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)

The cAMP responsiveness of the phosphoenolpyruvate carboxykinase (PEPCK) gene promoter is mediated by a cAMP response unit, which includes three CCAAT/enhancer-binding protein (C/EBPs) sites, and a cAMP response element (CRE). Because both the CRE-binding protein and several C/EBP isoforms can to bind to the CRE with similar affinity, a variety of transcription factor bindings arrays in the cAMP response unit are possible that may affect the protein kinase A (PKA) responsivity of the promoter. To explore this issue, we have designed PEPCK promoter variants that have the native cis-elements within the cAMP response unit replaced with one or more LexA- and/or GAL4-binding sites. We also engineered the corresponding C/EBP and CRE-binding protein chimeras, which have their basic region leucine zipper domains replaced with LexA or GAL4 DNA-binding domains. Using this approach, we have reconstituted the PKA responsiveness of permissive PEPCK promoters in hepatoma cells and have characterized the PKA responsivity of the promoter under defined transcription factor occupancy patterns. Furthermore, analysis of deletion mutants of C/EBPalpha indicated that the domains that mediate its constitutive and PKA-inducible activities vary depending on which cis-element it occupies on the PEPCK promoter. These results suggest that promoter context may influence which domains within a transcription factor are employed to mediate transactivation.
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PMID:Different transcription factor binding arrays modulate the cAMP responsivity of the phosphoenolpyruvate carboxykinase gene promoter. 1223 88

C(4) phosphoenolpyruvate carboxylase (PEPCase: EC 4.1.1.31) is subjected to in vivo regulatory phosphorylation by a light up-regulated, calcium-independent protein kinase. Salt stress greatly enhanced phosphoenolpyruvate carboxylase-kinase (PEPCase-k) activity in leaves of Sorghum. The increase in PEPCase-k anticipated the time course of proline accumulation thereby suggesting that water stress was not involved in the kinase response to salt. Moreover, osmotic stress seemed not to be the main factor implicated, as demonstrated by the lack of effect when water availability was restricted by mannitol. In contrast, LiCl (at a concentration of 10 mM in short-term treatment of both excised leaves and whole plants) mimicked the effects of 172 mM NaCl salt-acclimation, indicating that the rise in PEPCase-k activity resulted primarily from the ionic stress. Both NaCl and LiCl treatments increased the activity of a Ca(2+)-independent, 35 kDa kinase, as demonstrated by an in-gel phosphorylation experiment. Short-term treatment of excised leaves with NaCl or LiCl partially reproduces the effects of whole plant treatments. Finally, salinization also increased PEPCase-k activity and the phosphorylation state of PEPCase in darkened Sorghum leaves. This fact, together with increased malate production during the dark period, suggests a shift towards mixed C(4) and crassulacean acid metabolism types of photosynthesis in response to salt stress.
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PMID:Characterization of salt stress-enhanced phosphoenolpyruvate carboxylase kinase activity in leaves of Sorghum vulgare: independence from osmotic stress, involvement of ion toxicity and significance of dark phosphorylation. 1256 7

A number of therapeutic targets are currently under investigation for inhibition of hepatic glucose production with small molecules. Antagonists of the glucagon receptor, glycogen phosphorylase, 11-beta-hydroxysteroid dehydrogenase-1 and fructose 1,6-bisphosphatase are, or have been, under evaluation in human clinical trials. Other strategies, including glucocorticoid receptor antagonists and carnitine palmitoyltransferase inhibitors, are supported by proof of principle studies in man as well as rodents. Several potential targets including glucose-6-phosphatase, glucose-6-phosphatase translocase, glycogen synthase kinase-3, adenosine receptor 2B antagonists, phosphoenolpyruvate carboxykinase and pyruvate dehydrogenase kinase, have been validated by compounds that are effective in animal models. Other targets like PGC-1a and CREB have initial validation support but no medicinal chemistry has been reported.
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PMID:Potential drug targets and progress towards pharmacologic inhibition of hepatic glucose production. 1257 Jul 14

The CDPK-SnRK superfamily consists of seven types of serine-threonine protein kinases: calcium-dependent protein kinase (CDPKs), CDPK-related kinases (CRKs), phosphoenolpyruvate carboxylase kinases (PPCKs), PEP carboxylase kinase-related kinases (PEPRKs), calmodulin-dependent protein kinases (CaMKs), calcium and calmodulin-dependent protein kinases (CCaMKs), and SnRKs. Within this superfamily, individual isoforms and subfamilies contain distinct regulatory domains, subcellular targeting information, and substrate specificities. Our analysis of the Arabidopsis genome identified 34 CDPKs, eight CRKs, two PPCKs, two PEPRKs, and 38 SnRKs. No definitive examples were found for a CCaMK similar to those previously identified in lily (Lilium longiflorum) and tobacco (Nicotiana tabacum) or for a CaMK similar to those in animals or yeast. CDPKs are present in plants and a specific subgroup of protists, but CRKs, PPCKs, PEPRKs, and two of the SnRK subgroups have been found only in plants. CDPKs and at least one SnRK have been implicated in decoding calcium signals in Arabidopsis. Analysis of intron placements supports the hypothesis that CDPKs, CRKs, PPCKs and PEPRKs have a common evolutionary origin; however there are no conserved intron positions between these kinases and the SnRK subgroup. CDPKs and SnRKs are found on all five Arabidopsis chromosomes. The presence of closely related kinases in regions of the genome known to have arisen by genome duplication indicates that these kinases probably arose by divergence from common ancestors. The PlantsP database provides a resource of continuously updated information on protein kinases from Arabidopsis and other plants.
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PMID:The Arabidopsis CDPK-SnRK superfamily of protein kinases. 1280 96

Higher plant phosphoenolpyruvate carboxylase (PEPC) is subject to in vivo phosphorylation of a regulatory serine located in the N-terminal domain of the protein. Studies using synthetic peptide substrates and mutated phosphorylation domain photosynthetic PEPC (C4 PEPC) suggested that the interaction of phosphoenolpyruvate carboxylase kinase (PEPCk) with its target was not restricted to this domain. However, no further information was available as to where PEPCk-C4 PEPC interactions take place. In this work, we have studied the possible interaction of the conserved 19-amino acid C-terminal sequence of sorghum (Sorghum vulgare Pers cv Tamaran) C4 PEPC with PEPCk. In reconstituted assays, a C-terminal synthetic peptide containing this sequence (peptide C19) was found to inhibit the phosphorylation reaction by the partially purified Ca2+-independent PEPCk (50% inhibition of initial activity = 230 microm). This effect was highly specific because peptide C19 did not alter C4 PEPC phosphorylation by either a partially purified sorghum leaf Ca2+-dependent protein kinase or the catalytic subunit of mammalian protein kinase A. In addition, the Ca2+-independent PEPCk was partially but significantly retained in affinity chromatography using a peptide C19 agarose column. Because peptide C15 (peptide C19 lacking the last four amino acids, QNTG) also inhibited C4 PEPC phosphorylation, it was concluded that the amino acid sequence downstream from the QNTG motif was responsible for the inhibitory effect. Specific antibodies raised against peptide C19 revealed that native C4 PEPC could be in two different conformational states. The results are discussed in relation with the reported crystal structure of the bacterial (Escherichia coli) and plant (maize [Zea mays]) enzymes.
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PMID:A conserved 19-amino acid synthetic peptide from the carboxy terminus of phosphoenolpyruvate carboxylase inhibits the in vitro phosphorylation of the enzyme by the calcium-independent phosphoenolpyruvate carboxylase kinase. 1280 37

The genome of the budding yeast (Saccharomyces cerevisiae) provides an important paradigm for transgenomic comparisons with other eukaryotic species. Here, we report a systematic comparison of the protein kinases of yeast (119 kinases) and a reference plant Arabidopsis (1,019 kinases). Using a whole-protein-based, hierarchical clustering approach, the complete set of protein kinases from both species were clustered. We validated our clustering by three observations: (a) clustering pattern of functional orthologs proven in genetic complementation experiments, (b) consistency with reported classifications of yeast kinases, and (c) consistency with the biochemical properties of those Arabidopsis kinases already experimentally characterized. The clustering pattern identified no overlap between yeast kinases and the receptor-like kinases (RLKs) of Arabidopsis. Ten more kinase families were found to be specific for one of the two species. Among them, the calcium-dependent protein kinase and phosphoenolpyruvate carboxylase kinase families are specific for plants, whereas the Ca(2+)/calmodulin-dependent protein kinase and provirus insertion in mouse-like kinase families were found only in yeast and animals. Three yeast kinase families, nitrogen permease reactivator/halotolerance-5), polyamine transport kinase, and negative regulator of sexual conjugation and meiosis, are absent in both plants and animals. The majority of yeast kinase families (21 of 26) display Arabidopsis counterparts, and all are mapped into Arabidopsis families of intracellular kinases that are not related to RLKs. Representatives from 11 of the common families (54 kinases from Arabidopsis and 17 from yeast) share an extremely high degree of similarity (blast E value < 10(-80)), suggesting the likelihood of orthologous functions. Selective expansion of yeast kinase families was observed in Arabidopsis. This is most evident for yeast genes CBK1, HRR25, and SNF1 and the kinase family S6K. Reduction of kinase families was also observed, as in the case of the NEK-like family. The distinguishing features between the two sets of kinases are the selective expansion of yeast families and the generation of a limited number of new kinase families for new functionality in Arabidopsis, most notably, the Arabidopsis RLKs that constitute important components of plant intercellular communication apparatus.
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PMID:Systematic trans-genomic comparison of protein kinases between Arabidopsis and Saccharomyces cerevisiae. 1291 70

To assess the role of insulin receptor (IR) substrate (IRS)-2 in insulin action and resistance in the liver, immortalized neonatal hepatocyte cell lines have been generated from IRS-2(-/-), IRS-2(+/-), and wild-type mice. These cells maintained the expression of the differentiated liver markers albumin and carbamoyl phosphate synthetase, as well as bear a high number of IRs. The lack of IRS-2 did not result in enhanced IRS-1 tyrosine phosphorylation or IRS-1-associated phosphatidylinositol (PI) 3-kinase activity on insulin stimulation. Total insulin-induced PI 3-kinase activity was decreased by 50% in IRS-2(-/-) hepatocytes, but the translocation of PI-3,4,5-trisphosphate to the plasma membrane in these cells was almost completely abolished. Downstream PI 3-kinase, activation of Akt, glycogen synthase kinase (GSK)-3 (alpha and beta isoforms), Foxo1, and atypical protein kinase C were blunted in insulin-stimulated IRS-2(-/-) cells. Reconstitution of IRS-2(-/-) hepatocytes with adenoviral IRS-2 restored activation of these pathways, demonstrating that IRS-2 is essential for functional insulin signaling in hepatocytes. Insulin induced a marked glycogen synthase activity in wild-type and heterozygous primary hepatocytes; interestingly, this response was absent in IRS-2(-/-) cells but was rescued by infection with adenoviral IRS-2. Regarding gluconeogenesis, the induction of phosphoenolpyruvate carboxykinase and glucose 6-phosphatase by dibutyryl cAMP and dexamethasone was observed in primary hepatocytes of all genotypes. However, insulin was not able to suppress gluconeogenic gene expression in primary hepatocytes lacking IRS-2, but when IRS-2 signaling was reconstituted, these cells recovered this response to insulin. Suppression of gluconeogenic gene expression in IRS-2-deficient primary hepatocytes was also restored by infection with dominant negative Delta 256Foxo1.
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PMID:Molecular mechanisms of insulin resistance in IRS-2-deficient hepatocytes. 1294 62

In mesophyll cells (MC) of Digitaria sanguinalis, the C(4)-phosphoenolpyruvate carboxylase (C(4)-PEPC) initiating the photosynthetic pathway is controlled by a complex light-dependent phosphorylation process. We showed previously that the transduction cascade involves the phosphoinositide pathway and a Ca(2+)-dependent step, which precedes the upregulation of the PEPC kinase (PEPCk). We have now further characterized the cascade component requiring Ca(2+). A Ca(2+)-dependent protein kinase that shows several characteristics of the conventional type of mammalian protein kinase C (PKC) was detected in protein extracts from mesophyll cell protoplasts (MCPs). It catalyzed the in vitro phosphorylation of the C1-peptide PKC substrate and was markedly inhibited by a PKC-specific pseudosubstrate domain. However, it was only modestly activated by the phospholipids phosphatidylserine and lysophosphatidylcholine, while choline, oleyl acetylglycerol, phosphatidylinositol, and the phorbol ester phorbol 12-myristate 13-acetate did not show any effect. Nevertheless, its activity was found to be associated with a polypeptide of 75kDa that was recognized by a PKC antibody raised against the C-terminus of rabbit PKCbeta II. In addition, this protein kinase was also inhibited by the Ca(2+)-dependent protein kinase (CDPK)/PKC inhibitors W7, H7, and staurosporine. Surprisingly, it was found to be phosphorylated in dark-adapted MCPs, albeit to a low extent, and this did not change during protoplast induction by light. W7, H7, and staurosporine were shown to markedly inhibit C(4)-PEPC phosphorylation in light-treated MCPs. These results support the view that this protein kinase is a good candidate to represent the Ca(2+)-activated component of the C(4)-PEPC phosphorylation cascade.
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PMID:A Ca(2+)-dependent protein kinase with characteristics of protein kinase C in leaves and mesophyll cell protoplasts from Digitaria sanguinalis: possible involvement in the C(4)-phosphoenolpyruvate carboxylase phosphorylation cascade. 1473 23

It appears that low amounts of fructose improve, whereas increased concentrations impair glucose tolerance and hepatic glucose metabolism. In this study, we compared directly the effects of low vs. high portal vein fructose concentrations on hepatic glucose metabolism in rats, using glucose-6-phosphatase gene expression as an endpoint. In the control group (C; n = 7), pancreatic clamps were performed using somatostatin and replacement of insulin such that basal glucose levels were maintained. In the experimental groups (n = 8/group), hyperglycemic, hyperinsulinemic pancreatic clamps were performed in which glucose (G) or glucose + fructose was infused into a jejunal vein. Fructose was infused to achieve either low (F1; <0.3 mmol/L) or high (F2; >1.0 mmol/L) portal vein concentrations. Total sugar load to the liver was equalized among the 3 experimental groups. Compared with C, liver glucose-6-phosphatase catalytic subunit mRNA was reduced by approximately 55% in G and F1, whereas it was increased approximately 180% in F2. F2 did not differentially affect glucose-6-phosphate translocase or phosphoenolpyruvate carboxykinase mRNA levels in liver, nor kidney glucose-6-phosphatase catalytic subunit mRNA. Livers from the F2 group were characterized by an accumulation of pentose phosphate intermediates and reduced phosphorylation of glycogen synthase kinase-3 (active form). However, in separate studies (n = 5/group), the infusion of a glycogen synthase kinase-3 inhibitor did not prevent the effects of F2 on glucose-6-phosphatase gene expression. We hypothesize that elevated fructose concentrations, similar to levels achieved after ingestion of sucrose- or fructose-enriched meals, initiate signals within the liver that elicit selective changes in hepatic gene expression.
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PMID:An acute increase in fructose concentration increases hepatic glucose-6-phosphatase mRNA via mechanisms that are independent of glycogen synthase kinase-3 in rats. 1498 44

Phosphatidic acid (PA) is emerging as an important lipid signalling molecule. In plants, it is implicated in various stress-signalling pathways and is formed in response to wounding, osmotic stress, cold stress, pathogen elicitors, Nod factors, ethylene and abscisic acid. How PA exerts its effects is still unknown, mainly because of the lack of characterized PA targets. In an approach to isolate such targets we have used PA-affinity chromatography. Several PA-binding proteins were present in the soluble fraction of tomato and Arabidopsis cells. Using mass spectrometric analysis, several of these proteins, including Hsp90, 14-3-3 proteins, an SnRK2 serine/threonine protein kinase and the PP2A regulatory subunit RCN1 could be identified. As an example, the binding of one major PA-binding protein, phosphoenolpyruvate carboxylase (PEPC), was characterized further. Competition experiments with different phospholipids confirmed specificity for PA. Hypo-osmotic treatment of the cells increased the amount of PEPC that bound the PA beads without increasing the absolute amount of PEPC. This suggests that PEPC's affinity for PA had increased. The work shows that PA-affinity chromatography/mass spectrometry is an effective way to isolate and identify PA-binding proteins from plants.
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PMID:Isolation and identification of phosphatidic acid targets from plants. 1527 72


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