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Molecular Endocrinology 19(4):833–842 Copyright © 2005 by The Endocrine Society doi: 10.1210/me.2004-0486

MINIREVIEW

Mechanisms of Estrogen Receptor Signaling: Convergence of Genomic and Nongenomic Actions on Target Genes Linda Bjo¨rnstro¨m and Maria Sjo¨berg Department of Cell and Molecular Biology, Karolinska Institutet, SE-171 77 Stockholm, Sweden Estrogen receptors (ERs) act by regulating transcriptional processes. The classical mechanism of ER action involves estrogen binding to receptors in the nucleus, after which the receptors dimerize and bind to specific response elements known as estrogen response elements (EREs) located in the promoters of target genes. However, ERs can also regulate gene expression without directly binding to DNA. This occurs through protein-protein interactions with other DNA-binding transcription factors in the nucleus. In addition, membraneassociated ERs mediate nongenomic actions of estrogens, which can lead both to altered functions of proteins in the cytoplasm and to regulation of gene expression. The latter two mechanisms of ER action enable a broader range of genes to be

regulated than the range that can be regulated by the classical mechanism of ER action alone. This review surveys our knowledge about the molecular mechanism by which ERs regulate the expression of genes that do not contain EREs, and it gives examples of the ways in which the genomic and nongenomic actions of ERs on target genes converge. Genomic and nongenomic actions of ERs that do not depend on EREs influence the physiology of many target tissues, and thus, increasing our understanding of the molecular mechanisms behind these actions is highly relevant for the development of novel drugs that target specific receptor actions. (Molecular Endocrinology 19: 833–842, 2005)

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dimerize and bind to specific response elements known as estrogen response elements (EREs) located in the promoters of target genes (1). Hormone binding also induces a conformational change within the ligand binding domain of the receptors, and this conformational change allows coactivator proteins to be recruited (2). However, evidence for signaling pathways that deviate from this classical model has emerged, and it is now accepted that ERs can regulate gene expression by a number of distinct mechanisms. Around one third of the genes in humans that are regulated by ERs do not contain ERE-like sequences (3). The molecular mechanisms by which ERs regulate transcription at alternative response elements are not fully understood but are becoming increasingly clear. ERs can regulate gene expression without binding directly to DNA by modulating the function of other classes of transcription factors through protein-protein interactions in the nucleus (4). The interaction of ERs with the activator protein 1 (AP-1) transcription factor complex is a typical example of such ERE-independent genomic actions. In addition, a number of estrogen-responsive genes that lack EREs contain ERE half-sites, or binding sites for the orphan nuclear hormone receptor SF-1 [SF-1 response elements (SFREs)] that serve as direct ER binding sites (3). ER␣, but not ER␤, is able to bind to SFREs (5).

STROGENS ARE STEROID hormones that regulate growth, differentiation, and function in a broad range of target tissues in the human body. The most potent and dominant estrogen in humans is 17␤estradiol, but lower levels of the estrogens estrone and estriol are also present. The biological effects of estrogens are mediated through estrogen receptor (ER) ␣ and ␤, which are members of a large superfamily of nuclear receptors. These receptors act as ligandactivated transcription factors. The classical mechanism of ER action involves estrogen binding to receptors in the nucleus, after which the receptors First Published Online February 3, 2005 Abbreviations: AF, Activation function; AP-1, activator protein 1; C/EBP␤, CCAAT/enhancer binding protein ␤; CRE, cAMP response element; CREB, CRE binding protein; DBD, DNA binding domain; EGF, epidermal growth factor; eNOS, endothelial nitric oxide synthase; ER, estrogen receptor; ERE, estrogen response element; LDL, low-density lipoprotein; LDL-R, LDL receptor; NF-␬␤, nuclear factor ␬␤; PI, phosphoinositol; SERM, selective ER modulators; SRE, serum response element; STAT, signal transducer and activator of transcription. Molecular Endocrinology is published monthly by The Endocrine Society (http://www.endo-society.org), the foremost professional society serving the endocrine community. 833

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Estrogens exert some of their effects through the action of ERs on gene expression, but a number of other effects of estrogens are so rapid that they cannot depend on the activation of RNA and protein synthesis. These actions are known as nongenomic actions and are believed to be mediated through membraneassociated ERs. The actions are frequently associated with the activation of various protein-kinase cascades (6). However, nongenomic actions of estrogens may indirectly influence gene expression, through the activation of signal transduction pathways that eventually act on target transcription factors. The functions of many transcription factors, including AP-1, are regulated through protein kinase-mediated phosphorylation, and these transcription factors may thus be targets of nongenomic actions of estrogens. This signaling pathway can be referred to as nongenomic-togenomic signaling, and it provides for a mechanism, distinct from protein-protein interactions in the nucleus, by which ERs can modulate the functions of transcription factors, and thus regulate the expression of genes that do not contain EREs. This review surveys our knowledge about the molecular mechanisms by which ERs regulate transcription at alternative response elements. We give examples that show that genomic and nongenomic actions of ERs on target genes converge, and we briefly dis-

Bjo¨rnstro¨m and Sjo¨berg • Minireview

cuss the responses of ERs to synthetic ligands as a possibility of designing drugs that target specific receptor actions.

ERE-INDEPENDENT GENOMIC ACTIONS A number of studies have shown that ERs can regulate transcription without binding directly to DNA. The receptors in such cases are tethered through proteinprotein interactions to a transcription factor complex that contacts the DNA, as illustrated in Fig. 1. ERs regulate by this mechanism the expression of a large number of estrogen-responsive genes that do not contain EREs. The mechanism is commonly used by members of the nuclear receptor superfamily and is often referred to as transcriptional cross talk (4). Several genes are activated by 17␤-estradiol through the interaction of ERs with Fos and Jun proteins at AP-1 binding sites. Such genes include those for ovalbumin (7), IGF-I (8), collagenase (9), and cyclin D1 (10, 11). Transcription that depends on AP-1 is also repressed by ERs activated by 17␤-estradiol (12–14). One gene that is repressed by 17␤-estradiol through ER-AP-1 complexes is the choline acetyltransferase gene (15). ERs interact with AP-1 proteins in a com-

Fig. 1. Schematic Illustration of ER Signaling Mechanisms 1. Classical mechanism of ER action. Nuclear E2-ERs bind directly to EREs in target gene promoters. 2. ERE-independent genomic actions. Nuclear E2-ER complexes are tethered through protein-protein interactions to a transcription factor complex (TF) that contacts the target gene promoter. 3. Ligand-independent genomic actions. Growth factors (GF) activate protein-kinase cascades, leading to phosphorylation (P) and activation of nuclear ERs at EREs. 4. Nongenomic actions. Membrane E2-ER complexes activate protein-kinase cascades, leading to altered functions of proteins in the cytoplasm, e.g. activation of eNOS, or to regulation of gene expression through phosphorylation (P) and activation of a TF.


plex manner, and the involvement of ER activation function (AF) domains depends both on the receptor subtype and on the type of ligand (16). The activation of ER␣-AP-1 complexes that is induced by 17␤-estradiol requires the AF-2 domain of the receptor, which binds p160 coactivators and stabilizes in this way the formation of a multiprotein complex containing c-Jun, ER␣, and transcriptional coactivators at the promoter (17, 18). Genes that contain GC-rich promoter sequences are regulated in a similar manner through the interaction of ERs with the Sp1 transcription factor (19). Increasing numbers of genes are being found that are activated by 17␤-estradiol through ER-Sp1 complexes, including the LDL-R [low-density lipoprotein (LDL) receptor] (20), c-fos (21) and cyclin D1 (22) genes. The actions of ERs at Sp1 binding sites depend on the ligand, the cell type, and the receptor subtype (23), as is the case at for the actions of ERs at AP-1 binding sites (16). Repression of the IL-6 gene by 17␤-estradiol is mediated through the interaction of ERs with two transcription factors, nuclear factor ␬␤ (NF-␬B) and CCAAT/enhancer binding protein ␤ (C/EBP␤) (24, 25). In addition, it has been suggested that the repression of erythropoiesis by 17␤-estradiol involves the interaction of ERs with the GATA-1 transcription factor (26). ERs also regulate genes, such as the ␤-casein gene, that contain signal transducer and activator of transcription (STAT) 5 binding sites, but the outcome of the interaction of ERs with the STAT5 transcription factor is controversial. Some authors have suggested that STAT5-dependent transcription is repressed by ERs activated by 17␤-estradiol, and that this repression requires an intact ER AF-2 domain (27, 28). Other authors have suggested that the transcriptional activity of STAT5 is increased by ERs in a way that does not depend on the ER AF-2 domain (29). ERE-independent genomic actions that involve tethering of ERs to other DNA binding transcription factors do not require an interaction between the receptor and DNA, as is the case in the classical mechanism of ER action. However, the DNA binding domain (DBD) of the receptors is frequently involved, although intact DNA binding activity per se is not required (7–10, 13, 15, 25, 28, 29). Mutational analysis has revealed specific residues within the second zinc finger structure of the ER␤ DBD that discriminate between the classical mechanism of ER action and the modulation of AP-1 and STAT5 activities through tethering (14). This region is well conserved among nuclear receptors, and GR-mediated repression of AP-1 activity depends on residues within the second zinc finger structure of the GR DBD (30). The DBD may be required for proper protein-protein interactions or it may be involved in recruiting additional coregulator proteins to the promoter region.

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NONGENOMIC ACTIONS OF ESTROGENS Evidence is accumulating that estrogens exert nongenomic actions—actions that are too rapid to be accounted for by the activation of RNA and protein synthesis. Nongenomic actions are a common property of steroid hormones and are frequently associated with the activation of various protein-kinase cascades (6). The nongenomic actions of 17␤-estradiol that have been reported include the mobilization of intracellular calcium (31), and the stimulation of adenylate cyclase activity and cAMP production (32, 33). Activation of the MAPK signaling pathway by 17␤-estradiol has been extensively studied in several cell types, including breast cancer (34), endothelial (35), bone (36, 37), and neuroblastoma (38) cells. 17␤-Estradiol also activates the phosphoinositol (PI) 3-kinase signaling pathway in endothelial (35), breast cancer (39), and liver (40) cells. Some authors have suggested that the nongenomic actions of estrogens are mediated through a subpopulation of the classical ERs, ER␣ and ER␤, that is located at the plasma membrane (33, 41). Other authors disagree (42). In endothelial cells, as in other cell types, ERs have been found in caveolae where they activate endothelial nitric oxide synthase (eNOS) through protein kinase-mediated phosphorylation (43–45). Caveolae are specialized membrane invaginations enriched in the scaffold protein caveolin-1. Caveolae facilitate signal transduction by providing a location for various signaling molecules (46). The only membrane receptors for estrogens that have been found are the classical ERs, ER␣ and ER␤. However, an isoform of ER␣ that is alternatively spliced (of molecular mass 46 kDa) and that has a truncated N-terminal domain has been identified in endothelial cells (47). ERs do not contain a trans-membrane domain, and the ability of ER␣ to associate with the plasma membrane may be due to palmitoylation of the receptor (47, 48). The plasma membrane ERs exist as functional dimers when activated by estrogens (49). It appears that the ligand binding domain of the classical ER␣, when targeted to the plasma membrane, is sufficient for mediating nongenomic actions of estrogens (50). However, other functional domains of the receptors might contribute to the magnitude of these actions by participating in various protein-protein interactions. It is likely that the interaction of ERs with various scaffold or signaling molecules facilitates the activity. ERs at the plasma membrane associate with the scaffold protein caveolin-1 (44, 51), and with a variety of proximal signaling molecules such as G proteins (33, 52), Src kinase, and ras (34, 53), the p85␣ regulatory subunit of PI3-kinase (54) and Shc (55). Furthermore, the scaffold protein MNAR promotes the interaction of ER␣ activated by 17␤-estradiol with Src kinase, leading to an increase in Src kinase activity,

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and hence activation of the MAPK signaling pathway (56). Membrane ERs also activate membrane tyrosine kinase receptors in various cell types, including breast cancer cells. ER␣ activated by 17␤-estradiol interacts directly with the IGF-I receptor, leading to activation of the IGF-I receptor, and hence activation of the MAPK signaling pathway (57). ER␣ also interacts directly with ErbB2 (HER-2/neu) (58). In addition, ER␣ activated by 17␤-estradiol activates the epidermal growth factor (EGF) receptor by a mechanism that involves activation of G proteins, Src kinase, and matrix metalloproteinases, leading to an increase in MAPK and Akt (protein kinase B) activities (59). The studies described above suggest that the molecular mechanisms underlying the nongenomic actions of estrogens are specific for the cell type. The responses to estrogens may depend on a number of conditions, such as the set of signal transduction molecules and downstream targets present in the target cell, and thus, the responses are likely to be diverse.

NONGENOMIC-TO-GENOMIC SIGNALING: A NOVEL MODE OF REGULATING TRANSCRIPTION Signal transduction pathways may connect the nongenomic actions of estrogens to genomic responses. The functions of many transcription factors are regulated through protein kinase-mediated phosphorylation, and these transcription factors may thus be targets for nongenomic actions of estrogens, as illustrated in Fig. 1. This type of nongenomic-to-genomic signaling is a distinct mechanism by which ERs can regulate transcription at alternative response elements. This mechanism functions in addition to the ERE-independent genomic actions described above that involve protein-protein interactions in the nucleus. The transcription factors Elk-1 (60), C/EBP␤ (61), and CREB [cAMP response element (CRE) binding protein] (62) are all targets for phosphorylation by the MAPK signaling pathway. Two processes have been demonstrated in several cell types. These processes are the phosphorylation of Elk-1 that is induced by 17␤-estradiol, and the activation of serum response elements (SREs) by a mechanism that is dependent on ERs and that requires intact MAPK activity (55, 63–65). C/EBP␤ and CREB are phosphorylated after the MAPK signaling pathway has been activated by 17␤estradiol (65), and the phosphorylation of CREB leads to the expression of genes that contain CREs (65–67). CREB is a target for phosphorylation by the cAMP/ protein kinase A signaling pathway (68), in addition to its MAPK-mediated activation. Thus, CREB may also be activated by 17␤-estradiol through the stimulation of cAMP production. The transcriptional activity of the AP-1 transcription factor complex is regulated through MAPK-mediated phosphorylation (69). Activation of the MAPK signaling


pathway by 17␤-estradiol results in enhanced AP-1 DNA binding activity and transcriptional activation (40, 67, 70). In contrast, inhibition of the c-jun aminoterminal kinase signaling pathway by 17␤-estradiol results in repression of AP-1 activity (65). The latter observation agrees with previous results showing that the c-jun amino-terminal kinase signaling pathway, and hence AP-1 phosphorylation and transcriptional activation, are inhibited by other ligand-activated nuclear receptors (71, 72). The NF-␬B transcription factor complex is a target for phosphorylation by the Akt kinase (protein kinase B) (73, 74), and activation of the PI3-kinase/Akt signaling pathway by 17␤-estradiol leads to the expression of genes that contain NF-␬B binding sites (75). Tyrosine phosphorylation of the STAT family of transcription factors is required for their nuclear translocation and DNA binding activity (76). It has been suggested that additional serine phosphorylation modulates the maximal transcriptional activity of STATs (77). It has been reported that the phosphorylation of endogenous STAT3 and STAT5 on tyrosine and serine residues is induced by 17␤-estradiol in endothelial cells, and that STAT-dependent transcription, by a mechanism that is dependent on ERs and that requires intact MAPK, PI3-kinase, and Src kinase activities, is activated by 17␤-estradiol (78). These observations disagree with a previous study showing that the tyrosine phosphorylation and DNA binding activity of overexpressed STAT5 is decreased by ERs in COS-1 cells, both in the presence and in the absence of 17␤-estradiol (79). The discrepancies between these studies may be explained by differences in experimental conditions, such as the use of different types of cells and the analysis of endogenous or overexpressed proteins. It is important to note that ER␣ and ER␤ are also targets of phosphorylation by the MAPK signaling pathway (80, 81). Thus, nongenomic actions of estrogens may modulate the functions of ERs themselves, and in this way augment the classical mechanism of ER action. In addition, phosphorylation of the steroid receptor coactivator-3 is induced by 17␤-estradiol, leading to increased transcriptional potency of ERs (82). In the absence of estrogens, other signaling pathways, such as the IGF-I and EGF signaling pathways, can modulate the functions of ERs through phosphorylation of the receptors on certain residues (83), as illustrated in Fig. 1.

CONVERGENCE OF GENOMIC AND NONGENOMIC ACTIONS ON TARGET GENES The studies described above suggest that ERs can modulate the functions of transcription factors, and in this way regulate gene expression, by at least two distinct mechanisms. These mechanisms are proteinprotein interactions in the nucleus and activation of


signal transduction pathways at the plasma membrane. These mechanisms function in addition to the classical mechanism of ER action that is extensively reviewed elsewhere (1). Natural gene promoters are often regulated by a large number of transcription factors that bind to distinct regulatory sites. Thus, the possible convergence of genomic and nongenomic actions at multiple response elements provides an extremely fine degree of control for the regulation of transcription by ERs, as illustrated in Fig. 2. Cyclin D1 is a well-defined target for estrogens in breast cancer cells, and it is important for the progression of cells through the G1 phase of the cell cycle (84, 85). The cyclin D1 promoter is complex and contains binding sites for several transcription factors, but no ERE-like sequences have been identified (86). It has been suggested that activation of the cyclin D1 gene by 17␤-estradiol results mainly from the interaction of ERs with the Sp1 transcription factor at GC-rich promoter sequences (22), and with c-Jun/ATF-2 heterodimers at a variant CRE (10). However, it has also been reported that activation of the CRE by 17␤estradiol results from activation of the cAMP/protein kinase A signaling pathway (22), suggesting that nongenomic actions are involved. In addition, the cyclin D1 gene is activated by the association of ER␣ that


has been activated by 17␤-estradiol with Src kinase and the p85␣ regulatory subunit of PI3-kinase in breast cancer cells (39). In liver (40) and endometrial (87) cells, activation of the cyclin D1 gene by 17␤estradiol is mediated through the AP-1 binding site, and this activation requires intact MAPK activity (40). Furthermore, opposing actions of ER␣ and ER␤ at the cyclin D1 promoter have been observed (11), adding to the complexity of cyclin D1 gene regulation by ERs. The cyclin D1 promoter also contains binding sites for STAT5 and NF-␬B, and these may be targets for ERs through both genomic and nongenomic actions. Regulation of the immediate early c-fos gene by ERs is also mediated through both genomic and nongenomic actions. The interaction of ERs with the Sp1 transcription factor at GC-rich promoter sequences results in activation of the c-fos gene by 17␤-estradiol (21), and this activation also results from activation of the MAPK and PI3-kinase signaling pathways (64, 88). Thus, the nongenomic actions involve activation of both MAPK and PI3-kinase signaling pathways, which target the Elk-1 and the SRF transcription factors, respectively. These transcription factors bind to the SRE located in the c-fos promoter. The c-fos promoter also contains a sis-inducible element recognized by STATs and a cAMP response element, and these may

Fig. 2. Schematic Illustration of How Genomic and Nongenomic Actions of ERs on a Target Gene Promoter May Converge Nuclear E2-ER complexes bind to EREs, and to transcription factor complexes, e.g. AP-1, STATs, ATF-2 (activation transcription factor 2)/c-Jun, Sp1, and NF-␬B, that are bound to their cognate DNA binding sites. Membrane E2-ER complexes activate protein-kinase cascades, leading to phosphorylation (P) of target transcription factors, e.g. AP-1, STATs, Elk-1, SRF (serum response factor), CREB, and NF-␬B. The phosphorylation results in their transcriptional activation and/or in modulation of the transcriptional activities of ER-AP-1, ER-STAT, ER-Sp1, and ER-NF-␬B complexes at the promoter. Protein-kinase cascades also target ERs themselves and steroid receptor coactivator coactivators, resulting in enhanced transcriptional activity of ERs at EREs. The distinct actions of ERs at multiple response elements provide an extremely fine degree of control for the regulation of target gene transcription.

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be targets for ERs through both genomic and nongenomic actions. Estrogens increase the clearance of LDL and they lower plasma LDL cholesterol levels in postmenopausal women, and these may be the mechanisms by which estrogens reduce the risk of cardiovascular disease (89). The LDL-R gene is activated by 17␤-estradiol in the liver, although the LDL-R promoter does not contain any ERE-like sequences (90). It has been reported that the interaction of ERs with the Sp1 transcription factor at GC-rich promoter sequences gives rise to activation of the LDL-R gene by 17␤-estradiol (20). In addition, tyrosine-kinase activity is required for the 17␤-estradiol-induced activation of the LDL-R gene (91), suggesting that nongenomic actions are involved. The tyrosine-kinase pathway may be connected with Sp1-dependent transcription. The LDL-R promoter also contains an SRE and this may be the target for ERs through nongenomic actions that involve activation of the MAPK and PI3-kinase signaling pathways. Some estrogen-responsive genes contain a combination of direct ER binding sites and binding sites to which ERs indirectly associate through tethering. One such gene is the vascular endothelial growth factor gene that contains a variant ERE that specifically binds ERs activated by 17␤-estradiol, and G/GC-rich sequences that bind ER-Sp1 and ER-Sp3 complexes


(92, 93). Furthermore, a number of genes that contain ERE half-sites in proximity to Sp1 binding sites, both of which must be occupied for maximal activation, are activated by 17␤-estradiol. Such genes include those for retinoic acid receptor ␣ (94), TGF␣ (95), and progesterone receptor (96). It is evident that the regulation of gene expression by ERs is a multifactorial process, involving both genomic and nongenomic actions that often converge at certain response elements located in the promoters of target genes. The final gene responses, however, may depend on a number of conditions such as the combination of transcription factors bound to a specific gene promoter, the cellular localization of ERs, the expression levels of various coregulator proteins and signal transduction components, and the nature of extracellular stimuli. These variables are highly specific for the cell type. Thus, estrogens may use different signaling pathways depending on the cellular context, and in this way evoke distinct gene responses in different types of target cells, as illustrated in Fig. 3.

RESPONSES TO SYNTHETIC LIGANDS ER actions at alternative response elements have agonist and antagonist profiles that differ from the pro-

Fig. 3. Schematic Illustration of How ERs May Regulate Transcription at a Putative Target Gene Promoter in Different Types of Target Cells The signaling pathways used, i.e. ER actions at EREs or ERE-independent genomic and nongenomic actions of ERs, depend on the cellular localization of ERs, and in this way, the magnitude of the gene response evoked by E2 may differ between target cells.


files of the classical mechanism of ER action. The different responses to synthetic ligands may enable the use of such compounds to dissociate between distinct signaling pathways, and hence between gene responses. It has been suggested that estrogen-responsive genes containing AP-1 binding sites contribute to the difference in response that particular tissues make to selective ER modulators (SERMs). These differences, it is suggested, arise through differences in the activation of ER␣ and ER␤ in the nucleus (97). Differential activation of ER␣-Sp1 and of ER␤-Sp1 complexes at GC-rich promoters takes place also in the presence of SERMs (98). More recent studies, however, suggest that some of the responses to SERMs are mediated through nongenomic actions, which subsequently lead to genomic responses (70, 99). It may be possible, therefore, to develop of novel tissue-specific drugs that modulate the function of ERs at specific cellular locations. It has been claimed that estren, a synthetic estrogen, exerts only nongenomic actions (50). However, estren also affects ER actions in the nucleus (70), and it activates transcription at EREs, although the transcriptional potency of estren is lower that of 17␤estradiol (100). The full estrogen antagonist ICI 182,780, on the other hand, is unable to exert nongenomic actions mediated through classical receptors, and this antagonist blocks completely the nongenomic actions of estrogens (40, 50, 55, 63–65, 67, 78). However, ICI 182,780 is a potent agonist to both ER␣ and ER␤ when the receptors are tethered to the AP-1 (13, 14, 97), Sp1 (98, 101), and STAT5 (29) transcription factors in the nucleus. Thus, not all signaling pathways are inhibited by this antagonist. CONCLUDING REMARKS It is evident that many genes are regulated by ERs and that these genes are of two types: those that contain EREs and those that do not. The latter genes contain binding sites for a variety of heterogeneous transcription factors. ER actions at alternative response elements enable a broader range of genes to be regulated than the range that can be regulated by the classical mechanism of ER action alone. It is important to note that the nongenomic actions of estrogens target many of the transcription factors, such as AP-1, that participate in protein-protein interactions with ERs in the nucleus. The possible convergence of genomic and nongenomic actions on target genes is an attractive mechanism by which ERs can finely regulate gene expression. The relative contributions of genomic and nongenomic actions to certain gene responses in vivo, however, remain to be elucidated. The molecular mechanisms underlying the effects of estrogens are likely to be specific for the cell type, and thus, the gene responses are likely to be diverse.


It appears that different functional domains of ERs are involved in different receptor actions. Thus, naturally occurring splice variants such as ER␤cx (102), ER␤␦3 (103), and the 46-kDa ER␣ (47), although unable to regulate transcription at EREs, may act through alternative signaling pathways in their target tissues. If this is the case, it will have important biological significance. Further knowledge about the molecular mechanisms by which ERs regulate transcription at alternative response elements may enable drug design to target specific receptor actions. Drugs that have been designed to differently affect ER actions at specific cellular locations in a manner that depends on the cell type would widely expand the pharmacological possibilities open to physicians. Acknowledgments Received December 7, 2004. Accepted January 25, 2005. Address all correspondence and requests for reprints to: Maria Sjo¨berg, Department of Cell and Molecular Biology, Karolinska Institutet, SE-171 77 Stockholm, Sweden. E-mail: [email protected].

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