Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.25.1 (deoxyribonuclease)
 1,471 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have investigated the ability of dehydroepiandrosterone (DHEA) to alter the production of interleukin-2 (IL-2) and to bind to a specific binding complex in antiCD3 epsilon activated T cells. Binding activity correlated with the presence of a specific DHEA binding complex in the cytosol and nuclei of DHEA-responsive T-cell hybridomas, as well as in CD4+ and CD8+ cells isolated from peripheral lymph nodes of normal mice. Scatchard analysis determined that intact lymphocytes and cytosolic fractions contained high affinity binding for [3H]DHEA (approx. 2.6 nM) with 1000-7000 binding sites existing per cell. Five of the T-cell hybridomas tested both responded to DHEA treatment with increased production of IL-2 and also contained specific high affinity [3H]DHEA binding. Four additional T-cell hybridomas were found to contain no specific [3H]DHEA binding and were also unresponsive to DHEA influences on IL-2 production. Sucrose density gradients demonstrated a 3-4s [3H]DHEA binding complex in high salt and a 7-8s binding complex in low salt. Specific binding was inhibited by preincubation of the cytosol fractions with either trypsin or chymotrypsin, or by heating to 60 degrees C for 1 h (less than 15% of control). [3H]DHEA binding was unaffected by preincubation of the cytosol fractions with ribonuclease, deoxyribonuclease, or phospholipase A. The DHEA-protein complexes bound to DNA-cellulose with the amount of binding being slightly increased by preincubation at 25 degrees C as compared to 4 degrees C. As expected, [3H]DHEA binding was inhibited by the addition of unlabeled DHEA, but was also modestly inhibited by dihydrotestosterone and cortisol. Binding of DHEA was unaffected by progesterone, dexamethasone, estradiol, androsterone, DHEAS, and beta-etiocholanolone at all concentrations tested. DHEA was incapable of inhibiting the binding of [3H]DHT to the androgen receptor or [3H]dexamethasone to the glucocorticoid receptor. Collectively, these findings suggest that murine T cells contain a specific DHEA receptor. We believe that DHEA is a steroid hormone that is directly involved in the regulation of IL-2 production by both normal and some T-cell hybridomas.
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PMID:The presence of a dehydroepiandrosterone-specific receptor binding complex in murine T cells. 135 1

Glucocorticoid-induced lymphocyte cell death is a programmed process which is thought to involve the calcium-dependent degradation of DNA into multiples of 180 basepairs, characteristic of internucleosomal degradation. We have used the glucocorticoid-sensitive mouse lymphoma cell line S49.1 [wild-type (wt)] and the glucocorticoid-resistant cell line S49.22r (nt-) to evaluate the role of both glucocorticoid receptors and calcium in the regulation of internucleosomal DNA degradation and expression of calcium-dependent deoxyribonuclease activity. DNA was isolated from untreated (control) and dexamethasone (dex)-treated viable cells and analyzed for internucleosomal DNA degradation by agarose gel electrophoresis, followed by ethidium bromide staining. Glucocorticoid treatment resulted in substantial internucleosomal DNA degradation in wt cells, but not in nt- cells. This effect was inhibited by coincubation of cells with dex and the glucocorticoid receptor antagonist RU486. In contrast to the glucocorticoid response, administration of either of two calcium ionophores, ionomycin or A23187, produced internucleosomal degradation of DNA in both wt and nt- cells, although the latter were less sensitive to ionophore treatment. Interestingly, A23187 treatment also resulted in a loss of cell viability in HeLa S3 cells, a cell line that does not exhibit glucocorticoid-induced apoptosis. No internucleosomal DNA degradation was detected in HeLa S3 cells killed by A23187. To determine whether similar nucleases are associated with this internucleosomal DNA degradation resulting from both glucocorticoid and calcium ionophore treatment, 0.3 M NaCl nuclear protein extracts were prepared from control and treated cells and analyzed for protein composition or nuclease activity. To assay for nuclease activity, nuclear extracts were electrophoresed in sodium dodecyl sulfate-polyacrylamide gels impregnated with [32P]DNA. Nuclease activity was detected by removal of sodium dodecyl sulfate from the gel, activation with calcium, and subsequent visualization of the loss of [32P]DNA by autoradiography. Dex treatment of wt cells resulted in the appearance of several proteins within the mol wt range of 12-18 kDa, only one of which (16-18 kDa) exhibited calcium-dependent nuclease activity. The appearance of these proteins in nuclear extracts was inhibited by coincubation of glucocorticoid-treated cells with RU 486. Glucocorticoid treatment did not result in the appearance of nuclease activity in nuclear extracts from nt- cells. Interestingly, A23187 or ionomycin treatment resulted in an increase in activity of the 16- to 18-kDa nuclease in both wt and nt- cells. These findings indicate that both glucocorticoid receptors and calcium may share common features in the regulation of apoptosis in lymphoid cells.
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PMID:Similar actions of glucocorticoids and calcium on the regulation of apoptosis in S49 cells. 194 10

Binding of highly purified glucocorticoid receptor complexes to nuclear matrix was evaluated. Extraction of purified nuclei with 2M potassium chloride and brief deoxyribonuclease digestion leaves a matrix structure containing 1% of nuclear DNA and 6-12% of nuclear proteins. The nuclear matrix retained two binding sites for receptor complexes, a high affinity, low capacity site and a low affinity, high capacity site. These sites have affinities and capacities consistent with those reported for binding of these complexes to intact nuclei. More extensive deoxyribonuclease treatment of the matrix resulted in a marked reduction of high affinity complex binding. Furthermore, the DNA binding form of the receptor complex but not the unactivated receptor complex bound to DNA fibers anchored to nuclear matrix as visualized by 18 nm gold particle receptor complexes. The data suggest that the nuclear matrix is the major site for coordinating glucocorticoid hormone action in the nucleus.
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PMID:The nuclear matrix is the site of glucocorticoid receptor complex action in the nucleus. 372 32

We have isolated epithelial cell clusters from mammary glands of pregnant and lactating rats by collagenase-hyaluronidase-deoxyribonuclease digestion, followed by Ficoll density-gradient centrifugation. Clusters of greater than 90% viable cells were identified by light microscopy as essentially devoid of other cell types; the integrity of their subcellular organelles verified by electron microscopy. Binding characteristics of the synthetic glucocorticoid [3H]dexamethasone were studied in cytosols prepared from isolated cell clusters. Cytosols from both pregnant and lactating rats bound [3H]dexamethasone with high affinity to a single class of low capacity binding sites. In both types of cytosol the dissociation constant (Kd 4 degrees C approximately/nM) of the binding was similar; the number of sites per cell in lactating rats was approximately double that in pregnant rats. The specificity of binding was typical of a classical glucocorticoid receptor, with a hierarchy of affinity by competition studies dexamethasone greater than progesterone greater than aldosterone much much greater than testosterone = estradiol. In particular, no difference in progesterone affinity for these glucocorticoid receptors was seen between pregnancy and lactation. This suggests that reported differences in inhibitory action of progesterone, pregnancy versus post-partum, are not glucocorticoid-receptor mediated.
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PMID:Glucocorticoid receptors in epithelial cells isolated from the mammary glands of pregnant and lactating rats. 705 35

The glucocorticoid receptor (GR) is a ligand-activated transcription factor that induces expression of many genes. The GR has been useful for understanding how chromatin structure regulates steroid-induced transcription in model systems. However, the effect of glucocorticoids on chromatin structure has been examined on few endogenous mammalian promoters. We investigated the effect of glucocorticoids on the in vivo chromatin structure of the glucocorticoid-responsive I kappa B alpha gene promoter, the inhibitor of the ubiquitous transcription factor, nuclear factor kappa B (NF kappa B). Glucocorticoids inhibit NF kappa B activity in some tissues by elevating the levels of I kappa B alpha. We found that glucocorticoids activated the I kappa B alpha promoter in human T47D/A1-2 cells containing the GR. We then investigated the chromatin structure of the I kappa B alpha promoter in the absence and presence of glucocorticoids with the use of micrococcal nuclease, restriction enzyme, and deoxyribonuclease (DNaseI) analyses. In untreated cells, the promoter assembles into regularly positioned nucleosomes, and glucocorticoid treatment did not alter nucleosomal position. Restriction enzyme accessiblity studies indicated that the I kappa B alpha promoter is assembled as phased nucleosomes that adopt an "open" chromatin architecture in the absence of hormone. However, glucocorticoids may be required for transcription factor binding, because DNaseI footprinting studies suggested that regulatory factors bind to the promoter upon glucocorticoid treatment.
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PMID:Glucocorticoid receptor activation of the I kappa B alpha promoter within chromatin. 1169 73

At least three promoters (1A, 1B, and 1C) control the expression of mRNA transcripts for the human glucocorticoid receptor (hGR) protein. An hGR 1A promoter/exon sequence (-218/+269) contains at least 12 deoxyribonuclease (DNase) I footprints that contain bound protein. Whereas four of these footprints (FP6, FP7, FP8, and FP11) contain bound hGR in protein-DNA complexes that are formed, only two (FP7 and FP11) appear to be important for the up-regulation of hGR 1A promoter/exon activity in T-lymphoblasts. Furthermore, the activity of these DNA elements depends upon the promoter context, leading to a redundant and complex regulation of expression of the hGR 1A promoter/exon. FP7 appears to be required for hormonal responsiveness in the absence of upstream sequences (+41/+191), whereas the hormonal responsiveness of FP11 requires a functional, adjacent FP12 DNA sequence. FP12 contains overlapping binding sites for the protooncogene transcription factors c-Myb and c-Ets. It seems likely that binding of either c-Myb or c-Ets to FP12 is necessary for the direct or indirect binding of the hGR to FP11 (a nonconsensus glucocorticoid response element), and the resultant steroid-responsiveness of the hGR 1A promoter/exon sequence. We propose that the identity of the accessory transcription factor bound to FP12 (c-Myb or c-Ets) may determine the nature of regulation (positive or negative) of hGR gene expression by hormone, and that this may be important for hormone-induced apoptosis in T cell acute lymphoblastic leukemia.
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PMID:Steroid-responsive sequences in the human glucocorticoid receptor gene 1A promoter. 1504 98