Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.1 (protein kinase)
 81,284 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Inositol 2,4,5-trisphosphate irreversibly activated capacitative calcium entry in Xenopus oocytes, whereas guanosine thiotriphosphate (GTP[S]) and AIF4- only activated capacitative calcium entry transiently. Both GTP[S] and AIF4- inhibited capacitative calcium entry activated by thapsigargin pretreatment, but guanosine thiodiphosphate (GDP[S]), inositol 2,4,5-trisphosphate and dibutyryl cyclic GMP did not affect capacitative calcium entry. This suggests the involvement of heterotrimeric GTP-binding proteins in the regulation of capacitative calcium entry. Activation of protein kinase C or cyclic-AMP-dependent protein kinase had profound effects on capacitative calcium entry, which were consistent with the hypothesis that the effects of GTP[S] and AIF4- on capacitative calcium entry may be mediated via heterotrimeric GTP-binding protein stimulation of kinases. Further evidence for this hypothesis was derived from the result that the effects of GTP[S] on calcium entry could be inhibited by the application of the protein kinase inhibitor staurosporine.
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PMID:G-protein regulation of capacitative calcium entry may be mediated by protein kinases A and C in Xenopus oocytes. 774 94

In addition to inhibiting the proteolytic activity of the matrix metalloproteinases, tissue inhibitors of metalloproteinases (TIMPs) promote the growth of cells in the absence of other exogenous growth factors. TIMP-2 stimulates the proliferation of fibrosarcoma (HT-1080) cells and normal dermal fibroblasts (Hs68) in a dose-dependent manner. This response is evident as early as 2 h and persists up to 48 h after treatment with recombinant TIMP-2 (rTIMP-2). The specificity of this response is demonstrated by the ability of affinity-purified polyclonal anti-TIMP-2 antibodies to ablate TIMP-2 mitogenesis and by the lack of response to TIMP-1. This response is also blocked by the presence of an adenylate cyclase inhibitor, 9-(tetrahydro-2-furyl)adenine (SQ22536). Although SQ22536 did not affect untreated fibroblasts or fibrosarcoma cells, this inhibitor completely abrogates the proliferative response induced by rTIMP-2. Treatment of these cells with rTIMP-2 also stimulates the production of cAMP in a time-dependent manner that differs for the two cell lines. Moreover, treatment of purified cell membranes with rTIMP-2 suppresses cholera toxin-mediated ADP-ribosylation of the GTP-binding protein, Gs alpha subunit. These results indicate that the alpha beta gamma heterotrimer is dissociated by treatment with rTIMP-2, which may facilitate the Gs alpha-mediated activation of adenylate cyclase and subsequent production of cAMP. Since cAMP binds to the regulatory subunit of cAMP-dependent protein kinase and activates kinase activity, we evaluated how treatment with rTIMP-2 affected both these parameters. We demonstrate in this report that the cAMP produced in response to treatment with rTIMP-2 binds to the type I regulatory subunit of cAMP-dependent protein kinase and stimulates kinase activity. These results are the first demonstration that TIMP-2 directly activates adenylate cyclase to produce cAMP, which increases cAMP-dependent protein kinase activity, resulting in stimulation of fibroblast mitogenesis.
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PMID:Tissue inhibitor of metalloproteinase-2 stimulates fibroblast proliferation via a cAMP-dependent mechanism. 776 48

1. Whole-cell recordings were made from striatal neurones obtained from neonatal rats and maintained in primary cultures. The effects of dopamine D1 receptor activation were studied on the voltage-gated sodium current. 2. Bath application of a specific D1 agonist, SKF38393 (1 microM), reduced the neuronal excitability recorded in current-clamp by increasing the threshold for generation of action potentials. 3. In voltage-clamp recordings, SKF38393 (1 microM) reversibly reduced the peak amplitude of the sodium current by 37.8 +/- 4.95%. This effect was reversed by the D1 antagonist SCH23390 and was blocked by the intracellular loading of GDP-beta-S (2 mM) suggesting GTP-binding protein involvement. 4. The D1 agonist reduced the peak amplitude of the sodium current without significantly affecting (i) the voltage dependence of the current-voltage relationship, (ii) the voltage dependence of the steady-state activation and inactivation, (iii) the kinetics of the time-dependent inactivation, and (iv) the kinetics of recovery from inactivation. 5. The peak amplitude of the sodium current was progressively reduced by intracellular loading of cyclic AMP-dependent protein kinase (100 U ml-1). 6. Diffusion of a specific peptide inhibitor of the cyclic AMP-dependent protein kinase (PKI; 10 microM) into the cytosol of neurones blocked the effect of the D1 agonist on the sodium current amplitude. 7. These results demonstrate that dopamine acting at the D1 receptor reduces the amplitude of the sodium current without modifying its voltage- and time-dependent properties. This effect involves activation of the cyclic AMP-dependent protein kinase and results in a depression of the striatal neuronal excitability by increasing the threshold for generation of action potentials.
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PMID:Dopamine D1 receptor modulates the voltage-gated sodium current in rat striatal neurones through a protein kinase A. 777 43

Subcellular fractions were prepared from human platelet membranes by sucrose density gradient centrifugation and the localization of a low M(r) GTP-binding protein, rap1 protein (Rap1) was analysed by immunoblotting using a specific antibody. Rap1, which has been purified from human platelets, was found to be located in plasma membrane and alpha-granule fractions in resting platelets. Treatment of isolated alpha-granules with pronase led to proteolysis of Rap1, indicating that this protein is exposed to the cytoplasmic face of the granules. Degranulation of alpha-granules consists of translocation and subsequent fusion of the granules with the open canalicular system. Activation of this process by thrombin induced the redistribution of Rap1 on the alpha-granules to plasma membranes. On the other hand, Rap1 is known to be phosphorylated by cyclic AMP-dependent protein kinase (A-kinase) in vitro and in vivo. In intact human platelets, phosphorylation of Rap1 by A-kinase in response to prostaglandin E1 (PGE1) was observed only in Rap1 localized in plasma membranes and not on alpha-granules, although Rap1 was phosphorylated in a cell-free system when plasma membranes and alpha-granule membranes were exposed to A-kinase as substrates. These results strongly suggest that Rap1 in plasma membranes and the protein on alpha-granules are regulated by different mechanisms, and have different functions.
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PMID:A low M(r) GTP-binding protein, Rap1, in human platelets: localization, translocation and phosphorylation by cyclic AMP-dependent protein kinase. 778 83

We have recently identified a new member of the Ras/GTPase superfamily termed Rad which has unique sequence features and is overexpressed in the skeletal muscle of humans with type II diabetes (Reynet, C., and Kahn, C. R. (1993) Science, 262, 1441-1444). When expressed in bacteria as a glutathione S-transferase fusion protein, Rad bound [alpha-32P]GTP quickly and saturably. Binding was specific for guanine nucleotides and displayed unique magnesium dependence such that both GTP and GDP binding were optimal at relatively high Mg2+ concentrations (1-10 mM). Rad had low intrinsic GTPase activity which was greatly enhanced by a GTPase-activating protein (GAP) activity present in various tissues and cell lines. Several known GAPs had no stimulatory effect toward Rad. Conversion of Ser to Asn at position 66 in Rad (equivalent to position 12 in Ras) resulted in a total loss of GTP binding. Mutation of Pro61 (equivalent to Gly12 in Ras) or Gln109 (equivalent to Gln61 in Ras) had no effect on Rad GTPase activity, whereas creation of a double mutation at these positions resulted in exceptionally high intrinsic GTPase activity. In vitro, Rad was phosphorylated by the catalytic subunit of cAMP-dependent protein kinase (PK). Phosphopeptide mapping indicated two PKA phosphorylation sites near the COOH terminus. Rad also co-precipitated a serine/threonine kinase activity from extracts of various tissues and cell lines which catalyzed phosphorylation on Rad but was not inhibited by PKA inhibitor. Thus, Rad is a GTP-binding protein and a GTPase which has some structure/function similarities to Ras, but displays unique features. Rad may also be phosphorylated on serine/threonine residues by PKA and other kinases, as well as regulated by its own GAP which is present in many tissues and cell types.
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PMID:Characterization of Rad, a new member of Ras/GTPase superfamily, and its regulation by a unique GTPase-activating protein (GAP)-like activity. 787 54

Several independent studies indicate that synthetic inhibitors of cyclic-3',5'-nucleotide phosphodiesterase (PDE) isozymes, especially inhibitors of PDE-IV, are potent agents which suppress generation of reactive oxygen metabolites (ROM) by NADPH oxidase in leukocytes. Recent studies also show that NADPH oxidase is present in all cell types populating glomeruli. In view of this, we investigated PDE isozymes and their relation to ROM in isolated rat glomeruli. Glomeruli have the capacity to hydrolyze cAMP by isozymes PDE-II, PDE-III and PDE-IV, whereas cGMP is hydrolyzed by PDE-I and PDE-V. Inhibitor of PDE-IV rolipram inhibited significantly (cca 40 to 50%) ROM generation in response to stimulation by phorbol myristate acetate (PMA). Inhibitor of PDE-III cilostamide had only minor suppressive effects and inhibitors of other PDE isozymes did not influence ROM generation. Rolipram (3 microM) suppressed ROM generation without detectable increase in cAMP content. Incubation of glomeruli with forskolin, which increased cAMP content in glomeruli tenfold, inhibited ROM generation to a similar degree as rolipram. The suppression of ROM generation by rolipram was prevented by Rp-cAMPS, a specific inhibitor of protein kinase A (PKA) activity. Further, incubation of glomeruli with rolipram elicited marked in situ activation of PKA (+ 100%), as documented by increase in the (-cAMP/+cAMP) PKA activity ratio. We suggest that selective inhibitor of PDE-IV rolipram acted via the cAMP-signaling pathway and suppressed ROM generation possibly via phosphorylating ras-type GTP-binding protein component of NADPH oxidase and thereby blocking assembly of functional NADPH oxidase complex.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Formation of reactive oxygen metabolites in glomeruli is suppressed by inhibition of cAMP phosphodiesterase isozyme type IV. 793 46

LTE1 belongs to the CDC25 family that encodes a guanine nucleotide exchange factor for GTP-binding proteins of the ras family. Previously we have shown that LTE1 is essential for termination of M phase at low temperatures. We have identified TEM1 as a gene that, when present on a multicopy plasmid, suppresses the cold-sensitive phenotype of lte1. Sequence analysis of TEM1 and GTP-binding analysis of the gene product revealed that TEM1 encodes a novel low-molecular-weight GTP-binding protein. The defect of TEM1 was lethal, and the tem1-defective cells were arrested at telophase with high H1-kinase activity under restrictive conditions, indicating that TEM1 is required to exit from M phase. The defect of TEM1 was suppressed by a high dose of CDC15, which encodes a protein kinase homologous to mitogen-activated protein kinase kinase kinases. The genetic interaction among LTE1, TEM1, and CDC15 indicates that they cooperatively play an essential role for termination of M phase.
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PMID:The yeast TEM1 gene, which encodes a GTP-binding protein, is involved in termination of M phase. 793 62

The adenylate cyclase system has been implicated in sweet taste transduction. The purpose of this study was to determine whether application of modulators of the adenylate cyclase system to the tongue alters sweet taste responses. Integrated chorda tympani (CT) recordings were made in gerbils to sweet tastants before and after a 4-min application of four types of modulators of the adenylate cyclase system. The four types of modulators tested were: a) NaF, a compound that promotes dissociation of GTP-binding protein; b) forskolin, a powerful stimulant of adenylate cyclase; c) 8-bromoadenosine 3' :5'-cyclic monophosphate sodium salt (8BrcAMP) and N6,2'-O-dibutyryladenosine 3' :5'-cyclic monophosphate sodium salt (DBcAMP), two membrane permeable forms of cAMP; and d) 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine dihydrochloride (H-7) and N-(2-[methylamino]ethyl)-5-isoquinolinesulfonamide dihydrochloride) (H-8), which are protein kinase inhibitors. The sweet compounds tested were: sucrose (30 mM and 100 mM), glucose (300 mM), fructose (300 mM), maltitol (150 mM and 300 mM), mannitol (300 mM and 500 mM), sodium saccharin (10 mM), D-tryptophan (6.5 mM), dulcin (0.88 mM, 1.75 mM, and 3.5 mM), and stevioside (0.55 mM and 1.1 mM). NaCl (30 mM and 100 mM) and KCl (300 mM and 500 mM) were used as control stimuli. The main findings were as follows. Application of NaF (20 mM) for 4 min as a rinse significantly enhanced all of the sweet compounds by at least 23%, except for 10 mM sodium saccharin and 6.5 mM D-tryptophan, while all control compounds were suppressed.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Effect of modulators of the adenylate cyclase system on sweet electrophysiological taste responses in gerbil. 797 6

We examined the regulation of an amiloride-sensitive sodium conductance expressed in human B lymphoid cells. This conductance was activated by two independent pathways, one involving cyclic adenylyl monophosphate (cAMP)-dependent protein kinase and the other involving a pertussis toxin-sensitive G-protein. Cholera toxin, presumably by increasing cellular cAMP, and pertussis toxin, which ADP-ribosylates certain GTP-binding proteins, both independently increased the amiloride-sensitive sodium conductance. Simultaneous treatment with both toxins, however, failed to increase the sodium conductance, implying that a single set of sodium channels was being affected by both toxins. In cells preactivated with pertussis toxin, 8-chlorophenylthio-cAMP inhibited the activated sodium conductance back to the basal level. Thus, cyclic AMP-dependent pathways can either activate or inhibit amiloride-sensitive sodium channels, depending upon the activation state of a pertussis toxin-sensitive GTP-binding protein. These findings support a hypothesis for the regulation of amiloride-sensitive sodium channels which incorporates the independent effects of cholera and pertussis toxins, and in which cyclic AMP can play a dual role in the regulation of channel activity.
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PMID:G-proteins modulate amiloride-sensitive sodium channels. 802 31

Many growth factors whose receptors are protein tyrosine kinases stimulate the MAP kinase pathway by activating first the GTP-binding protein Ras and then the protein kinase p74raf-1. p74raf-1 phosphorylates and activates MAP kinase kinase (MAPKK). To understand the mechanism of activation of MAPKK, we have identified Ser217 and Ser221 of MAPKK1 as the sites phosphorylated by p74raf-1. This represents the first characterization of sites phosphorylated by this proto-oncogene product. Ser217 and Ser221 lie in a region of the catalytic domain where the activating phosphorylation sites of several other protein kinases are located. Among MAPKK family members, this region is the most conserved, suggesting that all members of the family are activated by the phosphorylation of these sites. A 'kinase-dead' MAPKK1 mutant was phosphorylated at the same residues as the wild-type enzyme, establishing that both sites are phosphorylated directly by p74raf-1, and not by autophosphorylation. Only the diphosphorylated form of MAPKK1 (phosphorylated at both Ser217 and Ser221) was detected, even when the stoichiometry of phosphorylation by p74raf-1 was low, indicating that phosphorylation of one of these sites is rate limiting, phosphorylation of the second then occurring extremely rapidly. Ser217 and Ser221 were both phosphorylated in vivo within minutes when PC12 cells were stimulated with nerve growth factor. Analysis of MAPKK1 mutants in which either Ser217 or Ser221 were changed to glutamic acid, and the finding that inactivation of maximally activated MAPKK1 required the dephosphorylation of both serines, shows that phosphorylation of either residue is sufficient for maximal activation.
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PMID:Identification of the sites in MAP kinase kinase-1 phosphorylated by p74raf-1. 815


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