A GSK3β Phosphorylation Site in Axin Modulates Interaction with β-Catenin and Tcf-Mediated Gene Expression

Department of Genetics and Development, College of Physicians and Surgeons, Columbia University, 701 West 168th Street, New York, New York, 10032, USA.
Biochemical and Biophysical Research Communications (Impact Factor: 2.3). 01/2000; 266(1):28-35. DOI: 10.1006/bbrc.1999.1760
Source: PubMed


Upon binding of a Wnt to its receptor, GSK3beta is inhibited through an unknown mechanism involving Dishevelled (Dsh), resulting in the dephosphorylation and stabilization of beta-catenin, which translocates to the nucleus and interacts with Lef/Tcf transcription factors to activate target gene expression. Axin is a scaffold protein which binds beta-catenin and GSK3beta (as well as several other proteins) and thus promotes the phosphorylation of beta-catenin. Here we report that Axin is phosphorylated on Ser and Thr residues in several regions in vivo, while only one region (amino acids 600-672) is efficiently phosphorylated by GSK3beta in vitro. Site-directed mutagenesis, together with in vitro and in vivo phosphorylation assays, demonstrates that Axin residues T609 and S614 are physiological GSK3beta targets. Substitutions for one or more of these residues, which lie within a beta-catenin binding site, reduce the ability of Axin to modulate Wnt-induced signaling in a Lef/Tcf reporter assay. These amino acid substitutions also reduce the binding between Axin and beta-catenin. We propose a model in which inhibition of GSK3beta activity upon Wnt signaling leads to the dephosphorylation of GSK3beta sites in Axin, resulting in the release of beta-catenin from the phosphorylation complex.

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    • "Axin, as a crucial component of canonical Wnt pathway, is also under the regulation of phosphorylation/dephosphorylation. Without Wnt stimulation, Axin is phosphorylated, which enhances its binding affinity with β-catenin, leading to stabilization of Axin (Jho et al., 1999; Yamamoto et al., 1999); upon Wnt stimulation , Axin is dephosphorylated, leading to a less effective binding with β-catenin and consequently Axin degradation (Strovel et al., 2000; Willert et al., 1999). GSK3 is most likely the key kinase that phosphorylates Axin and facilitates its function in " destruction complex " . "
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    ABSTRACT: The Wnt signaling pathway plays crucial roles during embryonic development, whose aberration is implicated in a variety of human cancers. Axin, a key component of canonical Wnt pathway, plays dual roles in modulating Wnt signaling: on one hand, Axin scaffolds the "β-catenin destruction complex" to promote β-catenin degradation and therefore inhibits the Wnt signal transduction; on the other hand, Axin interacts with LRP5/6 and facilitates the recruitment of GSK3 to the plasma membrane to promote LRP5/6 phosphorylation and Wnt signaling. The differential assemblies of Axin with these two distinct complexes have to be tightly controlled for appropriate transduction of the "on" or "off" Wnt signal. So far, there are multiple mechanisms revealed in the regulation of Axin activity, such as post-transcriptional modulation, homo/hetero-polymerization and auto-inhibition. These mechanisms may work cooperatively to modulate the function of Axin, thereby playing an important role in controlling the canonical Wnt signaling. In this review, we will focus on the recent progresses regarding the regulation of Axin function in canonical Wnt signaling.
    Protein & Cell 01/2014; 5(3). DOI:10.1007/s13238-014-0019-2 · 3.25 Impact Factor
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    • ".. Milk protein Donella-Deana et al. (1983) Protein phosphatase-1 G-subunit S38xxxS42xxxS46 Signaling/protein dephosphorylation Dent et al. (1989) Protein phosphatase inhibitor-2 T72xxxS86 Signaling/protein dephosphorylation Aitken et al. (1984), DePaoli-Roach (1984) Axin S322xxxS326xxxS330 Wnt pathway, development, cancer Ikeda et al. (1998), Jho et al. (1999), Yamamoto et al. (1999) "
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    ABSTRACT: Glycogen synthase kinase-3 (GSK-3) is a widely expressed and highly conserved serine/threonine protein kinase encoded in mammals by two genes that generate two related proteins: GSK-3α and GSK-3β. GSK-3 is active in cells under resting conditions and is primarily regulated through inhibition or diversion of its activity. While GSK-3 is one of the few protein kinases that can be inactivated by phosphorylation, the mechanisms of GSK-3 regulation are more varied and not fully understood. Precise control appears to be achieved by a combination of phosphorylation, localization, and sequestration by a number of GSK-3-binding proteins. GSK-3 lies downstream of several major signaling pathways including the phosphatidylinositol 3' kinase pathway, the Wnt pathway, Hedgehog signaling and Notch. Specific pools of GSK-3, which differ in intracellular localization, binding partner affinity, and relative amount are differentially sensitized to several distinct signaling pathways and these sequestration mechanisms contribute to pathway insulation and signal specificity. Dysregulation of signaling pathways involving GSK-3 is associated with the pathogenesis of numerous neurological and psychiatric disorders and there are data suggesting GSK-3 isoform-selective roles in several of these. Here, we review the current knowledge of GSK-3 regulation and targets and discuss the various animal models that have been employed to dissect the functions of GSK-3 in brain development and function through the use of conventional or conditional knockout mice as well as transgenic mice. These studies have revealed fundamental roles for these protein kinases in memory, behavior, and neuronal fate determination and provide insights into possible therapeutic interventions.
    Frontiers in Molecular Neuroscience 11/2011; 4:40. DOI:10.3389/fnmol.2011.00040 · 4.08 Impact Factor
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    • "A fraction of GSK-3 is complexed with the scaffolding proteins adenomatous polyposis coli (APC) and Axin1 (or Axin2), and efficiently phosphorylates β-catenin [ " primed " by casein kinase-1 (CK1)] on a series of N-terminal domain residues earmarking it for polyubiquitination and proteasomal degradation (Doble and Woodgett, 2003). GSK-3 also phosphorylates APC and Axin, increasing their affinities for β-catenin (Hoeflich et al., 2000; Ikeda et al., 1998b; Jho et al., 1999; Yamamoto et al., 1999). Activation of Wnt/β-catenin signalling requires two types of receptors. "
    Embryonic Stem Cells - Basic Biology to Bioengineering, 09/2011; , ISBN: 978-953-307-278-4
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