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:3.1.4.1 (phosphodiesterase)
18,767 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Mammalian phosphodiesterase types 3 and 4 (PDE3 and PDE4) hydrolyze cAMP and are essential for the regulation of this intracellular second messenger in many cell types. Whereas these enzymes share structural and biochemical similarities, each can be distinguished by its sensitivity to isozyme-specific inhibitors. By using a series of chimeric enzymes, we have localized the region of PDE4 that confers sensitivity to selective inhibitors. This inhibitor specificity domain lies within a short sequence at the carboxyl terminus of the catalytic domain of the protein, consistent with the competitive nature of inhibition by these compounds. Surprisingly, the identified region also includes some of the most highly conserved residues among PDE isoforms. A yeast-based expression system was used for the isolation and characterization of mutations within this area that confer resistance to the PDE4-specific inhibitor rolipram. Analysis of these mutants indicated that both conserved and unique residues are required for isoform-specific inhibitor sensitivity. In some cases, combined point mutations contribute synergistically to the reduction of sensitivity (suppression of IC50). We also report that several mutations display differential sensitivity changes with respect to distinct structural classes of inhibitors.
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PMID:Identification of inhibitor specificity determinants in a mammalian phosphodiesterase. 998 24

The aim of this study was to investigate the effects of IL-10, a cell permeable analogue of cyclic AMP, dibutyryl-cAMP (db-cAMP), modulators of intracellular cyclic AMP such as phosphodiesterase (PDE) inhibitors and a beta 2-adrenoceptor agonist, salmeterol, on pulmonary inflammation following acute lung injury induced by endotoxin exposure in rats. Pulmonary inflammation was induced in adult Wistar rats by a 60-min exposure to endotoxin (lipopolysaccharide, LPS, 100 micrograms/mL). 4 h later bronchoalveolar lavage (BAL) was performed. The PDE inhibitors, rolipram (3 and 5 mg/kg) and theophylline (30 and 100 mg/kg) inhibited neutrophil recruitment, TNF-alpha release and cellular activation in BAL. Salmeterol (0.5 mg/mL) and IL-10 (0.1 microgram) only inhibit TNF-alpha increase in the BAL fluid and db-AMPc (2.5 micrograms/rat) was ineffective. The present data show that the selective PDE4 inhibitor, rolipram, and the non-selective PDE inhibitor, theophylline, markedly reduced the pulmonary inflammation associated with acute lung injury in the rat. These effects may be mediated in part by IL-10 rather than by cyclic AMP, as demonstrated by the potent inhibitory activity of exogenous IL-10 on the increase in TNF-alpha release in BAL fluid of rats exposed to LPS.
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PMID:Effects of interleukin-10 and modulators of cyclic AMP formation on endotoxin-induced inflammation in rat lung. 1002 94

We have previously shown that mitogenic activation of human PBMC rapidly increases both the intracellular phosphatidic acid (PA) level and cyclic nucleotide phosphodiesterase (PDE) activity, with time-course responses, suggesting a causative relationship between the two events. PA also directly stimulated cAMP-PDE activity in acellular systems. Thus the mitogenic properties of PA night be due to its ability to lower the level of cAMP, a negative effector of lymphocyte activation, through PDE activation. In this study, human PBMC were stimulated either with the mitogenic lectin ConA, the anti-CD3 mAb OKT3, or the phorbol ester TPA. All three agonists increased the radiolabeled PA level and the PA mass in treated cells and simultaneously increased cytosolic and particulate cAMP- and cGMP-PDE activities, with significant positive correlations between PA accumulation and PDE activities. Furthermore, the ConA-induced PDE activation was dose-dependently reduced by treatment of PBMC with the diacylglycerol-kinase inhibitor R59022. This compound also dose-dependently lowered the PA level and inhibited the proliferative response to ConA. In addition, TPA-induced PDE activation was totally abolished by ethanol, which strongly reduced PA accumulation in response to the phorbol ester. These data suggest that PA increase may be linked to mitogen-induced PDE activation. Experiments performed in the presence of rolipram indicated that ConA and TPA stimulated both the rolipram-sensitive PDE4 and the rolipram-insensitive PDE activities, OKT3 being more active on PDE4. All three agonists stimulated the cGMP-specific PDE5. These results suggest that PA is an important component of the mechanisms that maintain a low level of cyclic nucleotides, which is a prerequisite for an optimal lymphoproliferative response.
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PMID:Relationships between phosphatidic acid and cyclic nucleotide phosphodiesterases in activated human blood mononuclear cells. 1008 May 43

The increase in cellular cAMP concentration during 10-min incubation of rat hepatocytes with glucagon or forskolin was enhanced markedly when the hepatocytes had been cultured for several hours with herbimycin A. This effect of herbimycin was accompanied by inhibition of tyrosine-phosphorylation of cellular proteins including alpha-tubulin, antagonized by coaddition of Na3VO4 plus H2O2, which also antagonized the herbimycin-induced tyrosine phosphorylation, and overcome by the addition to the 10-min incubation medium of a certain inhibitor of cAMP phosphodiesterase (PDE), which caused a huge accumulation of cAMP. The effective PDE inhibitors were 4-[3-(cyclopentyloxy)-4-methoxyphenyl]-2-pyrrolidinone (rolipram) and 4-(3-butyloxy-4-methoxyphenyl)-2-imidazolidinone (Ro-20-1724, a PDE4 inhibitor), in addition to 3-isobutyl-1-methylxanthine (a nonselective inhibitor). Rapid breakdown of the once-accumulated cAMP in cultured hepatocytes during the subsequent incubation without PDE inhibitors was progressively prevented when the concentration of herbimycin was increased from 0.3 to 10 microM during prior culture. This effect of herbimycin to inhibit PDE activity in intact cells was abolished by coaddition of a microtubule-disrupting agent, either colchicine or vinblastine, into the culture, but remained unchanged if the vinblastine-containing medium was further supplemented with taxol, a microtubule-stabilizing agent, which by itself mimicked the effect of herbimycin. None of these agents, which thus affected PDE activity in intact cells, inhibited the PDE activity assayable in the cell lysates. The taxol-like and vinblastine-suppressible action of herbimycin to stimulate microtubular assembly was antagonized by Na3VO4/H2O2, as confirmed by confocal microscopic images of the cells stained with fluorescein-bound anti-(alpha-tubulin). Thus, 4-h culture of hepatocytes with herbimycin inhibits phosphorylation of the C-terminal tyrosine residue of alpha-tubulin, thereby stimulating formation of a microtubular network which is responsible for the inhibition of PDE4 in the intact cells by an unknown mechanism.
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PMID:Decreases in cAMP phosphodiesterase activity in hepatocytes cultured with herbimycin A due to cellular microtubule polymerization related to inhibition of tyrosine phosphorylation of alpha-tubulin. 1009 74

In light of the important role of the second messengers cAMP and cGMP in the mechanism of relaxation in the human myometrium, specific regulation of the phosphodiesterase (PDE) enzymatic system responsible for cyclic nucleotide inactivation is essential. We previously identified in the human myometrium PDE4 cAMP-specific PDE as by far the most abundant isoform. Here we have studied the expression patterns of mRNAs for the four cloned human PDE4 genes in the myometria of pregnant and non-pregnant women. Concurrent expression of the PDE4A, 4B, 4C and 4D genes is demonstrated. We found that the PDE4D transcripts are the most prominently expressed. PDE4A and PDE4B mRNAs also are markedly abundant, whereas lower expression is observed for PDE4C mRNAs. Interestingly, we showed that transcripts of PDE4B2 are more abundant in the myometria of pregnant women than in non-pregnant women, whereas no difference between the two tissues was detected for PDE4A, 4C and 4D mRNAs. Cultured human myometrial cells, which present a high level of PDE4 activity and express the four PDE4 mRNA subtypes, provide us with an appropriate model to further evaluate whether the level of expression of the PDE 4B2 mRNA subtype is under hormonal regulation.
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PMID:Effect of pregnancy on PDE4 cAMP-specific phosphodiesterase messenger ribonucleic acid expression in human myometrium. 1020 42

The cAMP-specific phosphodiesterase (PDE) HSPDE 4A4B(pde46) selectively bound SH3 domains of SRC family tyrosyl kinases. Such an interaction profoundly changed the inhibition of PDE4 activity caused by the PDE4-selective inhibitor rolipram and mimicked the enhanced rolipram inhibition seen for particulate, compared with cytosolic pde46 expressed in COS7 cells. Particulate pde46 co-localized with LYN kinase in COS7 cells. The unique N-terminal and LR2 regions of pde46 contained the sites for SH3 binding. Altered rolipram inhibition was triggered by SH3 domain interaction with the LR2 region. Purified LYN SH3 and human PDE4A LR2 could be co-immunoprecipitated, indicating a direct interaction. Protein kinase A-phosphorylated pde46 remained able to bind LYN SH3. pde46 was found to be associated with SRC kinase in the cytosol of COS1 cells, leading to aberrant kinetics of rolipram inhibition. It is suggested that pde46 may be associated with SRC family tyrosyl kinases in intact cells and that the ensuing SH3 domain interaction with the LR2 region of pde46 alters the conformation of the PDE catalytic unit, as detected by altered rolipram inhibition. Interaction between pde46 and SRC family tyrosyl kinases highlights a potentially novel regulatory system and point of signaling system cross-talk.
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PMID:Association with the SRC family tyrosyl kinase LYN triggers a conformational change in the catalytic region of human cAMP-specific phosphodiesterase HSPDE4A4B. Consequences for rolipram inhibition. 1020 97

The WD-repeat protein receptor for activated C-kinase (RACK1) was identified by its interaction with the cyclic AMP-specific phosphodiesterase (PDE4) isoform PDE4D5 in a yeast two-hybrid screen. The interaction was confirmed by co-immunoprecipitation of native RACK1 and PDE4D5 from COS7, HEK293, 3T3-F442A, and SK-N-SH cell lines. The interaction was unaffected by stimulation of the cells with the phorbol ester phorbol 2-myristate 3-acetate. PDE4D5 did not interact with two other WD-repeat proteins, beta'-coatomer protein and Gsbeta, in two-hybrid tests. RACK1 did not interact with other PDE4D isoforms or with known PDE4A, PDE4B, and PDE4C isoforms. PDE4D5 and RACK1 interacted with high affinity (Ka approximately 7 nM) [corrected] when they were expressed and purified from Escherichia coli, demonstrating that the interaction does not require intermediate proteins. The binding of the E. coli-expressed proteins did not alter the kinetics of cAMP hydrolysis by PDE4D5 but caused a 3-4-fold change in its sensitivity to inhibition by the PDE4 selective inhibitor rolipram. The subcellular distributions of RACK1 and PDE4D5 were extremely similar, with the major amount of both proteins (70%) in the high speed supernatant (S2) fraction. Analysis of constructs with specific deletions or single amino acid mutations in PDE4D5 demonstrated that a small cluster of amino acids in the unique amino-terminal region of PDE4D5 was necessary for its interaction with RACK1. We suggest that RACK1 may act as a scaffold protein to recruit PDE4D5 and other proteins into a signaling complex.
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PMID:The RACK1 signaling scaffold protein selectively interacts with the cAMP-specific phosphodiesterase PDE4D5 isoform. 1032 91

The effects of family selective inhibitors of phosphodiesterase (PDEI, PDE2, PDE3, PDE4, and PDE5) on the behavior of rats under either a differential-reinforcement-of-low-rate (DRL) 72-s schedule or a variable-interval (VI) 30-s schedule were determined; previous work has shown that antidepressant drugs increase reinforcement rate under long DRL schedules. The PDE4-selective inhibitor rolipram (0.03-0.1 mg/kg) reduced response rate and increased reinforcement rate under the DRL schedule in a dose-dependent manner; similar effects were observed with the tricyclic antidepressant drug desipramine (3-10 mg/kg). Both of these drugs produced biphasic effects on behavior maintained under the VI schedule, increasing response rate at the lower doses tested (rolipram: 0.003 mg/kg; desipramine: 0.03 mg/kg) and decreasing response rate at higher doses (rolipram: 0.1 mg/kg; desipramine: 0.3-18 mg/kg). Of the other PDE inhibitors tested, only the PDE5-selective inhibitor zaprinast (10 mg/kg) produced an antidepressant-like effect on DRL behavior. However, in contrast to the biphasic effects of rolipram and desipramine on VI behavior, zaprinast produced monotonic decreases in response rate (10-30 mg/kg). The PDE2-selective inhibitor trequinsin produced biphasic effects on response rate under the VI schedule, increasing rates at low doses (3-5.6 mg/kg) and decreasing rates at higher doses (18-30 mg/kg). Trequinsin also reduced response rate under the DRL schedule (30 mg/kg); however, the reduction in response rate was not accompanied by increased reinforcement rate. The PDE3-selective inhibitor milrinone (1-10 mg/kg) tended to increase response rates under both schedules while the PDE1-selective inhibitor vinpocetine did not affect behavior at the dose range tested (1-30 mg/kg). These findings suggest that inhibition of PDE4 results in a rather unique pattern of behavioral effects, most notably an antidepressant-like effect on DRL behavior. It remains to be determined if a similar effect produced by zaprinast also implicates PDE5 in the mediation of antidepressant activity or represents an effect of this drug on PDE4 activity at high doses.
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PMID:Behavioral effects of family-selective inhibitors of cyclic nucleotide phosphodiesterases. 1034 May 40

We described the development of a recombinant cell-based system for the characterisation of phosphodiesterase (PDE) 4 isoforms and the evaluation of inhibitors. The Chinese hamster ovary (CHO) cell, which was found to have a low endogenous PDE4 background and no beta-adrenergic receptors (beta-AR), was transiently transfected with beta-AR and various PDE4 isoforms which were expressed as functionally coupled molecules. From correlations of elevation of adenosine 3',5'-cyclic monophosphate in situ and the inhibition of catalytic activity in vitro with the various PDE4 isoforms, it was apparent that PDE4A4, 4B2, 4C2, 4D2, and 4D3 all adopted a high-affinity binding conformation (i.e. expressed the high-affinity rolipram binding site) in the CHO cell, whereas PDE4A330 was expressed in a low-affinity conformation in situ. This gives the opportunity of using this system to screen and optimise inhibitors against a low-affinity conformation of PDE4 in situ and use a high-affinity conformation of PDE4 as a counterscreen, as inhibitor activity against this conformer has been linked with undesirable side effects. This system could also be utilised to screen inhibitors against various PDE4 isoforms in isolation against a low endogenous PDE background in situ for isoform-selective inhibitors.
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PMID:Development of a recombinant cell-based system for the characterisation of phosphodiesterase 4 isoforms and evaluation of inhibitors. 1035 58

The phosphodiesterase activity in the HT4.7 neural cell line was pharmacologically characterized, and phosphodiesterase isozyme 4 (PDE4) was found to be the predominant isozyme. The Km for cAMP was 1-2 microM, indicative of a "low Km" phosphodiesterase, and the activity was inhibited by PDE4-selective inhibitors rolipram and Ro20-1724, but not PDE3- or PDE2-selective inhibitors. Calcium, calmodulin, and cGMP, regulators of PDE1, PDE2, and PDE3, had no effect on cAMP hydrolysis. The protein tyrosine kinase inhibitor, genistein, inhibited HT4.7 cAMP phosphodiesterase activity by 85-95% with an IC50 of 4 microM; whereas daidzein, an inactive structural analog of genistein, had little effect on phosphodiesterase activity. This is a common pharmacological criterion used to implicate the regulation by a tyrosine kinase. However, genistein still inhibited phosphodiesterase activity with a mixed pattern of inhibition even when ion-exchange chromatography was used to partially purify phosphodiesterase away from the tyrosine kinase activity. Moreover, tyrphostin 51, another tyrosine kinase inhibitor, was found to also inhibit partially purified phosphodiesterase activity noncompetitively. These data suggest that HT4.7 phosphodiesterase activity is dominated by PDE4 and can be regulated by genistein and tyrphostin 51 by a tyrosine kinase-independent mechanism.
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PMID:Tyrosine kinase-independent inhibition of cyclic-AMP phosphodiesterase by genistein and tyrphostin 51. 1035 87


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