Human Endothelial Cell Storage Granules

Russell, Fraser D.; Skepper, Jeremy N.; Davenport, Anthony P.

doi:10.1161/01.res.83.3.314

Cited by 88 publications

(24 citation statements)

References 42 publications

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“…Following removal of the signal peptides, the inactive intermediate big ET is formed consisting of 38 -41 amino acids. ECE then cleaves big ET between Trp 21 and Val 22 /Ile 22 to generate the mature active 21-amino acid protein (1,5). The importance of this cleavage site is demonstrated by the fact that the vasoconstricting activity of either ET-1-(1-20) amino acids or ET-1-(1-22) amino acids is three orders of magnitude weaker than ET-1-(1-21) amino acids (6).…”

Section: Endothelin-converting Enzyme (Ece)mentioning

confidence: 99%

Constitutive Phosphorylation of Human Endothelin-converting Enzyme-1 Isoforms

MacLeod¹,

Husain

Gage

et al. 2002

Journal of Biological Chemistry

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To investigate the phosphorylation of human endothelin-converting enzyme-1 (hECE-1) and identify potential residues involved, both in vivo and in vitro phosphorylation labeling assays of hECE-1 isoforms were performed in combination with site-directed mutagenesis and mass spectrometric analyses. Initial studies found that endogenous hECE-1 was constitutively phosphorylated in a primary endothelial cell line. The four known isoforms of hECE-1 expressed in this cell line (1a, 1b, 1c, and 1d) were then cloned by reverse transcription-PCR to determine which isoform(s) may be phosphorylated. The isoforms differ only in the first portion of their short aminoterminal cytoplasmic domains whereas their transmembrane domains and ectodomains of the proteins are identical. Isoforms 1b, 1c, and 1d but not 1a, were constitutively phosphorylated in vivo when expressed in Chinese hamster ovary cells and casein kinase I readily phosphorylated the immunopurified isoforms in vitro. Site-directed mutagenesis established that two conserved serine residues, Ser 18 and Ser 20, (numbering based on isoform 1c) form at least one phosphorylation site in these three isoforms. Mutant forms of 1b, 1c, and 1d were constructed in which a single alanine was introduced at either serine residue and a double mutant for each isoform was constructed as well in which both serines were replaced with alanine. Phosphorylation of the single mutants was greatly reduced and was nearly abolished in the double mutants in both in vivo and in vitro labeling assays. Analysis by MALDI-MS of 32 P-labeled proteolytic peptides derived from wild type 1c and the 1c mutants supported both Ser 18 and Ser 20 as phosphorylated residues. These data demonstrate the first finding that hECE-1 is constitutively phosphorylated within its cytoplasmic domain in an isoform-specific manner.

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Section: Endothelin-converting Enzyme (Ece)mentioning

confidence: 99%

Constitutive Phosphorylation of Human Endothelin-converting Enzyme-1 Isoforms

MacLeod¹,

Husain

Gage

et al. 2002

Journal of Biological Chemistry

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“…Recent studies suggest that transport through vesicular channels and not transcytosis is dominant in the rat aorta [9]. In addition to molecules transported from the luminal to the basal surface or back, products synthesized in the endothelial cells such as adenosine triphosphate [10], big endothelin [11, 12], endothelin [11, 13, 14], von Willebrand factor [12, 15, 16], platelet-derived growth factor (PDGF) and other growth factors [17, 18] are stored in membrane-bound vesicles and secreted in response to specific stimuli. Thrombin localization and internalization into such vesicles were also described in cultured human umbilical vein endothelial cells [19].…”

Section: Introductionmentioning

confidence: 99%

“…Thrombin localization and internalization into such vesicles were also described in cultured human umbilical vein endothelial cells [19]. Additionally, endothelin-converting enzyme [12,20,21,22] and nitric oxide synthase [23] were reported to be linked to the vesicular membrane.…”

Section: Introductionmentioning

confidence: 99%

Selective Suppression of an Endothelin and Platelet-Derived Growth Factor Containing Vesicular System in Endothelium of Rat Saphenous Vein by Long-Term Orthostasis

et al. 2005

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Electron-dense vesicles were observed in rat vascular endothelium. The purpose of this study was to characterize their content(s), venous-arterial distribution and response to chronic orthostatic stress in extremity vessels. Saphenous and brachial vessels – saphenous vein (SV), saphenous artery (SA), brachial vein, brachial artery – were prepared for electron microscopy to quantitate the vesicle area within the endothelium following immunohistochemical and immunocytochemical identification. The effect of long-term orthostasis was assessed by exposure to head-up tilt for 2 weeks. The vesicular area in relation to the total cross-sectional area of the endothelial cells in the SV and SA of normal and confined control groups was 3.88 ± 0.38 versus 0.89 ± 0.06% (p < 0.05) and 4.92 ± 0.25 versus 1.09 ± 0.47% (p < 0.05), respectively. Head-up tilt suppressed the vesicle content of the SV to 2.26 ± 0.39% (p < 0.05), but it remained low in the SA (1.29 ± 0.45%), brachial vein (0.45 ± 0.12%) and brachial artery (0.59 ± 0.17%). Endothelin and platelet-derived growth factor, but not acidic phosphatase activity or lipid content, could be identified in the vesicles. Plasma endothelin levels were unchanged. We conclude that dense vesicles in the endothelium of extremity vessels are not cell degradation products. They may represent a vesicular secretory or storage system for endothelin and platelet-derived growth factor which participates in regional vascular adaptation to long-term orthostatic load.

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“…Extensive work aimed to define the intracellular localization of ECE-1 has been done in endothelial cells (21,35,36). Most of these studies were however non relevant to isoform specificity due to the lack of antibodies that can distinguish between ECE-1b, 1c, and 1d.…”

mentioning

confidence: 99%

Heterodimerization of Endothelin-converting Enzyme-1 Isoforms Regulates the Subcellular Distribution of This Metalloprotease

Muller

Barret

Etienne

et al. 2003

Journal of Biological Chemistry

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Endothelin-converting enzyme (ECE) is a membrane metalloprotease that generates endothelin from its direct precursor big endothelin. Four isoforms of ECE-1 are produced from a single gene through the use of alternate promoters. These isoforms share the same extracellular catalytic domain and contain unique cytosolic tails, which results in their specific subcellular targeting. We investigated the distribution of ECE-1 isoforms in transfected AtT-20 neuroendocrine cells. Whereas ECE-1a and 1c were present at the plasma membrane, ECE-1b and ECE-1d were retained inside the cells. We found that both intracellular isoforms were concentrated in the endosomal system: ECE-1d in recycling endosomes, and ECE-1b in late endosomes/multivesicular bodies. Leucine-based motifs were involved in the intracellular retention of these isoforms, and the targeting of ECE-1b to the degradation pathway required an additional signal in the N terminus. The concentration of ECE-1 isoforms in the endosomal system suggested new functions for these enzymes. Potential novel functions include redistribution of other isoforms through direct interaction. We have showed that ECE-1 isoforms could heterodimerize, and that in such heterodimers the ECE-1b targeting signal was dominant. Interaction of a plasma membrane isoform with ECE-1b resulted in its intracellular localization and decreased its extracellular activity. These data demonstrated that the targeting signals specific for ECE-1b constitute a regulatory domain per se that could modulate the localization and the activity of other isoforms.Endothelins (ET 1 -1, ET-2, and ET-3) are 21-residue peptides derived from three distinct genes (1). They are pleiotropic factors that play an important role in the regulation of the cardiovascular and endocrine systems (for review see Refs. 2 and 3). Endothelins are also crucial developmental factors as demonstrated by the targeted disruption of the genes coding for the precursors of ET-1 and ET-3, and of the genes coding for their receptors, ETA and ETB (4 -7). In addition, the expression of endothelin is associated with many pathological processes and with tumor growth (2, 8 -10). In order to fulfill such a wide spectrum of physiological functions, endothelins act through autocrine and paracrine mechanisms. Their biosynthesis thus requires tight local control. Endothelins are synthesized in the endoplasmic reticulum as precursors that undergo a two-step proteolytic maturation. Pro-endothelins are first processed at conserved multibasic sites by furin or a furin-like enzyme, in order to release an intermediate called big endothelin (big-ET) (11, 12), which is devoid of biological activity (13). Big-ET is then processed by endothelin converting enzyme (ECE) at a Trp-Val/Ile bond, which releases the biologically active peptide. This latter proteolytic step can occur in the extracellular medium and in the secretory pathway, so that cells secrete either big-ETs alone or together with endothelins (14, 15, 16). Endothelial cells co-express the precursor, the c...

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Human Endothelial Cell Storage Granules

Cited by 88 publications

References 42 publications

Constitutive Phosphorylation of Human Endothelin-converting Enzyme-1 Isoforms

Constitutive Phosphorylation of Human Endothelin-converting Enzyme-1 Isoforms

Selective Suppression of an Endothelin and Platelet-Derived Growth Factor Containing Vesicular System in Endothelium of Rat Saphenous Vein by Long-Term Orthostasis

Heterodimerization of Endothelin-converting Enzyme-1 Isoforms Regulates the Subcellular Distribution of This Metalloprotease

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