An abundant transcript induced in differentiating human endothelial cells encodes a polypeptide with structural similarities to G-protein-coupled receptors.

T Hla,T Maciag

doi:10.1016/s0021-9258(19)38849-0

Abstract

The tumor promoter phorbol 12-myristate 13-acetate (PMA) inhibits the growth of human endothelial cells and induces differentiation into capillary-like, tubular structures. We have isolated cDNA clones induced by PMA in the presence of cycloheximide and report the characterization of a novel immediate-early cDNA clone, termed edg-1, from human endothelial cells. The 3-kilobase edg-1 transcript is rapidly induced when endothelial cells are treated with PMA and superinduced in the presence of cycloheximide. While superinduction is due, at least in part, to the stabilization of the edg-1 transcript, nuclear run-on analysis demonstrates that the transcription of edg-1 is stimulated by PMA. Although the edg-1 transcript is very abundant in endothelial cells, transcripts related to human edg-1 are also detected at lower levels in vascular smooth muscle cells, fibroblasts, melanocytes, and cells of epithelioid origin. The deduced polypeptide sequence of edg-1 contains seven transmembrane domains with significant structural similarities to G-protein-coupled receptors (GPRs). Although the identity of the ligand for edg-1 is presently unknown, the structure of edg-1 polypeptide strongly implies that the edg-1 translation product is an inducible endothelial cell GPR. Since GPRs are involved in diverse biological processes such as signal transduction, cell proliferation, and differentiation, the characterization of human edg-1 as a highly inducible and abundant endothelial cell GPR suggest that it may be involved in the processes that regulate the differentiation of endothelial cells.

Highlights

13-acetate (PMA) inhibits the growth of human endothelial cells and induces differentiation into capillary-like, tubular structures
Poly(A+) RNA (10 fig) from human umbilical vein endothelial cells (HUVEC) exposed to phorbol 12-myristate 13-acetate (PMA), and chx was converted to double-stranded cDNA and cloned into the
To obtain cDNAs representing mRNAs induced by PMA in HUVEC, we performed differential hybridization with cDNA probes derived from quiescent HUVEC either treated or not treated with 20 rig/ml PMA and 5 rg/ml cycloheximide for 4 h

Summary

Laboratory for the Biomedical

13-acetate (PMA) inhibits the growth of human endothelial cells and induces differentiation into capillary-like, tubular structures. Polypeptide cytokines, such as interleukin-1, tumor necrosis factor-a, y-interferon, transforming growth factor-p as well as phorbol 12-myristate 13-acetate (PMA)l inhibit the growth of endothelial cells and promote the phenotypic transition from a nonpolar, cobblestone monolayer phenotype into a polar, elongated, fibroblast-like phenotype (3-S) This phenotype is similar to the morphology of endothelial cells during the early phases of the endothelial cell differentiation pathway in vitro [9]. PMA-inducible immediate-early transcript from human umbilical vein endothelial cells (HUVEC) We tentatively name this transcript as endothelial differentiation gene (edg)-1. Structural features of the deduced polypeptide of the edg-1 transcript suggest that it may be a novel member of the Gprotein-coupled receptor family These results implicate the involvement of G-protein-coupled receptors in the regulation of the early stage of human endothelial cell differentiation in vitro

EXPERIMENTAL PROCEDURES

RESULTS AND DISCUSSION

SF AHMBE