Angiotensin II (ATII)-inducible Platelet-derived Growth Factor A-chain Gene Expression Is p42/44 Extracellular Signal-regulated Kinase-1/2 and Egr-1-dependent and Mediated via the ATII Type 1 but Not Type 2 Receptor

Abstract

Angiotensin II (ATII) and platelet-derived growth factor (PDGF) are two vasoconstrictors implicated in the maintenance of normal vascular homeostasis. PDGF Achain levels increase in cultured vascular smooth muscle cells (SMCs) exposed to ATII. The molecular mechanisms underlying this induction are not known. We used transient transfection analysis to show that ATII can increase reporter gene activity driven by fragments of the PDGF-A promoter bearing recognition elements for the transcription factor, Egr-1. Nuclear run-off experiments indicate that ATII induces Egr-1 expression at the level of transcription. Gel shift and supershift studies show that Egr-1 protein accumulates in the nuclei of SMCs exposed to ATII and binds to the proximal region of the PDGF-A promoter in a specific, time-dependent manner. ATII induced extracellular-signal regulated kinase (p42/44 ERK) activity as did phorbol 12-myristate 13-acetate. The specific MEK1/2 inhibitor, PD98059, suppressedbothPDGF-AandEgr-1endogenousandpromoter-dependent expression inducible by ATII. The ATII type 1 receptor (AT1) antagonist, Losartan, inhibited ATII-induction of p42/44 ERK, as well as Egr-1 and PDGF-A, whereas neither PD123319, an AT2 receptor antagonist, nor wortmannin, an inhibitor of phosphatidylinositol 3-kinase and c-Jun N-terminal kinase, had any effect. ATII-induction of Egr-1 and PDGF-A was blocked by SIN-1, a NO donor. In addition, this pathway was blocked by overexpression of NO synthase. Collectively, these findings demonstrate that ATII activation of the PDGF-A promoter is mediated via the MEK/ERK/Egr-1 pathway and AT1 receptor and that this process is antagonized by NO.Angiotensin II (ATII), 1 a peptide hormone with potent vasoconstrictor activity, has long been implicated in the pathobiology of hypertension. In vascular smooth muscle cells (SMCs), ATII stimulates protein synthesis (1), cellular hypertrophy (2-5), migration (6), extracellular matrix synthesis (7,8), and the activation of a large number of transcription factors. These include Jak/STAT (9), Ets-1 (10), SRF (11), MHox (11), c-Jun (12, 13), JunB (12), and c-Fos (14). ATII is produced in the vessel wall by the actions of renin, which converts angiotensinogen to ATI, which is then cleaved to ATII by angiotensin-converting enzyme. Two ATII receptor subtypes have been described, AT1 and AT2. Signal transduction through G-protein-coupled AT1 receptors involves phospholipase C, phospholipase A 2 , phospholipase D, adenylate cyclase, and the release of intracellular calcium (reviewed in Refs. 15 and 16). The AT1 receptor also regulates neointimal thickening after mechanical injury to the rat carotid artery wall and ATII infusion (17). AT2 receptor signaling is less well understood, but evidence suggests that this receptor is involved in growth inhibition (18), Bcl-2 dephosphorylation (19), and apoptosis (20, 21). Platelet-derived growth factor (PDGF) consists of an A-chain and B-chain held together in homo-or heterodimeric configuration by disulfide bonds (reviewed in Refs. 22 and 2...

Opposing Actions of Stat1 and Stat6 on IL-13-Induced Up-Regulation of Early Growth Response-1 and Platelet-Derived Growth Factor Ligands in Pulmonary Fibroblasts

“…Several studies have shown that ERK regulates Egr-1-mediated PDGF-A and PDGF-C expression (19,32,33). We found that inhibition of the ERK signaling pathway with the MEK inhibitor PD98059 did not reduce IL-13-stimulated Egr-1 production but instead increased Stat6 phosphorylation and resulted in increased Egr-1 protein expression (Fig.…”

Regulation of Inducible Heparanase Gene Transcription in Activated T Cells by Early Growth Response 1

“…We have shown this region binds Egr1, and we propose that it also binds Sp1 in our T cell model. In support of this, the C3 complex observed in our EMSA studies in unstimulated cell nuclear extracts is likely to be Sp1 as follows: (i) it has been shown previously that Sp1 binds to this sequence of the HPSE promoter in thyroid tumor cells (34); (ii) the C3 complex is lost when the GGCG motif is mutated; and (iii) previous studies using a consensus Egr1-binding oligonucleotide in EMSA have identified a C1 complex that contains Sp1 (37), and this complex exhibits a similar electrophoretic mobility to the C3 complex identified in our studies. Significantly, Sp1 has been shown to compete with Egr1 for binding to promoter elements.…”

“…Nuclear extraction was then performed as described previously (37). Nuclear extracts were incubated for 30 min at room temperature with 10 4 to 1.5 ϫ 10 4 cps of 32 P-labeled double-stranded oligonucleotides in a 20-l reaction volume containing 10 mM Tris-HCl, pH 8.0, 50 mM MgCl 2 , 1 mM EDTA, 1 mM dithiothreitol, 5% glycerol, 5% sucrose, 1 mM phenylmethylsulfonyl fluoride, 1 g of poly(dI⅐dC)-poly(dI⅐dC), and 1 g of salmon sperm DNA.…”

Mechanism of Erythropoietin Regulation by Angiotensin II

“…AT 1 R has been demonstrated to signal through a variety of downstream molecules, including GTP-binding proteins, SHPs, and ERK1/2 MAPK (Day et al, 1999;Godeny et al, 2007;Olson et al, 2008). We examined the activation of EPO gene expression in the presence of inhibitors of kinase and phosphatase pathways.…”