Gene expression network regulated by DNA methylation and microRNA during microcystin-leucine arginine induced malignant transformation in human hepatocyte L02 cells
Gene expression network regulated by DNA methylation and microRNA during microcystin-leucine arginine induced malignant transformation in human hepatocyte L02 cells
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微囊藻毒素-亮氨酸精氨酸诱导人肝细胞L02细胞恶性转化过程中DNA甲基化和microRNA调控的基因表达网络
DOI：
10.1016/j.toxlet.2018.03.003
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发表时间：
2018-06-01
影响因子：
3.5
通讯作者：
Liu, Jin-Yi
中科院分区：
文献类型：
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作者：
Chen, Hong-Qiang;Zhao, Ji;Liu, Jin-Yi
微囊藻毒素（MC）是一种环七肽化合物，可导致肝癌的发生。然而，潜在的表观遗传调控机制在很大程度上是未知的。本研究采用微囊藻毒素-LR（L：赖氨酸，R：精氨酸，MC-LR）诱导人肝细胞株L02恶性转化。通过高通量测序检测基因表达谱、microRNA（miRNA）和DNA甲基化。与对照组相比，MC-LR处理组有826个基因和187个miRNAs的表达发生显著变化。DNA甲基化序列分析显示，在基因启动子或编码DNA序列（CDS）中有2592个CpG位点发生了差异甲基化，而DNA甲基转移酶3 α（DNMT 3a）和DNA甲基转移酶3 β（DNMT 3b）表达显著上调。功能分析和京都基因与基因组百科全书（KEGG）通路分析表明，显著改变的mRNA和microRNA主要参与了肿瘤的形成、增殖、侵袭、迁移和代谢。miRNA-mRNA网络和mRNA-mRNA网络分析显示hsa-miR-320 a、hsa-miR-331- 3 p、hsa-miR-26 a-5 p、hsa-miR-196 a-5 p、hsa-miR-221- 3 p、卷曲螺旋结构域180（CCDC 180）、黑色素瘤抗原基因家族成员D1（MAGED 1）、跨膜4-结构域A7（MS 4A 7）、类肝素蛋白1（HEPHL 1）、BH3（Bcl-2同源3）样基序，细胞死亡诱导物（BLID），基质金属肽酶13（MMP 13），鸟苷酸结合蛋白5（GBP 5），脂肪生成调节因子（ADIRF），含β-H2 O2同源2结构域1（FHDC 1），蛋白激酶cAMP依赖性II型调节亚基β（PRKAR 2B），非选择性的Nodium渗漏通道（NALCN），肌球蛋白轻链激酶3（myosin light chain kinase 3，MYLK 3）、表皮生长因子受体（epidermal growth factor receptor，EGFR）和锌指蛋白704（zinc finger protein 704，ZNF 704）是MC-LR诱导L02细胞恶性转化的关键基因。此外，我们还发现MYLK 3、EGFR和ZNF 704的表达受DNA甲基化和miRNAs的调控，这些基因影响细胞周期和细胞分裂。我们的研究表明，由DNA甲基化和miRNA调控的特征性基因改变可能在环境MC-LR诱导的肝癌发生中起重要作用。
Microcystin (MC) is a cyclic heptapeptide compound which could lead to the development of hepatocellular carcinoma. However, the underlying epigenetic regulation mechanism is largely unknown. In this study, microcystin-LR (L: lysine, R: arginine, MC-LR) was used to induce the malignant transformation of human hepatocyte L02 cell line. The profile of gene expression, microRNA (miRNA) and DNA methylation were detected through high-throughput sequencing. Compared with control group, the expression of 826 genes and 187 miRNAs changed significantly in MC-LR treated group. DNA methylation sequencing analysis showed that 2592 CpG sites differentially methylated in promoter or the coding DNA sequence (CDS) of genes, while DNA methyltransferase 3 alpha (DNMT3a) and DNA methyltransferase 3 beta (DNMT3b) were dramatically up-regulated. Functional analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis showed that significantly changed mRNAs and microRNAs were mainly involved in the formation of cancer, proliferation, invasion, migration and metabolism. MiRNA-mRNA network and mRNA-mRNA network analysis showed that hsa-miR-320a, hsa-miR-331-3p, hsa-miR-26a-5p, hsa-miR-196a-5p, hsa-miR-221-3p, coiled-coil domain containing 180 (CCDC180), melanoma antigen gene family member D1 (MAGED1), membrane spanning 4-domains A7 (MS4A7), hephaestin like 1 (HEPHL1), BH3 (Bcl-2 homology 3)-like motif containing, cell death inducer (BLID), matrix metallopeptidase 13 (MMP13), guanylate binding protein 5 (GBP5), adipogenesis regulatory factor (ADIRF), formin homology 2 domain containing 1 (FHDC1), protein kinase CAMP-dependent type II regulatory subunit beta (PRKAR2B), nodium leak channel, non-selective (NALCN), myosin light chain kinase 3 (MYLK3), epidermal growth factor receptor (EGFR) and zinc finger protein 704 (ZNF704) were key miRNAs and genes in the malignant transformation induced by MC-LR in L02 cells. Moreover, we found that expression of MYLK3, EGFR and ZNF704 were regulated by DNA methylation and miRNAs, and these genes affected the cell cycle and cell division. Our study suggested that characteristic gene alterations regulated by DNA methylation and miRNA could play an important role in environmental MC-LR induced hepatic carcinogenesis.