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:2.7.11.1 (protein kinase)
81,284 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Astroglial cells synthesize and release endozepines, neuropeptides that are related to the octadecaneuropeptide ODN. Glial cells also express PACAP/VIP receptors. We have investigated the possible effect of PACAP on the release of ODN-like immunoreactivity (ODN-LI) by cultured rat astrocytes. Administration of PACAP27 and PACAP38 induced a concentration-dependent increase in secretion of ODN-LI whereas VIP was approximately 1000-fold less potent. The maximum effect of PACAP38 occurred after 5 min, then gradually declined during the next 10 min. The stimulatory effects of PACAP and VIP were abrogated by the PACAP antagonist PACAP6-38. PACAP38 stimulated cAMP formation, activated polyphosphoinositide turnover, and provoked calcium mobilization from IP3-sensitive pools. The PKA inhibitor H89 suppressed PACAP-induced secretion of ODN-LI, whereas PLC inhibitor U73122 and the PKC inhibitor chelerythrine had no effect. In contrast, U73122 restored the stimulatory action of PACAP on ODN-LI release and cAMP formation during prolonged (15 min) incubation with the peptide, and this effect was prevented by PMA. The present results demonstrate that PACAP stimulates endozepine release through activation of PAC1 receptors coupled to the AC/PKA pathway. Our data also show that activation of the PLC/PKC pathway down-regulates the effect of PACAP on endozepine release.
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PMID:Pituitary adenylate cyclase-activating polypeptide (PACAP) stimulates endozepine release from cultured rat astrocytes via a PKA-dependent mechanism. 1252 8

1. Optimal dipeptide and peptidomimetic drug transport across the intestinal mucosal surface is dependent upon the co-operative functional activity of the di/tripeptide transporter hPepT1 and the Na(+)/H(+) exchanger NHE3. The ability of the anti-absorptive enteric neuropeptide VIP (vasoactive intestinal peptide) to modulate dipeptide uptake was determined using human intestinal (Caco-2) epithelial cell monolayers. 2. Uptake of glycylsarcosine (Gly-Sar) across the apical membrane of Caco-2 cell monolayers is inhibited by basolateral exposure to either VIP, pituitary adenylate cyclase-activating polypeptide (PACAP), or the VPAC(1) receptor agonist [(11,22,28)Ala]-VIP. Inhibition of Gly-Sar uptake is observed only in the presence of extracellular Na(+). Reverse-transcription polymerase chain reaction (RT-PCR) demonstrates that VPAC(1) mRNA is expressed in Caco-2 cells whereas VPAC(2) mRNA is not detected. 3. The VIP-induced inhibition of Gly-Sar uptake is abolished in the presence of the protein kinase A (PKA) inhibitor H-89 (N-[2-(p-bromocinnamylamino)ethyl]-5-isoquinolinesulfonamide.2HCl). 4. (22)Na(+) uptake across the apical membrane is inhibited by the selective NHE3 inhibitor S1611. Experiments with BCECF [2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein]-loaded Caco-2 cells demonstrate that VIP reduces the NHE3-dependent recovery of intracellular pH (pH(i)) after dipeptide-induced acidification. Western blot of Caco-2 cell protein demonstrates expression of the NHE regulatory factor NHERF1 (expression of which is thought to be required for PKA-mediated inhibition of NHE3). 5. VIP has no effect on Gly-Sar uptake in the presence of S1611 suggesting that VIP and S1611 both modulate dipeptide uptake via the same mechanism. 6. These observations demonstrate that VIP (and PACAP) modulate activity of the H(+)/dipeptide transporter hPepT1 in a Na(+)-dependent manner consistent with the modulation being indirect through inhibition of NHE3.
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PMID:Inhibition of intestinal dipeptide transport by the neuropeptide VIP is an anti-absorptive effect via the VPAC1 receptor in a human enterocyte-like cell line (Caco-2). 1259 10

The sphingolipid metabolites, ceramides, are critical mediators of the cellular stress response and play an important role in the control of programmed cell death. In particular, ceramides have been shown to induce apoptosis of cerebellar granule cells. We show that pituitary adenylate cyclase-activating polypeptide (PACAP) prevents C2-ceramide-induced apoptosis. The neuroprotective effect of PACAP was dose-dependent and blocked by its antagonist, PACAP6-38, whereas the PACAP-related peptide VIP was inactive. The effect of PACAP on cell survival was mimicked by dibutyryl-cAMP (dbcAMP) and forskolin and prevented by the MEK inhibitor U0126, indicating that both the adenylyl-cyclase and MAP-kinase pathways contribute to the neuroprotective action of the peptide. C2-ceramide and PACAP induced opposite effects on phosphorylated forms of ERK and JNK without affecting the total amounts of ERK and JNK, suggesting that a balance between these two MAP-kinases is critical for the cell survival/death decision. The effect of PACAP on ERK phosphorylation was blocked by U0126, but was not affected by H89 or chelerythrine indicating that PACAP activates ERK through a PKA- and PKC-independent mechanism. C2-ceramide induced a time-dependent activation of caspase-3, enhanced the amount of cleaved caspase-3 and stimulated the DNA fragmentation process, while PACAP strongly inhibited the C2-ceramide-induced activation of caspase-3, reduced the expression of cleaved caspase-3 and blocked DNA fragmentation. Taken together, the present results show that C2-ceramide induces apoptosis of cerebellar granule cells through a mechanism involving activation of caspase-3. Our data also demonstrate that PACAP is a potent inhibitor of C2-ceramide-induced apoptosis.
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PMID:Pituitary adenylate cyclase-activating polypeptide prevents C2-ceramide-induced apoptosis of cerebellar granule cells. 1269 97

The role of RhoA in myosin light-chain (MLC)(20) dephosphorylation and smooth muscle relaxation by PKA and PKG was examined in freshly dispersed and cultured smooth muscle cells expressing wild-type RhoA, constitutively active Rho(V14), and phosphorylation site-deficient Rho(A188). Activators of PKA (5,6-dichloro-1-beta-ribofuranosyl benzimidazole 3',5'-cyclic monophosphothionate, Sp-isomer; cBIMPS) or PKG [8-(4-chlorophenylthio)guanosine 3',5'-cyclic monophosphate (8-pCPT-cGMP), sodium nitroprusside (SNP)] or both PKA and PKG (VIP) induced phosphorylation of constitutively active Rho(V14) and agonist (ACh)- or GTPgammaS-stimulated wild-type RhoA but not Rho(A188). Phosphorylation was accompanied by translocation of membrane-bound wild-type RhoA and Rho(V14) to the cytosol and complete inhibition of ACh-stimulated Rho kinase and phospholipase D activities, RhoA/Rho kinase association, MLC(20) phosphorylation, and sustained muscle contraction. Each of these events was blocked depending on the agent used, by the PKG inhibitor KT5823 or the PKA inhibitor myristoylated PKI. Inhibitors were used at a concentration (1 microM) previously shown by direct measurement of kinase activity to selectively inhibit the corresponding kinase. In muscle cells overexpressing the active phosphorylation site-deficient mutant Rho(A188), MLC(20) phosphorylation was partly inhibited by SNP, VIP, cBIMPS, and 8-pCPT-cGMP, suggesting the existence of an independent inhibitory mechanism downstream of RhoA. Results demonstrate that dephosphorylation of MLC(20) and smooth muscle relaxation are preferentially mediated by PKG- and PKA-dependent phosphorylation and inactivation of RhoA.
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PMID:Inhibition of sustained smooth muscle contraction by PKA and PKG preferentially mediated by phosphorylation of RhoA. 1273 49

Circadian rhythmicity in mammals is generated by a pair of nuclei in the anterior hypothalamus known as the suprachiasmatic nuclei (SCN), whose neurons express a variety of neuropeptides that are thought to play an important role in the circadian timing system. To evaluate the influence of VIP on inhibitory synaptic transmission between SCN neurons, we used whole cell patch-clamp recording in an acute brain slice preparation of mouse SCN. Baseline spontaneous GABAergic inhibitory postsynaptic currents (IPSCs) varied significantly between regions and across phases, with a greater frequency of IPSCs observed in the dorsomedial region during the early night. Bath-applied VIP caused a significant increase in the frequency of spontaneous inhibitory postsynaptic currents (sIPSC) in a reversible and dose-dependent manner with no effect on the mean amplitude or kinetic parameters. The effect of VIP was widespread throughout the SCN and observed in both ventrolateral (VL) and dorsomedial (DM) regions. In the presence of tetrodotoxin, VIP increased the frequency of miniature IPSCs without affecting the mean magnitude or kinetic parameters. The magnitude of the enhancement by VIP was significantly larger during the day than during the night. Pretreatment with the VIP-PACAP receptor antagonist [Ac-Tyr1, D-Phe2]-GHRF 1-29 or the selective VPAC2 receptor antagonist PG 99-465 completely blocked the VIP-induced enhancement. The effect of VIP appears to be mediated by a cAMP/PKA-dependent mechanism as forskolin mimics, while the PKA antagonist H-89 blocks the observed enhancement of GABA currents. Our data suggest that VIP activates presynaptic VPAC2 receptors to regulate inhibitory synaptic transmission within the SCN and that this effect varies from day to night.
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PMID:Regulation of inhibitory synaptic transmission by vasoactive intestinal peptide (VIP) in the mouse suprachiasmatic nucleus. 1296 76

Novel neurotrophin-1/B cell stimulating factor-3 (NNT-1/BSF-3) is a gp130 cytokine potently stimulating corticotroph proopiomelanocortin gene expression and ACTH secretion by a Janus kinase-signal transducer and activator of transcription (JAK-STAT)-dependent mechanism. In the current study, we examined the regulation of NNT-1/BSF-3 mRNA expression in murine pituitary folliculostellate TtT/GF cells using Northern blot technique. A 5- to 9-fold and a 4- to 7-fold induction in NNT-1/BSF-3 mRNA expression was observed between 2 and 6 h stimulation with the protein kinase C (PKC) stimulus phorbol-12-myristate-13-acetate (100 nm) and the protein kinase A (PKA) stimulus Bu(2)cAMP (5 mm), respectively. Pituitary adenylate cyclase-activating polypeptide (PACAP-38, 50 nm) and vasoactive intestinal peptide (VIP, 50 nm) also stimulated NNT-1/BSF-3 mRNA expression 5- to 9-fold between 2 and 6 h. Preincubation with PKC and PKA inhibitors such as H-7 (20 microm), GF109203X (50 microm), and H-89 (50 microm) decreased the stimulatory effects of PACAP and VIP. Both PACAP-38 and VIP also rapidly induced ERK1/2 phosphorylation and their stimulatory effect on NNT-1/BSF-3 mRNA expression was reduced by the MAPK kinase/ERK kinase (MEK) inhibitor U0126 (10 microm). Dexamethasone (10(-7) m) was a potent inhibitor of phorbol-12-myristate-13-acetate-induced NNT-1/BSF-3 expression. RT-PCR analysis demonstrated TtT/GF cells to express the short and the hop variant but not the hip variant of the PACAP-1 receptor (PAC1-R). In addition, TtT/GF cells express the VIP/PACAP-2 receptor (VPAC2-R). In summary, NNT-1/BSF-3 is expressed in pituitary folliculostellate TtT/GF cells and induced by PKC-, PKA-, and ERK1/2-dependent mechanisms. The novel gp130 cytokine NNT-1/BSF-3 derived from folliculostellate cells might act as a paracrine neuroimmunoendocrine modulator of pituitary corticotroph function.
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PMID:Expression of novel neurotrophin-1/B-cell stimulating factor-3 (NNT-1/BSF-3) in murine pituitary folliculostellate TtT/GF cells: pituitary adenylate cyclase-activating polypeptide and vasoactive intestinal peptide-induced stimulation of NNT-1/BSF-3 is mediated by protein kinase A, protein kinase C, and extracellular-signal-regulated kinase1/2 pathways. 1460 1

We have previously shown that Gq protein-coupled receptor (GqPCR) agonists stimulate epidermal growth factor receptor (EGFr) transactivation and activation of mitogen-activated protein kinases (MAPK) in colonic epithelial cells. This constitutes a mechanism by which Cl- secretory responses to GqPCR agonists are limited. In the present study we examined a possible role for the EGFr in regulating Cl- secretion stimulated by agonists that act through GsPCRs. All experiments were performed using monolayers of T84 colonic epithelial cells grown on permeable supports. Protein phosphorylation and protein-protein interactions were analyzed by immunoprecipitation and Western blotting. Cl- secretion was measured as changes in short-circuit current (DeltaIsc) across voltage-clamped T84 cells. The GsPCR agonist, vasoactive intestinal polypeptide (VIP; 100 nM), rapidly stimulated EGFr phosphorylation in T84 cells. This effect was mimicked by a cell-permeant analog of cAMP, Bt2cAMP/AM (3 microM), and was attenuated by the protein kinase A (PKA) inhibitor, H-89 (20 microM). The EGFr inhibitor, tyrphostin AG1478 (1 microM), inhibited both Bt2cAMP/AM-stimulated EGFr phosphorylation and Isc responses. VIP and Bt2cAMP/AM both stimulated ERK MAPK phosphorylation and recruitment of the p85 subunit of phosphatidylinositol 3-kinase (PI3K) to the EGFr in a tyrphostin AG1478-sensitive manner. The PI3K inhibitor, wortmannin (50 nM), but not the ERK inhibitor, PD 98059 (20 microM), attenuated Bt2cAMP/AM-stimulated secretory responses. We conclude that GsPCR agonists rapidly transactivate the EGFr in T84 cells by a signaling pathway involving cAMP and PKA. Through a mechanism that likely involves PI3K, transactivation of the EGFr is required for the full expression of cAMP-dependent Cl- secretory responses.
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PMID:Gs protein-coupled receptor agonists induce transactivation of the epidermal growth factor receptor in T84 cells: implications for epithelial secretory responses. 1466 Jun 4

1. In the human airway epithelium, VIP/PACAP receptors are distributed in nerve fibers and in epithelial cells but their role in transepithelial ion transport have not been reported. Here, we show that human bronchial epithelial Calu-3 cells expressed the VPAC(1) receptor subtype which shares similar high affinity for VIP and PACAP-27. 2. The stoichiometric binding parameters characterizing the (125)I-VIP and (125)I-PACAP-27 binding to these receptors were determined. 3. We found that VIP (EC(50) approximately 7.6 nM) and PACAP-27 (EC(50) approximately 10 nM) stimulated glibenclamide-sensitive and DIDS-insensitive iodide efflux in Calu-3 cells. 4. The protein kinase A (PKA) inhibitor, H-89 and the protein kinase C (PKC) inhibitor, chelerythrine chloride prevented activation by both peptides demonstrating that PKA and PKC are part of the signaling pathway. This profile corresponds to the pharmacological signature of CFTR. 5. In the cystic fibrosis airway epithelial IB3-1 cell lacking functional CFTR but expressing VPAC(1) receptors, neither VIP, PACAP-27 nor forskolin stimulated chloride transport. 6. Ussing chamber experiments demonstrated stimulation of CFTR-dependent short-circuit currents by VIP or PACAP-27 applied to the basolateral but not to the apical side of Calu-3 cells monolayers. 7. This study shows the stimulation in human bronchial epithelial cells of CFTR-dependent chloride secretion following activation by VIP and PACAP-27 of basolateral VPAC(1) receptors.
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PMID:Activation of VPAC1 receptors by VIP and PACAP-27 in human bronchial epithelial cells induces CFTR-dependent chloride secretion. 1474 18

Pituitary adenylate cyclase activating polypeptides (PACAP) and PAC1 receptor signaling have diverse roles in central and peripheral nervous system development and function. In recent microarray analyses for PACAP and PAC1 receptor modulation of neuronal transcripts, the mRNA of Homer 1a (H1a), which encodes the noncrosslinking and immediate early gene product isoform of Homer, was identified to be strongly upregulated in superior cervical ganglion (SCG) sympathetic neurons. Given the prominent roles of Homer in synaptogenesis, synaptic protein complex assembly and receptor/channel signaling, we have examined the ability for PACAP to induce H1a expression in sympathetic, cortical and hippocampal neurons to evaluate more comprehensively the roles of PACAP in synaptic function. In both central and peripheral neuronal cultures, PACAP peptides increased transiently H1a transcript levels approximately 3.5- to 6-fold. From real-time quantitative PCR measurements, the temporal patterns of PACAP-mediated H1a mRNA induction among the different neuronal cultures appeared similar although the onset of sympathetic H1a transcript expression appeared protracted. The increase in H1a transcripts was accompanied by increases in H1a protein levels. Comparative studies with VIP and PACAP(6-38) antagonist demonstrated that the PACAP effects reflected PAC1 receptor activation and signaling. The PAC1 receptor isoforms expressed in central and peripheral neurons can engage diverse intracellular second messenger systems, and studies using selective signaling pathway inhibitors demonstrated that the cyclic AMP/PKA and MEK/ERK cascades are principal mediators of the PACAP-mediated H1a induction response. In modulating H1a transcript and protein expression, these studies may implicate broad roles for PACAP and PAC1 receptor signaling in synaptic development and plasticity.
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PMID:Pituitary adenylate cyclase activating polypeptide and PAC1 receptor signaling increase Homer 1a expression in central and peripheral neurons. 1551

A H(+)-coupled amino acid transporter has been characterised functionally at the brush border membrane of the human intestinal cell line Caco-2. This carrier, hPAT1 (human Proton-coupled Amino acid Transporter 1) or SLC36A1, has been identified recently at the molecular level and hPAT1 protein is localised to the brush border membrane of human small intestine. hPAT1 transports both amino acids (e.g., beta-alanine) and therapeutic agents (e.g., D-cycloserine). In human Caco-2 cells, hPAT1 function (H(+)/amino acid symport) is associated with a decrease in intracellular pH (pH(i)), which selectively activates the Na(+)/H(+) exchanger NHE3, and thus maintains pH(i) and the driving force for hPAT1 function (the H(+) electrochemical gradient). This study provides the first evidence for regulation of hPAT1 function. Activation of the cAMP/protein kinase A pathway in Caco-2 cell monolayers either using pharmacological tools (forskolin, 8-br-cAMP, [(11,22,28)Ala]VIP) or physiological activators (the neuropeptides VIP and PACAP) inhibited hPAT1 function (beta-alanine uptake) at the apical membrane. Under conditions where NHE3 is inactive (the absence of Na(+), apical pH 5.5, the presence of the NHE3 inhibitor S1611) no regulation of beta-alanine uptake is observed. Forskolin and VIP inhibit pH(i) recovery (NHE3 function) from beta-alanine-induced intracellular acidification. Immunocytochemistry localises NHERF1 (NHE3 regulatory factor 1) to the apical portion of Caco-2 cells where it will interact with NHE3 and allow PKA-mediated phosphorylation of NHE3. In conclusion, we have shown that amino acid uptake via hPAT1 is inhibited by activators of the cAMP pathway indirectly through inhibition of NHE3 activity.
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PMID:Indirect regulation of the intestinal H+-coupled amino acid transporter hPAT1 (SLC36A1). 1575 24


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