Glucocorticoid receptors bound to the antagonist RU486 are not downregulated despite their capacity to interact in vitro with defined gene regions

Rajpert, Ewa J.; Lemaigre, Frédéric P.; Eliard, P H; Place, Martine; Lafontaine, D A; Economidis, Ioannis V.; Belayew, Alexandra; Martial, Joseph; Rousseau, Guy

doi:10.1016/0022-4731(87)90001-x

Cited by 48 publications

(14 citation statements)

References 27 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…One piece of this puzzle is the question of whether or not ligands modulate the interactions between GR and specific DNA recognition sites. Methods including gel-shifts, gradient sedimentation, and footprinting have been used to measure the allosteric effects of ligands on the interactions between GR and DNA in vitro; however, these studies have led to contradictory results (12)(13)(14)(15)(16)(17). Importantly, each observation is distinct with respect to the choice of ligand, DNA promoter site, and method of detection.…”

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

Allosteric Effects of Dexamethasone and RU486 on Glucocorticoid Receptor-DNA Interactions

Pandit

Geissler

Harris

et al. 2002

Journal of Biological Chemistry

View full text Add to dashboard Cite

The glucocorticoid receptor (GR) is a DNA-binding protein that can regulate the transcription of a large number of genes in a ligand-dependent fashion. Although much progress has been made on the mechanism of transcriptional regulation by GR, a potential allosteric effect of GR-binding ligands on specific GR-DNA interactions is controversial. In this study, gel-shift methods are used to measure the effects of a classical agonist dexamethasone and a prototypical antagonist RU486 on the in vitro interactions of GR with DNA substrates, which contain glucocorticoid response elements (GREs) from promoters of GR-regulated genes. These studies show that cell extracts containing human GR bind specifically and with high affinity to GREs in the absence of ligand. An agonist dexamethasone and antagonist RU486 do not affect the affinity of GR for DNA but subtly alter the electrophoretic mobility of the GR-DNA complex. Importantly, the dissociation rate of GR from DNA increases as a function of the concentration of GRE-containing DNA. At a fixed DNA concentration, dexamethasone-bound GR dissociates from DNA significantly faster than does ligand-free GR or RU486-bound GR. These results are consistent with a model for transcriptional activation in which a dynamic complex is formed between agonist-bound GR and DNA. The glucocorticoid receptor (GR)1 is a member of the steroid hormone receptor family of transcription factors that regulates a large number of genes in a hormone-or ligand-dependent manner. GR is predominantly cytosolic and requires association with heat shock proteins for ligand binding, a feature that has complicated in vitro structural studies of GR (1-6). The molecular mechanism by which hormones and small molecule ligands regulate GR-mediated gene expression is not clearly understood. A favored model is that hormone binding induces dissociation of heat shock proteins from the ligand-bound GR complex, which then translocates to the nucleus for sequencespecific DNA binding to GREs (7-10). However, the underlying mechanism for the expression of agonist versus antagonist activity for the steroid-bound GR-DNA complex is not clear.Attempts to study the allosteric effects of hormones on GR-DNA interactions in vitro have led to contradictory observations. In general, studies indicate that hormones are not required for stable GR binding to DNA in vitro (11-16). Specifically, complexes of agonist triamcinolone acetonide and antagonist RU486 bound to GR have a comparable affinity for glucocorticoid response elements from different promoters (14 -16). In contrast to these studies done on asymmetric GR binding sites, gel-shift experiments reported with GR and an artificial palindromic GRE suggest that ligands directly affect GR-DNA binding (17). The agonist dexamethasone enhances GR-DNA binding severalfold, whereas antagonists including RU486 have no effect on GR-DNA binding. However, this observation is contested by a report that also implements gelshift methods with a palindromic GRE substrate but uses in vitro transcribed...

show abstract

Section: Discussionmentioning

confidence: 99%

“…In general, studies indicate that hormones are not required for stable GR binding to DNA in vitro (11)(12)(13)(14)(15)(16). Specifically, complexes of agonist triamcinolone acetonide and antagonist RU486 bound to GR have a comparable affinity for glucocorticoid response elements from different promoters (14 -16).…”

mentioning

confidence: 99%

Allosteric Effects of Dexamethasone and RU486 on Glucocorticoid Receptor-DNA Interactions

Pandit

Geissler

Harris

et al. 2002

Journal of Biological Chemistry

View full text Add to dashboard Cite

show abstract

“…We used a protocol developed in the laboratories of S. Akana and Dr. Mary Dallman at the University of California-San Francisco (2,29). After the binding of either Cort or Mif to the GR, the ligand-receptor complex translocates to the nucleus (14,38). With the use of a rabbit anti-GR antibody (PA1-511, Affinity Bioreagents) that has higher affinity for the occupied than for the unoccupied GR, dark punctate nuclear staining is observed when either Mif or Cort is bound to the receptor (i.e., in the presence of ligand), whereas in the absence of ligand the staining is weaker and more diffuse.…”

Section: Methodsmentioning

confidence: 99%

Glucocorticoids act in the dorsal hindbrain to increase arterial pressure

Scheuer¹,

Bechtold²,

Shank³

et al. 2004

American Journal of Physiology-Heart and Circulatory Physiology

View full text Add to dashboard Cite

are present at a high density in the nucleus of the solitary tract (NTS), an area of the dorsal hindbrain (DHB) that is critical for blood pressure regulation. However, whether these receptors play any role in the regulation of blood pressure is unknown. We tested the hypothesis that glucocorticoids act in the DHB to increase arterial pressure using two experimental strategies. In one approach, we implanted pellets of corticosterone (Cort) or sham pellets onto the DHB over the NTS. Compared with rats with sham pellets, rats with DHB Cort pellets had an increased (P Ͻ 0.05) mean arterial pressure (111 Ϯ 2 vs. 104 Ϯ 1 mmHg) and heart rate (355 Ϯ 9 vs. 326 Ϯ 5 beats/min) after 4 days. In the second approach, we implanted subcutaneous Cort pellets to increase the systemic Cort concentration and then subsequently implanted pellets of the GR antagonist mifepristone (Mif; previously RU-38486) or sham pellets onto the DHB. Two days of DHB Mif treatment reduced (P Ͻ 0.05) mean arterial pressure in those rats with elevated plasma Cort levels (118 Ϯ 2 vs. 108 Ϯ 1 mmHg for sham vs. Mif DHB pellets). Cort and Mif pellets placed on the dura had no effects on arterial pressure or heart rate, ruling out systemic cardiovascular effects of the steroids. DHB Cort treatment had no effects on plasma Cort concentration or adrenal weight, indicating that the contents of the DHB Cort pellet did not diffuse into the systemic circulation or into the forebrain areas that regulate plasma Cort concentration in concentrations sufficient to produce physiological effects. Immunohistochemistry for the occupied GRs demonstrated that the Cort and Mif from the DHB pellets were delivered to the DHB with minimal diffusion to the ventral hindbrain or forebrain. We conclude that glucocorticoids act in the DHB to increase arterial pressure.corticosterone; nucleus of the solitary tract; brain; hypertension GLUCOCORTICOIDS ARE IMPORTANT for the maintenance of normal baseline arterial pressure in both rats and humans. Adrenal insufficiency or blockade of type II glucocorticoid receptors (GRs) can lower blood pressure (18,22,61). The effects of adrenalectomy to lower blood pressure can be reversed by glucocorticoid, but not mineralocorticoid, replacement (61). It is also established that elevated glucocorticoids produce doserelated increases in arterial pressure, and sufficiently high levels of glucocorticoids, due to either exogenous administration or endogenous overproduction, cause hypertension in both experimental animals and humans (22,43,44,54). Long-term administration of glucocorticoids, commonly prescribed because of their anti-inflammatory and immunosupressive therapeutic effects, results in a 20% incidence of iatrogenic hypertension (23). There is also increasing evidence that prolonged mild elevations in glucocorticoids or increased sensitivity to the actions of glucocorticoids promote the development and maintenance of essential hypertension. Several studies have reported elevations in plasma and/or urinary glucocorticoids in essential hyperte...

show abstract

“…Mif and corticosterone both bind to the GR and cause the subsequent translocation of the ligand-bound receptor from the cytoplasm to the nucleus (22,45). We previously used immunohistochemical identification of GR to demonstrate that 4 days of treatment with DHB Mif or corticosterone caused nuclear translocation of the GR within the DHB, but not in the ventral hindbrain or the forebrain (55).…”

Section: Assessment Of Brain Mif Levelsmentioning

confidence: 99%

Chronic blockade of hindbrain glucocorticoid receptors reduces blood pressure responses to novel stress and attenuates adaptation to repeated stress

Bechtold

Patel²,

Hochhaus³

et al. 2009

American Journal of Physiology-Regulatory, Integrative and Comp

View full text Add to dashboard Cite

Exogenous glucocorticoids act within the hindbrain to enhance the arterial pressure response to acute novel stress. Here we tested the hypothesis that endogenous glucocorticoids act at hindbrain glucocorticoid receptors (GR) to augment cardiovascular responses to restraint stress in a model of stress hyperreactivity, the borderline hypertensive rat (BHR). A 3-to 4-mg pellet of the GR antagonist mifepristone (Mif) was implanted over the dorsal hindbrain (DHB) in WistarKyoto (WKY) and BHRs. Control pellets consisted of either sham DHB or subcutaneous Mif pellets. Rats were either subjected to repeated restraint stress (chronic stress) or only handled (acute stress) for 3-4 wk, then all rats were stressed on the final day of the experiment. BHR showed limited adaptation of the arterial pressure response to restraint, and DHB Mif significantly (P Յ 0.05) attenuated the arterial pressure response to restraint in both acutely and chronically stressed BHR. In contrast, WKY exhibited a substantial adaptation of the pressure response to repeated restraint that was significantly reversed by DHB Mif. DHB Mif and chronic stress each significantly increased baseline plasma corticosterone concentration and adrenal weight and reduced the corticosterone response to stress in all rats. We conclude that endogenous corticosterone acts via hindbrain GR to enhance the arterial pressure response to stress in BHR, but to promote the adaptation of the arterial pressure response to stress in normotensive rats. Endogenous corticosterone also acts in the hindbrain to restrain corticosterone at rest but to maintain the corticosterone response to stress in both BHR and WKY rats. corticosterone; brain; nucleus of the solitary tract; hypothalamicpituitary-adrenal axis; chronic stress EXAGGERATED CARDIOVASCULAR responses to acute stress and chronic stress increase the risk for hypertension and cardiovascular disease (1,3,13,15,31,38,42,58,68). Acute stress rapidly increases blood pressure, heart rate, plasma glucocorticoid concentration, and blood glucose levels (8), while chronic or repeated stress is associated with increases in baseline blood pressure and glucocorticoids (17). Chronically elevated glucocorticoids increase morbidity and mortality from cardiovascular disease and alterations in the glucocorticoid receptor (GR), and in glucocorticoid metabolizing enzymes are associated with hypertension and cardiovascular disease in humans (28,30,35,36,39,48,49,61, 65,67,70,71). Thus, chronic stress-induced elevations in glucocorticoids likely contribute to the adverse effects of stress on cardiovascular health.The mechanisms by which glucocorticoids modulate cardiovascular stress responses are not fully understood. A review by Sapolsky et al. (52) concluded that glucocorticoids act permissively to enhance the arterial pressure response to many physical stressors, in part by supporting the peripheral effects of catecholamines. Other studies indicate that chronic, systemic elevations in glucocorticoids enhance cardiovascular and catecholamine re...

show abstract

Glucocorticoid receptors bound to the antagonist RU486 are not downregulated despite their capacity to interact in vitro with defined gene regions

Cited by 48 publications

References 27 publications

Allosteric Effects of Dexamethasone and RU486 on Glucocorticoid Receptor-DNA Interactions

Allosteric Effects of Dexamethasone and RU486 on Glucocorticoid Receptor-DNA Interactions

Glucocorticoids act in the dorsal hindbrain to increase arterial pressure

Chronic blockade of hindbrain glucocorticoid receptors reduces blood pressure responses to novel stress and attenuates adaptation to repeated stress

Contact Info

Product

Resources

About