Epigenetic modulation of glycoprotein VI gene expression by DNA methylation
Epigenetic modulation of glycoprotein VI gene expression by DNA methylation
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DOI：
10.1016/j.lfs.2019.117103
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发表时间：
2020-01-15
期刊：
影响因子：
6.1
通讯作者：
Wu, Hong
中科院分区：
文献类型：
--
作者：
Gao, Shuibo;Han, Yongjun;Wu, Hong
目的：糖蛋白VI （GPVI）是一种重要的血小板膜受体。GPVI在冠心病患者血小板膜上的表达升高。DNA甲基化是最常见的复制后和转录前修饰之一，在基因表达调控中起着关键作用。在这里，我们旨在揭示甲基化如何调节GPVI的表达。主要方法：采用焦磷酸测序法检测冠心病患者白细胞GPVI启动子区是否低甲基化。采用qRT-PCR和Western blot检测冠心病患者GPVI的表达。采用体外荧光素酶法分析GPVI启动子区甲基化对其转录活性的调节作用。流式细胞术检测p -选择素在血小板样颗粒中的表达，Western blot检测SYK磷酸化水平。主要发现：我们发现冠心病患者白细胞中GPVI启动子区域低甲基化，并且在mRNA和蛋白水平上GPVI表达升高。我们还发现GPVI启动子区域的高甲基化抑制了Dami细胞中-322至+75、-539至+75和-937至+75区域GPVI的表达。此外，数据显示甲基化或去甲基化调节GPVI在Dami细胞中的表达和血小板样颗粒活化。意义：综上所述，这些结果表明DNA甲基化调节GPVI的表达，GPVI基因启动子区域的CpG甲基化水平可能是冠心病的生物标志物。
Aims: Glycoprotein VI (GPVI) is an important platelet membrane receptor. The expression of GPVI on platelet membranes is increased in patients with coronary heart disease (CHD). DNA methylation is one of the most common post-replication and pre-transcriptional modifications and plays a critical role in the regulation of gene expression. Here, we aimed to reveal how methylation regulates GPVI expression.Main methods: Pyrosequencing was used to determine whether the GPVI promoter region in leukocytes from CHD patients is hypomethylated. The expressions of GPVI in CHD patients were detected using qRT-PCR and Western blot. The effect of methylation of the GPVI promoter region on regulating its transcriptional activity was analyzed using in vitro luciferase assay. The expression of P-selectin in platelet-like particles was determined using flow cytometry, and SYK phosphorylation was observed using Western blot.Key findings: We found that the GPVI promoter region in leukocytes from CHD patients was hypomethylated and the expression of GPVI at the mRNA and protein level was elevated in CHD patients. We also found that the hypermethylation of GPVI promoter region inhibited the expression of GPVI in the -322 to +75, -539 to +75, and -937 to +75 regions in Dami cells. Moreover, the data showed that the methylation or demethylation regulated the GPVI expression and platelet-like particle activation in Dami cells.Significance: Taken together, these results indicate that DNA methylation regulates GPVI expression and that CpG methylation levels in the promoter region of the GPVI gene may be a biomarker of CHD.