scholarly journals SMAD1 and SMAD5 Expression Is Coordinately Regulated by FLI1 and GATA2 during Endothelial Development

2015 ◽  
Vol 35 (12) ◽  
pp. 2165-2172 ◽  
Author(s):  
Jonathon Marks-Bluth ◽  
Anchit Khanna ◽  
Vashe Chandrakanthan ◽  
Julie Thoms ◽  
Thomas Bee ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Selective Advantage ◽  
Regulatory Elements ◽  
Bmp Signaling ◽  
Tissue Specific ◽  
Morphogenetic Protein ◽  
Smad Signaling Pathway ◽  
Endothelial Development

The bone morphogenetic protein (BMP)/SMAD signaling pathway is a critical regulator of angiogenic sprouting and is involved in vascular development in the embryo. SMAD1 and SMAD5, the core mediators of BMP signaling, are vital for this activity, yet little is known about their transcriptional regulation in endothelial cells. Here, we have integrated multispecies sequence conservation, tissue-specific chromatin,in vitroreporter assay, andin vivotransgenic data to identify and validateSmad1+63 and theSmad5promoter as tissue-specificcis-regulatory elements that are active in the developing endothelium. The activity of these elements in the endothelium was dependent on highly conserved ETS, GATA, and E-box motifs, and chromatin immunoprecipitation showed high levels of enrichment of FLI1, GATA2, and SCL at these sites in endothelial cell lines and E11 dorsal aortasin vivo. Knockdown of FLI1 and GATA2 but not SCL reduced the expression of SMAD1 and SMAD5 in endothelial cellsin vitro. In contrast, CD31+cKit−endothelial cells harvested from embryonic day 9 (E9) aorta-gonad-mesonephros (AGM) regions of GATA2 null embryos showed reducedSmad1but notSmad5transcript levels. This is suggestive of a degree ofin vivoselection where, in the case of reduced SMAD1 levels, endothelial cells with more robust SMAD5 expression have a selective advantage.

scholarly journals Fibrillin-1 and -2 differentially modulate endogenous TGF-β and BMP bioavailability during bone formation

2010 ◽  
Vol 190 (6) ◽  
pp. 1107-1121 ◽  
Author(s):  
Harikiran Nistala ◽  
Sui Lee-Arteaga ◽  
Silvia Smaldone ◽  
Gabriella Siciliano ◽  
Luca Carta ◽  
...  
Keyword(s):  
Bone Formation ◽  
Transforming Growth Factor ◽  
Bone Mineralization ◽  
Bmp Signaling ◽  
Direct Role ◽  
Morphogenetic Protein ◽  
Fibrillin 1 ◽  
Osteoblast Maturation

Extracellular regulation of signaling by transforming growth factor (TGF)–β family members is emerging as a key aspect of organ formation and tissue remodeling. In this study, we demonstrate that fibrillin-1 and -2, the structural components of extracellular microfibrils, differentially regulate TGF-β and bone morphogenetic protein (BMP) bioavailability in bone. Fibrillin-2–null (Fbn2−/−) mice display a low bone mass phenotype that is associated with reduced bone formation in vivo and impaired osteoblast maturation in vitro. This Fbn2−/− phenotype is accounted for by improper activation of latent TGF-β that selectively blunts expression of osterix, the transcriptional regulator of osteoblast maturation, and collagen I, the structural template for bone mineralization. Cultured osteoblasts from Fbn1−/− mice exhibit improper latent TGF-β activation as well, but mature faster because of increased availability of otherwise matrix-bound BMPs. Additional in vitro evidence excludes a direct role of microfibrils in supporting mineral deposition. Together, these findings identify the extracellular microfibrils as critical regulators of bone formation through the modulation of endogenous TGF-β and BMP signaling.

The rat albumin promoter is composed of six distinct positive elements within 130 nucleotides

1989 ◽  
Vol 9 (11) ◽  
pp. 4750-4758
Author(s):  
P Herbomel ◽  
A Rollier ◽  
F Tronche ◽  
M O Ott ◽  
M Yaniv ◽  
...  
Keyword(s):  
Regulatory Elements ◽  
Negative Regulation ◽  
Base Pairs ◽  
Tissue Specific ◽  
Positive Elements ◽  
Specific Factors ◽  
Ccaat Box ◽  
Rat Hepatoma

No fewer than six different positive regulatory elements concentrated within 130 base pairs constitute the rat albumin promoter, which drives highly tissue specific transcription in rat hepatoma cells in culture. Inactivation of each element led to a decrease in transcriptional efficiency: from upstream to downstream, 3- to 4-fold for distal elements III and II, 15-fold for distal element I, and 50-fold for the CCAAT box and the proximal element (PE). Three of these elements, distal elements III and II and, more crucially, the PE, were found to be involved in the tissue-specific character of transcription, with an additional negative regulation possibly superimposed at the level of the PE. Finally, our mapping of these regulatory elements in vivo entirely coincided with footprint data obtained in vitro, thereby allowing the tentative assignment of specific factors to the effects observed in vivo.

scholarly journals Crossveinless 2 regulates bone morphogenetic protein 9 in human and mouse vascular endothelium

Blood ◽  
2012 ◽  
Vol 119 (21) ◽  
pp. 5037-5047 ◽  
Author(s):  
Yucheng Yao ◽  
Medet Jumabay ◽  
Albert Ly ◽  
Melina Radparvar ◽  
Anthony H. Wang ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Vascular Endothelium ◽  
Feedback Inhibition ◽  
Tube Formation ◽  
Bmp Signaling ◽  
Matrix Gla Protein ◽  
Morphogenetic Protein ◽  
Mutual Regulation ◽  
Bone Morphogenetic Protein 9

Abstract The importance of morphogenetic proteins (BMPs) and their antagonists in vascular development is increasingly being recognized. BMP-4 is essential for angiogenesis and is antagonized by matrix Gla protein (MGP) and crossveinless 2 (CV2), both induced by the activin receptor like-kinase 1 (ALK1) when stimulated by BMP-9. In this study, however, we show that CV2 preferentially binds and inhibits BMP-9 thereby providing strong feedback inhibition for BMP-9/ALK1 signaling rather than for BMP-4/ALK2 signaling. CV2 disrupts complex formation involving ALK2, ALK1, BMP-4, and BMP-9 required for the induction of both BMP antagonists. It also limits VEGF expression, proliferation, and tube formation in ALK1-expressing endothelial cells. In vivo, CV2 deficiency translates into a dysregulation of vascular BMP signaling, resulting in an abnormal endothelium with increased endothelial cellularity and expression of lineage markers for mature endothelial cells. Thus, mutual regulation by BMP-9 and CV2 is essential in regulating the development of the vascular endothelium.

scholarly journals Use of Bmp1/Tll1 Doubly Homozygous Null Mice and Proteomics To Identify and Validate In Vivo Substrates of Bone Morphogenetic Protein 1/Tolloid-Like Metalloproteinases

2003 ◽  
Vol 23 (13) ◽  
pp. 4428-4438 ◽  
Author(s):  
William N. Pappano ◽  
Barry M. Steiglitz ◽  
Ian C. Scott ◽  
Douglas R. Keene ◽  
Daniel S. Greenspan
Keyword(s):  
Bone Morphogenetic Protein ◽  
Residual Activity ◽  
Functional Redundancy ◽  
Bmp Signaling ◽  
Null Mouse ◽  
Genetic Redundancy ◽  
Morphogenetic Protein ◽  
Bone Morphogenetic Protein 1

ABSTRACT Bone morphogenetic protein 1 (BMP-1) and mammalian Tolloid (mTLD), two proteinases encoded by Bmp1, provide procollagen C-proteinase (pCP) activity that converts procollagens I to III into the major fibrous components of mammalian extracellular matrix (ECM). Yet, although Bmp1 −/− mice have aberrant collagen fibrils, they have residual pCP activity, indicative of genetic redundancy. Mammals possess two additional proteinases structurally similar to BMP-1 and mTLD: the genetically distinct mammalian Tolloid-like 1 (mTLL-1) and mTLL-2. Mice lacking the mTLL-1 gene Tll1 are embryonic lethal but have pCP activity levels similar to those of the wild type, suggesting that mTLL-1 might not be an in vivo pCP. In vitro studies have shown BMP-1 and mTLL-1 capable of cleaving Chordin, an extracellular antagonist of BMP signaling, suggesting that these proteases might also serve to modulate BMP signaling and to coordinate the latter with ECM formation. However, in vivo evidence of roles for BMP-1 and mTLL-1 in BMP signaling in mammals is lacking. To remove functional redundancy obscuring the in vivo functions of BMP-1-related proteases in mammals, we here characterize Bmp1 Tll1 doubly null mouse embryos. Although these appear morphologically indistinguishable from Tll1 −/− embryos, biochemical analysis of cells derived from doubly null embryos shows functional redundancy removed to an extent enabling us to demonstrate that (i) products of Bmp1 and Tll1 are responsible for in vivo cleavage of Chordin in mammals and (ii) mTLL-1 is an in vivo pCP that provides residual activity observed in Bmp1 −/− embryos. Removal of functional redundancy also enabled use of Bmp1 −/− Tll1 −/− cells in a proteomics approach for identifying novel substrates of Bmp1 and Tll1 products.

scholarly journals Immunohistologic and functional characterization of a vascular addressin involved in lymphocyte homing into peripheral lymph nodes.

1988 ◽  
Vol 107 (5) ◽  
pp. 1853-1862 ◽  
Author(s):  
P R Streeter ◽  
B T Rouse ◽  
E C Butcher
Keyword(s):  
Endothelial Cells ◽  
Lymph Node ◽  
Lymph Nodes ◽  
Peyer's Patches ◽  
Peripheral Lymph Node ◽  
Lymphocyte Homing ◽  
Tissue Specific ◽  
Peripheral Lymph

The tissue localization or "homing" of circulating lymphocytes is directed in part by specialized vessels that define sites of lymphocyte exit from the blood. In peripheral lymph nodes, mucosal lymphoid tissues (Peyer's patches and appendix), and sites of chronic inflammation, for example, lymphocytes leave the blood by adhering to and migrating between those endothelial cells lining postcapillary high endothelial venules (HEV). Functional analyses of lymphocyte interactions with HEV have shown the lymphocytes can discriminate between HEV in different tissues, indicating that HEV express tissue-specific determinants or address signals for lymphocyte recognition. We recently described such a tissue-specific "vascular addressin" that is selectively expressed by endothelial cells supporting lymphocyte extravasation into mucosal tissues and that appears to be required for mucosa-specific lymphocyte homing (Streeter, P. R., E. L. Berg, B. N. Rouse, R. F. Bargatze, and E. C. Butcher. 1988. Nature (Lond.). 331:41-46). Here we document the existence and tissue-specific distribution of a distinct HEV differentiation antigen. Defined by monoclonal antibody MECA-79, this antigen is expressed at high levels on the lumenal surface and in the cytoplasm of HEV in peripheral lymph nodes. By contrast, although MECA-79 stains many HEV in the mucosal Peyer's patches, expression in most cases is restricted to the perivascular or ablumenal aspect of these venules. In the small intestine lamina propria, a mucosa-associated site that supports the extravasation of lymphocytes, venules do not stain with MECA-79. Finally, we demonstrate that MECA-79 blocks binding of both normal lymphocytes and a peripheral lymph node-specific lymphoma to peripheral lymph node HEV in vitro and that it also inhibits normal lymphocyte homing to peripheral lymph nodes in vivo without significantly influencing lymphocyte interactions with Peyer's patch HEV in vitro or in vivo. Thus, MECA-79 defines a novel vascular addressin involved in directing lymphocyte homing to peripheral lymph nodes.

scholarly journals SMOC can act as both an antagonist and an expander of BMP signaling

eLife ◽  
2017 ◽  
Vol 6 ◽  
Author(s):  
J Terrig Thomas ◽  
D Eric Dollins ◽  
Kristin R Andrykovich ◽  
Tehyen Chu ◽  
Brian G Stultz ◽  
...  
Keyword(s):  
Calcium Binding ◽  
Mapk Signaling ◽  
Bmp Signaling ◽  
Mitogen Activated Protein Kinase ◽  
Matricellular Protein ◽  
Vitro Assay ◽  
Morphogenetic Protein ◽  
Mitogen Activated Protein

The matricellular protein SMOC (Secreted Modular Calcium binding protein) is conserved phylogenetically from vertebrates to arthropods. We showed previously that SMOC inhibits bone morphogenetic protein (BMP) signaling downstream of its receptor via activation of mitogen-activated protein kinase (MAPK) signaling. In contrast, the most prominent effect of the Drosophila orthologue, pentagone (pent), is expanding the range of BMP signaling during wing patterning. Using SMOC deletion constructs we found that SMOC-∆EC, lacking the extracellular calcium binding (EC) domain, inhibited BMP2 signaling, whereas SMOC-EC (EC domain only) enhanced BMP2 signaling. The SMOC-EC domain bound HSPGs with a similar affinity to BMP2 and could expand the range of BMP signaling in an in vitro assay by competition for HSPG-binding. Together with data from studies in vivo we propose a model to explain how these two activities contribute to the function of Pent in Drosophila wing development and SMOC in mammalian joint formation.

scholarly journals HNF1alpha is involved in tissue-specific regulation of CFTR gene expression

2004 ◽  
Vol 378 (3) ◽  
pp. 909-918 ◽  
Author(s):  
Nathalie MOUCHEL ◽  
Sytse A. HENSTRA ◽  
Victoria A. McCARTHY ◽  
Sarah H. WILLIAMS ◽  
Marios PHYLACTIDES ◽  
...  
Keyword(s):  
Gene Expression ◽  
Transcription Factor ◽  
Tissue Specificity ◽  
Regulatory Elements ◽  
Upstream Region ◽  
Mrna Levels ◽  
Tissue Specific ◽  
Specific Regulation

The CFTR (cystic fibrosis transmembrane conductance regulator) gene shows a complex pattern of expression with tissue-specific and temporal regulation. However, the genetic elements and transcription factors that control CFTR expression are largely unidentified. The CFTR promoter does not confer tissue specificity on gene expression, suggesting that there are regulatory elements outside the upstream region. Analysis of potential regulatory elements defined as DNase 1-hypersensitive sites within introns of the gene revealed multiple predicted binding sites for the HNF1α (hepatocyte nuclear factor 1α) transcription factor. HNF1α, which is expressed in many of the same epithelial cell types as CFTR and shows similar differentiation-dependent changes in gene expression, bound to these sites in vitro. Overexpression of heterologous HNF1α augmented CFTR transcription in vivo. In contrast, antisense inhibition of HNF1α transcription decreased the CFTR mRNA levels. Hnf1α knockout mice showed lower levels of CFTR mRNA in their small intestine in comparison with wild-type mice. This is the first report of a transcription factor, which confers tissue specificity on the expression of this important disease-associated gene.

scholarly journals The rat albumin promoter is composed of six distinct positive elements within 130 nucleotides.

1989 ◽  
Vol 9 (11) ◽  
pp. 4750-4758 ◽  
Author(s):  
P Herbomel ◽  
A Rollier ◽  
F Tronche ◽  
M O Ott ◽  
M Yaniv ◽  
...  
Keyword(s):  
Regulatory Elements ◽  
Negative Regulation ◽  
Base Pairs ◽  
Tissue Specific ◽  
Positive Elements ◽  
Specific Factors ◽  
Ccaat Box ◽  
Rat Hepatoma

No fewer than six different positive regulatory elements concentrated within 130 base pairs constitute the rat albumin promoter, which drives highly tissue specific transcription in rat hepatoma cells in culture. Inactivation of each element led to a decrease in transcriptional efficiency: from upstream to downstream, 3- to 4-fold for distal elements III and II, 15-fold for distal element I, and 50-fold for the CCAAT box and the proximal element (PE). Three of these elements, distal elements III and II and, more crucially, the PE, were found to be involved in the tissue-specific character of transcription, with an additional negative regulation possibly superimposed at the level of the PE. Finally, our mapping of these regulatory elements in vivo entirely coincided with footprint data obtained in vitro, thereby allowing the tentative assignment of specific factors to the effects observed in vivo.

Desmopressin (DDAVP) Enhances Platelet Adhesion to the Extracellular Matrix of Cultured Human Endothelial Cells through Increased Expression of Tissue Factor

1997 ◽  
Vol 77 (05) ◽  
pp. 0975-0980 ◽  
Author(s):  
Angel Gálvez ◽  
Goretti Gómez-Ortiz ◽  
Maribel Díaz-Ricart ◽  
Ginés Escolar ◽  
Rogelio González-Sarmiento ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Endothelial Cells ◽  
Tissue Factor ◽  
Platelet Adhesion ◽  
Umbilical Vein ◽  
Procoagulant Activity ◽  
Experimental Conditions ◽  
Chromogenic Assay

SummaryThe effect of desmopressin (DDAVP) on thrombogenicity, expression of tissue factor and procoagulant activity (PCA) of extracellular matrix (ECM) generated by human umbilical vein endothelial cells cultures (HUVEC), was studied under different experimental conditions. HUVEC were incubated with DDAVP (1, 5 and 30 ng/ml) and then detached from their ECM. The reactivity towards platelets of this ECM was tested in a perfusion system. Coverslips covered with DD A VP-treated ECMs were inserted in a parallel-plate chamber and exposed to normal blood anticoagulated with low molecular weight heparin (Fragmin®, 20 U/ml). Perfusions were run for 5 min at a shear rate of 800 s1. Deposition of platelets on ECMs was significantly increased with respect to control ECMs when DDAVP was used at 5 and 30 ng/ml (p <0.05 and p <0.01 respectively). The increase in platelet deposition was prevented by incubation of ECMs with an antibody against human tissue factor prior to perfusion. Immunofluorescence studies positively detected tissue factor antigen on DDAVP derived ECMs. A chromogenic assay performed under standardized conditions revealed a statistically significant increase in the procoagulant activity of the ECMs produced by ECs incubated with 30 ng/ml DDAVP (p <0.01 vs. control samples). Northern blot analysis revealed increased levels of tissue factor mRNA in extracts from ECs exposed to DDAVP. Our data indicate that DDAVP in vitro enhances platelet adhesion to the ECMs through increased expression of tissue factor. A similar increase in the expression of tissue factor might contribute to the in vivo hemostatic effect of DDAVP.

Epinephrine Induces von Willebrand Factor Release from Cultured Endothelial Cells: Involvement of Cyclic AMP-dependent Signalling in Exocytosis

1997 ◽  
Vol 77 (06) ◽  
pp. 1182-1188 ◽  
Author(s):  
Ulrich M Vischer ◽  
Claes B Wollheinn
Keyword(s):  
Endothelial Cells ◽  
Adenylate Cyclase ◽  
Von Willebrand Factor ◽  
Umbilical Vein ◽  
Free Calcium ◽  
Camp Content ◽  
Von Willebrand ◽  
Willebrand Factor

Summaryvon Willebrand factor (vWf) is released from endothelial cell storage granules after stimulation with thrombin, histamine and several other agents that induce an increase in cytosolic free calcium ([Ca2+]i). In vivo, epinephrine and the vasopressin analog DDAVP increase vWf plasma levels, although they are thought not to induce vWf release from endothelial cells in vitro. Since these agents act via a cAMP-dependent pathway in responsive cells, we examined the role of cAMP in vWf secretion from cultured human umbilical vein endothelial cells. vWf release increased by 50% in response to forskolin, which activates adenylate cyclase. The response to forskolin was much stronger when cAMP degradation was blocked with IBMX, an inhibitor of phosphodiesterases (+200%), whereas IBMX alone had no effect. vWf release could also be induced by the cAMP analogs dibutyryl-cAMP (+40%) and 8-bromo-cAMP (+25%); although their effect was weak, they clearly potentiated the response to thrombin. Epinephrine (together with IBMX) caused a small, dose-dependent increase in vWf release, maximal at 10-6 M (+50%), and also potentiated the response to thrombin. This effect is mediated by adenylate cyclase-coupled β-adrenergic receptors, since it is inhibited by propranolol and mimicked by isoproterenol. In contrast to thrombin, neither forskolin nor epinephrine caused an increase in [Ca2+]j as measured by fura-2 fluorescence. In addition, the effects of forskolin and thrombin were additive, suggesting that they act through distinct signaling pathways. We found a close correlation between cellular cAMP content and vWf release after stimulation with epinephrine and forskolin. These results demonstrate that cAMP-dependent signaling events are involved in the control of exocytosis from endothelial cells (an effect not mediated by an increase in [Ca2+]i) and provide an explanation for epinephrine-induced vWf release.

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