Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.1.27.1 (RNase)
16,360 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The effect of 12-O-tetradecanoylphorbol-13-acetate (TPA) on the regulation of the estrogen receptor (ER) was investigated in this study. After treatment with 100 nM TPA the concentration of receptor protein was measured using an enzyme immunoassay. By 24 h the receptor protein declined by about 80% from a level of approximately 236 fmol of ER/mg of protein in control cells to 50 fmol of ER/mg of protein in cells treated with TPA. Similar results were obtained with an estrogen receptor ligand binding assay. After removal of TPA, the level of ER returned to control values. 4-alpha-Phorbol, a compound related to TPA, had no effect on ER. The effects of TPA on ER expression appear to be mediated by activation of protein kinase C as H-7, an inhibitor of protein kinase C, blocks these effects. In addition to the effect on ER protein, TPA treatment also resulted in a decrease in the steady-state level of ER mRNA as determined by a RNase protection assay. The metabolic inhibitor cycloheximide was unable to prevent the TPA-induced decrease in ER mRNA. Transcription run-off experiments demonstrated that TPA had no effect on ER gene transcription. A half-life study demonstrated that TPA decreased ER mRNA half-life by a factor of 6 from approximately 4 h in control cells to 40 min in TPA-treated cells. These data suggest that the decline in ER expression is mediated by post-transcriptional destabilization of ER mRNA.
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PMID:Post-transcriptional destabilization of estrogen receptor mRNA in MCF-7 cells by 12-O-tetradecanoylphorbol-13-acetate. 191 23

We have analyzed human benign prostatic hyperplastic (BPH) tissue derived from eight radical prostatectomy specimens from patients with prostate cancer for the expression of the estrogen receptor (ER) messenger RNA. Four of the eight patients received a long-acting gonadotropin-releasing hormone agonist (GnRHa) for 4 months prior to surgery. An RNase protection assay utilizing six riboprobes spanning most of the ER protein-coding sequences demonstrated expression of the ER mRNA in human BPH tissue. A comparison of ER mRNA expression in four patients who had received 4 months pretreatment with the GnRHa vs. the four untreated patients suggested that there is upregulation of ER mRNA expression with the GnRHa treatment. The combined techniques of in situ hybridization and immunocytochemistry localized the ER mRNA expression to the prostatic basal epithelial cells and stroma. We conclude that ER mRNA is expressed in human BPH tissue and that this expression is modulated by treatment with a long-acting GnRH agonist.
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PMID:Estrogen receptor messenger RNA expression in human benign prostatic hyperplasia: detection, localization, and modulation with a long-acting gonadotropin-releasing hormone agonist. 753 25

The effects of long term treatment with 12-O-tetradecanoylphorbol-13-acetate (TPA) on estrogen receptor (ER) expression in the human breast cancer cell line, MCF-7, were studied. This study demonstrates that treatment of cells with the phorbol ester blocked estrogen receptor activity. Treatment of cells with 100 nM TPA resulted in an 80% decrease in the level of ER protein and a parallel decrease in ER mRNA and binding capacity. Following removal of TPA from the medium, the level of ER protein and mRNA returned to control values; however, the receptor failed to bind estradiol. These cells also failed to induce progesterone receptor in response to estradiol. In addition, TPA treatment blocked transcription from an estrogen response element in transient transfection assays and inhibited ER binding to its response element in a DNA mobility shift assay. The estrogen receptor in treated cells was recognized by two monoclonal anti-ER antibodies and was not quantitatively different from ER in control cells. RNase protection analysis failed to detect any qualitative changes in the ER mRNA transcript. Mixing experiments suggest that TPA induces/activates a factor which interacts with the ER to block binding of estradiol. The effects of TPA on ER levels and binding capacity were concentration-dependent. Low concentrations of TPA inhibited estradiol binding without a decrease in the level of protein, whereas higher concentrations were required to decrease the level of ER protein. The effects of TPA appear to be mediated by activation of protein kinase C since the protein kinase C inhibitors, H-7 and bryostatin, block the effects of TPA on estradiol induction of progesterone receptor. TPA treatment had no effect on the level or binding capacity of the glucocorticoid receptor, indicating that the effects are not universal for steroid receptors. These data demonstrate that activation of the protein kinase C signal transduction pathway modulates the estrogen receptor pathway. The long term effect of protein kinase C activation is to inhibit estrogen receptor function through induction/activation of a factor which interacts with the receptor.
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PMID:Effects of 12-O-tetradecanoylphorbol-13-acetate on estrogen receptor activity in MCF-7 cells. 755 63

We previously reported transiently elevated ER protein levels in the postnatal rat hippocampus suggesting that this brain region may be sensitive to estrogenic trophic and organizational influences during a 'critical period' of sexual differentiation. In order to examine whether alterations in ER gene expression underlie the ontogenetic pattern of the hippocampal ER, we examined ER mRNA levels over the early postnatal period and in adult rats. This was accomplished by both a highly quantitative RNase protection assay and in situ hybridization histochemistry. Hippocampal ER mRNA levels increased significantly (P < 0.005) between birth and postnatal day (PDN) 4 when peak concentrations were found and then declined by PND-10. Adult male hippocampal ER mRNA values were similar to those found in newborn and PND-10 animals but were significantly less (P < 0.05) than those observed on PND-4. Results from the in situ hybridization experiments correlated well with those from the RNase protection analysis. High levels of ER mRNA were present in the CA3 pyramidal layer with somewhat lower labeling intensities present in CA1 and the dentate gyrus of the PND-4 animal. In contrast, adult male animals demonstrated little hybridization throughout the hippocampus. Thus, the temporal pattern in ER mRNA levels in the hippocampus found in the present study correlates well with our previous developmental profile of the ER protein. These findings suggest that the ontogeny of ER in the hippocampus is regulated by alterations in ER gene expression in specific neuronal populations.
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PMID:Estrogen receptor mRNA alterations in the developing rat hippocampus. 760 32

Estrogen receptor (ER) expression by breast tumors is an important predictor of disease-free survival in breast cancer patients and, more importantly, is a strong predictor of response to endocrine therapy. Variant forms of the ER may play an important role in the loss of hormone responsiveness and the progression to hormone independence. We have examined a panel of human breast tumor cell lines, both ER-positive and ER-negative, and have identified an ER mRNA variant containing a deletion of exon 5 in the ER-negative BT-20 and ER-positive MCF-7 cell lines. This exon 5 deletion variant has been previously reported to be overexpressed in ER-negative/progesterone receptor-positive breast tumors. Using RNase protection analysis, we have found that the predominant ER transcript in the BT-20 cells is the exon 5 deletion variant, while the principal transcript in MCF-7 cells is the wild-type ER mRNA. The variant ER transcript is translated into a truncated receptor protein of approximately M(r) 42,000 when expressed in yeast and, more important, in breast tumor cells. This is the first demonstration of an exon 5 deletion variant ER protein. Functional analysis has shown that this variant ER possesses constitutive transcriptional regulatory activity with respect to an estrogen-regulated reporter gene construct in a yeast expression system. The presence of this ER variant in breast tumor cell lines, as well as breast tumor biopsies and uterine tissue, suggests that it is a naturally occurring variant that may arise by alternative splicing, and whose overexpression may be involved in the progression of breast tumors to a hormone-independent state.
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PMID:Expression of a constitutively active estrogen receptor variant in the estrogen receptor-negative BT-20 human breast cancer cell line. 826 6

Studies have shown that the pineal gland via its hormone, melatonin, induces the involution of male and female reproductive systems in seasonally reproducing animals. Melatonin has direct inhibitory effects on both hypothalamic and pituitary functions, which are also exquisitely sensitive to the feedback effects of estradiol. Since melatonin can modulate estrogen receptor (ER) expression in other tissues, immunocytochemical and ribonuclease protection analyses were used to examine the effects of 12 weeks of daily late afternoon injections of melatonin on ER protein and mRNA levels in the hypothalamus of Lak.LVG golden hamsters. Significant decreases in ER-immunoreactivity were noted in the medial preoptic area (MPOA) and bed nucleus of the stria terminalis (BNST) in response to melatonin, while other hypothalamic areas which express ER, e.g. the anterior hypothalamus, showed less dramatic changes. Hypothalamic ER mRNA was decreased in response to melatonin in both intact and ovariectomized animals by 25%. In intact, cycling female hamsters, there was a significant reduction in uterine weight after melatonin treatment. These results suggest that melatonin exerts its anti-reproductive effects in hamsters by modulating ER levels in neurons of the MPOA and BNST, thereby influencing steroid feedback mechanisms.
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PMID:Effects of melatonin on estrogen receptor expression in the forebrain of outbred (Lak.LVG) golden hamsters. 911 84

Environmental contaminants, such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), cause alterations in gene expression. In this study, we measured the regulation of estrogen receptor (ER) mRNA in female CD-1 mice by competitive RT-PCR. Previous work suggests that ER protein levels are affected by TCDD, but how this is regulated is uncertain. These studies found no significant changes in ER mRNA levels, but the methods used (Northern blot analysis and RNase protection assays) lack sensitivity for measuring the low levels of RNA transcript, such as ER mRNA. The method described here offers an excellent alternative for quantifying the changes in mRNA levels. Internal competitors were created with gene-specific primers for ER and beta-actin by PCR reactions at low annealing temperatures. For each sample, the mRNA levels of ER and beta-actin were determined. Using competitive RT-PCR, the relative changes in ER mRNA from TCDD-treated and control animals were determined after normalization with the levels of beta-actin mRNA. The ER mRNA from female CD-1 mice treated with TCDD (single dose 5 micrograms/kg, i.p., 4 days) was found to be significantly suppressed as compared with the vehicle control in all tissues examined. TCDD decreased ER mRNA in the liver (30.1%) as expected. However, the greatest effect was in the reproductive tissues, with a 64.2% reduction in ER mRNA in the ovary. This is the first demonstration that TCDD causes tissue-specific downregulation of ER mRNA. These effects may contribute to the tissue-specific toxicity of TCDD.
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PMID:Regulation of estrogen receptor mRNA by 2,3,7,8-tetrachlorodibenzo-p-dioxin as measured by competitive RT-PCR. 944 63

Unfolded proteins in the endoplasmic reticulum (ER) activate the ER transmembrane sensor Ire1 to trigger the unfolded protein response (UPR), a homeostatic signaling pathway that adjusts ER protein folding capacity according to need. Ire1 is a bifunctional enzyme, containing cytoplasmic kinase and RNase domains whose roles in signal transduction downstream of Ire1 are understood in some detail. By contrast, the question of how its ER-luminal domain (LD) senses unfolded proteins has remained an enigma. The 3.0-A crystal structure and consequent structure-guided functional analyses of the conserved core region of the LD (cLD) leads us to a proposal for the mechanism of response. cLD exhibits a unique protein fold and is sufficient to control Ire1 activation by unfolded proteins. Dimerization of cLD monomers across a large interface creates a shared central groove formed by alpha-helices that are situated on a beta-sheet floor. This groove is reminiscent of the peptide binding domains of major histocompatibility complexes (MHCs) in its gross architecture. Conserved amino acid side chains in Ire1 that face into the groove are shown to be important for UPR activation in that their mutation reduces the response. Mutational analyses suggest that further interaction between cLD dimers is required to form higher-order oligomers necessary for UPR activation. We propose that cLD directly binds unfolded proteins, which changes the quaternary association of the monomers in the membrane plane. The changes in the ER lumen in turn position Ire1 kinase domains in the cytoplasm optimally for autophosphorylation to initiate the UPR.
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PMID:On the mechanism of sensing unfolded protein in the endoplasmic reticulum. 1636 12

During endoplasmic reticulum (ER) stress, homeostatic signaling through the unfolded protein response (UPR) augments ER protein-folding capacity. If homeostasis is not restored, the UPR triggers apoptosis. We found that the ER transmembrane kinase/endoribonuclease (RNase) IRE1alpha is a key component of this apoptotic switch. ER stress induces IRE1alpha kinase autophosphorylation, activating the RNase to splice XBP1 mRNA and produce the homeostatic transcription factor XBP1s. Under ER stress--or forced autophosphorylation--IRE1alpha's RNase also causes endonucleolytic decay of many ER-localized mRNAs, including those encoding chaperones, as early events culminating in apoptosis. Using chemical genetics, we show that kinase inhibitors bypass autophosphorylation to activate the RNase by an alternate mode that enforces XBP1 splicing and averts mRNA decay and apoptosis. Alternate RNase activation by kinase-inhibited IRE1alpha can be reconstituted in vitro. We propose that divergent cell fates during ER stress hinge on a balance between IRE1alpha RNase outputs that can be tilted with kinase inhibitors to favor survival.
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PMID:IRE1alpha kinase activation modes control alternate endoribonuclease outputs to determine divergent cell fates. 1966 77

Accumulation of misfolded proteins in the endoplasmic reticulum (ER) triggers the unfolded protein response (UPR), an intracellular signaling pathway that adjusts the protein folding capacity of the ER according to need. If homeostasis in the ER protein folding environment cannot be reestablished, cells commit to apoptosis. The ER-resident transmembrane kinase-endoribonuclease inositol-requiring enzyme 1 (IRE1) is the best characterized UPR signal transduction molecule. In yeast, Ire1 oligomerizes upon activation in response to an accumulation of misfolded proteins in the ER. Here we show that the salient mechanistic features of IRE1 activation are conserved: mammalian IRE1 oligomerizes in the ER membrane and oligomerization correlates with the onset of IRE1 phosphorylation and RNase activity. Moreover, the kinase/RNase module of human IRE1 activates cooperatively in vitro, indicating that formation of oligomers larger than four IRE1 molecules takes place upon activation. High-order IRE1 oligomerization thus emerges as a conserved mechanism of IRE1 signaling. IRE1 signaling attenuates after prolonged ER stress. IRE1 then enters a refractive state even if ER stress remains unmitigated. Attenuation includes dissolution of IRE1 clusters, IRE1 dephosphorylation, and decline in endoribonuclease activity. Thus IRE1 activity is governed by a timer that may be important in switching the UPR from the initially cytoprotective phase to the apoptotic mode.
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PMID:Mammalian endoplasmic reticulum stress sensor IRE1 signals by dynamic clustering. 2079 50


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