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

Quantitative autoradiography was used to characterize and localize [3H]cGMP binding sites in the rat brain. [3H]cGMP binding was found to be pH-sensitive (with two optima at 7.4 and 5.0) and Mg2+-dependent. At pH 7.4, the binding was dependent on inclusion of the phosphodiesterase inhibitor IBMX. In contrast, at pH 5.0, IBMX had little effect on binding. The binding of [3H]cGMP was reversible and saturable with a Kd of 22 nM at pH 7.4 and 36 nM at pH 5.0. Bmax values were 172 fmol/mg at pH 7.4 and 462 fmol/mg at pH 5.0. [3H]cGMP binding was inhibited by cGMP and its analogues, with cGMP and cAMP being the most potent at pH 7.4 and cGMP and 8-Br-cGMP being the most potent at pH 5.0. Using an extracellular pH 7.4 buffer, the selective cGMP-dependent protein kinase (PKG) inhibitor Rp-8pCPT-cGMPS had very little effect on [3H]cGMP binding. In contrast, with a cytosolic pH 5.0 buffer, Rp-8pCPT-cGMPS displaced binding in the cerebellum. This indicates that PKG is localized in the cerebellum, and that the binding to PKG is favored under cytosolic conditions. Autoradiographic localization of [3H]cGMP binding sites revealed a heterogeneous distribution with the highest densities in the substantia nigra and interpeduncular nucleus. High densities were also observed in the basal ganglia, the medial habenular nucleus, the frontoparietal cortex, the lateral amygdaloid nucleus and the subiculum. It is concluded that the nature of [3H]cGMP binding is complex, with one site probably being related to cytosolic PKG mainly found in the cerebellum, and one site probably representing cGMP-stimulated phosphodiesterase mainly located in the forebrain.
 ...
PMID:Autoradiographic characterization of [3H]cGMP binding sites in the rat brain. 927 22

Glutamate, the neurotransmitter released by photoreceptors, excites horizontal cells and OFF-type bipolar cells by activating ionotropic receptors. This study investigated an additional action of glutamate in which it modulates a voltage-gated ion channel in horizontal cells. We find that glutamate and APB (2-amino-4-phosphonobutyrate) produce a delayed and moderately prolonged suppression of an inward rectifier current (IRK+). This effect is proposed to occur via an APB-sensitive metabotropic glutamate receptor (mGluR) because common agonists for the ionotropic or APB-insensitive mGluRs are ineffective and the APB-insensitive receptor antagonist alpha-methyl-4-carboxyphenylglycine (MCPG) does not block the actions of glutamate or APB. 8-Br-cGMP, 1-methyl-3-isobutylxanthine (IBMX), and atrial natriuretic peptide (ANP) but not 8-Br-cAMP mimic the suppression of IRK+. The effects of glutamate and APB are blocked by protein kinase inhibitors including Rp-8-pCPT-cGMPS, H-8, and H-7 as well as by ATPgammaS. We hypothesize that the APB receptor suppresses IRK+ via upregulation of cGMP and subsequent activation of a cGMP-dependent protein kinase. This pathway is likely regulated by an ATP-dependent phosphorylation. This is a novel signaling pathway for mGluRs and indicates that at least two distinct APB-activated pathways exist in the retina. Functionally, this APB receptor-mediated action found in horizontal cells would provide a means by which spatially restricted changes of glutamate, produced by local illumination of photoreceptors, could regulate IRK+ and consequently the response properties of these neurons. This would serve to adapt selectively retinal regions stimulated by small regions of the visual world.
 ...
PMID:Metabotropic glutamate receptor-mediated suppression of an inward rectifier current is linked via a cGMP cascade. 936 42

This study examines the involvement of hormones and neuromediators in the regulation of Na+, K+, Cl- cotransport in renal epithelial cells using Madin-Darby canine kidney cells with low transepithelial electrical resistance (194+/-47 Omega/cm2). In this cell line, Na+, K+, Cl- cotransport measured as bumetanide-sensitive 86Rb influx was inhibited up to 50-60% with agonists of P2-purinoceptors (ATP approximately ADP>UTP>AMP), slightly (15-30%) increased by activators of cAMP signaling (forskolin, 8-Br-cAMP) and was insensitive to activators of cGMP signaling (8-Br-cGMP, nitroprusside), EGF, angiotensin II, bradykinin, methacholine, propranolol, vasopressin, adenosine, dopamine and histamine. Thirty min of preincubation of MDCK cells with 0.1 microM PMA completely blocked the activity of Na+, K+, Cl- cotransport whereas down-regulation of this enzyme by 24 h of preincubation with 1 microM PMA activated Na+, K+, Cl- cotransport by 60% and abolished the effect of short-term treatment with PMA. Regulation of Na+, K+, Cl- cotransport by ATP was insensitive to down-regulation of PMA-sensitive isoforms of protein kinase C. In addition, an inhibitor of protein kinase activity, staurosporine, abolished the effect of 0.1 microM PMA but did not change inhibition of this carrier by ATP. Thus, these results show for the first time that P2-purinoceptors and PMA-sensitive isoforms of protein kinase C play a key role in the regulation of Na+, K+, Cl- cotransport in MDCK cells. These results also show that neither PMA- nor staurosporine-sensitive forms of protein kinase are involved in the inhibition of Na+, K+, Cl- cotransport by activators of P2-purinoceptors.
 ...
PMID:Complete inhibition of Na+, K+, Cl- cotransport in Madin-Darby canine kidney cells by PMA-sensitive protein kinase. 951 30

Cyclic nucleotide-gated (CNG) channels are expressed in many cell types in both the nervous system and nonexcitable tissues. In order to understand the roles of cGMP-gated channels, and to distinguish actions of cGMP mediated through CNG channels from those through cGMP-dependent protein kinase (G-kinase), several new cGMP analogs were tested for potency as CNG channel agonists. Using Xenopus oocytes expressing the rat rod cGMP-gated ion channel alpha-subunit, we showed that an analog containing a pCPT group at the 8-position, 8-pCPT-cGMP, was 80 times more potent than cGMP and 14 times more potent than 8-Br-cGMP. 8-pCPT-cGMP is the most potent CNG channel agonist so far described and also has the advantages of much better membrane permeability as well as much higher resistance to PDE-hydrolysis, as compared with 8-Br-cGMP. Modification of both 8-Br-cGMP and 8-pCPT-cGMP by introduction of a sulphur atom into the cyclic phosphate group gave smaller changes in agonist efficiency. Both Sp-8-Br-cGMPS and Sp-8-pCPT-cGMPS acted as agonists of CNG channels and are also G-kinase activators. In contrast, Rp-8-Br-cGMPS was a channel agonist, with an EC50 of 173.5 microM, but a G-kinase antagonist with a Ki of 4 microM. Finally, Rp-8-pCPT-cGMPS was a channel agonist and showed additional noncompetitive antagonist activity at higher concentrations. The results suggest that 8-pCPT-cGMPS is a highly potent photoreceptor CNG channel agonist with high membrane permeability and PDE-resistance and furthermore Rp-8-Br-cGMPS can be used to test whether the actions of cGMP are selectively mediated by CNG channels.
 ...
PMID:Substituted cGMP analogs can act as selective agonists of the rod photoreceptor cGMP-gated cation channel. 958 70

The actions of vasoactive intestinal polypeptide (VIP) on catecholamine secretion and changes in [Ca2+]i in single rat chromaffin cells were studied using amperometry and Indo-1. Application of VIP prior to acetylcholine (ACh) or co-application of VIP and ACh enhanced secretion by 94% and 153% respectively, compared to ACh alone. [Ca2+]i was increased by 17% when VIP was preapplied and by 73% upon co-application. Exposure to VIP before stimulation with 60 mM K+ enhanced secretion by 68%, but not [Ca2+]i. VIP application prior to DMPP and nicotine had no effect on [Ca2+]i, but increased [Ca2+]i signals to muscarine by 18%. VIP co-application potentiated only [Ca2+]i responses to muscarine, by 28%. The effect of VIP on muscarine-induced [Ca2+]i signals was mimicked by 8-Br-cAMP, and both were blocked by H-89, a protein kinase A inhibitor. Long-lasting increases in secretion accompanied by a sustained rise in [Ca2+]i to VIP alone were seen in 55% of cells. Removal of Ca2+ or addition of La3+ inhibited both responses, while L-, N- and P-type Ca2+ channel blockers were ineffective. SK&F 96365 inhibited VIP-induced secretion completely and rises in [Ca2+]i by 75%. Neither 8-Br-cAMP nor 8-Br-cGMP evoked responses similar to VIP alone. Thus in rat chromaffin cells, VIP acts both directly as a neurotransmitter in provoking sustained catecholamine secretion in a cAMP-independent manner, and also by enhancing ACh-induced secretion, via a cAMP-dependent action involving muscarinic receptors.
 ...
PMID:Vasoactive intestinal peptide potentiates and directly stimulates catecholamine secretion from rat adrenal chromaffin cells. 979 63

Native large conductance, voltage-dependent, and Ca2+-sensitive K+ channels are activated by cGMP-dependent protein kinase. Two possible mechanisms of kinase action have been proposed: 1) direct phosphorylation of the channel and 2) indirect via PKG-dependent activation of a phosphatase. To scrutinize the first possibility, at the molecular level, we used the human pore-forming alpha-subunit of the Ca2+-sensitive K+ channel, Hslo, and the alpha-isoform of cGMP-dependent protein kinase I. In cell-attached patches of oocytes co-expressing the Hslo channel and the kinase, 8-Br-cGMP significantly increased the macroscopic currents. This increase in current was due to an increase in the channel voltage sensitivity by approximately 20 mV and was reversed by alkaline phosphatase treatment after patch excision. In inside-out patches, however, the effect of purified kinase was negative in 12 of 13 patches. In contrast, and consistent with the intact cell experiments, purified kinase applied to the cytoplasmic side of reconstituted channels increased their open probability. This stimulatory effect was absent when heat-denatured kinase was used. Biochemical experiments show that the purified kinase incorporates gamma-33P into the immunopurified Hslo band of approximately 125 kDa. Furthermore, in vivo phosphorylation largely attenuates this labeling in back-phosphorylation experiments. These results demonstrate that the alpha-subunit of large conductance Ca2+-sensitive K+ channels is substrate for G-Ialpha kinase in vivo and support direct phosphorylation as a mechanism for PKG-Ialpha-induced activation of maxi-K channels.
 ...
PMID:The large conductance, voltage-dependent, and calcium-sensitive K+ channel, Hslo, is a target of cGMP-dependent protein kinase phosphorylation in vivo. 983 46

Studies in vitro have underestimated the importance of cGMP-dependent protein kinase (PKG) in the modulation of vascular smooth muscle cell (SMC) proliferation and apoptosis in vivo. This is attributable, in part, to a rapid decline in PKG levels as vascular SMC are passaged in culture. We used a recombinant adenovirus encoding PKG (Ad.PKG) to augment kinase activity in cultured rat pulmonary artery SMC (RPaSMC). Incubation of Ad. PKG-infected RPaSMC (multiplicity of infection = 200) with 8-Br-cGMP decreased serum-stimulated DNA synthesis by 85% and cell proliferation at day 5 by 74%. The effect of 8-Br-cGMP on DNA synthesis in Ad.PKG-infected RPaSMC was blocked by KT5823 (PKG inhibitor), but not by KT5720 (cAMP-dependent protein kinase inhibitor). A nitric oxide (NO) donor compound, S-nitrosoglutathione, at concentrations as low as 100 nM, inhibited DNA synthesis in Ad. PKG-infected RPaSMC, but not in uninfected cells or in cells infected with a control adenovirus. In addition, 8-Br-cGMP and S-nitrosoglutathione induced apoptosis in serum-deprived RPaSMC infected with Ad.PKG, but not in uninfected cells or in cells infected with a control adenovirus. These results demonstrate that modulation of PKG levels in vascular SMC can alter the sensitivity of these cells to NO and cGMP. Moreover, these observations suggest an important role for PKG in the regulation of vascular SMC proliferation and apoptosis by NO and cGMP.
 ...
PMID:Adenovirus-mediated gene transfer of cGMP-dependent protein kinase increases the sensitivity of cultured vascular smooth muscle cells to the antiproliferative and pro-apoptotic effects of nitric oxide/cGMP. 985 90

We investigated the effects of cGMP-elevating agents, including atrial natriuretic peptide (ANP), C-type natriuretic peptide (CNP) and sodium nitroprusside (SNP), on cGMP accumulation and on carbachol (CCh)-stimulated intracellular calcium ([Ca2+]i) mobilisation in SV-40 transformed cat iris sphincter smooth muscle (SV-CISM-2) cells and in primary cultured cat iris sphincter smooth muscle (CISM) cells. The stimulatory effects of the natriuretic peptides on cGMP production correlated well with their inhibitory effects on CCh-induced [Ca+1]i mobilisation, and these effects were significantly more pronounced in the SV-CISM-2 cells than in the CISM cells. Thus, ANP (1 microM) increased cGMP production in the SV-CISM-2 cells and CISM cells by 487- and 1.7-fold, respectively, and inhibited CCh-induced [Ca2+]i mobilisation by 95 and 3%, respectively. In the SV-CISM-2 cells, ANP and CNP dose dependently inhibited CCh-induced [Ca2+]i mobilisation with IC50 values of 156 and 412 nM, respectively, and dose dependently stimulated cGMP formation with EC50 values of 24 and 88 nM, respectively, suggesting that the inhibitory actions of the peptides are mediated through cGMP. Both ANP and CNP stimulated cGMP accumulation in a time-dependent manner. The potency of the cGMP-elevating agents were in the following order: ANP>>CNP>>SNP; these agents had no effect on cAMP accumulation. The inhibitory effects of the natriuretic peptides were mimicked by 8-Br-cGMP, a selective activator of cGMP-dependent protein kinase. LY83583, a soluble guanylyl cyclase inhibitor, significantly inhibited SNP-induced cGMP formation but had no effect on those of ANP and CNP. The basal activities of the guanylyl cyclase and the dissociation constant (Kd) and total receptor density (Bmax) values of the natriuretic peptide receptor for [125I]ANP binding were not significantly different between the two cell types. The cGMP system, as with the cAMP system, has a major inhibitory influence on the muscarinic responses in the iris sphincter smooth muscle cells, and SV-CISM-2 cells can serve as an excellent model for investigating the cross talk between cGMP and the Ca2+ signalling system.
 ...
PMID:Atrial natriuretic peptide provokes a dramatic increase in cyclic GMP formation and markedly inhibits muscarinic-stimulated Ca2+ mobilisation in SV-40 transformed cat iris sphincter smooth muscle (SV-CISM-2) cells. 1004 85

Atrial natriuretic peptide (ANP) and nitric oxide (NO) are key regulators of ion and water transport in the kidney. Here, we report that these cGMP-elevating hormones stimulate Ca2+ reabsorption via a novel mechanism specifically involving type II cGMP-dependent protein kinase (cGK II). ANP and the NO donor, sodium nitroprusside (SNP), markedly increased Ca2+ uptake in freshly immunodissected rabbit connecting tubules (CNT) and cortical collecting ducts (CCD). Although readily increasing cGMP, ANP and SNP did not affect Ca2+ and Na+ reabsorption in primary cultures of these segments. Immunoblot analysis demonstrated that cGK II, and not cGK I, was present in freshly isolated CNT and CCD but underwent a complete down-regulation during the primary cell culture. However, upon adenoviral reexpression of cGK II in primary cultures, ANP, SNP, and 8-Br-cGMP readily increased Ca2+ reabsorption. In contrast, no cGMP-dependent effect on electrogenic Na+ transport was observed. The membrane localization of cGK II proved to be crucial for its action, because a nonmyristoylated cGK II mutant that was shown to be localized in the cytosol failed to mediate ANP-stimulated Ca2+ transport. The Ca2+-regulatory function of cGK II appeared isotype-specific because no cGMP-mediated increase in Ca2+ transport was observed after expression of the cytosolic cGK Ibeta or a membrane-bound cGK II/Ibeta chimer. These results demonstrate that ANP- and NO-stimulated Ca2+ reabsorption requires membrane-targeted cGK II.
 ...
PMID:Atrial natriuretic peptide-stimulated Ca2+ reabsorption in rabbit kidney requires membrane-targeted, cGMP-dependent protein kinase type II. 1033 45

Melatonin, secreted nocturnally by the pineal gland, can bind to human benign prostate epithelial cells and attenuate their growth and viability. In the present study, melatonin binding and responses were explored in the human steroid-independent PC3 prostatic tumor cells. PC3 cells bound 125I-melatonin with low affinity (Kd ca. 0.9 nM) at high as well as low cell density. Melatonin enhanced cGMP and 3H-thymidine incorporation at low, but attenuated them at high cell density. In addition, melatonin inhibited cAMP at low, but augmented it at high cell density. These effects were associated with an increase in cell count at low- but not high-density cultures. Pertussis toxin treatment suppressed 125I-melatonin binding and ablated all the effects of melatonin on 3H-thymidine incorporation, cAMP, and cGMP at both cell densities. Cholera toxin treatment failed to block the effects of melatonin on 3H-thymidine incorporation, but prevented the modulation by melatonin of cAMP at low and cGMP at high cell density. The cGMP analog 8-Br-cGMP, inhibited melatonin's effects on 3H-thymidine incorporation at both cell densities. H89, a protein kinase A inhibitor, prevented melatonin's effects on 3H-thymidine incorporation at low but not high cell density. These results provide the first demonstration of direct interaction of melatonin with hormone-insensitive prostate tumor cells. The melatonin receptors in the PC3 cells are coupled to pertussis toxin-sensitive G proteins to induce cell density-dependent changes in cGMP, cAMP, and cell growth.
 ...
PMID:Melatonin receptors in PC3 human prostate tumor cells. 1034 Jul 23


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>