Endoglin expression in the endothelium is regulated by Fli-1, Erg, and Elf-1 acting on the promoter and a –8-kb enhancer

Abstract

Angiogenesis is critical to the growth and regeneration of tissue but is also a key component of tumor growth and chronic inflammatory disorders. Endoglin plays a key role in angiogenesis by modulating cellular responses to transforming growth factor-␤ (TGF-␤) signaling and is upregulated in proliferating endothelial cells. To gain insights into the transcriptional hierarchies that govern endoglin expression, we used a combination of comparative genomic, biochemical, and transgenic approaches. Both the promoter and a region 8 kb upstream of exon 1 were active in transfection assays in endothelial cells. In transgenic mice, the promoter directed low-level expression to a subset of endothelial cells. By contrast, inclusion of the ؊8 enhancer resulted in robust endothelial activity with additional staining in developing ear mesenchyme. Subsequent molecular analysis demonstrated that both the ؊8 enhancer and the promoter depend on conserved Ets sites, which were bound in endothelial cells in vivo by Fli-1, Erg, and Elf-1. This study therefore establishes the transcriptional framework within which endoglin functions during angiogenesis. IntroductionEndoglin is a transmembrane glycoprotein that is expressed predominantly in endothelial cells and is critical for the normal development of blood vessels. 1 Endoglin-null mice die in utero with defects in vascular remodeling, [2][3][4] and haploinsufficiency of the gene in humans is associated with type 1 hereditary hemorrhagic telangiectasia (HHT), an autosomal dominant disorder characterized by bleeding from vascular malformations in the skin, mucosa, and viscera (reviewed in Lebrin et al 5 and Abdalla and Letarte 6 ).Endoglin functions as an accessory receptor for members of the transforming growth factor-␤ (TGF-␤) superfamily of cytokines. 7 The TGF-␤ superfamily includes the TGF-␤ isoforms, nodal, activins, bone morphogenetic proteins (BMPs), and other related factors, which serve diverse functions during embryonic development and tissue homeostasis in the adult (reviewed in Massague 8 ). These cytokines assemble a primary receptor complex made up of type I and type II receptors on the surface of cells and control gene expression by receptor mediated phosphorylation of SMAD proteins. The type I receptors confer signal specificity by preferential phosphorylation of SMAD2/3 or SMAD1/5/8 isoforms, which broadly engage different binding partners and act on separate sets of target genes. Ancillary nonsignaling receptors such as endoglin regulate the pathway by selectively facilitating the binding of subsets of the TGF-␤ family members to the primary receptor complex. 9 Endoglin can also influence the cellular response to a particular cytokine by its preferential interaction with certain type I receptors. ALK1 is one such receptor, and mutations in this gene result in type 2 HHT, a clinical disorder that closely resembles that caused by mutations in the endoglin gene. 10 The endothelium is intimately involved in the development and progression of a number of pathologies, i...

PRH/HHex inhibits the migration of breast and prostate epithelial cells through direct transcriptional regulation of Endoglin

“…PRH also binds to sequences in the distal promoter near a previously characterised Enhancer (29,30) around 8kb upstream of the transcription start point. These regions both contain multiple copies of the core PRH binding site 5'TAAT'3.…”

“…This region contains three putative PRH binding sequences as defined by the presence of 5'TAAT3' motifs. Promoter sequences flanking a previously characterised enhancer (29,30) located 8kb upstream of first exon also contain multiple putative PRH binding sites. Primer pairs specific for promoter sequences from -7352 to -6914 and -8857 to -8398 (which contain 1 and 6 putative PRH binding sites respectively) show that PRH can also bind in these regions in PNT2-C2 cells (P3 and P4 in Fig.3D).…”

Dual requirement for the ETS transcription factors Fli-1 and Erg in hematopoietic stem cells and the megakaryocyte lineage

“…Although it is possible that they do so via distinct genetic pathways, published data suggest Fli-1 and Erg might regulate common target genes. Fli-1 and Erg have highly homologous DNA binding (Ets) domains, which recognize an identical Ets binding site (EBS) and in ChIP studies and reporter assays in cell lines, both are able to bind and transactivate promoters/enhancers of genes encoding the Ig heavy chain and endoglin, although it is not known whether binding of Fli-1 and Erg to regulator elements occurs simultaneously (15,21,22). Although current evidence suggests that Ets proteins bind DNA as monomers, the ability of Fli-1 and Erg to heterodimerize may serve as a mechanism to simultaneously target Erg and Fli-1 to distinct EBSs within the same region of common target genes, many of which have multiple EBSs.…”

A novel mode of enhancer evolution: The Tal1 stem cell enhancer recruited a MIR element to specifically boost its activity

“…The Tal1 enhancer has also been used to generate mouse models for inducible transgene expression, retroviral gene transfer, and Cre-mediated recombination as well as the development of acute myeloid leukemia mouse models (Murphy et al 2003;Eguchi et al 2005;Gothert et al 2005;Koschmieder et al 2005). At the molecular level, activity of the +19 element critically depends on binding sites for the Ets and GATA families of transcription factors and this combination of transcription factors constitutes a regulatory code employed by several other enhancers Pimanda et al 2006Pimanda et al , 2007aChan et al 2007;Landry et al 2008) that display similar expression patterns in transgenic assays.…”