下游存在与关键癌症途径相关基因（例如 FZD5）的显着富集。EBV 蛋白 LMP2A 通过 STAT3 的磷酸化导致 EHF 的上调。STAT3 敲低显示抑制 EBV 阳性 GC 细胞的细胞生长，而过表达 EHF 可挽救这种抑制作用。我们的数据强调了 EBV 感染通过增强子激活在胃肿瘤发生中的重要作用。
