Dataset Information

Localization of rat endothelin-converting enzyme to vascular endothelial cells and some secretory cells.

ABSTRACT: Endothelin is a potent vasoconstrictive peptide that is produced by vascular endothelial cells; it is formed from its precursor, big endothelin, by endothelin-converting enzyme (ECE). In this work, ECE was studied using specific monoclonal antibodies. In immunoblotting, ECE was estimated to be a 300 kDa protein on SDS/PAGE under non-reducing conditions, and 130 kDa under reducing conditions. Cross-linking experiments revealed that ECE is composed of two disulphide-linked subunits. Localization of ECE was studied at the cellular and subcellular levels in various rat tissues and cells. High-level expression of ECE was observed in membrane fractions of simian virus 40-transformed rat endothelial cells by immunoblotting, but the immunoreactive band was absent form aortic smooth muscle cells and cytosolic fractions of endothelial cells. In immunohistochemical analysis, ECE was found to be localized in the endothelial cells of the aorta, lung, kidney, liver and heart. Confocal immunofluorescent microscopy showed that most of the ECE in endothelial cells and cells transfected with ECE cDNA was clustered along the plasma membrane. Intact COS or CHO cells transfected with ECE cDNA rapidly and efficiently cleaved big endothelin-1 added to the culture medium. Thus endothelial cells express ECE on the plasma membrane and the active site of the enzyme faces outside the cells, i.e. it is an ectoenzyme. Other than endothelial cells, ECE was also present in some secretory cells. The enzyme was abundant in the adrenal gland, and localized in chromaffin cells. ECE was also highly condensed in pancreatic islet beta cells. It is concluded that ECE and endothelin may be involved in the regulated secretion of hormones.

SUBMITTER: Takahashi M

PROVIDER: S-EPMC1136050 | biostudies-other | 1995 Oct

REPOSITORIES: biostudies-other

ACCESS DATA

Similar Datasets

Project description:Glioblastoma (GBM) is the most common and aggressive type of brain tumor due to its elevated recurrence following treatments. This is mainly mediated by a subpopulation of cells with stemness traits termed glioblastoma stem-like cells (GSCs), which are extremely resistant to anti-neoplastic drugs. Thus, an advancement in the understanding of the molecular processes underlying GSC occurrence should contribute significantly towards progress in reducing aggressiveness. High levels of endothelin-converting enzyme-1 (ECE1), key for endothelin-1 (ET-1) peptide activation, have been linked to the malignant progression of GBM. There are four known isoforms of ECE1 that activate ET-1, which only differ in their cytoplasmic N-terminal sequences. Isoform ECE1c is phosphorylated at Ser-18 and Ser-20 by protein kinase CK2, which increases its stability and hence promotes aggressiveness traits in colon cancer cells. In order to study whether ECE1c exerts a malignant effect in GBM, we designed an ECE1c mutant by switching a putative ubiquitination lysine proximal to the phospho-serines Lys-6-to-Arg (i.e., K6R). This ECE1cK6R mutant was stably expressed in U87MG, T98G, and U251 GBM cells, and their behavior was compared to either mock or wild-type ECE1c-expressing clone cells. ECE1cK6R behaved as a highly stable protein in all cell lines, and its expression promoted self-renewal and the enrichment of a stem-like population characterized by enhanced neurospheroid formation, as well as increased expression of stem-like surface markers. These ECE1cK6R-derived GSC-like cells also displayed enhanced resistance to the GBM-related chemotherapy drugs temozolomide and gemcitabine and increased expression of the ABCG2 efflux pump. In addition, ECE1cK6R cells displayed enhanced metastasis-associated traits, such as the modulation of adhesion and the enhancement of cell migration and invasion. In conclusion, the acquisition of a GSC-like phenotype, together with heightened chemoresistance and invasiveness traits, allows us to suggest phospho-ECE1c as a novel marker for poor prognosis as well as a potential therapeutic target for GBM.

Dataset Information

Localization of rat endothelin-converting enzyme to vascular endothelial cells and some secretory cells.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets