Edited by: Richard Scott Jope, University of Alabama at Birmingham, USA
Reviewed by: Urs Albrecht, University of Fribourg, Switzerland; Hagit Eldar-Finkelman, Tel Aviv University, Israel
*Correspondence: Oksana Kaidanovich-Beilin, Samuel Lunenfeld Research Institute, Mount Sinai Hospital, 600 University Avenue, Room 983, Toronto, ON, Canada M5G 1X5. e-mail:
This is an open-access article subject to a non-exclusive license between the authors and Frontiers Media SA, which permits use, distribution and reproduction in other forums, provided the original authors and source are credited and other Frontiers conditions are complied with.
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.
Glycogen synthase kinase-3 (ATP:protein phosphotransferase, E.C. 2.7.1.37) is a serine/threonine protein kinase, belonging to the CMCG family of proline-directed kinases (Cyclin-dependent kinases (CDKs), Mitogen-activated protein kinases (MAPKs), Glycogen synthase kinases (GSKs), and CDK-like kinases (CLKs). GSK-3 is a monomeric, second messenger-independent protein kinase that was first discovered through its ability to activate the ATP–Mg-dependent form of type-1 protein phosphatase (“Factor A”) and to phosphorylate the key rate-limiting metabolic enzyme that catalyzes the last step of glycogen synthesis, glycogen synthase (GS; Embi et al.,
GSK-3 is a highly conserved protein kinase and has orthologs in plants, fungi, worms, flies, sea squirts, and vertebrates: isoenzymes from species as distant as flies and humans display more than 90% sequence similarity within the protein kinase domain (reviewed in Ali et al.,
GSK-3 is expressed ubiquitously and both gene products are found in virtually all mammalian tissues. The kinase is highly expressed in the brain (Woodgett,
In certain cell types of the brain, alternative splicing between exon 8 and 9 of GSK-3β leads to the generation of an additional “long” form containing a 13 amino acid insert within the catalytic domain (GSK-3β2; see Figure
An unusual feature of GSK-3 is that the kinase displays high activity in cells under resting/unstimulated conditions (Sutherland et al.,
GSK-3 is dual specificity kinase differentially regulated by tyrosine and serine/threonine phosphorylation. The activity of GSK-3 is positively regulated by phosphorylation on a “T loop” tyrosine residue within subdomain VIII (Tyr279 for GSK-3α and Tyr216 for GSK-3β; Hughes et al.,
From the crystal structure, it has been proposed that unphosphorylated Tyr276/Tyr216 act to block the access of primed substrates (as discussed below). Indeed, the structure of phosphorylated GSK-3β (Bax et al.,
In contrast to tyrosine phosphorylation, regulation of N-terminal serine phosphorylation is only conserved in GSK-3 homologs from mammals,
This inhibitory mechanism is induced by agonists such as neurotrophins and growth factors that activate protein kinases that act on the N-terminal domain of GSK-3 such as PKB/Akt, p90rsk, cyclic-AMP-dependent protein kinase, p70 S6 kinase, as well as regulators of phosphatase-1 (Sutherland et al.,
Growth factors, such as EGF and PDGF can also inhibit GSK-3 activity through the phosphatidylinositol 3′ kinase (PI3K) pathway (Stambolic and Woodgett,
Wnts are secreted glycolipoproteins that activate canonical and non-canonical (β-catenin independent) Wnt signaling cascades, which are essential for early embryonic patterning, cell fate, cellular polarity, cell movement, cell proliferation as well as adult homeostasis in both vertebrates and invertebrates (Logan and Nusse,
In cells a small fraction (<5–10%) of cellular GSK-3 is associated with a scaffolding protein termed Axin (Lee et al.,
Wnt ligand induces binding of the seven-pass transmembrane receptor Frizzled and the LRP5/6 co-receptor which leads to the recruitment of Dvl and induction of LRP 5/6 phosphorylation by GSK-3 and CK1. This creates a high affinity binding site for Axin (He et al.,
β-catenin is dephosphorylated primarily by protein phosphatase (PP) 2A (Su et al.,
Several molecular mechanisms have been proposed to explain how canonical Wnt signaling may interfere with GSK-3-dependent phosphorylation of β-catenin (reviewed in Kimelman and Xu,
Both mammalian isoforms of GSK-3 function equivalently in Wnt signaling and are entirely redundant (Doble et al.,
GSK-3 can also associate with other proteins, e.g., GSK-3-binding protein (GBP or FRAT; Li et al.,
In addition to binding proteins in the cytoplasm, there are differences in patterns of subcellular localization of the GSK-3 isozymes (Hoshi et al.,
The finding that GSK-3 acts downstream of multiple signaling pathways that have distinct effects on cells and tissues presents a conundrum. How might signal selectivity be achieved if a protein common to multiple pathways was a required intermediary? The elegant cellular solution to this is to fractionate GSK-3 between scaffolding proteins or other structures such that each system has its own population of GSK-3 molecules “assigned” to it. This effectively insulates the signals and requires that the GSK-3 subpopulations do not intermingle or exchange. It is still an open question why so many important pathways evolved with a common component, a subject of speculative commentary (McNeill and Woodgett,
The determination of the crystal structure of GSK-3β provided further insight into the molecular nature of the regulation of GSK-3 and its predilection for primed, pre-phosphorylated, substrates (Dajani et al.,
GSK-3 is one of only a handful of the over 500 known protein kinases that has a strong (500- to 1000-fold) preference for substrates that are already primed by phosphorylation at a proximal serine/threonine to the GSK-3 target residue (Thomas et al.,
The majority of GSK-3 substrates exhibit an absolute requirement for prior phosphorylation by another kinase at a “priming” residue located C-terminal to the site of subsequent phosphorylation by GSK-3 (Fiol et al.,
To prove that an
Substrate name | GSK-3′s phosphorylation site (in bold) |
Biological process/relevance to disease | Reference |
---|---|---|---|
ATP-citrate lyase | Fatty acid biosynthesis | Hughes et al. ( |
|
Glycogen synthase | Glycogen metabolism, diabetes | Rylatt et al. ( |
|
Pyruvate dehydrogenase | … |
Metabolism | Hoshi et al. ( |
Phosphocholine cytidylyltransferase (CTP) | C-terminus |
CDP-choline pathway | Cornell et al. ( |
κ-casein | …?… | Milk protein | Donella-Deana et al. ( |
Protein phosphatase-1 G-subunit | Signaling/protein dephosphorylation | Dent et al. ( |
|
Protein phosphatase inhibitor-2 | T72xxxS86 | Signaling/protein dephosphorylation | Aitken et al. ( |
Axin | Wnt pathway, development, cancer | Ikeda et al. ( |
|
Axil | Member of axin family | Yamamoto et al. ( |
|
Adenomatous polyposis coli (APC) | Wnt pathway, development, cancer | Rubinfeld et al. ( |
|
Cubitus interruptus/Gli | Hedgehog pathway, development, cancer | Jia et al. ( |
|
elF2B (Eukaryotic initiation factor 2B) | Growth, cancer | Welsh et al. ( |
|
Amyloid precursor protein (APP) | Neuronal function, Alzheimer’s disease | Aplin et al. ( |
|
Presenilin-1 | Alzheimer’s disease | Kirschenbaum et al. ( |
|
Heterogeneous nuclear ribonucleoprotein D | Transcriptional regulator | Tolnay et al. ( |
|
Phosphatase interactor targeting protein K (PITK) | PP-1 targeting subunit, modulates phosphorylation of hnRNP K | Kwiek et al. ( |
|
p21 CIP1 | Cell cycle, apoptosis | Rössig et al. ( |
|
Insulin receptor substrate 1 | Diabetes, growth, cancer | Eldar-Finkelman and Krebs ( |
|
Insulin receptor substrate 2 | Diabetes | Sharfi and Eldar-Finkelman ( |
|
P75 NGF receptor | …?… | Neurotrophin signaling | Taniuchi et al. ( |
Mcl-1 | Growth, apoptosis, cancer | Maurer et al. ( |
|
Cyclic-AMP-dependent protein kinase – RII subunit | cAMP pathway, hormonal responses | Hemmings et al. ( |
|
Cyclin D1 | Cell cycle, cancer | Diehl et al. ( |
|
Cyclin E | Cell cycle | Welcker et al. ( |
|
Myelin basic protein | Myelination of nerves in CNS | Yu and Yang ( |
|
Cry2 | Circadian rhythm | Harada et al. ( |
|
Per2 | …?… | Circadian rhythm | Kaladchibachi et al. ( |
Nucleoporin p62 | C-terminus |
Cell division | Miller et al. ( |
PTEN (phosphatase and tensin homolog) | Cell proliferation, migration and growth, cell survival | Al-Khouri et al. ( |
|
Lipoprotein receptor-related protein 6 | Wnt pathway, development., cancer | Zeng et al. ( |
|
TSC2 (tuberous sclerosis 2)/Tuberin | Tumor suppressor | Inoki et al. ( |
|
CDC25 (cell division cycle) | Cell cycle, cancer | Kang et al. ( |
|
RBL2/p130 | Growth, cell cycle, cancer | Litovchick et al. ( |
|
Voltage dependent anion channel | Apoptosis, cancer | Pastorino et al. ( |
|
CTPS (cytidine triphosphate synthetase | Cell growth, cancer | Higgins et al. ( |
|
Mdm2 (murine double minute 2) | Growth | Kulikov et al. ( |
|
Calcipressin/RCN1 (regulators of calcineurin) | Neuronal regulator of calcineurin, growth, Alzheimer’s disease, down syndrome | Hilioti et al. ( |
|
Gephyrin | GABA transmission, neuronal functions | Tyagarajan et al. ( |
|
Mixed lineage kinase-3 | c-Jun and p38 MAPK pathways, apoptosis, neurodegenerative disease | Mishra et al. ( |
|
OMA-1 | Oocyte maturation | Nishi and Lin ( |
|
p27Kip1 | Cell cycle regulator | Surjit and Lal ( |
|
Polypyrimidine track-binding protein-associated-splicing factor (PSF) | Alternative splicing, T cell function/activation | Heyd and Lynch ( |
|
Zinc finger CCHC domain-containing protein 8 (Zcchc8) | RNA metabolism | Gustafson et al. ( |
|
SC35 | Alternative splicing | Hernandez et al. ( |
|
DF3/MUC1 (high molecular weight mucin-like glycoprotein) | Wnt pathway, β-catenin, and E-cadherin complex | Li et al. ( |
|
Dynamin I | Endocytosis, neuronal function | Hong et al. ( |
|
Kinesin light chains (KLCs) | Axonal transport, mitosis, meiosis | Morfini et al. ( |
|
Microtubule-associated protein 1B | Neuronal functions | Lucas et al. ( |
|
Microtubule-associated protein 2C | Neuronal functions | Sánchez et al. ( |
|
Tau | Microtubule stabilization, neuronal functions, Alzheimer’s disease |
Hanger et al. ( |
|
Paxillin | Cell adhesion and migration | Cai et al. ( |
|
Collapsin response mediator protein 2 | Neuronal functions, axonal growth, neuronal polarity, Alzheimer’s disease | Cole et al. ( |
|
Collapsin response mediator protein 4 | Neuronal function, axonal growth | Cole et al. ( |
|
Neural cell adhesion protein (NCAM) | …?… | Cell–cell adhesion, neurite outgrowth, synaptic plasticity, learning, and memory | Mackie et al. ( |
Neurofilament L | Cell cytoskeleton, axonal growth, axonal diameter | Guan et al. ( |
|
Neurofilament M | |||
Neurofilament H | |||
Ninein | …?… | Centrosomal functions, brain development, tumorigenesis | Hong et al. ( |
Telokin kinase-related protein | Stabilization of smooth muscle myosin filaments | Krymsky et al. ( |
|
CLIP-associated protein 1 (CLASP 1) | … |
Neuronal functions | Wittmann and Waterman-Storer ( |
CLASP 2 (CLIP-associated protein 2) | Cell migration, neuronal function | Watanabe et al. ( |
|
Focal adhesion kinase | Cell cycle, survival, migration, cancer | Bianchi et al. ( |
|
Microtubule affinity-regulating kinase-2/PAR-1 | Neuronal function, axonal growth | Kosuga et al. ( |
|
Polycystin 2 | Growth, survival, polycystic kidney disease | Streets et al. ( |
|
Dystrophin | Cytoskeleton of muscle fibers | Michalak et al. ( |
|
Stathmin/oncoprotein 18 (STMN1) | Microtubule polymerization and dynamics | Moreno and Avila ( |
|
von Hippel–Lindau (VHL) | Oxygen sensor, tumor of CNS, kidney, eyes | Hergovich et al. ( |
|
β-catenin | Wnt pathway, development, cancer | Yost et al. ( |
|
δ-catenin | Cell adhesion | Oh et al. ( |
|
CCAAT/enhancer-binding protein αC/EBPα) | Cell proliferation, growth, differentiation | Ross et al. ( |
|
C/EBPβ (CCAAT/enhancer-binding protein β) | Cell proliferation, growth, differentiation | Tang et al. ( |
|
Cyclic AMP response element-binding protein (CREB) | Metabolism, neuronal function, memory formation, diabetes | Fiol et al. ( |
|
GATA4 | …2–116… |
Embryogenesis, myocardial differentiation and function | Morisco et al. ( |
Hypoxia-inducible factor-1 | Growth, cancer | Mottet et al. ( |
|
Heat shock factor-1 (HSF1) | Stress (heat) response | Chu et al. ( |
|
c-Myc | Growth, cancer, oncogenes | Saksela et al. ( |
|
L-myc | |||
N-myc downstream regulated gene 1 | Stress and hormone response, cell growth and differentiation, cancer | Murray et al. ( |
|
c-Jun, Jun B, Jun D | Growth and cancer | Boyle et al. ( |
|
c-Myb | Hematopoiesis, tumorigenesis. | Kitagawa et al. ( |
|
Nuclear factor of activated T cells c (NFATc) | SP2 domain |
Immune system response | Beals et al. ( |
Nuclear factor κB (NF-κB) |
Stress response, immune response, synaptic plasticity and memory, cancer, inflammation, autoimmune diseases | Demarchi et al. ( |
|
Notch 1C | …?… | Development, cell–cell communication, cancer. | Foltz et al. ( |
p53 | Cell cycle regulator, cancer. | Turenne and Price ( |
|
Snail | Epithelial to mesenchymal transition regulator | Zhou et al. ( |
|
Activator protein 1 (AP-1) | …?… | Differentiation, proliferation, apoptosis | de Groot et al. ( |
Glucocorticoid receptor | Stress and immune response, development, metabolism | Rogatsky et al. ( |
|
Microphthalmia-associated transcription factor | Melanocyte and osteoclast development | Takeda et al. ( |
|
NeuroD | …?… | Central nervous system development | Moore et al. ( |
BCL-3 | Growth and cancer | Viatour et al. ( |
|
Bmal1 | Circadian rhythm | Sahar et al. ( |
|
Rev-erb α | Circadian rhythm | Yin et al. ( |
|
Timeless | …?… | Circadian rhythm | Martinek et al. ( |
Clock | Circadian rhythm | Spengler et al. ( |
|
SMAD1 | …?… | Embryonic pattern formation, TGFβ signaling | Fuentealba et al. ( |
SMAD3 | TGFβ signaling, development, cancer | Guo et al. ( |
|
Neurogenin 2 (Ngn2) | Neuronal function, motor neurons, development | Ma et al. ( |
|
BCLAF1 (Bcl-2 interacting transcriptional repressor) | Apoptosis, cancer | Linding et al. ( |
|
Myocardin | Development, cardiac hypertrophy | Badorff et al. ( |
|
Histone H1.5 | Chromosome condensation | Happel et al. ( |
|
Nascent polypeptide associated complex | Transcriptional coactivator, bone development. | Quelo et al. ( |
|
Nuclear factor E2-related factor 2 | Antioxidant response, cell survival | Salazar et al. ( |
|
SKN-1 | Oxidative stress, detoxification | An et al. ( |
|
Sterol regulatory element-binding protein | Lipid and cholesterol metabolism | Sundqvist et al. ( |
|
MafA/RIPE3β1 | Regulates insulin gene expression in β cells of pancreas, pancreatic development | Han et al. ( |
With respect to biological processes, GSK-3 substrates may be classified into several groups of proteins/transcriptional factors/regulatory enzymes that have roles in processes such as metabolism, cellular architecture, gene expression, neurobiological processes, synaptogenesis, neurodevelopment, axonal growth and polarity, immune response, circadian rhythms, and neuronal/cellular survival (reviewed in Frame and Cohen,
Circadian (from the Latin
From a molecular standpoint, circadian rhythms are regulated by transcriptional and post-translational feedback loops generated by a set of interplaying “clock” proteins. The positive limb of the mammalian clock machinery is comprised of CLOCK and BMAL1, which are transcription factors that heterodimerize through their PAS domains and induce the expression of clock-controlled genes by binding to their promoters at E-boxes. Cryptochromes (Cry 1, Cry2) and Period genes (Per1, Per2, Per3) are clock-controlled genes that encode proteins that form the negative limb of the circadian machinery. PER and CRY proteins are classically thought to translocate into the nucleus to inhibit CLOCK:BMAL1 mediated transcription, thereby closing the negative feedback loop (reviewed in Sahar and Sassone-Corsi,
GSK-3 is expressed in the primary center of circadian rhythm regulation – the suprachiasmatic nucleus (SCN) of hypothalamus (Iitaka et al.,
Lithium has been shown to lengthen the period of circadian rhythms in a wide range of experimental systems, including unicellular organisms, insects, mice, and humans (Abe et al.,
GSK-3 has also been demonstrated to phosphorylate and regulate the stability of “core” circadian rhythm genes in mammals. GSK-3 together with another serine kinase, DYRK1A, phosphorylates CRY2 at Ser 557 and 553 (respectively) resulting in degradation of CRY2 (Harada et al.,
Several genetic approaches have been used to generate mutant mice for GSK-3: conventional knockouts and knock-ins (all tissues), conditional knockouts (tissue-specific), and transgenic mice (Table
Type of approach | Mouse design | Mouse name | Characterized by (reference) |
---|---|---|---|
Knockout | Deletion of exon 2 (ATP-binding loop) of GSK-3α | GSK-3α KO | MacAulay et al. ( |
Knockout | Deletion of exon 2 (ATP-binding loop) of GSK-3β | GSK-3β KO | Hoeflich et al. ( |
Knockout | Deletion of exon 2 (ATP-binding loop) of GSK-3β | GSK-3β HET | Hoeflich et al. ( |
Knock-in | Mutations GSK-3 αS21A, βS9A | GSK-3α, β [S21A,S9A] KI | McManus et al. ( |
Double shRNA knockdown | GSK-3α/β shRNA (shGSK-3-dh+/flox) × Nestin-Cre | Nestin-Cre/shGSK-3-dh+/Δ | Steuber-Buchberger et al. ( |
Conditional knockout | GSK-3α/GSK-3β flox/flox × Nestin-Cre | Nestin-GSK-3α + β KO | Kim et al. ( |
Double shRNA knockdown | GSK-3β shRNA | DG–GSK-3β knockdown (shRNA) | Omata et al. ( |
Dominant-negative (DN) GSK-3β | K85RGSK-3β × CamkII-tTA-Cre | DN-GSK-3β | Gomez-Sintes et al. ( |
Overexpression of GSK-3β | TetO GSK-3β × CamkII-tTA-Cre | Tet/GSK-3β | Lucas et al. ( |
Overexpression of constitutively active GSK-3β [S9A] | S9AGSK-3β in Thy-1 gene vector | GSK-3β [S9A] | Spittaels et al. ( |
Overexpression of GSK-3β | PrpGSK-3βL56 and PrpGSK-3βL64 | O’Brien et al. ( |
The first GSK-3 gene to be knocked out was GSK-3β (Hoeflich et al.,
GSK-3β heterozygous (HET) mice are viable, morphologically normal and have been tested extensively. These animals exhibit a lithium-mimetic, anti-depressant-like state (Beaulieu et al.,
In contrast to GSK-3β null mice, animals lacking GSK-3α are viable and exhibit improved insulin sensitivity and hepatic glycogen accumulation on the ICR background (MacAulay et al.,
Two studies have employed shRNA knockdown to suppress expression of the two GSK-3 genes in mouse brain. Nestin-Cre was employed to drive shRNA expression in the brain progenitor compartment by excising LoxP flanked transcriptional stop sites. This approach resulted in partial reduction of GSK-3α and β protein levels (60 and 50%, respectively) in whole brain lysate (Steuber-Buchberger et al.,
In an alternative approach, lentivirus-expressing short-hairpin RNAs targeting GSK-3β were injected bilaterally into the hippocampus to inactivate GSK-3β in the dentate gyrus (Omata et al.,
Dominant-negatively acting mutants interfere with the endogenous proteins by soaking up downstream targets or upstream regulators. This approach has been used to generate conditional transgenic expression of a dominant-negative (DN) form of GSK-3β in the brain (Gomez-Sintes et al.,
Overexpression of GSK-3β has been postulated to be embryonic lethal as viable transgenic animals show only modest levels of the exogenously engineered gene (Brownlees et al.,
Transgenic mice have also been generated that overexpress a mutant form of GSK-3β in which the inhibitory N-terminal phosphorylation site is mutated [S9A] (Spittaels et al.,
As mentioned above, mutation of the N-terminal phosphorylation sites of GSK-3 renders the protein kinase insensitive to inhibition by that mode of regulation (although the kinase remains sensitive to Wnt regulation, for example). Mice have been generated in which the phosphorylation sites of the endogenous alleles have been replaced by non-phosphorylatable alanine (GSK-αS21A, βS9A). Since serine phosphorylation of GSK-3 is increased by lithium, anti-psychotic drugs, anti-depressants, etc., this model is attractive to use for studying the mechanism of action of aforementioned drugs and related pathological conditions. GSK-αS21A, βS9A knock-in mice have normal development and growth, with no signs of metabolic abnormalities/insulin resistance (McManus et al.,
GSK-αS21A, βS9A knock-in mice exhibited increased susceptibility to hyperactivity and a heightened response to a novel environment (Polter et al.,
Alleles of GSK-3α and β have been generated in which exon 2 (containing essential residues for ATP-binding) are flanked by LoxP (flox) sites (Figure
Ten different GSK-3 animal models have been described to date (Table
Behavioral test | Type of test | GSK-3α KO | GSK-3α HET | GSK-3β HET | GSK-3αβ knock-in | GSK-3β [S9A] overexpression | DG–GSK-3β knockdown (shRNA) |
---|---|---|---|---|---|---|---|
Anxiety | EPM | ▲ In females only (1) | ▲ (2, 3) | Mild ▲ (6) | |||
O-maze | ▼ (11) | ||||||
Light–dark box | ≠ In males (1) | ||||||
Emotionality | Open field 5 min | ▲ (Both genders) (1) | |||||
Locomotor activity | Open field 30 min | ▼ (1) | ≠ (10) | ≠ (4, 5, 2) | ▲ (6, 11) | ▲ (8) | ≠ (12) |
Exploratory activity/curiosity | Open field 30 min | ▼ (1) | ▼ (2, 5) | ≠ (6) | ▲ (8) | ||
Light–dark box | ▲(11) | ||||||
Depression-like behavior | Learned helplessness | Stress-induced ▲ (6) | |||||
FST | ▼ (1) | ▼ (2, 4, 5) | ▲ (6), ▼ (11) | ▼ (8) | ▼ (12) | ||
TST | ▼ (1) | ▼(2, 4, 5) | ▲ (6) | ▼ (12) | |||
Stress reactivity | Stress response▲ (1), ▼corticosteroids (1) | General ▼ (11) ≠ (6), stress response ▲(6), ▲body weight, food, and fluid intake (11), ▼ (11) ≠ (6), corticosteroids, ▼ACTH (11) | ≠ Corticosteroids (8) |
Type of behavior | Type of the test | GSK-3α KO | GSK-3β HET | GSK-3αβ KI |
---|---|---|---|---|
Sociability and social novelty | Social interaction test | ▼ (1) | Impaired social preference to Str2 (7) | |
Aggression | Resident intruder | ▼ (1) | ▼ (Beaulieu et al., |
The differences in behavioral results between different GSK-3 animal models may be accounted for by the alternative design of the models, and/or by strain and gender differences, varying methodology, and animal house-keeping environment.
Well described and presented is effect of genetic GSK-3 manipulations on depressive-like phenotype in mice (Table
It is important to mention that different GSK-3 animal models have employed different “Cre” promoters. Activation of specific “Cre” recombinases may happen at different stages of embryogenesis (or after birth), thus may affect specific neuronal populations (post-mitotic or precursors), which may affect structure and function of adult brain. For example, dominant-negative GSK-3β and Tet/GSK-3β mice have been generated by using CamkIIα-Cre, compare to GSK-3β S9A mice which have been created by using Thy-1 gene promoter (Table
Moreover, there are different approaches have been used to generate mice with overexpression of GSK-3β gene. In all three models with overexpression of GSK-3β, different constructs for GSK-3β gene itself were used (Table
Comparison between GSK-3α KO and GSK-3α + β serine phosphorylation site KI mice revealed similar impaired sociability in both models, despite different genetic approaches being used. These data indicate that both the protein level of GSK-3α as well as serine phosphorylation of GSK-3 are important aspects for neuronal circuits responsible for social interaction.
Moreover, studying and analyzing genetic animal models may be used to make predictions about long-term usage of GSK-3 inhibitors (as therapeutic agents). For example, the well characterized GSK-3 inhibitor – lithium – has a diverse spectrum of effects after long-term treatment of patients, including tremor and death of Purkinje cells. Of note, similar changes in cerebellar structure and function were observed in GSK-3α KO and dominant-negative GSK-3β mice (Tables
Type of behavior | Type of the test | GSK-3α KO | GSK-3α HET | GSK-3β HET | GSK-3αβ KI | GSK-3β [S9A] overexpression | Tet/GSK-3β | DN–GSK-3β |
---|---|---|---|---|---|---|---|---|
Amphetamine response | OF + amp | ▲ | ▼ (4) | ▲ (6) | ▼ (16) | |||
Response to morphine | OF + morph | ▲ (23) | ||||||
Information processing | PPI/ASR | ▲ PPI (1) | ≠ (10) | ≠ (2, 5) | ▲ASR (8) | |||
LI | ▲ NPE in KO (1) | ≠ (10) | ||||||
Long-term memory | FC | ▼ (1) | Impaired memory reconsolidation (9) | ▲ (6), Enhanced emotion-associated memory | ||||
Passive avoidance | ≠ (1) | |||||||
LTP/LTD | abnormal LTD in the ventral hippocampus (6) | ▼ (20) | ||||||
Spatial memory | Impaired (22) | |||||||
Coordination, balance | Rotarod | ▼ (1) | ≠ (2, 5) | ▼ (16) | ||||
Motor learning | Rotarod | ≠ (1) | ≠ (2, 5) |
Type of analysis | Type of test | GSK-3α KO | Tet/GSK-3β | GSK-3αβ KI | GSK-3β [S9A] overexpression | Nestin-GSK-3αβ KO | DN–GSK-3β |
---|---|---|---|---|---|---|---|
Neuro genesis | BrdU [3H] thymidine incorporation | ▼ Proliferation (14) | ≠ Proliferation (8) | ▲ Proliferation (15), ▼ differentiation (15) | |||
Apoptosis necrosis | TUNEL, caspase IHC | ▲ Apoptosis (17) | ≠ Apoptosis (14) | ≠ Apoptosis (18), ≠ necrosis (18) | ▲ Apoptosis (16) | ||
Neuro anatomical changes | MRI | ▲ Cerebellum (1) | ▼ Cerebellum, cerebrum, hippocampus, cortex (18) | ||||
Histology | ▼ Dendrite length and surface area in the frontal cortex (12), ≠ in spine density (12), ≠ dendritic arborization (12), ▼Purkinje cells (1) | ▲ Microgliosis (17), ≠ tau fibrils (17, 22) | ▲ Neuronal density in cortex (18), ≠ number of cortical neurons (18), ▼ caliber of the proximal and distal part of the apical dendrites (18), ▼ size of the cell body of pyramidal neurons (18) | ▼ Tuj1, MAP2, SMI32, NeuN (15), ▲Nestin, Pax6 (15) | |||
Brain weight | ▲ Brain (1) | ▼ Brain and spinal cord (18, 8) | |||||
Biochemical molecular | Western blot RTPCR | ▲ pTau (17), ▼ nuclear β-catenin (17) | ▼ VEGF (14), ≠ BDNF (14) | ▼ MAP2 (18), ▲ pTau in old mice (19), ▼GSK-3α, ▼PPP2R3A and ▲Akt1 in striatum (8), ▲BDNF in hippocampus (8) | ▲ β-catenin, Axin, c-jun, Gli1, Gli2, Patched, Hes1, Hes5, NICD, c-myc, N-myc (15) | ▼pTau (16), ≠ β-catenin (16) |
Thus, comparative analysis of different animal models may be very informative, however critical and combinatory approach needs to be used to make correct interpretation and right conclusions.
To study the role of GSK-3β in the pathogenesis of Alzheimer’s disease, particularly with respect to the mechanism of tauopathy, double transgenic mice have been generated by inter-breeding mice overexpressing GSK-3β [S9A] with transgenic mice that overexpress the longest isoform of human protein tau (Spittaels et al.,
DISC1 (
GSK-3 acts as an important downstream component in the etiology of schizophrenia (reviewed in more detail elsewhere in this Special Topic series). There are several lines of evidence supporting the involvement of GSK-3 in the pathogenesis of schizophrenia. Polymorphisms in GSK-3 genes have been associated with schizophrenia (Souza et al.,
Several mouse models for Disc1 have been described (reviewed in Jaaro-Peled et al.,
An ENU-induced mutant of Disc1, Disc1-L100P, exhibits schizophrenia-related behaviors in mice (Clapcote et al.,
The emergence of sophisticated animal models with tissue and developmentally selective expression of GSK-3 has allowed direct assessment of the roles of this protein kinase in a variety of neurological processes and conditions. Clearly, the complexity of brain development and disease pathogenesis requires the use of animal models to examine the biological role of candidate components and with the numbers of candidate genes for neurological illness increasing, allows relatively rapid assessment of genetic interactions through inter-breeding of variant alleles.
While GSK-3 was first implicated in a neurological disorder in 1992 through its capacity to phosphorylate residues on Tau that are associated with neurofibrillary tangles in AD, the potential importance of this kinase in brain function and disease took off with the identification by Klein and Melton of GSK-3 as a direct target of lithium (Klein and Melton,
The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.
We thank members of the Woodgett lab (especially Charles Burger) for helpful discussions as well as Frankie Lee, Albert Wong, Tatiana Lipina, and John Roder. James Robert Woodgett is supported by an operating grant from the Canadian Institutes of Health Research (MOP74711).