1 Department of Neurology and Neurosurgery Brain Center Rudolf Magnus University Medical Center, Utrecht the Netherlands.
2 Hubrecht Institute (Royal Netherlands Academy of Arts and Sciences (KNAW)) University Medical Center, Utrecht the Netherlands.
J Am Heart Assoc. 2019 May 7;8(9):e011201. doi: 10.1161/JAHA.118.011201.
Background We previously showed that intracranial aneurysm ( IA )-associated single-nucleotide polymorphisms are enriched in promoters and putative enhancers identified in the human circle of Willis, on which IA s develop, suggesting a role for promoters and enhancers in IAs . We further investigated the role of putative enhancers in the pathogenesis of IA by identifying their potential target genes and validating their regulatory activity. Methods and Results Using our previously published circle of Willis chromatin immunoprecipitation and sequencing data, we selected 34 putative enhancers in IA -associated regions from genome-wide association studies. We then used a chromatin conformation capture technique to prioritize target genes and found that 15 putative enhancers interact with the promoters of 6 target genes: SOX 17, CDKN 2B, MTAP , CNNM 2, RPEL 1, and GATA 6. Subsequently, we assessed the activity of these putative enhancers in vivo in zebrafish embryos and confirmed activity for 8 putative enhancers. Last, we found that all 6 target genes are expressed in the circle of Willis, on the basis of RNA sequencing data and in situ hybridization. Furthermore, in situ hybridization showed that these genes are expressed in multiple cell types in the circle of Willis. Conclusions In 4 of 6 IA -associated genome-wide association study regions, we identified 8 putative enhancers that are active in vivo and interact with 6 nearby genes, suggesting that these genes are regulated by the identified putative enhancers. These genes, SOX 17, CDKN 2B, MTAP , CNNM 2, RPEL 1, and GATA 6, are therefore potential candidate genes involved in IA pathogenesis and should be studied using animal models in the future.
背景 我们之前的研究表明，颅内动脉瘤（IA）相关的单核苷酸多态性在人脑中动脉的启动子和潜在增强子中富集，IA 就是在这些部位发育的，这表明启动子和增强子在 IA 的发病机制中发挥作用。我们通过鉴定潜在增强子的潜在靶基因并验证其调控活性，进一步研究了潜在增强子在 IA 发病机制中的作用。
方法和结果 利用我们之前发表的大脑中动脉染色质免疫沉淀和测序数据，我们从全基因组关联研究中选择了 34 个与 IA 相关区域的潜在增强子。然后，我们使用染色质构象捕获技术来确定靶基因，并发现 15 个潜在增强子与 6 个靶基因的启动子相互作用：SOX17、CDKN2B、MTAP、CNNM2、RPEL1 和 GATA6。随后，我们在斑马鱼胚胎中评估了这些潜在增强子的体内活性，并证实了 8 个潜在增强子的活性。最后，我们根据 RNA 测序数据和原位杂交发现，所有 6 个靶基因都在大脑中动脉表达。此外，原位杂交显示这些基因在大脑中动脉的多种细胞类型中表达。
结论 在 4 个与 IA 相关的全基因组关联研究区域中，我们鉴定了 8 个具有体内活性并与 6 个邻近基因相互作用的潜在增强子，这表明这些基因受鉴定的潜在增强子调控。因此，这些基因 SOX17、CDKN2B、MTAP、CNNM2、RPEL1 和 GATA6 是潜在的与 IA 发病机制相关的候选基因，应在未来使用动物模型进行研究。
