An Intronic Flk1 Enhancer Directs Arterial-Specific Expression via RBPJ-Mediated Venous Repression.

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.

Becker PW ，Sacilotto N ，Nornes S ，Neal A ，Thomas MO ，Liu K ，Preece C ，Ratnayaka I ，Davies B ，Bou-Gharios G ，De Val S ... -  《-》

ETS factors are required but not sufficient for specific patterns of enhancer activity in different endothelial subtypes.

Correct vascular differentiation requires distinct patterns of gene expression in different subtypes of endothelial cells. Members of the ETS transcription factor family are essential for the transcriptional activation of arterial and angiogenesis-specific gene regulatory elements, leading to the hypothesis that they play lineage-defining roles in arterial and angiogenic differentiation directly downstream of VEGFA signalling. However, an alternative explanation is that ETS binding at enhancers and promoters is a general requirement for activation of many endothelial genes regardless of expression pattern, with subtype-specificity provided by additional factors. Here we use analysis of Ephb4 and Coup-TFII (Nr2f2) vein-specific enhancers to demonstrate that ETS factors are equally essential for vein, arterial and angiogenic-specific enhancer activity patterns. Further, we show that ETS factor binding at these vein-specific enhancers is enriched by VEGFA signalling, similar to that seen at arterial and angiogenic enhancers. However, while arterial and angiogenic enhancers can be activated by VEGFA in vivo, the Ephb4 and Coup-TFII venous enhancers are not, suggesting that the specificity of VEGFA-induced arterial and angiogenic enhancer activity occurs via non-ETS transcription factors. These results support a model in which ETS factors are not the primary regulators of specific patterns of gene expression in different endothelial subtypes.

Neal A ，Nornes S ，Louphrasitthiphol P ，Sacilotto N ，Preston MD ，Fleisinger L ，Payne S ，De Val S ... -  《-》

Convergence of Notch and beta-catenin signaling induces arterial fate in vascular progenitors.

Yamamizu K ，Matsunaga T ，Uosaki H ，Fukushima H ，Katayama S ，Hiraoka-Kanie M ，Mitani K ，Yamashita JK ... -  《-》

FoxH1 negatively modulates flk1 gene expression and vascular formation in zebrafish.

Choi J ，Dong L ，Ahn J ，Dao D ，Hammerschmidt M ，Chen JN ... -  《-》

Specification of arterial, venous, and lymphatic endothelial cells during embryonic development.

Kume T 《-》