Mechanisms of Estrogen Receptor Action in the Myocardium

Abstract

We have previously shown that the myocardium is a target tissue for estrogen. Here, we have identified rapid non-nuclear estrogen effects on the expression of the early growth response gene-1 (Egr-1) in cardiomyocytes. Egr-1 mRNA and protein were rapidly and strongly induced by estrogen in an estrogen receptordependent manner via the extracellular signal-regulated kinase, ERK1/2. A promoter analysis study of a 1.2-kilobase Egr-1 promoter fragment revealed that the serum response elements (SREs) but not the estrogen response elements or AP-1 sites are responsible for Egr-1 induction by estrogen, identifying a novel mechanism of estrogen receptor-dependent gene activation in the myocardium. Both estrogen receptor-␣ and -␤ induced the Egr-1 promoter via the SREs as well as an artificial promoter consisting of only five SREs in cardiomyocytes. Electrophoretic mobility shift assays showed that a protein complex containing serum response factor or an antigenically related protein was recruited to the SREs by estrogen treatment of primary cardiomyocytes. The recruitment of the protein complex was inhibited by the specific estrogen receptor antagonist ICI 182,780 as well as the MEK inhibitor PD 98059. Taken together, these results identify SREs as important promoter control elements for an estrogen receptordependent mechanism of gene activation in the myocardium.Gender-specific differences in the occurrence of cardiac disease and the protective role of estrogens in the heart have been established in numerous clinical studies (1, 2). Hormone replacement therapy may prevent the primary development of coronary artery disease in postmenopausal women (1) but have not been effective in secondary prevention trials (3). These findings call for a better understanding of the role and function of estrogen and estrogen-like substances in the heart. The vast majority of studies have concentrated on vascular hormone effects (4, 5). However, cardioprotection by estrogen is not necessarily restricted to the vasculature. We have previously shown that functional estrogen receptors in the myocardium regulate the expression of relevant target genes such as connexin 43 (6) and ␣-myosin heavy chain, one of the major contractile proteins in the heart (7).In addition to the classical genomic regulation of target gene expression, a new role for estrogens as mediators of rapid non-genomic effects has recently been identified. These rapid effects include the stimulation of nitric oxide release in vascular cells (8, 9) and the activation of Ras and Raf-1 kinase (10), resulting in the activation of ERK1/2 1 and the phosphorylation of transcription factors in both non-myocytes (11-15) and cardiomyocytes (16). In the present work we have for the first time analyzed in detail one of the mechanisms of rapid gene induction by estrogen in the myocardium by performing a detailed promoter analysis of a bona fide estrogen target gene identified by us, the early growth response gene 1 (Egr-1).Surprisingly, a cluster of serum response elements (SREs) turned o...

In vivo oestrogenic modulation of Egr1 and Pitx1 gene expression in female rat pituitary gland

“…The significance of SRE sites in oestrogen-induced early response gene transcriptional activity was also reported in ER-positive human breast carcinoma MCF-7 cells where both Egr1 (Chen et al 2004) and c-fos (Fos) promoter (Duan et al 2001) required SREdriven activation of intracellular ERK1/2 signalling and subsequent ELK1 phosphorylation. It has been suggested that rapid SRE-mediated Egr1 gene induction by both ERA (ESR1) and ERB (ESR2) represents a mechanism by which oestrogen might exert an array of effects through the modulation of Egr1 target gene expression (de Jager et al 2001). In this paper, however, we also report that prolonged 3-day oestradiol exposure led to an increased Egr1 mRNA expression in the anterior pituitary gland.…”

“…Indeed, a transient increase in Egr1 mRNA level was reported in the uterus of ovariectomised rats as early as possible within 2 h after oestradiol supplementation (Suva et al 1991), whereas in primary cardiomyocytes, Egr1 mRNA expression reached a maximum at 15 min after oestradiol stimulation. Moreover, co-transfected ERA (ESR1) and ERB (ESR2) rapidly enhanced Egr1 promoter activity in these cells and this effect required SREs but not ERE/AP1 site activation (de Jager et al 2001). The significance of SRE sites in oestrogen-induced early response gene transcriptional activity was also reported in ER-positive human breast carcinoma MCF-7 cells where both Egr1 (Chen et al 2004) and c-fos (Fos) promoter (Duan et al 2001) required SREdriven activation of intracellular ERK1/2 signalling and subsequent ELK1 phosphorylation.…”

“…As EGR1 protein accumulates, it regulates the transcription of secondarylike MAPK phosphatase 2 (Zhang et al 2001) and tertiarylike Lhb response genes (Call & Wolfe 2002). Several cis-acting regulatory sites were identified on the 5 0 -flanking Egr1 gene sequence: two oestrogen response element (ERE) half-palindromic sites, two AP1 sites located in an upstream promoter region, one cAMP response element (CRE), five functional serum response elements (SREs) encompassing the consensus sequence CC(A/T) 6 GG (de Jager et al 2001, Duan et al 2002 and multiple binding sites for ternary complex factors having the Ets consensus core sequence (Mayer et al 2008). The importance of EGR1 in the pituitary is emphasised by the consequences of Egr1 gene knockout in mice.…”

Genome‐wide analysis of changes in early gene expression induced by oestrogen

“…It is known that activation of EGR-1 is a non-nuclear action of ER (de Jager et al 2001) and that the IGF-I gene is not regulated through the oestrogen response element in its transcriptional regulatory region. Although EGR-1 and IGF-I were activated much more in wildtype cells, activation of these genes in receptor-deficient mice might reflect other activation mechanisms not directly related to ER.…”

The Role of MAPKs in B Cell Receptor-induced Down-regulation of Egr-1 in Immature B Lymphoma Cells

“…In cardiomyocytes, estrogen induces the rapid induction of Egr-1 mRNA through activation of ERK1/2 (24). In human monocytes, the rapid induction of Egr-1 by lipopolysaccharide is dependent on the Ras/Raf-1/MEK/ERK pathway (19).…”