Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.11 (AMPK)
 12,425 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Treatment of hamster cells in culture with the DNA alkylating agent N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) induces DNA polymerase beta (beta-pol) gene expression and cellular levels of the enzyme. Transcriptional activity of a cloned beta-pol promoter in transient expression assays is also stimulated. Among the requirements for these responses are methylation damage to genomic DNA, cellular cAMP-dependent protein kinase, and the ATF/CREB site of the cloned beta-pol promoter. In the present study, HeLa cell nuclear extract from MNNG-treated cells was much more active in an in vitro transcription assay than nuclear extract from normal cells. By using an oligonucleotide affinity column to deplete the nuclear extract of ATF/CREB, we showed that the difference was due to ATF/CREB activator. Purified ATF/CREB activator from MNNG-treated cells was approximately 10-fold more active than ATF/CREB purified from normal cells as a transcriptional activator for the depleted nuclear extract. ATF/CREB in the extract from normal cells is known to activate in vitro transcription by increasing the rate of promoter clearance [Narayan, S., Widen, S. G., Beard, W. A., & Wilson, S. H. (1994) J. Biol. Chem. 269, 12755-12763]. With ATF/CREB from MNNG-treated cells, the amount of preinitiation complex formed was much greater than with ATF/CREB from normal cells, and the kinetics of both the closed to open preinitiation complex isomerization and promoter clearance were altered. These results indicate that the mechanism of transcriptional activation secondary to DNA alkylation damage is recruitment of more preinitiation complex and alteration of the kinetic scheme of transcription initiation.
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PMID:DNA damage-induced transcriptional activation of a human DNA polymerase beta chimeric promoter: recruitment of preinitiation complex in vitro by ATF/CREB. 781 26

Temporal cellular events responsible for hormonal activation of responses mediated by the cAMP-dependent protein kinase (PKA) have been studied in living cells. By selectively perturbing molecular function of Gs, the catalytic subunit of PKA (C), or the nuclear factor CREB, in cells through microinjection of inhibitory agents specific for these molecules or activated forms of these molecules, we have obtained evidence for a requirement for the function of each of these molecules in the hormonal stimulation of cAMP-regulated genes. Moreover, by introducing fluorescently labeled PKA subunits into these cells as molecular tracers, or by immunofluorescence of C subunit, we have observed biological translocation of C subunit from the cytoplasm to the nucleus during transcriptional activation and a quenching of this by the inhibitor molecule, PKI. The implications of these cellular and molecular events in the signal transduction of hormonal responses are discussed.
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PMID:Signal transduction through the cAMP-dependent protein kinase. 793 49

We report that the small tumor (small-t) antigen of simian virus 40 (SV40) forms complexes with nuclear protein phosphatase 2A (PP2A) and regulates the phosphorylation and transcriptional transactivation function of the cyclic AMP (cAMP)-regulatory element binding protein (CREB). PP2A coimmunoprecipitated with small t from nuclear extracts from HepG2 cells expressing small t or from rat liver nuclear extracts to which recombinant small t was added. Protein phosphatase 1 was not detected in small-t immunoprecipitates. In HepG2 cells expressing small t, dibutyryl-cAMP (Bt2cAMP) stimulated the phosphorylation of CREB 65-fold, whereas CREB phosphorylation was stimulated only 5- to 8-fold by Bt2cAMP in cells not expressing small t. Small t also inhibited the dephosphorylation of cAMP-dependent protein kinase (PKA)-phosphorylated CREB in rat liver nuclear extracts. In cells expressing small t, Bt2cAMP-stimulated transcription from the phosphoenolpyruvate carboxykinase (PEPCK) gene promoter was enhanced over the level of transcription from the PEPCK promoter in cells not expressing small t. Small t also enhanced Bt2cAMP-stimulated transcription from a Gal4-responsive promoter in cells expressing a chimeric protein containing the Gal4 DNA-binding domain linked to the CREB transactivation domain. However, small t did not stimulate transcription either from a 5' deletion mutant of the PEPCK promoter that is not able to bind CREB or from the Gal4-responsive promoter in the absence of the Gal4-CREB protein. These data suggest that small t enhances Bt2cAMP-stimulated gene transcription by inhibiting the dephosphorylation of PKA-phosphorylated CREB by nuclear PP2A. These findings support previous observations that nuclear PP2A is the primary phosphatase that dephosphorylates PKA-phosphorylated CREB.
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PMID:Simian virus 40 small tumor antigen inhibits dephosphorylation of protein kinase A-phosphorylated CREB and regulates CREB transcriptional stimulation. 806 21

Phosphorylation of CREB (cyclic AMP [cAMP]- response element [CRE]-binding protein) by cAMP-dependent protein kinase (PKA) leads to the activation of many promoters containing CREs. In neurons and other cell types, CREB phosphorylation and activation of CRE-containing promoters can occur in response to elevated intracellular Ca2+. In cultured cells that normally lack this Ca2+ responsiveness, we confer Ca(2+)-mediated activation of a CRE-containing promoter by introducing an expression vector for Ca2+/calmodulin-dependent protein kinase type IV (CaMKIV). Activation could also be mediated directly by a constitutively active form of CaMKIV which is Ca2+ independent. The CaMKIV-mediated gene induction requires the activity of CREB/ATF family members but is independent of PKA activity. In contrast, transient expression of either a constitutively active or wild-type Ca2+/calmodulin-dependent protein kinase type II (CaMKII) fails to mediate the transactivation of the same CRE-containing reporter gene. Examination of the subcellular distribution of transiently expressed CaMKIV and CaMKII reveals that only CaMKIV enters the nucleus. Our results demonstrate that CaMKIV, which is expressed in neuronal, reproductive, and lymphoid tissues, may act as a mediator of Ca(2+)-dependent gene induction.
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PMID:Calcium/calmodulin-dependent protein kinase types II and IV differentially regulate CREB-dependent gene expression. 806 43

Hormones and cytokines regulate many cellular functions by activating the ubiquitous cAMP-dependent protein kinase (A kinase) system. Newly synthesized cAMP molecules bind to regulatory (R) subunits in A kinase holoenzymes, causing them to release their catalytic (C) subunits. These free C subunits then phosphorylate proteins until the cAMP level falls, whereupon the R subunits regain their affinity for free C subunits, and thus form inactive holoenzymes again. However if cAMP levels remain persistently elevated, many cells change their A kinase system. Some cells alter the rate of degradation of subunits, and some cells change the level or stability of the messages encoding subunits. Cellular behavior often changes if cAMP levels remain elevated: many cells differentiate, some cells proliferate, and some cells die, depending on the stage of the cell cycle. The two forms of A kinase holoenzyme (type I and type II) contain identical C subunits, but contain either an RI dimer or an RII dimer. In some tissues, type II holoenzyme is compartmentalized to subcellular organelles via specific anchoring proteins, whereas type I holoenzyme is generally cytosolic. Free RI subunits turn over more rapidly than free RII subunits in most cells, but all free subunits are degraded more rapidly than when they are associated together in holoenzymes. Free C subunits can phosphorylate a broad spectrum of proteins in both the cytoplasm and nucleus, depending on the type of cell, its state of differentiation, and the hormonal milieux. If free C subunit is microinjected into the cytoplasm of some intact cells, it migrates to the nucleus, whereas if free R subunit is microinjected, it remains in the cytoplasm. If both subunits are coinjected, R subunit blocks the nuclear migration of the C subunit. A major nuclear target for free C subunits is the CREB family of nuclear proteins, which bind to cAMP response elements (CREs) in the promoter regions of cAMP-responsive genes. Phosphorylation of CREB proteins alters their ability to form dimers and to interact with CREs. Many CREB proteins can be phosphorylated by other kinases as well, indicating this is one means by which cells coordinate cAMP- and non-cAMP-mediated gene responses. However, interactions between CREB and a number of other nuclear proteins with which they can dimerize, especially proteins whose levels are rapidly altered in response to hormones, provide an even higher degree of complexity of gene regulation than is possible from various kinases phosphorylating the different sites in CREB proteins.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:The ways in which hormones change cyclic adenosine 3',5'-monophosphate-dependent protein kinase subunits, and how such changes affect cell behavior. 826 10

The cAMP/cAMP-dependent protein kinase (A-kinase) and Ca2+/calmodulin-dependent protein kinase (Cam-kinase) signal transduction pathways are well known to regulate gene transcription, but this has not been demonstrated directly for the cGMP/cGMP-dependent protein kinase (G-kinase) signal transduction pathway. Here we report that transfection of G-kinase into G-kinase-deficient cells causes activation of the human c-fos promoter in a strictly cGMP-dependent manner. The effect of G-kinase appeared to be mediated by several sequence elements, most notably the serum response element (SRE), the AP-1 binding site (FAP), and the cAMP response element (CRE). The magnitude of G-kinase transactivation of the fos promoter was similar to that of A-kinase, but there were significant differences between G-kinase and A-kinase activation of single enhancer elements and of a chimeric Gal4-CREB transcription factor. Our results indicate that G-kinase transduces signals to the nucleus independently of A-kinase or Ca2+, although it may target some of the same transcription factors as A-kinase and Cam-kinase.
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PMID:Regulation of gene expression by cGMP-dependent protein kinase. Transactivation of the c-fos promoter. 861 18

Activity-mediated gene expression is thought to play an important role in many forms of neuronal plasticities. We have used pentylenetetrazol-induced seizure that produces synchronous and sustained neuronal activity as a model to examine the mechanism(s) of gene activation. The transcription factor CREB (Ca2+/cAMP response element-binding protein) is thought to be necessary for long-term memory formation both in invertebrates and vertebrates. When phosphorylated on Ser133 either by cAMP-dependent protein kinase and/or Ca2+/calmodulin-dependent protein kinases, CREB increases transcription of genes containing the CRE (cAMP response element) sequence. Using an antibody that detects Ser133-phosphorylated CREB protein, we show that CREB phosphorylation is maximal between 3 and 8 min after the onset of seizure activity and declines slowly both in the hippocampus and the cortex. The total amount of CREB protein did not change at the time points examined. The increased phosphorylation of CREB protein is preceded by an increase in the amount of cAMP, suggestive of cAMP-dependent protein kinase activation, in the hippocampus and activation of Ca2+/calmodulin-dependent protein kinases in the cortex. Subsequent to CREB phosphorylation, the expression of the CRE-containing gene, c-fos, and the AP-1 complexes (heterodimers of Fos and Jun family members) is increased. These findings support the role of CREB-mediated gene expression in activity-dependent neuronal plasticities.
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PMID:Neuronal activity increases the phosphorylation of the transcription factor cAMP response element-binding protein (CREB) in rat hippocampus and cortex. 866 77

Transcription of a number of eukaryotic genes is activated in response to an increase in the intracellular cAMP concentration. These genes stimulated by cAMP have a common promoter element, cAMP response element (CRE). The CRE is recognized by a CRE binding protein, CREB. The binding of CREB to CRE does not induce transcription. Activation of transcription requires the phosphorylation or CREB at Ser-133. In the case of the cAMP pathway, the activated catalytic subunit of cAMP-dependent protein kinase (PKA) translocates to the nucleus and phosphorylates Ser-133 of CREB. In the nervous system, signals transmitted across synapses are known to regulate gene expression in the post-synaptic cell. This process often involves membrane depolarization and subsequent amplification of intracellular Ca2+. The transcriptional activation induced by membrane depolarization and Ca2+ influx is mediated by a promoter element, called the Ca(2+)-responsive element (CaRE). Recent studies of c-fos and proenkephalin gene expression have shown that the CaRE is indistinguishable from a CRE. In this paper, we focus on the possible interactions between Ca2+ and the cAMP signaling pathways into the nucleus.
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PMID:[Transcriptional regulation by cAMP and calcium]. 872 Oct 7

cAMP/cAMP-dependent protein kinase (PKA) signaling pathway has been recently proposed to participate in both the late phase of long term potentiation in the hippocampus and in the late, protein synthesis-dependent phase of memory formation. Here we report that a late memory consolidation phase of an inhibitory avoidance learning is regulated by an hippocampal cAMP signaling pathway that is activated, at least in part, by D1/D5 receptors. Bilateral infusion of SKF 38393 (7.5 microg/side), a D1/D5 receptor agonist, into the CA1 region of the dorsal hippocampus, enhanced retention of a step-down inhibitory avoidance when given 3 or 6 h, but not immediately (0 h) or 9 h, after training. In contrast, full retrograde amnesia was obtained when SCH 23390 (0.5 microg/side), a D1/D5 receptor antagonist, was infused into the hippocampus 3 or 6 h after training. Intrahippocampal infusion of 8Br-cAMP (1.25 microg/side), or forskolin (0.5 microg/side), an activator of adenylyl cyclase, enhanced memory when given 3 or 6 h after training. KT5720 (0.5 microg/side), a specific inhibitor of PKA, hindered memory consolidation when given immediately or 3 or 6 h posttraining. Rats submitted to the avoidance task showed learning-specific increases in hippocampal 3H-SCH 23390 binding and in the endogenous levels of cAMP 3 and 6 h after training. In addition, PKA activity and P-CREB (phosphorylated form of cAMP responsive element binding protein) immunoreactivity increased in the hippocampus immediately and 3 and 6 h after training. Together, these findings suggest that the late phase of memory consolidation of an inhibitory avoidance is modulated cAMP/PKA signaling pathways in the hippocampus.
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PMID:Involvement of hippocampal cAMP/cAMP-dependent protein kinase signaling pathways in a late memory consolidation phase of aversively motivated learning in rats. 919 88

Recent studies from our laboratory (Endocrinology 136:4762-4768, 1995) demonstrating that the expression of cAMP-dependent nuclear transcription factor CREB (cAMP response element binding protein) is lost following ovulation in macaques has revealed a novel mechanism by which the cytoplasmic and nuclear actions the cAMP-protein kinase A (PKA) intracellular signaling system may be regulated independently. Implicit in this hypothesis is the assumption that PKA activity is maintained throughout the luteal phase of the menstrual cycle, yet to date there have been no published reports regarding PKA activity in the primate corpus luteum. PKA activity was assessed by the incorporation of 32P from radiolabeled ATP into a PKA-specific peptide substrate (kemptide) in the presence or absence of cAMP. Luteal cytosolic fractions were obtained from corpora lutea collected during the spontaneous luteal phase (days 3-5, 7-8, 10-11, 13-15, and postmenses) or obtained from animals on days 11 or 16 of the luteal phase after the animals received seven days of exogenous human CG (hCG) treatment. Examination of PKA activity in luteal slices from various aged CL maintained in short-term organ culture in the presence or absence of recombinant cynomolgus monkey LH was also performed. There were no significant differences in basal or cAMP-stimulated PKA activities in corpora lutea collected throughout the spontaneous luteal phase. Further, Western immunoblot analyses of the catalytic subunit of PKA (PKA C alpha) in corpora lutea collected throughout the luteal phase revealed immunoreactive protein bands with similar intensities. In vitro addition of recombinant cynomolgus LH and dibutyryl cAMP stimulated PKA activity in corpora lutea collected during the early, mid, and late luteal phases. In corpora lutea obtained from animals treated with hCG during the midluteal phase, basal PKA activity was decreased 65% as compared with untreated day 11 controls and in late luteal phase, hCG-exposed CL basal PKA activity was decreased 30% as compared with untreated day 16 controls. However, there were no measurable differences in cAMP-stimulated PKA activity in CL exposed to prior hCG treatment in vivo and Western immunoblot analyses for PKA C alpha in these tissues revealed immunoreactive protein bands that were comparable with corpora lutea collected from untreated animals. Further, immunoblot analyses for CREB in corpora lutea collected from hCG-treated animals revealed that CREB immunoreactivity remained undetectable following a treatment regimen with hCG that mimics early pregnancy. These results demonstrate that, although CREB expression ceases following ovulation, PKA activity is maintained throughout the luteal phase, which provides a mechanism by which the acute steroidogenic actions of LH may be separated from longer term trophic actions that may rely the transcriptional activity of CREB.
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PMID:Cyclic adenosine monophosphate signaling in the primate corpus luteum: maintenance of protein kinase A activity throughout the luteal phase of the menstrual cycle. 923


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