Department of Basic Medicine, Shantou University Medical College, Shantou 515041, China.
Department of Cardiovascular Sciences, National Institute for Health Research, Leicester Biomedical Research Centre, University of Leicester, Leicester LE3 9QP, UK.
Cells. 2023 Jun 21;12(13):1681. doi: 10.3390/cells12131681.
Genome-wide association studies have revealed an association between the genetic variant rs17514846 in the gene and coronary artery disease. We investigated the mechanism through which rs17514846 modulates expression. An analysis of isogenic monocytic cell lines showed that the cells of the rs17514846 A/A genotype expressed higher levels of than cells of the C/C genotype. Pyrosequencing showed that the cytosine (in a CpG motif) at the rs17514846 position on the C allele was methylated. Treatment with the DNA methylation inhibitor 5-aza-2'-deoxycytidine increased expression. An electrophoretic mobility super-shift assay with a probe corresponding to the DNA sequence at and around the rs17514846 position of the C allele detected DNA-protein complex bands that were altered by an anti-MeCP2 antibody. A chromatin immunoprecipitation assay with the anti-MeCP2 antibody showed an enrichment of the DNA sequence containing the rs17514846 site. siRNA-mediated knockdown of MeCP2 caused an increase in FURIN expression. Furthermore, MeCP2 knockdown increased monocyte migration and proliferation, and this effect was diminished by a FURIN inhibitor. The results of our study suggest that DNA methylation inhibits expression and that the coronary artery disease-predisposing variant rs17514846 modulates expression and monocyte migration via an allele-specific effect on DNA methylation.
全基因组关联研究揭示了基因中的遗传变异 rs17514846 与冠状动脉疾病之间的关联。我们研究了 rs17514846 调节表达的机制。对同基因单核细胞系的分析表明，rs17514846 A/A 基因型的细胞表达的水平高于 C/C 基因型的细胞。焦磷酸测序显示，C 等位基因上 rs17514846 位置的胞嘧啶（CpG 基序）发生了甲基化。用 DNA 甲基化抑制剂 5-氮杂-2'-脱氧胞苷处理可增加的表达。与 C 等位基因 rs17514846 位置的 DNA 序列相对应的探针的电泳迁移率超迁移实验显示，与抗 MeCP2 抗体结合的 DNA-蛋白质复合物带发生了改变。用抗 MeCP2 抗体进行的染色质免疫沉淀实验显示，含有 rs17514846 位点的 DNA 序列富集。用 siRNA 介导的 MeCP2 敲低导致 FURIN 表达增加。此外，MeCP2 敲低增加单核细胞迁移和增殖，而 FURIN 抑制剂可减弱这种作用。我们的研究结果表明，DNA 甲基化抑制表达，而冠心病易感变异 rs17514846 通过对 DNA 甲基化的等位基因特异性影响来调节表达和单核细胞迁移。
