Phosphatidylinositol 3′-Kinase and p70s6k Are Required for Insulin but Not Bisperoxovanadium 1,10-Phenanthroline (bpV(phen)) Inhibition of Insulin-like Growth Factor Binding Protein Gene Expression

Band, Christian J.; Posner, Barry I.

doi:10.1074/jbc.272.1.138

Cited by 56 publications

(51 citation statements)

References 71 publications

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“…Therefore, there is a PKB-independent regulation of this gene promoter that does not regulate the PEPCK or IGFBP1 gene promoters. This is consistent with several other studies that suggest that insulin regulates these three gene promoters by different mechanisms despite the existence of a related insulin response sequence in each (18,21,26,28,47). Nakae et al (32) have previously demonstrated a PKB-independent regulation of FOXO, a transcription factor closely associated with the insulin repression of these gene promoters.…”

Section: Discussionsupporting

confidence: 79%

“…However the importance of PKB and/or FOXO in the regulation of these genes has been questioned (18,28). For example, overexpression of dominant-negative PKB does not block insulin action on PEPCK (29) or G6Pase (22) and inhibitors of mTOR will block insulin regulation of IGFBP1 but not PKB or FOXO (21,26), while inhibitors of GSK3 (also downstream of PKB) will inhibit these genes without regulating FOXO activity (30,31). It is also suggested that insulin can regulate FOXO activity independently of PKB (32).…”

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confidence: 99%

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Characterization of a Protein Kinase B Inhibitor In Vitro and in Insulin-Treated Liver Cells

et al. 2007

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OBJECTIVE-Abnormal expression of the hepatic gluconeogenic genes (glucose-6-phosphatase [G6Pase] and PEPCK) contributes to hyperglycemia. These genes are repressed by insulin, but this process is defective in diabetic subjects. Protein kinase B (PKB) is implicated in this action of insulin. An inhibitor of PKB, Akt inhibitor (Akti)-1/2, was recently reported; however, the specificity and efficacy against insulin-induced PKB was not reported. Our aim was to characterize the specificity and efficacy of Akti-1/2 in cells exposed to insulin and then establish whether inhibition of PKB is sufficient to prevent regulation of hepatic gene expression by insulin.RESEARCH DESIGN AND METHODS-Akti-1/2 was assayed against 70 kinases in vitro and its ability to block PKB activation in cells exposed to insulin fully characterized.RESULTS-Akti-1/2 exhibits high selectivity toward PKB␣ and PKB␤. Complete inhibition of PKB activity is achieved in liver cells incubated with 1-10 mol/l Akti-1/2, and this blocks insulin regulation of PEPCK and G6Pase expression. Our data demonstrate that only 5-10% of maximal insulin-induced PKB is required to fully repress PEPCK and G6Pase expression. Finally, we demonstrate reduced insulin sensitivity of these gene promoters in cells exposed to submaximal concentrations of Akti-1/2; however, full repression of the genes can still be achieved by high concentrations of insulin.CONCLUSIONS-This work establishes the requirement for PKB activity in the insulin regulation of PEPCK, G6Pase, and a third insulin-regulated gene, IGF-binding protein-1 (IGFBP1); suggests a high degree of functional reserve; and identifies Akti-1/2 as a useful tool to delineate PKB function in the liver. Diabetes 56:2218-2227, 2007 P rotein kinase B (PKB) is a member of the AGC family of protein kinases (1-3). In mammals, there are three isoforms (PKB␣, PKB␤, and PKB␥) (1). PKB is activated following induction of phosphatidylinositol 3 (PI3) kinase activity and the resultant generation of the lipid second messengers PI 3,4,5 trisphosphate and PI 3,4 bisphosphate (4). These lipids bind to the PH domain of PKB, altering its conformation and permitting access to upstream protein kinases (5). Phosphoinositide-dependent protein kinase-1 phosphorylates PKB at Thr 308 (6), and a second phosphorylation (at Ser 473 ) occurs through the action of an alternative kinase, such as the rapamycin-insensitive mTOR complex 2 (TORC2) (7). Therefore, most growth factors, including platelet-derived growth factor, epidermal growth factor, and insulin, which are potent activators of PI3 kinase, also strongly induce PKB in cells.One of the first substrates of PKB to be characterized was GSK3, as part of the insulin signaling pathway that regulates glycogen metabolism (8). Since then, multiple potential substrates of PKB have been proposed including the proapoptotic protein Bad (9,10), the tuberous sclerosis complex (TSC)2 gene product (11), the Rab-GAP AS160 (12), proline-rich Akt substrate of 40 kDa (PRAS40) (13), and the key forkhead transcription ...

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Section: Discussionsupporting

confidence: 79%

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confidence: 99%

Characterization of a Protein Kinase B Inhibitor In Vitro and in Insulin-Treated Liver Cells

et al. 2007

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“…Immunoprecipitates were extensively washed, and the protein A-Sepharose pellets were resuspended in 50 μl of kinase assay buffer (20 mM Tris-HCl, pH 7.5, 100 mM NaCl, 0.5 mM EGTA) containing 0.5 mg/ml L-α-phosphatidylinositol (Avanti Polar Lipids, Inc., Alabaster, AL) and assayed for PI3-kinase activity as described previously (Band and Posner 1997).…”

Section: Pi3-kinase Activity Assaymentioning

confidence: 99%

Role of the PI3-kinase/mTor pathway in the regulation of the stearoyl CoA desaturase (SCD1) gene expression by insulin in liver

Mauvoisin

Rocque

Arfa

et al. 2007

J. Cell Commun. Signal.

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The stearoyl-CoA desaturase 1 (SCD1) catalyzes the synthesis of monounsaturated fatty acids. This enzyme is a critical control point regulating hepatic lipogenesis and lipid oxidation. Therefore SCD1 may be a potential therapeutic target in the treatment of obesity and metabolic syndrome. Regulation of SCD1 expression occurs primarily at the level of transcription. In the present study, we characterized the insulin response elements (IREs) and the insulin signaling pathway mediating the regulation of SCD1 gene transcription in liver. In chicken embryo hepatocytes (CEH) and HepG2 cells, insulin stimulates SCD1 promoter activity by 2.5 folds. This activation is mediated by two different IREs on the chicken promoter, one localized between −1,975 and −1,610 bp and one between −372 and −297 bp. The latter binds both NF-Y and SREBP-1 transcription factors in response to insulin. We also demonstrated that insulin induction of SCD1 gene expression and promoter activity is abolished by pre-incubation of cells with specific inhibitors of both PI3-kinase (LY294002) and mTor (Rapamycin) or by over-expression of a dominant negative mutant of PI3-kinase. The PI3-kinase and mTor pathway mediates the insulin response on both IREs. In summary, insulin activates SCD1 gene expression in liver via a signaling pathway that involves PI3-kinase and mTor and the downstream transcription factors NF-Y and SREBP-1.Sentence summary: Insulin regulates SCD1 gene expression via two different IREs. The most 3′ IRE is localized between −372 and −297 bp and binds the NF-Y and SREBP-1 transcription factors in response to insulin. PI3-kinase and mTor mediate the action of insulin on both IREs.

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“…For example, the mTOR inhibitor rapamycin blocks the regulation of glucocorticoid-induced IGFBP-1 expression by insulin (36). In addition, detailed characterization of the IGFBP-1 and PEPCK TIRE sequences suggests that FKHR can bind to mutant TIRE sequences that do not respond to insulin (37).…”

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confidence: 99%

Insulin Regulation of Insulin-like Growth Factor-binding Protein-1 Gene Expression Is Dependent on the Mammalian Target of Rapamycin, but Independent of Ribosomal S6 Kinase Activity

Patel

Lochhead

Rena

et al. 2002

Journal of Biological Chemistry

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Insulin inhibits the expression of the hepatic insulinlike growth factor-binding protein-1 (IGFBP-1) and glucose-6-phosphatase (G6Pase) genes. The signaling pathway that mediates these events requires the activation of phosphatidylinositol 3-kinase, whereas transfection studies have suggested an involvement of Akt (protein kinase B) and FKHR, a transcription factor regulated by Akt. We now demonstrate that insulin repression of endogenous IGFBP-1 gene transcription was blocked by rapamycin or by amino acid starvation. Rapamycin inhibited the mammalian target of rapamycin (mTOR) and the subsequent activation of p70/p85 S6 protein kinase-1 (S6K1) by insulin, whereas amino acid depletion prevented insulin induction of these signaling molecules. Importantly, we demonstrate that insulin regulation of the thymine-rich insulin response element of the IGFBP-1 promoter was also inhibited by rapamycin. However, sustained activation of S6K1 did not repress this promoter. In addition, rapamycin did not affect insulin regulation of G6Pase expression or Akt activation. WeproposethattheseobservationsindicatethatanmTORdependent, but S6K-independent mechanism regulates the suppression of IGFBP-1 (but not G6Pase) gene expression by insulin. Therefore, although the insulin-responsive sequence of the G6Pase gene promoter is related to that of the IGFBP-1 promoter, the signaling pathways that mediate suppression of these genes are distinct.

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Phosphatidylinositol 3′-Kinase and p70s6k Are Required for Insulin but Not Bisperoxovanadium 1,10-Phenanthroline (bpV(phen)) Inhibition of Insulin-like Growth Factor Binding Protein Gene Expression

Cited by 56 publications

References 71 publications

Characterization of a Protein Kinase B Inhibitor In Vitro and in Insulin-Treated Liver Cells

Characterization of a Protein Kinase B Inhibitor In Vitro and in Insulin-Treated Liver Cells

Role of the PI3-kinase/mTor pathway in the regulation of the stearoyl CoA desaturase (SCD1) gene expression by insulin in liver

Insulin Regulation of Insulin-like Growth Factor-binding Protein-1 Gene Expression Is Dependent on the Mammalian Target of Rapamycin, but Independent of Ribosomal S6 Kinase Activity

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