An Intronic Flk1 Enhancer Directs Arterial-Specific Expression via RBPJ-Mediated Venous Repression.
An Intronic Flk1 Enhancer Directs Arterial-Specific Expression via RBPJ-Mediated Venous Repression.
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DOI：
10.1161/atvbaha.116.307517
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发表时间：
2016-06
期刊：
影响因子：
--
通讯作者：
De Val S
中科院分区：
文献类型：
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作者：
Becker PW;Sacilotto N;Nornes S;Neal A;Thomas MO;Liu K;Preece C;Ratnayaka I;Davies B;Bou-Gharios G;De Val S
文本中提供了补充数字内容。血管内皮生长因子 (VEGF) 受体 Flk1 对于血管发育至关重要，但在内皮细胞中调节其表达的信号传导和转录途径仍不清楚。尽管之前的研究已经鉴定出 2 个 Flk1 调节增强子，但这些对于 Flk1 表达来说是可有可无的，表明额外的增强子有助于内皮细胞中的 Flk1 调节。在本研究中，我们试图鉴定有助于内皮细胞表达的 Flk1 增强子。 Flk1 基因第 10 个内含子 (Flk1in10) 的一个区域被鉴定为假定的增强子，并在小鼠和斑马鱼转基因模型中进行了测试。该区域强有力地指导动脉内皮细胞中的报告基因表达。通过结合转录因子结合位点的定向诱变和转录因子的基因沉默，我们发现所有内皮细胞中的 Flk1in10 增强子活性都需要 Gata 和 Ets 因子。此外，我们发现，通过 Notch 通路转录调节因子 Rbpj 抑制静脉内皮细胞中的基因表达，Flk1in10 增强子的活性仅限于动脉。这项研究展示了一种新的动静脉同一性获取机制，表明了 Notch 和 VEGF 信号通路之间的直接联系，并说明了顺式调控多样性如何允许有限的转录调控因子产生差异表达结果。
Supplemental Digital Content is available in the text. The vascular endothelial growth factor (VEGF) receptor Flk1 is essential for vascular development, but the signaling and transcriptional pathways by which its expression is regulated in endothelial cells remain unclear. Although previous studies have identified 2 Flk1 regulatory enhancers, these are dispensable for Flk1 expression, indicating that additional enhancers contribute to Flk1 regulation in endothelial cells. In the present study, we sought to identify Flk1 enhancers contributing to expression in endothelial cells. A region of the 10th intron of the Flk1 gene (Flk1in10) was identified as a putative enhancer and tested in mouse and zebrafish transgenic models. This region robustly directed reporter gene expression in arterial endothelial cells. Using a combination of targeted mutagenesis of transcription factor–binding sites and gene silencing of transcription factors, we found that Gata and Ets factors are required for Flk1in10 enhancer activity in all endothelial cells. Furthermore, we showed that activity of the Flk1in10 enhancer is restricted to arteries through repression of gene expression in venous endothelial cells by the Notch pathway transcriptional regulator Rbpj. This study demonstrates a novel mechanism of arterial–venous identity acquisition, indicates a direct link between the Notch and VEGF signaling pathways, and illustrates how cis-regulatory diversity permits differential expression outcomes from a limited repertoire of transcriptional regulators.