The glucocorticoid receptor (GR) activates transcription in certain glucocorticoid response element (GRE) contexts, and represses or displays no activity in others. We isolated point mutations in one GRE, plfG, at which GR activated transcription under conditions in which the wild-type element was inactive or conferred repression, implying that GREs may carry signals that are interpreted by bound receptors. Consistent with this notion, we identified a mutant rat GR, K461A, which activated transcription in all GRE contexts tested, implying that this residue is important in interpretation of GRE signals. In a yeast screen of 60,000 GR mutants for strong activation from plfG, all 13 mutants isolated contained substitutions at K461. This lysine residue is highly conserved in the zinc-binding region (ZBR) of the intracellular receptor (IR) superfamily; when it was mutated in MR and RARP, the resulting receptors similarly activated transcription at response elements that their wild-type counterparts repressed or were inactive. We suggest that IR response elements serve in part as signaling components, and that a critical lysine residue serves as an allosteric "lock" that restricts IRs to inactive or repressing configurations except in response element contexts that signal their conversion to transcriptional activators. Therefore, mutation of this residue produces altered receptors that activate in many or all response element contexts.[Key Words: Glucocorticoid receptor; transcriptional repression; DNA-binding domain; context-dependent activity; intracellular receptor] Received January 1, 1996; accepted in revised form March 27, 1996.Regulation of transcription typically commences with the specific binding of a regulatory protein to a response element close to the target promoter; the regulator also interacts with other regulatory factors, with the transcription machinery andlor chromatin components, therefore modulating the rate of transcription initiation (for review, see Sheldon and Reinberg 1995;Hori and Carey 1994; Tjian and Maniatis 1994). Numerous regulators have been shown to activate transcription in certain cell types and at certain promoters, and to repress in other settings (Meyer et al. 1980;Keleher et al. 1988;Pedersen et al. 1991). Therefore, it is apparent that the precise activity of a regulator can be determined largely by its context-cell type, promoter, response element, and extracellular signals. The mechanisms by which regulators sense and interpret their contexts and thereby display distinct activities in different settings remain to be determined.The glucocorticoid receptor (GR) is a member of the intracellular receptor (IR) superfamily, which encompasses a large group of transcriptional regulators (Tsai Present addresses: 2Department
The TATA box element is not only important for establishing basal levels of transcription, but it can also be used to modulate cell type or stage specific gene activity. In the case of the human osteocalcin gene, which is transcriptionally repressed by glucocorticoids, a specific binding element for the glucocorticoid receptor (GR) overlaps a noncanonical TATA box. In the present study, the relevance and function of the TATA element in glucocorticoid-mediated repression of the human osteocalcin gene was characterized. Mutating this noncanonical TATA box into a consensus TATA box within the context of the osteocalcin promoter greatly decreased hormone-dependent transcriptional repression by GR. TATA-binding protein (TBP) bound this mutated element much more strongly suggesting a physiologically relevant role for the weak osteocalcin TATA element in the regulation of this bone specific gene. The optimization of the putative transcription factor IIB recognition site did not affect the level of GR-mediated repression. Our results support a model wherein competitive DNA binding of GR and TBP for their overlapping sites explains conditional repression of the osteocalcin gene by glucocorticoids.Transcription of RNA polymerase II-dependent genes requires, in addition to the enzyme itself, a number of general factors that form a specific multiprotein complex near the transcription start site by interacting with basal promoter elements. The most well studied core promoter element is the TATA box, which is typically located 25-30 base pairs upstream of the transcription start site of many eukaryotic genes (1). The central step in formation of the preinitiation complex at TATA-containing promoters involves the binding of transcription factor IID (TFIID) 1 to the TATA box (2). TFIID consists of the TATA-binding protein (TBP) and a fairly large number of associated factors (3). The affinity of the TBP/TATA box interaction has been proposed to contribute to promoter strength in vivo and in vitro, and subsequent assembly of the other general transcription factors into a functional preinitiation complex is dependent upon this initial interaction at the TATA box (1, 4 -6).In addition to its role as a nucleation point for preinitiation complex assembly, the TATA box can also have an important function with respect to cell type or stage specific modulation of gene activity. In several reported cases, minor deviations from the consensus TATA box DNA sequence (TATA(A/T)A(A/T)A) are thought to be critical for proper regulation. For example, in the case of the rabbit uteroglobin gene, two factors, one cell type specific and the other ubiquitously expressed, have been proposed to facilitate the interaction of TBP with the weak TATA box, TACAAA, by binding the TACA site (7). Another example is the unusual inverted TTTATA sequence that is involved in the negative regulation of the bone sialoprotein gene (8).Several examples of repression of gene activity by competitive binding at the TATA box have also been postulated. In these cases, the...
The action of the glucocorticoid receptor (GR) on -casein gene transcription serves as a well-studied example of a case where the action of the GR is dependent on the activity of another transcription factor, STAT5. We have investigated the domain-requirement of the GR for this synergistic response in transfection experiments employing GR mutants and CV-1 or COS-7 cells. The results were influenced by the expression levels of the GR constructs. At low expression, STAT5-dependent transactivation by mutants of the GR DNA binding domain or N-terminal transactivation domain was impaired and the antiglucocorticoid RU486 exhibited a weak agonistic activity. When the N-terminal region of the GR was exchanged with the respective domain of the progesterone receptor, STAT5-dependent transactivation was reduced at low and high expression levels. Only at high expression levels did the GR exhibit the properties of a coactivator and enhanced STAT5 activity in the absence of a functional DNA binding domain and of GR binding sites in the proximal region of the -casein gene promoter. Furthermore, at high GR expression levels RU486 was nearly as efficient as dexamethasone in activating transcription via the STAT5 dependent -casein gene promoter. The results reconcile the controversial issue regarding the DNA binding-independent action of the GR together with STAT5 and provide evidence that the mode of action of the GR depends not only on the type of the particular promoter at which it acts but also on the concentration of the GR. GR DNA binding function appears to be mandatory for -casein gene expression in mammary epithelial cells, since the promoter function is completely dependent on the integrity of GR binding sites in the promoter.Modulation of gene expression by the glucocorticoid receptor (GR) involves a combination of several mechanisms such as modulation of chromatin structure (5, 27); binding to specific DNA response elements (24); interaction with sequence-specific transcription factors, coactivators, and corepressors; and ligand-dependent alterations in the balance of corepressors and coactivators bound to the receptor (20). The actual type of mechanism employed by the receptor strongly depends on the genes that are regulated and on the cellular context. There is a differential requirement for domains in the GR, depending on the prevalent mechanism utilized by the receptor. For instance, a specific subset of GR-regulated genes is affected in transgenic mice in which the wild-type GR is replaced by a mutant defective in dimerization (26). Since this mutant is strongly impaired in binding to palindromic canonical glucocorticoid response elements (GREs) (7), it can no longer regulate genes that contain functional GREs. Consistent with the observation that in most cases the binding of the GR to DNA is a prerequisite for transactivation but not for transrepression, transgenic mice expressing the dimerization mutant predominantly exhibit a defect in the expression of genes induced by glucocorticoids.One of the exception...
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.