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)

Inhibitor-1 becomes a potent inhibitor of protein phosphatase 1 when phosphorylated by cAMP-dependent protein kinase at Thr(35). Moreover, Ser(67) of inhibitor-1 serves as a substrate for cyclin-dependent kinase 5 in the brain. Here, we report that dephosphoinhibitor-1 but not phospho-Ser(67) inhibitor-1 was efficiently phosphorylated by protein kinase C at Ser(65) in vitro. In contrast, Ser(67) phosphorylation by cyclin-dependent kinase 5 was unaffected by phospho-Ser(65). Protein kinase C activation in striatal tissue resulted in the concomitant phosphorylation of inhibitor-1 at Ser(65) and Ser(67), but not Ser(65) alone. Selective pharmacological inhibition of protein phosphatase activity suggested that phospho-Ser(65) inhibitor-1 is dephosphorylated by protein phosphatase 1 in the striatum. In vitro studies confirmed these findings and suggested that phospho-Ser(67) protects phospho-Ser(65) inhibitor-1 from dephosphorylation by protein phosphatase 1 in vivo. Activation of group I metabotropic glutamate receptors resulted in the up-regulation of diphospho-Ser(65)/Ser(67) inhibitor-1 in this tissue. In contrast, the activation of N-methyl-d-aspartate-type ionotropic glutamate receptors opposed increases in striatal diphospho-Ser(65)/Ser(67) inhibitor-1 levels. Phosphomimetic mutation of Ser(65) and/or Ser(67) did not convert inhibitor-1 into a protein phosphatase 1 inhibitor. On the other hand, in vitro and in vivo studies suggested that diphospho-Ser(65)/Ser(67) inhibitor-1 is a poor substrate for cAMP-dependent protein kinase. These observations extend earlier studies regarding the function of phospho-Ser(67) and underscore the possibility that phosphorylation in this region of inhibitor-1 by multiple protein kinases may serve as an integrative signaling mechanism that governs the responsiveness of inhibitor-1 to cAMP-dependent protein kinase activation.
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PMID:Phosphorylation of protein phosphatase inhibitor-1 by protein kinase C. 1677 99

The circadian clock is regulated by a transcription/translation negative feedback loop. A key negative regulator of circadian rhythm in mammals is the PER2 (mammalian PERIOD 2) protein. Its daily degradation at the end of the night accompanies de-repression of transcription. CKI (casein kinase I ) has been identified as the kinase that phosphorylates PER2, targeting it for ubiquitin-mediated proteasomal degradation. We now report that PER2 degradation is also negatively regulated by PP1 (protein phosphatase 1)-mediated dephosphorylation. In Xenopus egg extract, PP1 inhibition by Inhibitor-2 accelerated mPER2 degradation. Co-immunoprecipitation experiments showed that PER2 bound to PP1c in transfected HEK-293 cells. PP1 immunoprecipitated from HEK-293 cells, mouse liver and mouse brain, dephosphorylated CKI-phosphorylated PER2, showing that PER2 is a substrate for mammalian endogenous PP1. Moreover, over-expression of the dominant negative form of PP1c, the D95N mutant, accelerated ubiquitin and proteasome-mediated degradation of PER2, and shortened the PER2 half-life in HEK-293 cells. Over-expression of the PP1 inhibitors, protein phosphatase 1 holoenzyme inhibitor-1 and Inhibitor-2, confirmed these results. Thus PP1 regulates PER2 stability and is therefore a candidate to regulate mammalian circadian rhythms.
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PMID:Protein phosphatase 1 regulates the stability of the circadian protein PER2. 1695 25

The ubiquitously expressed protein glycogen synthase kinase-3 (GSK3) is constitutively active, however its activity is markedly diminished following phosphorylation of Ser21 of GSK3alpha and Ser9 of GSK3beta. Although several kinases are known to phosphorylate Ser21/9 of GSK3, for example Akt, relatively much less is known about the mechanisms that cause the dephosphorylation of GSK3 at Ser21/9. In the present study KCl-induced plasma membrane depolarization of SH-SY5Y cells, which increases intracellular calcium concentrations caused a transient decrease in the phosphorylation of Akt at Thr308 and Ser473, and GSK3 at Ser21/9. Overexpression of the selective protein phosphatase-1 inhibitor protein, inhibitor-2, increased basal GSK3 phosphorylation at Ser21/9 and significantly blocked the KCl-induced dephosphorylation of GSK3beta, but not GSK3alpha. The phosphorylation of Akt was not affected by the overexpression of inhibitor-2. GSK3 activity is known to affect sarcoplasmic/endoplasmic reticulum calcium ATPase 2 (SERCA2) levels. Overexpression of inhibitor-2 or treatment of cells with the GSK3 inhibitors lithium and SB216763 increased the levels of SERCA2. These results indicate that the protein phosphatase-1/inhibitor-2 complex differentially regulates GSK3 dephosphorylation induced by KCl and that GSK3 activity regulates SERCA2 levels.
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PMID:The protein phosphatase-1/inhibitor-2 complex differentially regulates GSK3 dephosphorylation and increases sarcoplasmic/endoplasmic reticulum calcium ATPase 2 levels. 1698 14

Human and experimental heart failure is characterized by increases in type-1 protein phosphatase activity, which may be partially attributed to inactivation of its endogenous regulator, protein phosphatase inhibitor-1. Inhibitor-1 represents a nodal integrator of two major second messenger pathways, adenosine 3',5'-cyclic monophosphate (cAMP) and calcium, which mediate its phosphorylation at threonine 35 and serine 67, respectively. Here, using recombinant inhibitor-1 wild-type and mutated proteins, we identified a novel phosphorylation site in inhibitor-1, threonine 75. This phosphoamino acid was phosphorylated in vitro by protein kinase Calpha independently and to the same extent as serine 67, the previous protein kinase Calpha-identified site. Generation of specific antibodies for the phosphorylated and dephosphorylated threonine 75 revealed that this site is phosphorylated in rat and dog hearts. Adenoviral-mediated expression of the constitutively phosphorylated threonine 75 inhibitor-1 in isolated myocytes was associated with specific stimulation of type-1 protein phosphatase activity and marked inhibition of the sarcoplasmic calcium pump affinity for calcium, resulting in depressed contractility. Thus, phosphorylation of inhibitor-1 at threonine 75 represents a new mechanism of cardiac contractility regulation, partially through the alteration of sarcoplasmic reticulum calcium transport activity.
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PMID:Identification of a novel phosphorylation site in protein phosphatase inhibitor-1 as a negative regulator of cardiac function. 1704 26

The second messenger cyclic adenosine monophosphate (cAMP) plays a pivotal role in axonal growth and guidance, but its downstream mechanisms remain elusive. In this study, we report that type II protein kinase A (PKA) is highly enriched in growth cone filopodia, and this spatial localization enables the coupling of cAMP signaling to its specific effectors to regulate guidance responses. Disrupting the localization of PKA to filopodia impairs cAMP-mediated growth cone attraction and prevents the switching of repulsive responses to attraction by elevated cAMP. Our data further show that PKA targets protein phosphatase-1 (PP1) through the phosphorylation of a regulatory protein inhibitor-1 (I-1) to promote growth cone attraction. Finally, we find that I-1 and PP1 mediate growth cone repulsion induced by myelin-associated glycoprotein. These findings demonstrate that the spatial localization of type II PKA to growth cone filopodia plays an important role in the regulation of growth cone motility and guidance by cAMP.
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PMID:Spatial targeting of type II protein kinase A to filopodia mediates the regulation of growth cone guidance by cAMP. 1720 Apr 17

The liver has enormous regenerative capacity such that, after partial hepatectomy, hepatocytes rapidly replicate to restore liver mass, thus providing a context for studying in vivo mechanisms of cell growth regulation. Bax inhibitor-1 (BI-1) is an evolutionarily conserved endoplasmic reticulum (ER) protein that suppresses cell death. Interestingly, the BI-1 protein has been shown to regulate Ca(2+) handling by the ER similar to antiapoptotic Bcl-2 family proteins. Effects on cell cycle entry by Bcl-2 family proteins have been described, prompting us to explore whether bi-1-deficient mice display alterations in the in vivo regulation of cell cycle entry using a model of liver regeneration. Accordingly, we compared bi-1(+/+) and bi-1(-/-) mice subjected to partial hepatectomy with respect to the kinetics of liver regeneration and molecular events associated with hepatocyte proliferation. We found that bi-1 deficiency accelerates liver regeneration after partial hepatectomy. Regenerating hepatocytes in bi-1(-/-) mice enter cell cycle faster, as documented by more rapid incorporation of deoxynucleotides, associated with earlier increases in cyclin D1, cyclin D3, cyclin-dependent kinase (Cdk) 2, and Cdk4 protein levels, more rapid hyperphosphorylation of retinoblastoma protein, and faster degradation of p27(Kip1). Dephosphorylation and nuclear translocation of nuclear factor of activated T cells 1 (NFAT1), a substrate of the Ca(2+)-sensitive phosphatase calcineurin, were also accelerated following partial hepatectomy in BI-1-deficient hepatocytes. These findings therefore reveal additional similarities between BI-1 and Bcl-2 family proteins, showing a role for BI-1 in regulating cell proliferation in vivo, in addition to its previously described actions as a regulator of cell survival.
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PMID:Mice lacking bi-1 gene show accelerated liver regeneration. 1730 82

The depressed function of failing hearts has been partially attributed to increased protein phosphatase-1 through its impaired regulation by inhibitor-1. Phosphorylation of inhibitor-1 at Thr35 by PKA results in potent inhibition of protein phosphatase-1 activity, while phosphorylation at Ser67 or Thr75 by PKC attenuates the inhibitory activity. To examine the functional role of dual-site (Ser67, Thr75) phosphorylation of inhibitor-1 by PKC, the constitutively phosphorylated Ser67 (S67D) and/or Thr75 (T75D) human inhibitor-1 forms were expressed in adult cardiomyocytes. Expression of either single or double phosphorylated inhibitor-1 was associated with similar decreases in cardiac contractility, indicating that maximal inhibition can be elicited by each of these sites alone and that their inhibitory effects are not additive. Notably, activation of the cAMP pathway could only partially reverse the depressed contractile parameters. Accordingly, protein phosphatase-1 activity remained elevated, phosphorylation of phospholamban at Ser16 was decreased, and the EC(50) values of the sarcoplasmic reticulum calcium transport system were higher compared with controls. Thus phosphorylation of Ser67 and/or Thr75 in inhibitor-1 may mitigate the stimulatory effects of the cAMP pathway, resulting in compromised cardiac function.
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PMID:Phosphorylation of human inhibitor-1 at Ser67 and/or Thr75 attenuates stimulatory effects of protein kinase A signaling in cardiac myocytes. 1741 10

Inhibitor-1alpha is one of the isoforms of human protein phosphatase inhibitor-1. It is a product of alternative splicing of inhibitor-1 gene and lacks 51 internal amino acids from residue 84 to 134 of inhibitor-1. Here we have characterized the structural and biochemical properties of inhibitor-1alpha. Structural analysis of recombinant inhibitor-1alpha by NMR spectroscopy revealed that inhibitor-1alpha adopts a predominantly random coil conformation. Excluding the region from residue 84 to 134 of inhibitor-1, the structural features of inhibitor-1 and inhibitor-1alpha are almost the same as each other. The IC(50) value of inhibitor-1alpha in inhibition of Protein phosphatase-1 (PP1) is comparable to that of inhibitor-1, indicating that inhibitor-1alpha is a potent inhibitor of PP1 when Thr-35 is phosphorylated by PKA. For phosphorylation by PKA and dephosphorylation by protein phosphatase-1, -2A, and -2B, the measured kinetic parameters of inhibitor-1alpha are very close to those of inhibitor-1. Taken together, these results suggest that inhibitor-1alpha preserves the structure of inhibitor-1, the PP1 inhibitory activity and the functional specificities toward phosphorylation by PKA and dephosphorylation by protein phosphatase-1, -2A, and -2B.
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PMID:Structural and biochemical characterization of inhibitor-1alpha. 1751 Sep 62

Plasminogen activator inhibitor-1 (PAI-1) controls the regulation of the fibrinolytic system in blood by inhibiting both urokinase-type and tissue-type plasminogen activators. Enhanced levels of PAI-1 are related to pathological conditions associated with hypoxia or hyperinsulinemia. In this study, we investigated the regulation of PAI-1 expression by glucagon and the cAMP/PKA/CREB signalling pathway in the liver. Stimulation of the cAMP/PKA/CREB signalling cascade by starvation in vivo or glucagon in vitro induced PAI-1 gene expression in liver. Furthermore, this response was associated with enhanced phosphorylation of CREB. By using EMSAs we found that three promoter elements, the HRE2, E-box 4 and E-box 5, were able to bind CREB but only the HRE2 and E5 appeared to be functionally active. Reporter gene assays confirmed that cAMP induced PAI-1 gene transcription via the same element in both human and rat promoters. Interestingly, although the HRE2 was involved, the glucagon/cAMP pathway had no influence on hypoxia-inducible factor-1 (HIF-1) mRNA and protein levels. Thus, CREB binding to the HIF-1 responsive elements in PAI-1 promoter mediates the glucagon effect in the liver.
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PMID:CREB binding to the hypoxia-inducible factor-1 responsive elements in the plasminogen activator inhibitor-1 promoter mediates the glucagon effect. 1772 10

The specificities of 65 compounds reported to be relatively specific inhibitors of protein kinases have been profiled against a panel of 70-80 protein kinases. On the basis of this information, the effects of compounds that we have studied in cells and other data in the literature, we recommend the use of the following small-molecule inhibitors: SB 203580/SB202190 and BIRB 0796 to be used in parallel to assess the physiological roles of p38 MAPK (mitogen-activated protein kinase) isoforms, PI-103 and wortmannin to be used in parallel to inhibit phosphatidylinositol (phosphoinositide) 3-kinases, PP1 or PP2 to be used in parallel with Src-I1 (Src inhibitor-1) to inhibit Src family members; PD 184352 or PD 0325901 to inhibit MKK1 (MAPK kinase-1) or MKK1 plus MKK5, Akt-I-1/2 to inhibit the activation of PKB (protein kinase B/Akt), rapamycin to inhibit TORC1 [mTOR (mammalian target of rapamycin)-raptor (regulatory associated protein of mTOR) complex], CT 99021 to inhibit GSK3 (glycogen synthase kinase 3), BI-D1870 and SL0101 or FMK (fluoromethylketone) to be used in parallel to inhibit RSK (ribosomal S6 kinase), D4476 to inhibit CK1 (casein kinase 1), VX680 to inhibit Aurora kinases, and roscovitine as a pan-CDK (cyclin-dependent kinase) inhibitor. We have also identified harmine as a potent and specific inhibitor of DYRK1A (dual-specificity tyrosine-phosphorylated and -regulated kinase 1A) in vitro. The results have further emphasized the need for considerable caution in using small-molecule inhibitors of protein kinases to assess the physiological roles of these enzymes. Despite being used widely, many of the compounds that we analysed were too non-specific for useful conclusions to be made, other than to exclude the involvement of particular protein kinases in cellular processes.
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PMID:The selectivity of protein kinase inhibitors: a further update. 1785 Feb 14


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