Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Combined quantitative polymerase chain reaction (PCR) and cytosolic binding assay techniques are used to measure mineralocorticoid receptor (MR) and glucocorticoid receptor (GR) mRNA, Kd, and Bmax in various rat central nervous system (CNS) regions, namely amygdala, hypothalamus, hippocampus, cortex, pituitary, and cervical, thoracic, and lumbar spinal cord. Two internal standards (i.s.) cDNA were cloned for quantitative PCR purposes. The i.s. templates differed from the respective wild-type (wt) templates for a single base-pair mutation introduced by PCR that generated a unique restriction site, thus allowing amplification products arising from coamplification of wt and i.s. to be distinguished. Results show that cerebellum, which displayed average Bmax values for both receptors, contained the highest level of MR and GR mRNA. Hippocampus also had a high level of MR mRNA. Low mRNA content was found in the hypothalamus for MR and GR as well as in the cortex for GR. High Bmax values for both MR and GR were found in the lumbar spinal cord, despite a modest mRNA content. The lowest Bmax values were found in the cortex for both receptors. It is, therefore, concluded that mRNA content and Bmax are not closely correlated in the rat CNS. These data suggest a differential regulation of various adrenocorticoid receptor isoforms. Moreover, this quantitative PCR method is very sensitive and can be used to assay small amounts of material in order to obtain absolute measurements of mRNA expression.
J Mol Neurosci 1997 Aug
PMID:Distribution of adrenocorticoid receptors in the rat CNS measured by competitive PCR and cytosolic binding. 935 22

Human embryonic kidney (K293) cells transfected with a mouse mammary tumor virus (MMTV) promoter-luciferase reporter construct (pHH-Luc) were utilized to investigate the potential effects of trans-retinoic acid (tRA), either by itself or in combination with glucocorticoid (GC) hormones, on a well-characterized, GC-sensitive transcriptional response. tRA or the synthetic GC hormone dexamethasone induced transcription from the MMTV promoter in a dose-dependent manner, with 1 micromol tRA and 1 micromol dexamethasone alone causing a four- to six-fold and a 40-fold induction of basal transcription, respectively. Simultaneous treatment with 1 micromol dexamethasone and 1 micromol tRA resulted in a synergistic transcriptional response that was 120-fold higher than basal level and 2.5 times the predicted response, based on a simple additive effect of both agonists. tRA does not appear to mediate this synergistic transcriptional response by enhancing GC receptor (GR) binding capacity, affinity, or nuclear translocation. tRA was unable to potentiate GC-induced transcriptional activity from a minimal GC response element (GRE), and GC were unable to potentiate tRA-induced transcriptional activity from a minimal retinoic acid response element (RARE). These data rule out direct protein-protein interactions between GC and retinoid receptors as a mechanism for the observed synergism. tRA also synergized with aldosterone-induced, mineralocorticoid receptor (MR)-mediated, transcriptional activation of the MMTV promoter, resulting in a response that was 1.7 times the predicted additive response. The MMTV GRE located between -187 and -165 was required for GC-induced and synergistic activation of the MMTV promoter, whereas sequences located within -151 to +5 were sufficient for tRA-induced transcription from the MMTV promoter. Mutation of a consensus RARE half-site (CCAAGT) identified at position -65 to -60 within the MMTV-LTR did not affect either tRA-induced transcriptional activation or synergism with GC. We propose that the tRA-induced transcriptional response from the MMTV promoter, as well as synergism with GC, may be mediated by the activation or induction of a factor(s) that either directly binds to the MMTV promoter or indirectly stabilizes binding of another transcription factor to these sequences.
J Steroid Biochem Mol Biol 1997 Jun
PMID:Trans-retinoic acid and glucocorticoids synergistically induce transcription from the mouse mammary tumor virus promoter in human embryonic kidney cells. 939 48

In addition to mineralocorticoid and glucocorticoid receptors, a third category of corticosteroid binding sites has been described in the kidney, the Type III binding protein. This intracellular binder has high affinity for corticosterone, but binds neither aldosterone nor synthetic glucocorticoids. Based on similarities in steroid specificity and kinetic parameters, we hypothesized that these corticosterone binding sites belong to the type 2 isoform of 11 beta-hydroxysteroid dehydrogenase (11 beta-HSD2). The goal of this study was to express the recombinant rabbit 11 beta-HSD2 in mammalian cells and test if such cells acquire both NAD-dependent 11 beta-HSD2 activity as well as high affinity corticosterone binding sites. Stably transfected CHO cell lines expressed high, NAD-dependent, unidirectional 11 beta-HSD2 activity. At the same time, the transfected cells also acquired a large number of corticosterone-specific binding sites (1.21 +/- 0.3 x 10[6]), whereas non-transfected cells had no corticosterone binding above background. The Kd for corticosterone was 25 +/- 8 nM. Neither the glucocorticoid receptor (GR) agonists dexamethasone and RU 28362 nor the mineralocorticoid receptor (MR) agonist aldosterone bound to these sites. The steroid specificity of the binding sites, as determined by competing [3H]corticosterone with unlabeled steroids, is identical to that of 11 beta-HSD2: corticosterone >> 11-hydroxyprogesterone > carbenoxolone > 11 dehydrocorticosterone > cortisol > progesterone approximately DOC >>> DEX > RU 28362 - aldosterone. These results strongly suggest that the previously described high affinity corticosterone binding sites are 11 beta-HSD2. Thus, though Type III binding sites are not corticosteroid receptors as originally thought, they play an important role in regulating the activity of both mineralocorticoid- and glucocorticoid receptors.
Mol Cell Endocrinol 1997 Nov 15
PMID:11 beta-hydroxysteroid dehydrogenase-2 is a high affinity corticosterone-binding protein. 942 59

The glucocorticoid (GR) and the mineralocorticoid (MR) receptor mediate corticosteroid actions in the mammalian brain. Here, we report the sequence and distribution of both receptor subtype mRNAs in the central nervous system of the tree shrew Tupaia belangeri, a non-rodent mammal, phylogenetically located between insectivores and primates. The specific glucocorticoid and mineralocorticoid receptor cDNAs were cloned, employing polymerase chain reaction (PCR) based methods. The GR cDNA and MR cDNA encode the 776-amino acid (aa) and 977-aa receptor, respectively. Comparisons of both GR and MR with corresponding cDNA-sequences of other species revealed the highest homology to the human equivalents (GR: 90%, MR: 89% nucleotide sequence identity of the coding regions). The localization of GR and MR mRNA in tree shrew brain was investigated by in situ hybridization using 35S-labeled riboprobes. The GR mRNA is widely distributed throughout all observed brain areas, with high signal intensities in the dentate gyrus, piriform cortex, cerebellum, anterior pituitary, subfornical organ and pineal gland. Whereas, moderate expression of GR mRNA was noted in region CA1 of the hippocampus, region CA3 displayed only low signal intensity. MR mRNA hybridization is mainly restricted to the strongly labeled hippocampal formation, but in contrast to the localization pattern found in rat, higher signal intensities are detected in field CA1 than in CA3. These data indicate that both GR and MR mRNAs are highly expressed in tree shrew brain with a species-specific expression pattern.
Brain Res Mol Brain Res 1998 Apr
PMID:Cloning of glucocorticoid receptor and mineralocorticoid receptor cDNA and gene expression in the central nervous system of the tree shrew (Tupaia belangeri). 958 28

We have examined the human mineralocorticoid receptor gene and the genes encoding the three subunits of the human amiloride-sensitive epithelial sodium channel. Eight new common polymorphisms were identified in these genes which may be useful in genotyping and linkage analysis.
J Steroid Biochem Mol Biol 1998 Mar
PMID:Common polymorphisms in genes encoding the human mineralocorticoid receptor and the human amiloride-sensitive sodium channel. 961 22

The effects of aldosterone are mediated by the mineralocorticoid receptor (MR), a ligand-dependent transcription factor. We investigated the structural determinants for ligand binding to the receptor using a series of human MR (hMR) deletion mutants. These proteins were produced in vitro in rabbit reticulocyte lysate and analyzed for their ability to bind agonists, antagonists, and the heat shock protein hsp90, which is a prerequisite for ligand binding to hMR. Studies on N terminus-truncated hMRs showed that the ligand-binding domain (LBD: amino acids 734-984) has a lower affinity for aldosterone than the entire receptor [dissociation constant (Kd) 2.9 vs. 0.47 nM] and does not interact with hsp90. Addition of the five-amino acid sequence (729-733) upstream from the LBD is necessary for interaction with hsp90, but a larger region is needed for high aldosterone affinity. Deletions at the C-terminal end of the hMR greatly reduced both agonist and antagonist binding: deletion of the last three amino acids reduced the affinity for aldosterone to 1/20 that of the entire protein, and deletion of the last four amino acids completely abolished binding, although the interaction with hsp90 was not affected. These effects can be explained by misfolding of the receptor, since limited proteolysis assays showed that deletions at the C-terminal end of hMR affect the accessibility of the cleavage sites within the DNA-binding domain and the N-terminal part of the hinge region to trypsin. Thus, our results support the idea that a short sequence upstream of the LBD is essential for the interaction of hMR with hsp90 and that the C terminus of hMR and hsp90 are both essential for folding of the receptor in a high-affinity hormone-binding state.
Mol Endocrinol 1998 Jun
PMID:Folding requirements of the ligand-binding domain of the human mineralocorticoid receptor. 962 61

11Beta-hydroxysteroid dehydrogenase (11beta-HSD) is thought to confer aldosterone specificity to mineralocorticoid target cells by protecting the mineralocorticoid receptor (MR) from occupancy by endogenous glucocorticoids. In aldosterone target cells the type 2 11beta-HSD is present, which, in contrast to the type 1 11beta-HSD, has very high affinity for its substrate, is unidirectional and prefers NAD as cofactor. cDNAs encoding 11beta-HSD2 have been recently cloned from different species, and the cell-specific expression of its mRNA and protein were determined. 11Beta-HSD2 is expressed in every aldosterone target tissue. Northern analysis revealed that the rabbit 11beta-HSD2 is expressed at high levels in the renal collecting duct and at much lower levels in the colon. RT-PCR experiments demonstrated that 11beta-HSD2 mRNA is present only in aldosterone target cells within the kidney. We determined the subcellular localization of the rabbit 11beta-HSD2 using a chimera encoding 11beta-HSD2 and the green fluorescent protein (GFP). This construct was stably transfected into CHO and MDCK cells. The expressed 11beta-HSD2/GFP protein retained high enzymatic activity, and its characteristics were undistinguishable from those of the native enzyme. The intracellular localization of this protein was determined by fluorescence microscopy. 11Beta-HSD2-associated fluorescence was observed as a reticular network over the cytoplasm whereas the plasma membrane and the nucleus were negative, suggesting endoplasmic reticulum (ER) localization. Co-staining with markers for ER proteins, the Golgi membrane, mitochondria and nucleus confirmed that 11beta-HSD2 is localized exclusively to the ER. To determine what structural motifs are responsible for the ER localization, we generated deletion mutants missing the C-terminal 42 and 118 amino acids, and fused them to GFP. Similarly as with the intact 11beta-HSD2, these mutants localized exclusively to the ER. Both C-terminal deletion mutants completely lost dehydrogenase activity, independently whether activity was determined in intact cells or homogenates. These results indicate that 11beta-HSD2 has a novel ER retrieval signal which is not localized to the C-terminal region. In addition, the C-terminal 118 amino acids are essential for NAD-dependent 11beta-HSD activity.
J Steroid Biochem Mol Biol 1998 Apr
PMID:The role of 11beta-hydroxysteroid dehydrogenase in steroid hormone specificity. 969 85

11 Beta-hydroxysteroid dehydrogenase (11 beta-HSD) catalyses the interconversion of active corticosterone and inert 11-dehydrocorticosterone. The recently discovered type 2 isozyme (11 beta-HSD-2) is a high affinity, NAD-dependent, exclusive 11 beta-dehydrogenase, which rapidly inactivates glucocorticoids. Thus the enzyme generates aldosterone-selectivity for intrinsically non-selective mineralocorticoid receptors in vivo as well as excluding glucocorticoids from glucocorticoid receptors, the latter being particularly important during development. Aldosterone exerts selective central effects upon salt appetite and blood pressure whilst glucocorticoids have potent effects upon postnatal neurogenesis and brain remodelling. We examined 11 beta-HSD-2 expression during postnatal ontogeny and in adult rat brain. High 11 beta-HSD-2 mRNA expression was found specifically in the postnatal thalamus and the external granule cell layer of the cerebellum. Expression peaked at the end of the first postnatal week and declined rapidly thereafter. Postnatal brain showed considerable activity of high affinity 11 beta-HSD-2 which paralleled expression of 11 beta-HSD-2 messenger ribonucleic acid (mRNA). Adult brain showed high 11 beta-HSD-2 mRNA expression limited to the subcommissural organ, with lower expression in the ventromedial nucleus of the hypothalamus, amygdala, locus coeruleus and nucleus tractus solitarius. These discrete areas are compatible with proposed selective central actions of aldosterone on blood pressure (subcommissural organ, nucleus tractus solitarius) and salt appetite (ventromedial nucleus, amygdala). In contrast, early postnatal 11 beta-HSD-2 coincides with glucocorticoid receptor rather than mineralocorticoid receptor expression, and areas of expression are among the regions where glucocorticoids have been demonstrated to have profound effects upon neuronal division, growth and maturation.
Brain Res Mol Brain Res 1998 Oct 30
PMID:11 Beta-hydroxysteroid dehydrogenase type 2 in the postnatal and adult rat brain. 979 98

We have studied the immediate and long-term effects of high doses of corticosterone (CORT) on mRNA expression and binding properties of mineralocorticoid receptor and glucocorticoid receptor in the hippocampus and spinal cord of rats. Animals were treated with corticosterone (10 mg/d subcutaneously) for 21 consecutive days, and mineralocorticoid and glucocorticoid receptors were studied either 24 h or 2 wk after the last injection. Major results show that corticosterone treatment reduces mineralocorticoid and glucocorticoid receptor maximum binding capacity (Bmax) in both the hippocampus and spinal cord and that this reduction is partially reversed after cessation of treatment. With respect to mRNA expression, in the hippocampus recovery after cessation of treatment is complete. By contrast, in the spinal cord, mineralocorticoid receptor mRNA expression is irreversibly increased after treatment, but the glucocorticoid receptor mRNA level remains unaffected during and after treatment. Thus, these data suggest the presence of distinct regulatory mechanisms for adrenocorticoid receptors in rat brain and spinal cord, in response to long-term exposure to high levels of circulating corticosterone and after recovery from treatment.
J Mol Neurosci 1998 Aug
PMID:Corticosterone treatment differentially affects adrenocorticoid receptors expression and binding in the hippocampus and spinal cord of the rat. 982 89

The binding of aldosterone (ALDO) to the mineralocorticoid receptor (MR) induces a conformational change of the protein referred to as 'transformation'. This feature can be evidenced in vivo by the capacity of the MR to interact with chromatin, and in vitro by the ability of the MR to bind to DNA strands or to shift the sedimentation coefficient (S) to lower values. The transformation process allows MR to work as a transcription factor after interacting with specific sequences of DNA. The signal transduction pathway for the MR transformation remains unknown. As a first step towards elucidating the mechanism of steroid-dependent MR transformation, we asked if the MR-signaling pathway is affected by the phosphorylation status of the MR-heterocomplex, and how that pathway may be regulated. Incubation of preformed [3H]ALDO-MR complex with bovine intestinal alkaline phosphatase led to an increase in the rate of MR-transformation (measured as 9.4-5.4S shift). This alkaline phosphatase-dependent MR transformation was inhibited by the specific alkaline phosphatase-type inhibitor levamisole, and was not evident in incubations performed with acid phosphatases. A direct correlation between the DNA-cellulose binding capacity of the [3H]ALDO-MR complex and the percentage of transformed 5.4S MR form was also observed. When rat kidney cytosol was incubated in the absence of both exogenous phosphatase and stabilizing agents (such as molybdate or vanadate), MR transformation also took place, in a time- and temperature-dependent process. In contrast with the inhibitory effect observed upon alkaline phosphatase-promoted transformation, levamisole was unable to inhibit the endogenous transforming activity of MR, suggesting that an endogenous phosphatase other than those which belong to the alkaline-type may be responsible for that transformation. Tautomycin, a polyketide produced by the soil bacteria Streptomyces which inhibits serine/threonine phosphatases of the PP1/PP2A subgroup, was able to inhibit the endogenous phosphatase activity in a concentration-dependent form (Ki(app)=7.35 nM). These results support the idea that the endogenous renal activity involved in the regulation of rat kidney MR transformation may be a protein phosphatase which belongs to the PP1/PP2A subgroup.
Mol Cell Endocrinol 1998 Sep 25
PMID:Tautomycin inhibits phosphatase-dependent transformation of the rat kidney mineralocorticoid receptor. 986 32


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