Epstein-Barr virus-positive gastric cancer involves enhancer activation through activating transcription factor 3
Epstein-Barr virus-positive gastric cancer involves enhancer activation through activating transcription factor 3
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DOI：
10.1111/cas.14370
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发表时间：
2020-03-20
期刊：
影响因子：
5.7
通讯作者：
Kaneda, Atsushi
中科院分区：
文献类型：
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作者：
Asakawa, Yuta;Okabe, Atsushi;Kaneda, Atsushi
EB 病毒 (EBV) 与特定形式的胃癌 (GC) 相关。我们之前表明，EBV 感染胃上皮细胞会诱导启动子区域异常 DNA 高甲基化和抑癌基因沉默。我们对胃细胞 EBV 感染期间的转录组和表观基因组变化进行了综合分析，以研究可能有助于肿瘤发生的增强子区域和相关转录因子 (TF) 的激活。甲醛辅助调控元件分离 (FAIRE) 测序 (-seq) 数据显示，在 EBV 感染的 MKN7 细胞 (MKN7_EB) 中，假定的 H3K4me1(+) H3K4me3(-) 增强子中有 19 992 个开放染色质区域，其中 10 260 个区域显示 H3K27ac 增加。基序分析显示候选转录因子，例如激活转录因子3 (ATF3)，可能与这些激活的增强子结合。由于 EBV 因素，包括 EBV 确定的核抗原 1 (EBNA1)、EBV 编码的 RNA 1 和潜伏膜蛋白 2A，ATF3 在 MKN7_EB 中显着上调。突变体 EBNA1 的表达减少了 EBV 基因组的拷贝数，导致 ATF3 表达相对下调。 Epstein-Barr 病毒也感染了正常胃上皮细胞 GES1，证实了 ATF3 的上调。对 ATF3 结合位点的染色质免疫沉淀序列分析和对 ATF3 敲低的 MKN7_EB 的 RNA 序列分析揭示了 ATF3 激活增强子靶向的 96 个基因，这些基因与癌症标志相关，例如逃避生长抑制剂。与MKN7相比，这96个ATF3靶基因在MKN7_EB中显着上调，并且当ATF3在EBV阳性GC细胞SNU719和NCC24中被敲低时显着下调。在 EBV 感染的 MKN7、SNU719 和 NCC24 细胞中敲低 ATF3 均会通过凋亡细胞的增加导致细胞生长显着减少。这些表明通过 ATF3 的增强子激活可能有助于 EBV 阳性 GC 的肿瘤发生。
Epstein-Barr virus (EBV) is associated with particular forms of gastric cancer (GC). We previously showed that EBV infection into gastric epithelial cells induced aberrant DNA hypermethylation in promoter regions and silencing of tumor suppressor genes. We here undertook integrated analyses of transcriptome and epigenome alteration during EBV infection in gastric cells, to investigate activation of enhancer regions and related transcription factors (TFs) that could contribute to tumorigenesis. Formaldehyde-assisted isolation of regulatory elements (FAIRE) sequencing (-seq) data revealed 19 992 open chromatin regions in putative H3K4me1(+) H3K4me3(-) enhancers in EBV-infected MKN7 cells (MKN7_EB), with 10 260 regions showing increase of H3K27ac. Motif analysis showed candidate TFs, eg activating transcription factor 3 (ATF3), to possibly bind to these activated enhancers. ATF3 was considerably upregulated in MKN7_EB due to EBV factors including EBV-determined nuclear antigen 1 (EBNA1), EBV-encoded RNA 1, and latent membrane protein 2A. Expression of mutant EBNA1 decreased copy number of the EBV genome, resulting in relative downregulation of ATF3 expression. Epstein-Barr virus was also infected into normal gastric epithelial cells, GES1, confirming upregulation of ATF3. Chromatin immunoprecipitation-seq analysis on ATF3 binding sites and RNA-seq analysis on ATF3 knocked-down MKN7_EB revealed 96 genes targeted by ATF3-activating enhancers, which are related with cancer hallmarks, eg evading growth suppressors. These 96 ATF3 target genes were significantly upregulated in MKN7_EB compared with MKN7 and significantly downregulated when ATF3 was knocked down in EBV-positive GC cells SNU719 and NCC24. Knockdown of ATF3 in EBV-infected MKN7, SNU719, and NCC24 cells all led to significant decrease of cellular growth through an increase of apoptotic cells. These indicate that enhancer activation though ATF3 might contribute to tumorigenesis of EBV-positive GC.