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:3.1.3.16 (calcineurin)
17,112 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The effects of okadaic acid (OA), a protein phosphatase inhibitor, on transcriptional enhancement activity of rat glucocorticoid receptor (GR) were examined in transiently transfected cells. In the absence of hormone, GRs expressed in CV-1 and COS-1 fibroblasts were capable of enhancing transcription from cotransfected chloramphenicol acetyltransferase reporter plasmids in response to OA treatment. Synergistic enhancement resulted from combined hormone and OA treatment. The effects of OA on GR-mediated enhancement required the presence of linked glucocorticoid response elements and were observed with reporter plasmids that contained different promoters and glucocorticoid response elements. Since OA did not affect nuclear translocation of the receptor, enhancement mediated by unliganded GR was most likely accounted for by the accumulation of some unliganded GRs within nuclei of transfected CV-1 and COS-1 cells. Deletion of individual GR transactivation domains and point mutations within DNA- and hormone-binding domains severely reduced the response of receptors to OA, although some mutant receptors retained the capacity to elicit a synergistic response when exposed to OA and hormone. The effects of OA on transcriptional enhancement did not appear to correlate with major changes in GR phosphorylation, as visualized by two-dimensional tryptic mapping of in vivo 32P-labeled GRs. Thus, phosphorylation of various components of the GR signal transduction pathway, and not necessarily the receptor itself, may influence its transcriptional enhancement activity.
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PMID:Effects of okadaic acid, a protein phosphatase inhibitor, on glucocorticoid receptor-mediated enhancement. 131 Jul 97

We have used okadaic acid (OA), a cell-permeable inhibitor of serine/threonine protein phosphatase types 1 (PP-1) and 2A (PP-2A), to demonstrate that the subcellular distribution of glucocorticoid receptor (GR) in rat fibroblasts is influenced by its phosphorylation state. Nuclear GRs in OA-treated cells retain transcriptional enhancement activity. Nuclear import or export of hormone agonist-bound GRs is not affected by OA. However, a dose of OA that fully inhibits PP-2A and partially inhibits PP-1, but not a lower dose that only partially inhibits PP-2A, leads to inefficient nuclear retention of agonist-bound GRs, and their redistribution into the cytoplasm. These receptors appear to be trapped in the cytoplasmic compartment and are unable to recycle (i.e. reenter the nucleus). Addition of OA during different steps of GR recycling demonstrates that OA must be present during nuclear export of GRs to block GR recycling. A direct role for PP-1 and/or PP-2A in GR recycling is suggested by site-specific hyperphosphorylation of GRs in vivo during OA inhibition of recycling. These are the same sites that undergo in vitro site-specific dephosphorylation by PP-1 and PP-2A. The block in GR recycling that results from inhibition of PP-1 and/or PP-2A resembles effects previously observed in v-mos-transformed rat fibroblasts. Interestingly, OA inhibition of PP-2A in v-mos-transformed cells leads to the reversal of oncoprotein effects on GR recycling and retention of receptors within the nuclear compartment. We propose that GR recycling is influenced by the activities of distinct protein phosphatases (PP-1 and/or PP-2A), and that the interference of this pathway observed in v-mos-transformed cells may be the result of effects of the oncoprotein on the phosphatases or a specific subset of their targets.
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PMID:Protein phosphatase types 1 and/or 2A regulate nucleocytoplasmic shuttling of glucocorticoid receptors. 166 12

We have stably introduced expression vectors for the glucocorticoid receptor and a sensitive, hormone-responsive reporter (mouse mammary tumor virus-luciferase) into a human breast carcinoma-derived cell line. Employing this cell line, we have conducted a detailed examination of the induction of glucocorticoid-regulated genes and the phosphorylation of glucocorticoid receptor following pharmacologic manipulation of cell signaling pathways. The hormone response can be enhanced from 2 to 10-fold by activators of protein kinase A, protein kinase C, and inhibitors of protein phosphatase. Forskolin and 8-bromoadenosine 3':5'-cyclic monophosphate (BrcAMP), but not BrcGMP, enhance the hormone effect, yet surprisingly, phosphodiesterase inhibitors, isobutylmethylxanthine and Ro20-1724, strongly inhibit hormone-mediated induction of the reporter gene. These treatments do not alter cellular receptor content, dexamethasone binding, nor hormone-mediated receptor down-regulation. Tryptic peptide analysis of 32P-labeled receptor reveals that neither BrcAMP, isobutylmethylxanthine, nor the tumor promoter and protein kinase C activator, 12-O-tetradecanoyl-phorbol-13-acetate, detectably alter the state of glucocorticoid receptor phosphorylation. The only agent which alters receptor phosphorylation is the protein phosphatase inhibitor okadaic acid, but only at concentrations higher than required for maximum effects on glucocorticoid receptor transactivation. We propose that these effectors do not modify receptor directly but alter its interaction with transcription complexes.
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PMID:Modulation of cell signaling pathways can enhance or impair glucocorticoid-induced gene expression without altering the state of receptor phosphorylation. 769 81

Glucocorticoids (GC) inhibit IL-2 gene transcription by interfering with the binding of the nuclear factor activator protein-1 on the IL-2 promoter. Calcineurin, a Ca2+/calmodulin-dependent protein phosphatase, is an essential component of the T cell antigen receptor signal transduction pathway leading to IL-2 gene transcription. Therefore, we have asked whether this phosphatase may also be regulated by GC. Jurkat T cells were cotransfected with plasmids containing the intact IL-2 promoter or its NF-AT and Oct-1 motifs, and a deletion mutant (delta CaM-AI) of calcineurin known to have Ca(2+)-independent constitutive phosphatase activity. Cotransfection of IL-2 promoter with delta CaM-AI allowed the activation of IL-2 promoter in the presence of phorbol ester alone. Under these conditions dexamethasone (Dex; 10(-6) M) inhibited IL-2 promoter activation by 50-60%. The inhibitory effect of Dex was specific, as demonstrated by experiments using an unrelated promoter (simian virus 40) and estradiol. Furthermore, it was completely reversed in the presence of excess amounts of the glucocorticoid antagonist RU 486, which suggests that it is mediated through the glucocorticoid receptor. Overexpression of calcineurin via delta CaM-AI in Jurkat cells decreased their apparent sensitivity to Dex (approximately 5-fold increase in IC50). Similar results were obtained with the NF-AT and Oct-1 constructs, which are also known to be activated by calcineurin. Thus, in addition to their known inhibitory effects on activator protein-1, GC also inhibit calcineurin-dependent pathways for T cell activation.
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PMID:Glucocorticoids regulate calcineurin-dependent trans-activating pathways for interleukin-2 gene transcription in human T lymphocytes. 776 70

The synthetic glucocorticoid dexamethasone regulates tight junction permeability resulting in an increased transepithelial electrical resistance (TER) of cultured 31EG4 mammary epithelial cells. Inhibition of cellular type 1 and type 2A protein phosphatase activity by okadaic acid reduced the TER of dexamethasone-treated monolayers of 31EG4 cells to basal levels within 24 h. Coincident with the increase in tight junction permeability, immunofluorescence revealed that okadaic acid caused a partial cellular redistribution of the ZO-1 tight junction-associated protein. The potent glucocorticoid antagonist RU486 had no effect on TER or ZO-1 distribution, indicating that the effects of okadaic acid are not a result of disrupting glucocorticoid receptor function. Immunoprecipitation of 32P-labeled cells and V8 protease peptide mapping demonstrated that dexamethasone did not alter ZO-1 phosphorylation. However, consistent with the changes in TER, dexamethasone induced a 2.3-fold stimulation in ZO-1 protein levels which was reduced to 73% of basal levels by okadaic acid. No effects on ZO-1 transcript levels were observed. Monolayers grown in the presence of glucocorticoids had only 28% less junction density and 16.5% more linear junction/cell, which cannot account for the observed increases of TER and ZO-1 protein levels. Taken together, our results have shown that a disruption of phosphorylation/dephosphorylation activity overrides the glucocorticoid regulation of tight junction permeability in 31EG4 mammary cells.
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PMID:Relationship of serine/threonine phosphorylation/dephosphorylation signaling to glucocorticoid regulation of tight junction permeability and ZO-1 distribution in nontransformed mammary epithelial cells. 820 10

Glucocorticoids induce apoptosis in murine T cell hybridomas. It was inhibited by okadaic acid and calyculin A, potent inhibitors of protein phosphatase 1 and 2A, but not by 1-norokadaone, a structural analog of okadaic acid without phosphatase inhibitory activity. The inhibitory effect of okadaic acid was significant even when it was added 9 h after the start of the culture. Okadaic acid did not prevent either the translocation of glucocorticoid receptor from the cytoplasm to the nucleus or the induction of luciferase activity in the T cell hybridoma transfected with a plasmid containing the luciferase gene under the control of glucocorticoid response elements. These results indicate that protein dephosphorylation is an essential step for glucocorticoid-induced apoptosis in T cell hybridomas, and that the step is at the late stage of the apoptotic process.
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PMID:Okadaic acid inhibits glucocorticoid-induced apoptosis in T cell hybridomas at its late stage. 826 31

The immunosuppressants cyclosporin A (CyA), FK-506, and rapamycin (RAP) have multiple actions on target cells that appear to be mediated by interaction of drug-binding protein complexes. Both FK-506 and CyA, but not RAP, inhibit the Ca2(+)-dependent phosphatase, calcineurin, and in so doing have been found to inhibit Na(+)-K(+)-ATPase activity in various nephron segments. Of interest, FK-506 and RAP, but not CyA, are bound by the steroid receptor-associated FK-506-binding heat shock protein of 56 kDa, HSP56. To determine the physiological effect of this interaction on a steroid-mediated phenomenon, the effect of these agents on steroid-mediated Na+ transport in A6 cells was investigated. Aldosterone stimulation of Na+ transport and Na(+)-K(+)-ATPase activity are significantly inhibited by prolonged incubation with FK-506 and RAP. Although CyA inhibits basal Na(+)-K(+)-ATPase activity, it has no effect on aldosterone-induced Na+ transport or the aldosterone-induced increase in Na(+)-K(+)-ATPase activity. FK-506 inhibits the aldosterone-induced synthesis of G alpha i-3 protein but has no effect on glucocorticoid receptor number as quantified by Western blotting. The results suggest that FK-506 and RAP inhibit steroid-mediated Na+ transport at some pretranslational site. The common interaction of these agents with the steroid receptor-associated HSP56 might account for these findings.
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PMID:FK-506 and rapamycin but not cyclosporin inhibit aldosterone-stimulated sodium transport in A6 cells. 876 46

Increased production of prostaglandins by the gestational tissues is pivotal for the initiation and maintenance of human labour. A major source of prostaglandins in the pregnant human uterus is the amnion membrane, which synthesizes increased amounts of prostaglandin E2 (PGE2) at parturition. We have found that the activity of prostaglandin endoperoxide H2 synthase (PGHS), the enzyme catalyzing the committing step of prostanoid biosynthesis, increases significantly in the amnion at term and preterm labour, and also prior to the onset of clinical labour at term. Furthermore, the abundance of the mRNA encoding the inducible PGHS-2 isoenzyme was higher in the amnion after spontaneous delivery that before labour. The level of the constitutive PGHS-1 mRNA remained unchanged. In addition, we found a significant positive correlation between PGHS activity and the level of PGHS-2 mRNA, but not of PGHS-1 mRNA, in the individual tissue samples, also indicating that PGHS-2 was selectively induced in the amnion membrane at labour. The regulation of PGHS expression by agonists was studied using primary cultures of amnion cells. Glucocorticoid treatment enhanced the activity of PGHS and the level of PGHS-2 mRNA in the cultured cells, without affecting PGHS-1 mRNA abundance. The stimulation was glucocorticoid specific and was blocked by the glucocorticoid receptor antagonist RU486, suggesting that it was mediated by the glucocorticoid receptor. Inhibition of protein synthesis did not block the accumulation of PGHS-2 mRNA showing that the steroid acted directly, without inducing an intervening protein. Protein kinase C activator and protein phosphatase inhibitor compounds and epidermal growth factor also promoted PGHS-2 mRNA expression, demonstrating the involvement of protein kinase dependent mechanisms in PGHS-2 regulation. However, the role of these effectors in the in vivo control of PGHS-2 expression remains to be determined.
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PMID:Regulation of prostaglandin endoperoxide H2 synthase in term human gestational tissues. 904 57

FK506 is a new FDA-approved immunosuppressant used for prevention of allograft rejection in, for example, liver and kidney transplantations. FK506 is inactive by itself and requires binding to an FK506 binding protein-12 (FKBP-12), or immunophilin, for activation. In this regard, FK506 is analogous to cyclosporin A, which must bind to its immunophilin (cyclophilin A) to display activity. This FK506-FKBP complex inhibits the activity of the serine/threonine protein phosphatase 2B (calcineurin), the basis for the immunosuppressant action of FK506. The discovery that immunophilins are also present in the nervous system introduces a new level of complexity in the regulation of neuronal function. Two important calcineurin targets in brain are the growth-associated protein GAP-43 and nitric oxide (NO) synthase (NOS). This review focuses on studies showing that systemic administration of FK506 dose-dependently speeds nerve regeneration and functional recovery in rats following a sciatic-nerve crush injury. The effect appears to result from an increased rate of axonal regeneration. The nerve regenerative property of this class of agents is separate from their immunosuppressant action because FK506-related compounds that bind to FKBP-12 but do not inhibit calcineurin are also able to increase nerve regeneration. Thus, FK506's ability to increase nerve regeneration arises via a calcineurin-independent mechanism (i.e., one not involving an increase in GAP-43 phosphorylation). Possible mechanisms of action are discussed in relation to known actions of FKBPs: the interaction of FKBP-12 with two Ca2+ release-channels (the ryanodine and inositol 1,4,5-triphosphate receptors) which is disrupted by FK506, thereby increasing Ca2+ flux; the type 1 receptor for the transforming growth factor-beta (TGF-beta 1), which stimulates nerve growth factor (NGF) synthesis by glial cells, and is a natural ligand for FKBP-12; and the immunophilin FKBP-52/FKBP-59, which has also been identified as a heat-shock protein (HSP-56) and is a component of the nontransformed glucocorticoid receptor. Taken together, studies of FK506 indicate broad functional roles for the immunophilins in the nervous system. Both calcineurin-dependent (e.g., neuroprotection via reduced NO formation) and calcineurin-independent mechanisms (i.e., nerve regeneration) need to be invoked to explain the many different neuronal effects of FK506. This suggests that multiple immunophilins mediate FK506's neuronal effects. Novel, nonimmunosuppressant ligands for FKBPs may represent important new drugs for the treatment of a variety of neurological disorders.
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PMID:FK506 and the role of immunophilins in nerve regeneration. 945 3

Ligand-induced glucocorticoid receptor (GR) activation has recently been linked to the inhibition of cell proliferation via the transcriptional induction of p21(WAF1/Cip1), which functions as a universal inhibitor of cyclin-dependent protein kinases. Herein, we identify a Ser/Thr protein phosphatase (PP5) that promotes cellular proliferation by inhibiting both glucocorticoid and p53-mediated signaling pathways leading to p21(WAF1/Cip1)-mediated growth arrest. The suppression of PP5 expression (1) markedly increases the association of GR with its cognate DNA-binding sequence, (2) induces GR transcriptional activity without the addition of hormone, and (3) increases dexamethasone-mediated induction of GR reporter activity to a level that is approximately 10 times greater than the maximal response obtainable in the presence of PP5. PP5 has no apparent effect on the binding of hormone to the GR, and dexamethasone-mediated growth arrest correlates with an increase in p53 phosphorylation. Comparative studies in p53-wild-type, p53-defective, and p53-deficient cell lines indicate that either (1) p53 participates in GR-mediated induction of p21(WAF1/Cip1), with the hyperphosphorylation of basal p53 induced by glucocorticoids sufficient for the propagation of an antiproliferative response when PP5 expression is inhibited, or (2) PP5 acts where p53-mediated and GR-induced signaling networks converge to regulate the transcriptional induction of p21(WAF1/Cip1). Thus, aberrant PP5 expression may have an additive effect on the development of human cancers by promoting cell proliferation via the inhibition of a GR-induced antiproliferative signaling cascade, and facilitating neoplastic transformation via the inhibition of a growth-arresting p53-mediated response that guards against genomic instability.
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PMID:Ser/Thr protein phosphatase type 5 (PP5) is a negative regulator of glucocorticoid receptor-mediated growth arrest. 1041 57


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