FRAT-2 Preferentially Increases Glycogen Synthase Kinase 3β-mediated Phosphorylation of Primed Sites, Which Results in Enhanced Tau Phosphorylation

Stoothoff, William H.; Cho, Jae‐Hyeon; McDonald, Roy P.; Johnson, Gail V.W.

doi:10.1074/jbc.m410061200

Cited by 19 publications

(26 citation statements)

References 45 publications

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“…Ser 188 conforms to consensus motifs for phosphorylation by GSK-3, PKA, and PKC, and all of these kinases phosphorylated Ser 188 in vitro. Recently, both FRAT1 and FRAT2 were shown to be phosphorylated in cells (23), and GSK-3 was found to phosphorylate FRAT2 in vitro (32). Our results indicate that GSK-3 phospho-FIGURE 9.…”

Section: Discussionsupporting

confidence: 56%

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FRAT1, a Substrate-specific Regulator of Glycogen Synthase Kinase-3 Activity, Is a Cellular Substrate of Protein Kinase A

Hagen

Cross

Culbert

et al. 2006

Journal of Biological Chemistry

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FRAT1, like itsXenopus homolog glycogen synthase kinase-3 (GSK-3)-binding protein, is known to inhibit GSK-3-mediated phosphorylation of ␤-catenin. It is currently unknown how FRAT-GSK-3-binding protein activity toward GSK-3 is regulated. FRAT1 has recently been shown to be a phosphoprotein in vivo; however, the responsible kinase(s) have not been determined. In this study, we identified Ser 188 as a phosphorylated residue in FRAT1. The identity of the kinase that catalyzes Ser 188 phosphorylation and the significance of this phosphorylation to FRAT1 function were investigated. Protein kinase A (PKA) was found to phosphorylate Ser 188 in vitro as well as in intact cells. Importantly, activation of endogenous cAMP-coupled ␤-adrenergic receptors with norepinephrine stimulated the phosphorylation of FRAT1 at Ser 188 . GSK-3 was also able to phosphorylate FRAT1 at Ser 188 and other residues in vitro or when overexpressed in intact cells. In contrast, endogenous GSK-3 did not lead to significant FRAT1 phosphorylation in cells, suggesting that GSK-3 is not a major FRAT1 kinase in vivo. Phosphorylation of Ser188 by PKA inhibited the ability of FRAT1 to activate ␤-catenin-dependent transcription. In conclusion, PKA phosphorylates FRAT1 in vitro as well as in intact cells and may play a role in regulating the inhibitory activity of FRAT1 toward GSK-3.

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Section: Discussionsupporting

confidence: 56%

“…2, both PKA and PKC phosphorylated wild type Myc-FRAT1. In contrast, no 32 P incorporation into the S188A 188 with human and mouse orthologs and paralogs. Conserved residues are shown in boldface type.…”

Section: Sermentioning

confidence: 94%

FRAT1, a Substrate-specific Regulator of Glycogen Synthase Kinase-3 Activity, Is a Cellular Substrate of Protein Kinase A

Hagen

Cross

Culbert

et al. 2006

Journal of Biological Chemistry

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“…After washing IgG beads containing the immunomobilized GSK3␤ four times with phosphate-buffered saline containing 350 mM NaCl (high salt) and 0.5% Nonidet P-40, the beads were washed two additional times with GSK3 kinase buffer (20 mM Tris-Cl, pH 7.5, 5 mM MgCl 2 , and 1 mM dithiothreitol). GSK3␤ activity assays were performed as described using phosphoglycogen synthase peptide 2 or recombinant tau as substrates (37) with the exception that GSK3␤ activity was normalized to GSK␤ levels in the sample, and the data were expressed as percentages of control (GFP alone). The same immunoprecipitation was performed to detect phospho-Ser 9 GSK3␤ (Cell Signaling) in each condition.…”

Section: Methodsmentioning

confidence: 99%

“…have provided evidence that GSK3-binding proteins, such as FRAT-1 and FRAT-2, which are the proteins involved in the Wnt canonical pathway, affect GSK3␤-mediated tau phosphorylation (23,37,39). Since LRP6 is a GSK3-binding protein that attenuates its activity in in vitro assays, we next examined the effects of LRP6 on GSK3-mediated FIGURE 5.…”

Section: The C Terminus Of Lrp6 Inhibits Gsk3␤-mediated Phosphorylatimentioning

confidence: 99%

The Low Density Lipoprotein Receptor-related Protein 6 Interacts with Glycogen Synthase Kinase 3 and Attenuates Activity

Dolan

Johnson

2006

Journal of Biological Chemistry

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117

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Glycogen synthase kinase 3 (GSK3) is a widely expressed Ser/Thr protein kinase that phosphorylates numerous substrates. This large number of substrates requires precise and specific regulation of GSK3 activity, which is achieved by a combination of phosphorylation, localization, and interactions with GSK3-binding proteins. Members of the Wnt canonical pathway have been shown to influence GSK3 activity. Through a yeast two-hybrid screen, we identified the Wnt canonical pathway co-receptor protein low density lipoprotein receptor-related protein 6 (LRP6) as a GSK3-binding protein. The interaction between the C terminus of LRP6 and GSK3 was also confirmed by in vitro GST pull-down assays and in situ coimmunoprecipitation assays. In vitro assays using immunoprecipitated proteins demonstrated that the C terminus of LRP6 significantly attenuated the activity of GSK3␤. In situ, LRP6 significantly decreased GSK3␤-mediated phosphorylation of tau at both primed and unprimed sites. Finally, it was also demonstrated that GSK3␤ phosphorylates the PPP(S/T)P motifs in the C terminus of LRP6. This is the first identification of a direct interaction between LRP6 and GSK3, which results in an attenuation of GSK3 activity. Glycogen synthase kinase 3 (GSK3)2 is a widely expressed protein kinase with high expression in the brain, and specifically within neurons (for a review, see Ref. 1). GSK3 is a unique Ser/Thr protein kinase that phosphorylates both primed (target Ser/Thr is 4 amino acids N-terminal to a prephosphorylated Ser/Thr) and unprimed (target Ser/Thr is flanked by a Pro) substrates (for a review, see Ref.2). A screen of a rat brain cDNA library revealed that GSK3 is encoded by two independent genes, GSK3␣ and GSK3␤, with molecular masses of 51 and 47 kDa, respectively (3). The two genes display 85% overall sequence identity, which is even higher in the catalytic domain (93%). In the brain, although GSK3␣ mRNA level is higher than GSK3␤, GSK3␤ protein level is higher (4). The poor relationship between transcription and translation in some tissues indicates that these two isoforms are subject to differential regulation, but little is known about the isoform-specific functions.More than 40 proteins have been reported to be phosphorylated by GSK3, including over a dozen transcription factors (reviewed in Ref.2). This large number of substrates illustrates the great potential of GSK3 to affect many cellular functions and suggests that the activity of GSK3 must be carefully regulated by individual mechanisms for each substrate. Although the mechanisms regulating GSK3 are not fully understood, precise control appears to be achieved by a combination of phosphorylation, localization, and interactions with GSK3-binding proteins (reviewed in Ref.2). Protein complexes that contain GSK3 are of major importance in regulating its actions. The best characterized of these complexes is involved in the Wnt canonical pathway, where GSK3-binding proteins control access to the GSK3 substrate, ␤-catenin, and generate a high degree of specifici...

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“…11 Moreover, in contrast to reports that FRAT1 prevents phosphorylation of Tau by GSK3, 29,53 it was recently observed that FRAT2 might in fact promote the GSK3-dependent phosphorylation of primed sites on Tau. 54 Thus, although it remains an attractive hypothesis that Frat allows the discrimination of primed and unprimed GSK3 substrates, this should be further investigated now that the regulation of β-catenin is understood in more detail.…”

Section: Towards Specific Inhibitionmentioning

confidence: 99%

Re-Evaluating the Role of Frat in Wnt-Signal Transduction

Amerongen

Berns

2005

Cell Cycle

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FRAT-2 Preferentially Increases Glycogen Synthase Kinase 3β-mediated Phosphorylation of Primed Sites, Which Results in Enhanced Tau Phosphorylation

Cited by 19 publications

References 45 publications

FRAT1, a Substrate-specific Regulator of Glycogen Synthase Kinase-3 Activity, Is a Cellular Substrate of Protein Kinase A

FRAT1, a Substrate-specific Regulator of Glycogen Synthase Kinase-3 Activity, Is a Cellular Substrate of Protein Kinase A

The Low Density Lipoprotein Receptor-related Protein 6 Interacts with Glycogen Synthase Kinase 3 and Attenuates Activity

Re-Evaluating the Role of Frat in Wnt-Signal Transduction

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