Figure 1. GSK3 signaling pathway
Glycogen synthase kinase 3s are serine/threonine kinases which were originally found as key regulatory enzymes in glucose metabolism. Mammals express two GSK3 isoforms, one is GSK3α (51kDa) and the other is GSK3β (47kDa). They are encoded by different genes, which are about 85% homologous to each other, with 95% identity in the kinase domains. There are two key functional domains of GSK3, a primed-substrate binding domain that recruits substrates to GSK3, and a kinase domain that phosphorylates the substrates. The former domain provides a binding site for major GSK3 substrates (Figure 2A, based on PDB ID: 4nm), those primed by prephosphorylation. The most common target for phosphorylation by GSK3 is the pre-phosphorylated sequence, S/T-X-X-X-S/T(P), where GSK3 phosphorylates a serine/threonine four residues N-terminal to a pre-phosphorylated serine/threonine (Figure 2B, based on PDB ID: 4nm).
Figure 2. Function domains of GSK3β.
GSK3 has an unconventional character of a constitutively active kinase, the substrate of which typically needs to be pre-phosphorylated by another kinase, and it is inhibited rather than activated. It is one of the few signaling mediators that play important roles in a variety of signaling pathways, including those activated by Wnts, hedgehog and G protein-coupled ligands. The answer to GSK3’s actions may be the multiple regulatory mechanisms available to arrange its substrate-specific actions. We categorized these crucial mechanisms to include the regulatory phosphorylation of GSK3 itself, the regulation of substrate availability, the subcellular localization of GSK3 and its substrates, and the incorporation of GSK3 into protein complexes. GSK3 involves in almost every aspect of cellular signaling and is involved in an unparalleled number of disease processes.
The Wnt family of secretory glycoproteins is one of the major families of developmental signaling molecules. They were initially characterized for their roles in regulating embryonic development and tumorigenesis. Studies in the past years have implicated Wnt signaling in a diverse range of physiological and pathophysiological processes, like bone development, angiogenesis, vasculature remodeling, myogenesis, adipogenesis, stem cell renewal and differentiation, and lipid and glucose metabolism.
In the absence of Wnt, many proteins, including AXIN, adenomatous polyposis coli (APC), GSK3, casein kinase Ia (CKIa), and β-catenin, form a complex (β-catenin destruction complex). Among them, β-catenin is phosphorylated by CKIa and GSK3. This phosphorylation event targets β-catenin for proteasome-mediated proteolytic degradation. When Wnt proteins bind the cell-surface receptors, frizzled (FZD) and lipoprotein receptor-related protein (LRP) 5/6, GSK3-dependent β-catenin phosphorylation is suppressed through a mechanism that requires the scaffold protein dishevelled (DVL), and β-catenin is stabilized. Stabilized β-catenin enters the nucleus then interacts with transcriptional regulators- lymphoid enhancing factor-1 (LEF1) and T cell factors (TCFs), to activate gene transcription.
Insulin signaling pathway
Insulin promotes the conversion of glucose to glycogen in skeletal muscle by stimulating glucose uptake and activating glycogen synthase. Insulin activates glycogen synthase by inducing its dephosphorylation at a cluster of C-terminal residues (Ser641, Ser645, Ser649 and Ser653), which are phosphorylated by glycogen synthase kinase-3α (GSK-3α) and GSK-3β. In muscle, insulin is thought to stimulate the dephosphorylation of glycogen synthase at these residues by inducing the inactivation of GSK-3α and GSK-3β via phosphorylation of an N-terminal Ser residue (Ser21 in GSK3α and Ser9 in GSK-3β), which is catalysed by Akt and reversed by the muscle glycogen-associated protein phosphatase-1. Consistent with this model of regulation, small cell-permeable inhibitors of GSK3 with diverse structures stimulate glycogen synthase activity in cell lines or skeletal muscle. Type 2 diabetes is the first disease condition associated with GSK-3β, due to its negative regulation of several aspects of insulin signaling pathway. In this pathway 3-phosphoinositide-dependent protein kinase activates AKT which in turn, inactivates GSK-3β. This inactivation of GSK3-β leads to the dephosphorylation and activation of glycogen synthase which helps glycogen synthesis.
For stem cell maintenance and tissue patterning in embryos, there are key signaling regulators named Hedgehogs (Hhs) which activate mutations in the pathways that increase GlI transcriptional activity and are causal in a diversity of cancers.
Desert hedgehog (DHH), Indian hedgehog (IHH) and sonic hedgehog (SHH) are three Hh genes. These Hhs function as ligands for the 12- pass transmembrane receptor patched (PTCH1) which controls many developmental signaling events. Hh ligands bind their receptor PTCH1. This will cause its internalization and degradation and release of Smoothened (SMO), a GPCR. SMO then promotes the dissociation of a suppressor of fused (SUFU)- glioma-associated oncogene homologue (GLI) complex. This makes the transcription factors GLI1 and GLI2 to translocate to the nucleus and ultimately activate the transcription of certain genes. In contrast, GLI3 is degraded during this process as it normally works as a repressor. GLI3 can be phosphorylated by PKA, GSK-3 and CK1 which results in GLI3 becoming a transcriptional repressor. Activated GLI proteins stimulate the transcription of Hh target genes, including: GLI1, GLI2, PTCH1, CCND1, IGF2, MYCN, and BCL2. In some studies IGF-2 has been determined to subsequently activate the PI3K/PTEN/Akt/mTOR pathway and influence proliferation and survival as well as inhibit GSK-3β. The Hh pathway is regulated by GSK-3. This pathway is inhibited when GSK-3 is suppressed. GSK-3β is a binding partner of SUFU. SUFU suppresses GLI activity by sequestering GLI in the cytoplasm. When GSK-3 phosphorylates SUFU, Hh pathway activity is increased. Activated Hh pathway stimulates GSK-3β to phosphorylate SUFU which induces the dissociation of SUFU from GLI3. GSK-3β regulates the Hh pathway by phosphorylating GLI. The Hh pathway activates the GLI transcription factor by suppressing the function of SUFU. SUFU recruits GSK-3β into a trimolecular complex of GLI3/SUFU/GSK-3β. GSK- 3β then phosphorylates GLI3 which results in a SUFU/ GLI3 transcriptional repressor. Shh then dissociates the GLI3/SUFU/GSK-3β complex from GLI3 which prevents GLI3 processing and results in GLI3 becoming a transcriptional activator instead of a transcriptional repressor.
Interactions with G protein-coupled signal transduction
Dopaminergic D2 receptor activation induces the association of β-arrestin, Akt, GSK3, and protein phosphatase 2A (PP2A). This facilitates PP2A-mediated dephosphorylation of Akt and GSK3, deactivating Akt and activating GSK3. GSK3 induces the formation of this complex, which provides a mechanism for GSK3 to induce its own activation. Self-activation of GSK3 is also exemplified by its phosphorylation of the protein phosphatase 1 (PP1) inhibitor I-2, resulting in increased PP1 activity, which dephosphorylates the inhibitory serine-phosphorylation of GSK3 to increase GSK3 activity. Activated serotonin (5HT) 2A receptor reduces serine-phosphorylation of GSK3, thereby increasing its activity. Activation of 5HT1A and Cholinergic muscarinic receptors increases the inhibitory serine-phosphorylation of GSK3. GSK3 promotes 5HT1B receptor-mediated activation of the heterotrimeric G protein, Gi, which inhibits cyclic AMP production.
Relationship with diseases
With its actual participation in a number of signaling pathways associated with disease pathology, GSK3 is being considered as a therapeutic target in many disorders.
Psychiatric diseases are likely to be one of the major classes that are amenable to GSK3 inhibitor therapeutics. Patients with bipolar disorder are already being effectively treated with lithium and it is no doubt that lithium is a GSK3 inhibitor. Although the therapeutic mechanism of action of lithium remains unclear, substantial evidence indicates that inhibition of GSK3 is a key component of lithium’s emotional stability.
GSK3 is likely to be therapeutic for a number of neurological disorders. The evidence is particularly strong for Alzheimer’s disease, which has been well discussed in multiple reviews. In Alzheimer’s disease, GSK3 has been proved to promote every major pathological process, including amyloid β peptide production and tau phosphorylation, which leads to the two hallmark pathologies of Alzheimer’s disease, amyloid plaques and neurofibrillary tangles, respectively. GSK3 inhibitors improve many cognitive functions in rodent models of Alzheimer’s disease, which is a key outcome since dementia is the defining characteristic of Alzheimer’s disease.
Diseases involving inflammation in the periphery have also been shown to benefit from the anti-inflammatory effects of GSK3 inhibitors. Interestingly, administration of GSK3 inhibitors was effective to afford mice protection (survival) from an otherwise lethal dose of the inflammatory stimulant lipopolysaccharide used to model sepsis. Administration of GSK3 inhibitors also reduced inflammation and pathology in rodent models of asthma, arthritis, colitis, and peritonitis. Thus, GSK3 inhibitors provide an effective intervention for all inflammatory diseases in which they have been tested in rodent models
|1.||Wu D, Pan W. GSK3: a multifaceted kinase in Wnt signaling. Trends in biochemical sciences. 2010;35(3):161-168.|
|2.||Hur E-M, Zhou F-Q. GSK3 signaling in neural development. Nature reviews Neuroscience. 2010;11(8):539-551.|
|3.||McCubrey JA, Steelman LS, Bertrand FE, et al. GSK-3 as potential target for therapeutic intervention in cancer. Oncotarget. 2014;5(10):2881-2911.|
|4.||Beurel E, Grieco SF, Jope RS. Glycogen synthase kinase-3 (GSK3): regulation, actions, and diseases. Pharmacology & therapeutics. 2015;0:114-131.|
|5.||Riobó NA, Lu K, Ai X, et al, Emerson CP. Phosphoinositide 3-kinase and Akt are essential for Sonic Hedgehog signaling. Proceedings of the National Academy of Sciences of the United States of America. 2006;103(12):4505-4510.|
|6.||Phukan S, Babu V, Kannoji A, et al. GSK3β: role in therapeutic landscape and development of modulators. British Journal of Pharmacology. 2010;160(1):1-19.|
|7.||McManus EJ, Sakamoto K, Armit LJ, et al. Role that phosphorylation of GSK3 plays in insulin and Wnt signalling defined by knockin analysis. The EMBO Journal. 2005;24(8):1571-1583.|