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, membrane-associated 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.

The retinoid X receptor (RXR) is a nuclear receptor that functions as a ligand-activated transcription factor. Little is known about the ligands that activate RXR in vivo. Here, we identified a factor in brain tissue from adult mice that activates RXR in cell-based assays. Purification and analysis of the factor by mass spectrometry revealed that it is docosahexaenoic acid (DHA), a long-chain polyunsaturated fatty acid that is highly enriched in the adult mammalian brain. Previous work has shown that DHA is essential for brain maturation, and deficiency of DHA in both rodents and humans leads to impaired spatial learning and other abnormalities. These data suggest that DHA may influence neural function through activation of an RXR signaling pathway.

Thyroid hormones and their receptors (TRs) have critical functions in development. Here we show that a chicken TR beta cDNA clone encodes a receptor with a novel, short N‐terminal domain. In vitro‐expressed TR beta protein bound thyroid hormone with similar affinity as the chicken TR alpha. Comparison of expression of TR alpha and TR beta mRNAs throughout chicken development until 3 weeks post‐hatching revealed ubiquitous expression of TR alpha mRNAs (in 14 different tissues) with some variations in levels, from early embryonic stages. In contast, expression of TR beta mRNA was restricted, occurring notably in brain, eye, lung, yolk sac and kidney, and was subject to striking developmental control, especially in brain where levels increased 30‐fold upon hatching. Levels also sharply increased in late embryonic lung, but were relatively high earlier in embryonic eye and yolk sac. RNase protection analyses detected no obvious mRNAs for alpha and beta TRs with variant C‐termini as demonstrated previously for the rat TR alpha gene. The data suggest a general role for TR alpha and specific developmental functions for TR beta, and that thyroid‐dependent development involves temporal and tissue‐specific expression of the TR beta gene.

17Beta-estradiol-activated estrogen receptor alpha (ERalpha) and beta (ERbeta) are able to induce transcriptional activation of signal transducer and activator of transcription (Stat)-regulated promoters via cytoplasmic signal transduction pathways. Stat5 and Stat3 are required for promoter induction, which correlates with cytoplasmic sublocalization of ERs and is independent of intact coactivator binding sites and DNA-binding domains. In endothelial cells, Stat5 and Stat3 are rapidly phosphorylated on both tyrosine and serine residues in response to 17beta-estradiol, and nuclear translocation is subsequently induced. 17Beta-estradiol-induced transactivation of a Stat-regulated promoter requires at least three different signal transduction pathways, including MAPK, Src-kinase, and phosphatidylinositol-3-kinase activities. In conclusion, this work identifies a novel pathway involving an agonist-bound ER-activated phosphorylation cascade, resulting in nuclear transcriptional activation of target transcription factors. These findings reveal novel targets for the development of drugs that modulate a nongenomic-to-genomic ER-dependent mechanism.

Bone cells' early responses to estrogen and mechanical strain were investigated in the ROS 17/2.8 cell line. Immunoblotting with antiphosphorylated estrogen receptor ␣ (ER-␣) antibody showed that when these cells were exposed for 10 minutes to estrogen (10 ؊8 M) or a single period of cyclic dynamic strain (peak 3400 ⑀, 1Hz, 600 cycles), there was an increase in the intensity of a 66-kDa band, indicating phosphorylation of ser

The oestrogen receptor is a member of the nuclear receptor family of transcription factors which, on binding the steroid hormone 17β‐oestradiol, interacts with co‐activator proteins and stimulates gene expression. Replacement of a single tyrosine in the hormone‐binding domain generated activated forms of the receptor which stimulated transcription in the absence of hormone. This increased activation is related to a decrease in hydrophobicity and a reduction in size of the side chain of the amino acid with which the tyrosine is replaced. Ligand‐independent, in common with ligand‐dependent transcriptional activation, requires an amphipathic α‐helix at the C‐terminus of the ligand‐binding domain which is essential for the interaction of the receptor with a number of potential co‐activator proteins. In contrast to the wild‐type protein, constitutively active receptors were able to bind both the receptor‐interacting protein RIP‐140 and the steroid receptor co‐activator SRC‐1 in a ligandindependent manner, although in the case of SRC‐1 this was only evident when the receptors were pre‐bound to DNA. We propose, therefore, that this tyrosine is required to maintain the receptor in a transcriptionally inactive state in the absence of hormone. Modification of this residue may generate a conformational change in the ligand‐binding domain of the receptor to form an interacting surface which allows the recruitment of co‐activators independent of hormone binding. This suggests that this tyrosine may be a target for a different signalling pathway which forms an alternative mechanism of activating oestrogen receptor‐mediated transcription.

Transcriptional activation by nuclear receptors (NRs) involves the concerted action of coactivators, chromatin components, and the basal transcription machinery. Crucial NR coactivators, which target primarily the conserved ligand-regulated activation (AF-2) domain, include p160 family members, such as TIF2, as well as p160-associated coactivators, such as CBP/p300. Because these coactivators possess intrinsic histone acetyltransferase activity, they are believed to function mainly by regulating chromatin-dependent transcriptional activation. Recent evidence suggests the existence of an additional NR coactivator complex, referred to as the thyroid hormone receptor-associated protein (TRAP) complex, which may function more directly as a bridging complex to the basal transcription machinery. TRAP220, the 220-kDa NR-binding subunit of the complex, has been identified in independent studies using both biochemical and genetic approaches. In light of the functional differences identified between p160 and TRAP coactivator complexes in NR activation, we have attempted to compare interaction and functional characteristics of TIF 2 and TRAP220. Our findings imply that competition between the NR-binding subunits of distinct coactivator complexes may act as a putative regulatory step in establishing either a sequential activation cascade or the formation of independent coactivator complexes. Nuclear hormone and orphan receptors (NRs)1 comprise a large family of transcription factors and participate in multiple aspects of development and homeostasis of higher eucaryotic organisms but also in deregulation of normal cellular functions (for review, see Refs. 1 and 2). They can be categorized into different subfamilies according to characteristics such as nature of ligand, DNA response element, or oligomerization status. Usually, steroid hormone receptors, which mainly form homodimers, are distinguished from a large diverse subfamily of receptors for nonsteroid ligands, such as thyroid hormone (TR), retinoids (retinoic acid receptor and retinoid X receptor (RXR)), and eicosanoids (peroxisome proliferator-activated receptor (PPAR)), as well as many orphan receptors, for which ligands have not been identified yet or do not exist. Unlike steroid receptors, most of these NRs function as heterodimers with RXR and thus represent highly dynamic transcription factor complexes due to the association of two receptor subunits with distinct structural and functional features (3-6). The majority of NRs utilizes two distinct domains for transcription activation, located in the N and C termini, respectively: a constitutive AF-1 and a ligand-regulated AF-2 as part of the multifunctional ligand-binding domain (LBD). NRs function in concert with multiple transcriptional cofactors, including basal transcription factors, corepressors, and coactivators (for review, see Refs. 7 and 8). Substantial progress in structural and functional analysis has allowed a more detailed understanding of interactions between the AF-2 domain and associated cofactors a...

Having used the cingulate cortex to demonstrate the validity of our methods for detecting hitherto unrecognized oestrogen receptor alpha (ERalpha)-immunoreactive neurones, we have now employed immunoprecipitation and double-label immunohistochemistry to investigate whether the ERalpha protein is present in gonadotrophin-releasing hormone (GnRH)-containing cells. The immortalized GnRH cell line GT1-7 and GnRH neurones within the rat preoptic area were found to possess ERalpha-immunoreactivity (ERalpha-IR). These observations indicate that oestrogen may regulate the synthesis and release of GnRH by direct actions on GnRH neurones.