Identification of Kinase-Phosphatase Signaling Modules Composed of p70 S6 Kinase-Protein Phosphatase 2A (PP2A) and p21-activated Kinase-PP2A

Rs, Westphal; Coffee, R. Lane; Marotta, A.; Sl, Pelech; Be, Wadzinski

doi:10.1074/jbc.274.2.687

Cited by 190 publications

(133 citation statements)

References 49 publications

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“…The knockdown experiments revealed that PP2A is the only member of this subfamily that is likely to play a direct role in dephosphorylation of Thr398. A direct role for PP2A in dephosphorylating S6K is consistent with data showing that the catalytic subunit of PP2A can be isolated in complexes with mammalian S6K [20,21] and previous experiments showing that knockdown of PP2A in Drosophila S2 leads to enhanced basal dS6K phosphorylation [18]. While the data are consistent with a direct role of PP2A in dephosphorylation of Thr398, the effect of PP2A dsRNA could also be due to altered phosphorylation of another component of the dTOR pathway.…”

Section: Discussionsupporting

confidence: 90%

“…In vitro fractionation and enzymatic characterization indicate that S6K is dephosphorylated by PP2A [19]. The catalytic subunit of PP2A can be isolated in a complex with S6K following cross-linking of soluble brain extracts [20] or by immunoprecipitation of S6K from Jurkat T cell lysates [21]. Dephosphorylation of S6K following amino acid starvation, rapamycin treatment, or cell stress is blocked by inhibitors with selectivity for the PP2A subfamily [21,22].…”

Section: Introductionmentioning

confidence: 99%

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Functional analysis of the PP2A subfamily of protein phosphatases in regulating Drosophila S6 kinase

Bielinski¹,

Mumby²

2007

Experimental Cell Research

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Phosphorylation and activation of ribosomal S6 protein kinase is an important link in the regulation of cell size by the target of rapamycin (TOR) protein kinase. A combination of selective inhibition and RNA interference were used to test the roles of members of the PP2A subfamily of protein phosphatases in dephosphorylation of Drosophila S6 kinase (dS6K). Treatment of Drosophila Schneider 2 cells with calyculin A, a selective inhibitor of PP2A-like phosphatases, resulted in a 7-fold increase in the basal level of dS6K phosphorylation at the TOR phosphorylation site (Thr398) and blocked dephosphorylation following inactivation of TOR by amino acid starvation or rapamycin treatment. Knockdown of the PP2A catalytic subunit increased basal dS6K phosphorylation and inhibited dephosphorylation induced by amino acid withdrawal. In contrast, depletion of the catalytic subunits of the other two members of the subfamily did not enhance dS6K phosphorylation. Knockdown of PP4 caused a 20% decrease in dS6K phosphorylation and knockdown of PP6 had no effect. Knockdown of the Drosophila B56-2 subunit resulted in enhanced dephosphorylation of dS6K following removal of amino acids. In contrast, knockdown of the homologs of the other PP2A regulatory subunits had no effects. Knockdown of the Drosophila homolog of the PP2A/PP4/PP6 interaction protein α4/Tap42 did not affect S6K phosphorylation, but did induce apoptosis. These results indicate that PP2A, but not other members of this subfamily, is likely to be a major S6K phosphatase in intact cells and is consistent with an important role for this phosphatase in the TOR pathway.

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Section: Discussionsupporting

confidence: 90%

Section: Introductionmentioning

confidence: 99%

Functional analysis of the PP2A subfamily of protein phosphatases in regulating Drosophila S6 kinase

Bielinski¹,

Mumby²

2007

Experimental Cell Research

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“…For example, studies by Heriche et al [12] indicate that casein kinase 2α associates with PP-2A and causes activation of PP-2A. A recent study by Westphal et al [43] indicates that PP-2A forms complexes with p70 S6 kinase, p21-activated kinase-1 (PAK-1) and p21-activated kinase-3 (PAK-3) in i o. The physiological significance of these associations is, however, unknown.…”

Section: Discussionmentioning

confidence: 99%

Role of Janus kinase-2 in insulin-mediated phosphorylation and inactivation of protein phosphatase-2A and its impact on upstream insulin signalling components

Begum

Ragolia

1999

Biochemical Journal

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Our recent studies indicate that insulin rapidly inactivates serine/threonine protein phosphatase-2A (PP-2A) by increasing tyrosine phosphorylation on the catalytic subunit. The exact mechanism of PP-2A inactivation by insulin in vivo is unclear. The Janus kinase (JAK) family of non-receptor protein tyrosine kinases constitute a novel type of signal-transduction pathway which is activated in response to a wide variety of polypeptide ligands, including insulin. In this study we investigated the potential role of JAK-2 in insulin-mediated tyrosine phosphorylation and inactivation of PP-2A using the rat skeletal muscle cell line L6. Co-immunoprecipitation studies revealed that PP-2A is associated with JAK-2 in the basal state. Insulin treatment did not alter JAK-2 association with PP-2A, but did increase JAK-2-mediated tyrosine phosphorylation of the PP-2A catalytic subunit and therefore inhibited PP-2A enzymic activity. Furthermore, PP-2A is associated with phosphoinositide 3-kinase (PI-3K) in the basal state and insulin treatment increases the catalytic activity of PI-3K bound to PP-2A. Pretreatment with AG-490, a specific JAK-2 inhibitor, and SpcAMP, a cAMP agonist, prevented the insulin-mediated increase in (i) JAK-2 kinase activity, (ii) PP-2A tyrosine phosphorylation, (iii) PP-2A inactivation and restored the enzyme activity to control levels, and (iv) PP-2A and JAK-2-associated PI-3K activity. These observations, together with the fact that insulin rapidly activates JAK-2 in L6 cells, and that this is accompanied by an increase in tyrosine phosphorylation of PP-2A in JAK-2 immunoprecipitates, suggest that insulin controls the activation status of PP-2A by tyrosine phosphorylation via JAK-2. PP-2A inactivation may result in an amplification of insulin-generated signals at the level of PI-3K.

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“…Thus, support for the second mechanism has been demonstrated recently in studies using phosphatase-specific inhibitors, which identified protein-serine/ threonine phosphatase 2A (PP2A) as a likely p70 S6K -specific phosphatase. 25,26 Cellular function of p70 S6K …”

Section: Regulation Of P70 S6kmentioning

confidence: 99%

Cellular function of p70^S6K: A role in regulating cell motility

Berven

Crouch

2000

Immunol Cell Biol

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The 70 kDa ribosomal S6 kinase (p70 S6K ) is activated through the phosphoinositide (PI) 3-kinase-regulated pathway (Fig. 1). This pathway is responsible for the generation of inositol lipids, which are key mediators of intracellular signalling. Multiple isoforms of PI 3-kinase have been identified and classed according to substrate specificity and structure. 1 Class I PI 3-kinase is involved in receptor-induced hormonal responses and the mechanism of activation of this enzyme differs depending on the particular extracellular stimuli. For tyrosine kinase-coupled receptor systems, p110 catalytic subunits α, β and δ are stimulated through interaction with the p85 adaptor subunit, which binds selectively through its SH2 domain to phosphorylated tyrosines on the activated receptor. 2 For G-protein-coupled receptor systems, the signalling for PI 3-kinase activation is less clear and several mechanisms have been suggested. A number of reports have identified a p110γ catalytic subunit, which is activated through binding either directly to the βγ subunits of activated trimeric G-proteins 3 or through interaction with a p101 adaptor protein. 4,5 Other studies have suggested alternative mechanisms involving G-protein-mediated activation of receptor tyrosine kinase activity 6 or through the p110β isoform, which can also be activated directly by G-protein βγ subunits. 7 For both receptor systems, PI 3-kinase catalyses the phosphorylation of phosphoinositides at the 3′-OH position. For hormone-induced responses in vivo, phosphatidylinositol 4,5-bisphosphate (PI-4,5P 2 ) is the primary substrate for PI 3-kinase, leading to the production of phosphatidylinositol 3,4,5-trisphosphate (PI-3,4,5P 3 ). Phosphatidylinositol 3,4,5P 3 is also rapidly dephosphorylated by 5′-phosphatases, giving rise to the production of phosphatidylinositol 3,4-bisphosphate (PI-3,4P 2 ). 8 Although the mechanism regulating the relative levels of these two phospholipids is complex, both are thought to act as second messengers in cellular responses such as mitosis, apoptosis, membrane trafficking, motility, differentiation and oncogenic transformation by targeting certain pleckstrin homology (PH) domain-containing proteins to the plasma membrane. 9 Two isoforms of p70 S6K have been identified: a 70 kDa cytoplasmic form and a 85 kDa nuclear form. The sequence of the longer form includes a 23 amino acid nuclear localization sequence at the amino terminus. The molecular sequences of these two isoforms are otherwise the same and can be jointly referred to as p70 S6K or S6K1. Recently, a second functional homologue, S6K2, has been identified. This second S6 kinase shows similar sensitivity to rapamycin and PI 3-kinase inhibitors and is upregulated in p70 S6K1 -deficient mice. 10,11 The identification of S6K2 has raised the possibility that some functional responses of S6K1 may be caused by activation of S6K2 or even other unidentified S6 kinases.Activation of p70 S6K occurs through a complex series of phosphorylation events on eight or more serine or threoni...

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Identification of Kinase-Phosphatase Signaling Modules Composed of p70 S6 Kinase-Protein Phosphatase 2A (PP2A) and p21-activated Kinase-PP2A

Cited by 190 publications

References 49 publications

Functional analysis of the PP2A subfamily of protein phosphatases in regulating Drosophila S6 kinase

Functional analysis of the PP2A subfamily of protein phosphatases in regulating Drosophila S6 kinase

Role of Janus kinase-2 in insulin-mediated phosphorylation and inactivation of protein phosphatase-2A and its impact on upstream insulin signalling components

Cellular function of p70^S6K: A role in regulating cell motility

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Identification of Kinase-Phosphatase Signaling Modules Composed of p70 S6 Kinase-Protein Phosphatase 2A (PP2A) and p21-activated Kinase-PP2A

Cited by 190 publications

References 49 publications

Functional analysis of the PP2A subfamily of protein phosphatases in regulating Drosophila S6 kinase

Functional analysis of the PP2A subfamily of protein phosphatases in regulating Drosophila S6 kinase

Role of Janus kinase-2 in insulin-mediated phosphorylation and inactivation of protein phosphatase-2A and its impact on upstream insulin signalling components

Cellular function of p70S6K: A role in regulating cell motility

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Cellular function of p70^S6K: A role in regulating cell motility