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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Activation of protein kinase B (PKB) by growth factors and hormones has been demonstrated to proceed via phosphatidylinositol 3-kinase (PI3-kinase). In this report, we show that PKB can also be activated by PKA (cyclic AMP [cAMP]-dependent protein kinase) through a PI3-kinase-independent pathway. Although this activation required phosphorylation of PKB, PKB is not likely to be a physiological substrate of PKA since a mutation in the sole PKA consensus phosphorylation site of PKB did not abolish PKA-induced activation of PKB. In addition, mechanistically, this activation was different from that of growth factors since it did not require phosphorylation of the S473 residue, which is essential for full PKB activation induced by insulin. These data were supported by the fact that mutation of residue S473 of PKB to alanine did not prevent it from being activated by forskolin. Moreover, phosphopeptide maps of overexpressed PKB from COS cells showed differences between insulin- and forskolin-stimulated cells that pointed to distinct activation mechanisms of PKB depending on whether insulin or cAMP was used. We looked at events downstream of PKB and found that PKA activation of PKB led to the phosphorylation and inhibition of glycogen synthase kinase-3 (GSK-3) activity, a known in vivo substrate of PKB. Overexpression of a dominant negative PKB led to the loss of inhibition of GSK-3 in both insulin- and forskolin-treated cells, demonstrating that PKB was responsible for this inhibition in both cases. Finally, we show by confocal microscopy that forskolin, similar to insulin, was able to induce translocation of PKB to the plasma membrane. This process was inhibited by high concentrations of wortmannin (300 nM), suggesting that forskolin-induced PKB movement may require phospholipids, which are probably not generated by class I or class III PI3-kinase. However, high concentrations of wortmannin did not abolish PKB activation, which demonstrates that translocation per se is not important for PKA-induced PKB activation.
Mol Cell Biol 1999 Jul
PMID:Mechanism of protein kinase B activation by cyclic AMP-dependent protein kinase. 1037 49

Axin is a recently identified protein encoded by the fused locus in mice that is required for normal vertebrate axis formation. We have defined a 25-amino-acid sequence in axin that comprises the glycogen synthase kinase 3beta (GSK-3beta) interaction domain (GID). In contrast to full-length axin, which has been shown to antagonize Wnt signaling, the GID inhibits GSK-3beta in vivo and activates Wnt signaling. Similarly, mutants of axin lacking key regulatory domains such as the RGS domain, which is required for interaction with the adenomatous polyposis coli protein, bind and inhibit GSK-3beta in vivo, suggesting that these domains are critical for proper regulation of GSK-3beta activity. We have identified a novel self-interaction domain in axin and have shown that formation of an axin regulatory complex in vivo is critical for axis formation and GSK-3beta activity. Based on these data, we propose that the axin complex may directly regulate GSK-3beta enzymatic activity in vivo. These observations also demonstrate that alternative inhibitors of GSK-3beta can mimic the effect of lithium in developing Xenopus embryos.
Mol Cell Biol 1999 Oct
PMID:Regulation of glycogen synthase kinase 3beta and downstream Wnt signaling by axin. 1049 Jun 50

Integrin-mediated interactions of cells with components of the extracellular matrix regulate cell survival, cell proliferation, cell differentiation, and cell migration. Some of these physiological responses are regulated via activation of transcription factors such as activator protein 1 (AP-1). Integrin-linked kinase (ILK) is an ankyrin repeat containing serine-threonine protein kinase whose activity is rapidly and transiently stimulated by cell-fibronectin interactions as well as by insulin stimulation. ILK activates protein kinase B and inhibits the glycogen synthase kinase 3 (GSK-3) activity in a phosphatidylinositol-3-kinase (PI 3-kinase)-dependent manner. We now show that cell adhesion to fibronectin results in a rapid and transient stimulation of AP-1 activity. At the same time, the kinase activity of ILK is stimulated whereas that of GSK-3 is inhibited. This fibronectin-dependent activation of AP-1 activity is inhibited in a dose-dependent manner if the cells are transfected with wild-type GSK-3, and also by inhibitors of PI 3-kinase. Stable or transient overexpression of ILK results in a stimulation of AP-1 activity which is inhibited by cotransfection with wild-type GSK-3 and kinase-deficient ILK. Transient transfection of ILK in HEK-293 cells stimulates complex formation between an AP-1 consensus oligonucleotide and nuclear proteins containing c-jun. The formation of this complex is inhibited by cotransfection with active GSK-3 or kinase-deficient ILK, suggesting that ILK may regulate AP-1 activation by inhibiting GSK-3, which has previously been shown to be a negative regulator of AP-1. In the presence of serum, ILK has no effect on the phosphorylation of Ser-73 in the N-terminal transactivation domain of c-jun. These results demonstrate a novel signaling pathway for the adhesion-mediated stimulation of AP-1 transcriptional activity involving ILK and GSK-3 and the subsequent regulation of the c-jun-DNA interaction.
Mol Cell Biol 1999 Nov
PMID:Cell-extracellular matrix interactions stimulate the AP-1 transcription factor in an integrin-linked kinase- and glycogen synthase kinase 3-dependent manner. 1052 30

Yeast has three A kinase catalytic subunits, which have greater than 75% identity and are encoded by the TPK genes (TPK1, TPK2, and TPK3) [Toda, T., Cameron, S., Sass, P., Zoller, M. & Wigler, M. (1987) Cell 50, 277-287]. Although they are redundant for viability, the three A kinases are not redundant for pseudohyphal growth [Robertson, L. S. & Fink, G. R. (1998) Proc. Natl. Acad. Sci. USA 95, 13783-13787; Pan, X. & Heitman, J. (1999) Mol. Cell. Biol. 19, 4874-4887]; Tpk2, but not Tpk1 or Tpk3, is required for pseudohyphal growth. Genome-wide transcriptional profiling has revealed unique signatures for each of the three A kinases leading to the identification of additional functional diversity among these proteins. Tpk2 negatively regulates genes involved in iron uptake and positively regulates genes involved in trehalose degradation and water homeostasis. Tpk1 is required for the derepression of branched chain amino acid biosynthesis genes that seem to have a second role in the maintenance of iron levels and DNA stability within mitochondria. The fact that TPK2 mutants grow better than wild types on nonfermentable carbon sources and on media deficient in iron supports the unique role of Tpk2 in respiratory growth and carbon source use.
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PMID:The yeast A kinases differentially regulate iron uptake and respiratory function. 1081 93

Using recombinant human tau protein phosphorylated by a brain extract and the glycogen synthase kinase-3beta in the absence or the presence of heparin, we showed that phosphorylation-dependent antibody AD2 recognition only requires phosphorylated Ser-396. By the Spot multiple peptide synthesis method, we showed that Tyr-394, Ser(P)-396 and Pro-397 are critical for AD2 binding. A decrease in the binding of AD2 was observed with increasing phosphorylation of residues in the vicinity of Ser(P)-396.
Brain Res Mol Brain Res 2000 May 31
PMID:Binding specificity of monoclonal antibody AD2: influence of the phosphorylation state of tau. 1089 98

The yeast Saccharomyces cerevisiae has four genes, MCK1, MDS1 (RIM11), MRK1, and YOL128c, that encode glycogen synthase kinase 3 (GSK-3) homologs. The gsk-3 null mutant, in which these four genes are disrupted, shows temperature sensitivity, which is suppressed by the expression of mammalian GSK-3beta and by an osmotic stabilizer. Suppression of temperature sensitivity by an osmotic stabilizer is also observed in the bul1 bul2 double null mutant, and the temperature sensitivity of the bul1 bul2 double null mutant is suppressed by multiple copies of MCK1. We have screened rog mutants (revertants of gsk-3) which suppress the temperature sensitivity of the mck1 mds1 double null mutant and found that two of them, rog1 and rog2, also suppress the temperature sensitivity of the bul1 bul2 double null mutant. Bul1 and Bul2 have been reported to bind to Rsp5, a hect (for homologous to E6-associated-protein carboxyl terminus)-type ubiquitin ligase, but involvement of Bul1 and Bul2 in protein degradation has not been demonstrated. We find that Rog1, but not Rog2, is stabilized in the gsk-3 null and the bul1 bul2 double null mutants. Rog1 binds directly to Rsp5, and their interaction is dependent on GSK-3. Furthermore, Rog1 is stabilized in the npi1 mutant, in which RSP5 expression levels are reduced. These results suggest that yeast GSK-3 regulates the stability of Rog1 in cooperation with Bul1, Bul2, and Rsp5.
Mol Cell Biol 2000 Sep
PMID:Yeast glycogen synthase kinase 3 is involved in protein degradation in cooperation with Bul1, Bul2, and Rsp5. 1095 69

The effect of insulin on glycogen synthesis and key enzymes of glycogen metabolism, glycogen phosphorylase and glycogen synthase, was studied in HepG2 cells. Insulin stimulated glycogen synthesis 1.83-3.30 fold depending on insulin concentration in the medium. Insulin caused a maximum of 65% decrease in glycogen phosphorylase 'a' and 110% increase in glycogen synthase activities in 5 min. Although significant changes in enzyme activities were observed with as low as 0.5 nM insulin level, the maximum effects were observed with 100 nM insulin. There was a significant inverse correlation between activities of glycogen phosphorylase 'a' and glycogen synthase 'a' (R2= 0.66, p < 0.001). Addition of 30 mM glucose caused a decrease in phosphorylase 'a' activity in the absence of insulin and this effect was additive with insulin up to 10 nM concentration. The inactivation of phosphorylase 'a' by insulin was prevented by wortmannin and rapamycin but not by PD98059. The activation of glycogen synthase by insulin was prevented by wortmannin but not by PD98059 or rapamycin. In fact, PD98059 slightly stimulated glycogen synthase activation by insulin. Under these experimental conditions, insulin decreased glycogen synthase kinase-3beta activity by 30-50% and activated more than 4-fold particulate protein phosphatase- activity and 1.9-fold protein kinase B activity; changes in all of these enzyme activities were abolished by wortmannin. The inactivation of GSK-3beta and activation of PKB by insulin were associated with their phosphorylation and this was also reversed by wortmannin. The addition of protein phosphatase-1 inhibitors, okadaic acid and calyculin A, completely abolished the effects of insulin on both enzymes. These data suggest that stimulation of glycogen synthase by insulin in HepG2 cells is mediated through the PI-3 kinase pathway by activating PKB and PP-1G and inactivating GSK-3beta. On the other hand, inactivation of phosphorylase by insulin is mediated through the PI-3 kinase pathway involving a rapamycin-sensitive p70(s6k) and PP-1G. These experiments demonstrate that insulin regulates glycogen phosphorylase and glycogen synthase through (i) a common signaling pathway at least up to PI-3 kinase and bifurcates downstream and (ii) that PP-1 activity is essential for the effect of insulin.
Mol Cell Biochem 2000 Aug
PMID:Reciprocal regulation of glycogen phosphorylase and glycogen synthase by insulin involving phosphatidylinositol-3 kinase and protein phosphatase-1 in HepG2 cells. 1105 55

Three founder transgenic mice were generated with a 108 kb human genomic fragment containing the entire autosomal dominant polycystic kidney disease (ADPKD) gene, PKD1, plus the tuberous sclerosis gene, TSC2. Two lines were established (TPK1 and TPK3) each with approximately 30 copies of the transgene. Both lines produced full-length PKD1 mRNA and polycystin-1 protein that was developmentally regulated, similar to the endogenous pattern, with expression during renal embryogenesis and neonatal life, markedly reduced at the conclusion of renal development. Tuberin expression was limited to the brain. Transgenic animals from both lines (and the TPK2 founder animal) often displayed a renal cystic phenotype, typically consisting of multiple microcysts, mainly of glomerular origin. Hepatic cysts and bile duct proliferation, characteristic of ADPKD, were also seen. All animals with two copies of the transgenic chromosome developed cysts and, in total, 48 of the 100 transgenic animals displayed a cystic phenotype. To test the functionality of the transgene, animals were bred with the Pkd1(del34) knockout mouse. Both transgenic lines rescued the embryonically lethal Pkd1(del34/del34) phenotype, demonstrating that human polycystin-1 can complement for loss of the endogenous protein. The rescued animals were viable into adulthood, although more than half developed hepatic cystic disease in later life, similar to the phenotype of older Pkd1(del34/+) animals. The TPK mice have defined a minimal area that appropriately expresses human PKD1. Furthermore, this model indicates that over-expression of normal PKD1 can elicit a disease phenotype, suggesting that the level of polycystin-1 expression may be relevant in the human disease.
Hum Mol Genet 2000 Nov 01
PMID:A human PKD1 transgene generates functional polycystin-1 in mice and is associated with a cystic phenotype. 1106 21

Agents that elevate intracellular cyclic AMP (cAMP) levels promote neuronal survival in a manner independent of neurotrophic factors. Inhibitors of phosphatidylinositol 3 kinase and dominant-inactive mutants of the protein kinase Akt do not block the survival effects of cAMP, suggesting that another signaling pathway is involved. In this report, we demonstrate that elevation of intracellular cAMP levels in rat cerebellar granule neurons leads to phosphorylation and inhibition of glycogen synthase kinase 3beta (GSK-3beta). The increased phosphorylation of GSK-3beta by protein kinase A (PKA) occurs at serine 9, the same site phosphorylated by Akt. Purified PKA is able to phosphorylate recombinant GSK-3beta in vitro. Inhibitors of GSK-3 block apoptosis in these neurons, and transfection of neurons with a GSK-3beta mutant that cannot be phosphorylated interferes with the prosurvival effects of cAMP. These data suggest that activated PKA directly phosphorylates GSK-3beta and inhibits its apoptotic activity in neurons.
Mol Cell Biol 2000 Dec
PMID:Cyclic AMP promotes neuronal survival by phosphorylation of glycogen synthase kinase 3beta. 1109 86

Grb10 is a member of a family of adapter proteins that binds to tyrosine-phosphorylated receptors including the insulin receptor kinase (IRK). In this study recombinant adenovirus was used to over-express hGrb10zeta, a new Grb10 isoform, in primary rat hepatocytes and the consequences for insulin signaling were evaluated. Over-expression of hGrb10zeta resulted in 50% inhibition of insulin-stimulated IRK autophosphorylation and activation. Analysis of downstream events showed that hGrb10zeta over-expression specifically inhibits insulin-stimulated glycogen synthase (GS) activity and glycogen synthesis without affecting insulin-induced IRS1/2 phosphorylation, PI3-kinase activation, insulin like growth factor binding protein-1 (IGFBP-1) mRNA expression, and ERK1/2 MAP kinase activity. The classical pathway from PI3-kinase through Akt-PKB/GSK-3 leading to GS activation by insulin was also not affected by hGrb10zeta over-expression. These results indicate that hGrb10zeta inhibits a novel and presently unidentified insulin signaling pathway leading to GS activation in liver.
Mol Cell Endocrinol 2001 Feb 28
PMID:Specific inhibition by hGRB10zeta of insulin-induced glycogen synthase activation: evidence for a novel signaling pathway. 1122 74


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