Is there a role of the cyclin-dependent kinase 5 activator p25 in Alzheimer's disease?

Sporadic Alzheimer's disease is the leading cause of dementia, but the underlying molecular processes are still unknown. Several studies have observed an accumulation of the protein fragment p25 in sporadic Alzheimer's disease brain. p25 derives from proteolysis of p35, and overactivates the tau kinase cyclin-dependent kinase 5. Transgenic mice expressing high levels of p25 exhibit hyperphosphorylation of tau as seen in Alzheimer's disease, and neurodegeneration. In contrast, low-level p25 expression, less than half of endogenous p35 expression, has a sex-specific effect on hippocampal synaptic plasticity and improves spatial learning in female but not in male mice. Therefore, p25 formation may initially be a compensatory response for early learning deficits in Alzheimer's disease, but continued formation could contribute to detrimental changes in Alzheimer's disease.     

Improved reversal learning and altered fear conditioning in transgenic mice with regionally restricted p25 expression

Cleavage of the cyclin-dependent kinase 5 activator p35 generates the protein fragment p25, which accumulates in the forebrain of patients with Alzheimer's disease. Although p25 expression has been suggested to affect learning and memory, this hypothesis has not been tested to date. To investigate the role of p25 in hippocampus-dependent learning and memory we have generated transgenic mice expressing p25 preferentially in postnatal forebrain. p25 expression was highest in hippocampus where it averaged approximately 33% of endogenous p35 expression. This low level of p25 expression did not seem to result in hyperphosphorylation of tau, but increased the phosphorylation of neurofilament M and enhanced the expression of tau protein. These molecular changes did not correlate with neurodegeneration or motor abnormalities. In the Morris water maze the p25 mutants were normal in learning an initial platform location, but surprisingly reversal learning was improved when the platform position was changed. The p25 mutants were normal in contextual fear conditioning. However, when trained with a tone presentation the mutants showed reduced contextual conditioning and enhanced tone fear conditioning. We conclude that low p25 expression has pleiotropic effects on learning and memory. As p25 expression can improve learning and memory, p25 formation could be a compensatory mechanism for learning and memory deficits in Alzheimer's disease.     

Overexpression of p35 in Min6 pancreatic beta cells induces a stressed neuron-like apoptosis

Cdk5 activity has been implicated in brain development and the regulation of many neuronal processes. Recently, the expression of p35 and Cdk5 activity has been reported in pancreatic beta cells. Decreased Cdk5 activity enhanced glucose-stimulated insulin secretion. This suggests that Cdk5 may play an important role in the regulation of insulin secretion. To further understand how Cdk5 regulates insulin secretion in glucose-stimulated pancreatic β cells, we first confirmed the presence of a low level of p35 in pancreatic Min6 cells. Next, in a time-course experiment in high glucose (25 mM) we showed that endogenous p35 increased gradually accompanied by a 3-fold increase in Cdk5 activity by 16 h. Insulin secretion, however, doubled after 2 h followed by progressive downregulation, negatively correlated with Cdk5 activity. On the other hand, overexpression of p35 in these cells resulted in more than a three-fold increase in Cdk5 activity within 2 h coupled to a 50% reduction in insulin secretion in both high and low (3 mM) glucose. Most significantly, cells overexpressing p35, treated with high glucose for 4 h, showed induction of p25, the p35-derived truncated fragment which hyperactivates Cdk5 in neurons. As a result, insulin secretion was inhibited and cells became apoptotic. Roscovitine or co-infection of dominant negative Cdk5 (dnCdk5) with p35 increased insulin secretion and inhibited apoptosis. These results suggest that the model for deregulation and hyperactivation of Cdk5 in neurodegeneration may apply to the pathology seen in type 2 diabetes (T2DM). It is consistent with the view that Alzheimer's disease and T2DM are linked metabolically and pathologically by Cdk5 in a number of ways.     

Synaptic deficits are rescued in the p25/Cdk5 model of neurodegeneration by the reduction of β-secretase (BACE1)

Alzheimer's disease (AD) is the most common cause of dementia, and is characterized by memory loss and cognitive decline, as well as amyloid β (Aβ) accumulation, and progressive neurodegeneration. Cdk5 is a proline-directed serine/threonine kinase whose activation by the p25 protein has been implicated in a number of neurodegenerative disorders. The CK-p25 inducible mouse model exhibits progressive neuronal death, elevated Aβ, reduced synaptic plasticity, and impaired learning following p25 overexpression in forebrain neurons. Levels of Aβ, as well as the APP processing enzyme, β-secretase (BACE1), are also increased in CK-p25 mice. It is unknown what role increased Aβ plays in the cognitive and neurodegenerative phenotype of the CK-p25 mouse. In the current work, we restored Aβ levels in the CK-p25 mouse to those of wild-type mice via the partial genetic deletion of BACE1, allowing us to examine the Aβ-independent phenotype of this mouse model. We show that, in the CK-p25 mouse, normalization of Aβ levels led to a rescue of synaptic and cognitive deficits. Conversely, neuronal loss was not ameliorated. Our findings indicate that increases in p25/Cdk5 activity may mediate cognitive and synaptic impairment via an Aβ-dependent pathway in the CK-p25 mouse. These findings explore the impact of targeting Aβ production in a mouse model of neurodegeneration and cognitive impairment, and how this may translate into therapeutic approaches for sporadic AD.     

Recent evidence regarding a role for Cdk5 dysregulation in Alzheimer's disease

Based on a growing literature, cyclin-dependent kinase 5 (Cdk5) has been implicated in the pathological processes that contribute to neurodegeneration in Alzheimer's disease (AD). Cdk5 is ubiquitously expressed, but its activity is largely localized to post-mitotic neurons due to neuron-specific expression of its activators p35 and p39. Sufficient Cdk5 activity is critical to normal central nervous system development, as in its absence, neuronal migration and axonal path finding are deranged. Conversely, excessive and mislocalized Cdk5 activity appears to be detrimental to neuronal function. In fact, the pathological hallmarks of AD, beta-amyloid aggregates and neurofibrillary tangles, have been linked to Cdk5-mediated neuronal death. In this model, beta-amyloid is the toxic stimulus that disrupts intracellular calcium homeostasis, leading to activation of calpains, a family of calcium-dependent proteases. Calpain-mediated cleavage of p35, yields a truncated p25 fragment that possesses a longer half-life, lacks the necessary sequence targeting it to membranes, but retains the capacity to activate Cdk5. The resulting excessive and mislocalized Cdk5 activity targets tau as a substrate for hyperphosphorylation, which is a prerequisite of paired helical filament (PHF) formation. A number of recent reports, utilizing diverse methods, lend further support to this model of AD neurodegeneration, and several strategies for combating Cdk5 dysregulation have even been devised. However, the study of Cdk5 in AD is not without controversy, and questions remain regarding its role in the pathology. Herein, the most recent findings regarding this model are reviewed.     

TFP5/TP5 peptide provides neuroprotection in the MPTP model of Parkinson’s disease

Cyclin-dependent kinase 5 (Cdk5) is a member of the serine-threonine kinase family of cyclin-dependent kinases. Cdk5 is critical to normal mammalian nervous system development and plays important regulatory roles in multiple cellular functions. Recent evidence indicates that Cdk5 is inappropriately activated in sev-eral neurodegenerative conditions, including Parkinson’s disease (PD). PD is a chronic neurodegenerative disorder characterized by the loss of dopamine neurons in the substantia nigra, decreased striatal dopamine levels, and consequent extrapyramidal motor dysfunction. During neurotoxicity, p35 is cleaved to form p25. Binding of p25 with Cdk5 leads deregulation of Cdk5 resulting in number of neurodegenerative pathologies. To date, strategies to speciifcally inhibit Cdk5 hyperactivity have not been successful without affecting normal Cdk5 activity. Here we show that inhibition of p25/Cdk5 hyperactivation through TFP5/TP5, truncated 24-aa peptide derived from the Cdk5 activator p35 rescues nigrostriatal dopaminergic neurodegeneration induced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP/MPP+) in a mouse model of PD. TP5 pep-tide treatment also blocked dopamine depletion in the striatum and improved gait dysfunction after MPTP administration. The neuroprotective effect of TFP5/TP5 peptide is also associated with marked reduction in neuroinlfammation and apoptosis. Here we show inhibition of Cdk5/p25-hyperactivation by TFP5/TP5 pep-tide, which identiifes Cdk5/p25 as a potential therapeutic target to reduce neurodegeneration in PD.     

Expression of p25 impairs contextual learning but not latent inhibition in mice

The cyclin-dependent kinase 5 activator p25, which is derived from cleavage of p35, is thought to be formed in the brain of patients with Alzheimer's disease and schizophrenia. Female, but not male, transgenic mice expressing low levels of p25 have enhanced hippocampal long-term potentiation and improved spatial learning, raising the hypothesis that p25 may compensate for early learning deficits in Alzheimer's disease in a sex-dependent manner. Here, we show that low levels of p25 do not alter latent inhibition, a phenomenon that is impaired in patients with schizophrenia. We also demonstrate that contextual fear conditioning is impaired in female, but not in male, p25 transgenic mice. Thus, low levels of p25 are not always beneficial for learning as was previously hypothesized.     

AbetaPP induces cdk5-dependent tau hyperphosphorylation in transgenic mice Tg2576

Previous studies of Abeta-induced neuronal damage of hippocampal cells in culture have provided strong evidence that deregulation of the Cdk5/p35 kinase system is involved in the neurodegeneration pathway. Cdk5 inhibitors and antisense probes neuroprotected hippocampal cells against the neurotoxic action of Abeta. To further investigate the mechanisms underlying the participation of Cdk5 in neuronal degeneration, the transgenic mouse containing the Swedish mutations, Tg2576, was used as an animal model. Immunocytochemical studies using anti-Abeta(1-17) antibody evidenced the presence of labeled small-clustered core plaques in the hippocampus and cortex of 18-month-old transgenic mice brains. The loss of granular cells without a compressed appearance was detected in the vicinity of the cores in the dentate gyrus of the hippocampus. Immunostaining of Tg2576 brain sections with antibodies AT8, PHF1 and GFAP labeled punctuate dystrophic neurites in and around the amyloid core. Reactive astrogliosis around the plaques in the hippocampus was also observed. Studies at the molecular level showed differences in the expression of the truncated Cdk5 activator p25 in the transgenic animal, as compared with wild type controls. However no differences in Cdk5 levels were detected, thus corroborating previous cellular findings. Interestingly, hyperphosphorylated tau epitopes were substantially increased as assessed with the AT8 and PHF1 antibodies, in agreement with the observation of a p25 increase in the transgenic animal. These observations strongly suggest that the increased exposure of Alzheimer's type tau phosphoepitopes in the transgenic mice correlated with deregulation of Cdk5 leading to an increase in p25 levels. These studies also provide further evidence on the links between extraneuronal amyloid deposition and tau pathology.     

Sex-Specific Regulation of Fear Memory by Targeted Epigenetic Editing of Cdk5

Background

Sex differences in the expression and prevalence of trauma- and stress-related disorders have led to a growing interest in the sex-specific molecular and epigenetic mechanisms underlying these diseases. Cyclin-dependent kinase 5 (CDK5) is known to underlie both fear memory and stress behavior in male mice. Given our recent finding that targeted histone acetylation of Cdk5 regulates stress responsivity in male mice, we hypothesized that such a mechanism may be functionally relevant in female mice as well.

Amelioration of social isolation-triggered onset of early Alzheimer's disease-related cognitive deficit by N-acetylcysteine in a transgenic mouse model

Epidemiological study reveals that socially isolated persons have increased risk of developing Alzheimer's disease (AD). Whether this risk arises from an oxidative stress is unclear. Here we show that N-acetylcysteine (NAC), an anti-oxidant, is capable of preventing social isolation-induced accelerated impairment of contextual fear memory and rundown of hippocampal LTP in 3-month old APP/PS1 mice. Increased hippocampal levels of γ-secretase activity, Aβ-40 and Aβ-42 seen in the isolated APP/PS1 mice were reduced by chronic treatment of NAC. In addition, social isolation-induced increase in calpain activity and p25/p35 ratio concomitant with decrease in membrane-associated p35 and p35/Cdk5 activity was normalized by NAC. NAC pretreatment also reversed isolation-induced decrease in GluR1 Ser831 phosphorylation, surface expression of AMPARs and p35-GluR1-CaMKII interactions. These results suggest that NAC decreases γ-secretase activity resulting in the attenuation of Aβ production, calpain activity and conversion of p35 to p25 which stabilized p35-GluR1-CaMKII interactions and restored GluR1 and GluR2 surface expression. Our results indicate that NAC is effective in mouse models of AD and has translation potential for the human disorder.     

Cdk5: one of the links between senile plaques and neurofibrillary tangles?

The relationship between amyloid plaques and neurofibrillary tangles, the two pathologic hallmarks of Alzheimer's disease (AD), is an unknown and controversial subject. However, emerging evidence from genetic and biochemical studies suggests that accumulation of amyloid beta peptides may play a causative role in AD pathogenesis. This led to the amyloid hypothesis, which proposes that amyloid beta peptides disrupt neuronal metabolic and ionic homeostasis and cause aberrant activation of kinases and/or inhibition of phosphatases. The resulting alteration in kinase and phosphatase activities ultimately leads to hyperphosphorylation of tau and formation of neurofibrillary tangles. Cyclin-dependent kinase 5 (Cdk5) is a tau kinase whose activity is induced by amyloid beta peptides. Its deregulation may represent one of the signal transduction pathways that connect amyloid beta toxicity to tau hyperphosphorylation. This article reviews the functions and regulation of Cdk5. Evidence that suggests deregulation of Cdk5 activity in AD by virtue of calpain cleavage of its activator p35 to p25 will be discussed.     

Preferential targeting of p39-activated Cdk5 to Rac1-induced lamellipodia

Cdk5 is a member of the cyclin-dependent kinase (Cdk) family that plays a role in various neuronal activities including brain development, synaptic regulation, and neurodegeneration. Cdk5 requires the neuronal specific activators, p35 and p39 for subcellular compartmentalization. However, it is not known how active Cdk5 is recruited to F-actin cytoskeleton, which is a Cdk5 target. Here we found p35 and p39 localized to F-actin rich regions of the plasma membrane and investigated the underlying targeting mechanism in vitro by expressing them with Rho family GTPases in Neuro2A cells. Both p35 and p39 accumulated at the cell peripheral lamellipodia and perinuclear regions, where active Rac1 is localized. Interestingly, p35 and p39 displayed different localization patterns as p35 was found more at the perinuclear region and p39 was found more in peripheral lamellipodia. We then confirmed this distinct localization in primary hippocampal neurons. We also determined that the localization of p39 to lamellipodia requires myristoylation and Lys clusters within the N-terminal p10 region. Additionally, we found that p39–Cdk5, but not p35–Cdk5 suppressed lamellipodia formation by reducing Rac1 activity. These results suggest that p39–Cdk5 has a dominant role in Rac1-dependent lamellipodial activity.     

N-acetylcysteine prevents beta-amyloid toxicity by a stimulatory effect on p35/cyclin-dependent kinase 5 activity in cultured cortical neurons

Although previous studies have indicated that the neuroprotective effect of N-acetylcysteine (NAC) required activation of the Ras-extracellular-signal-regulated kinase (ERK) pathway, the detailed mechanisms and signal cascades leading to activation ERK are not clear. In the present study, we investigated the effect of NAC on A beta(25-35)-induced neuronal death. Pretreatment of neurons with NAC 1 hr before application of A beta prevented A beta-mediated cell death. NAC increased cyclin-dependent kinase 5 (Cdk5) phosphorylation, an effect that was blocked by Cdk5 inhibitor. The neuroprotective effect of NAC was significantly attenuated by Cdk5 inhibitors or in neurons transfected with Cdk5 or p35 small interfering RNA (siRNA). Conversely, pretreatment of neurons with the calpain inhibitors calpeptin or MDL28170 enhanced the neuroprotective effect of NAC. A beta(25-35) caused a significant decrease in the level of p35, with a concomitant increase in p25, which was completely prevented by NAC. This effect of NAC was blocked by the Cdk5 inhibitors roscovitine and butyrolactone. In addition, NAC increased Cdk5/p35 kinase activity but reduced Cdk5 kinase activity. A beta(25-35) treatment decreased phosphorylated levels of ERK, which could be reversed by NAC. The effect of NAC was completely blocked by Cdk5 inhibitors. NAC reversed the A beta(25-35)-induced decrease in the expression of Bcl-2, which could be blocked by the MAPK kinase (MEK) inhibitor or Cdk5 inhibitors. These results suggest that NAC-mediated neuroprotection against A beta toxicity is likely mediated by the p35/Cdk5-ERKs-Bcl-2 signal pathway.     

p35/Cdk5 pathway mediates soluble amyloid-beta peptide-induced tau phosphorylation in vitro

Alzheimer's disease (AD) is pathologically characterized by deposition of amyloid-beta peptides (Abeta) as senile plaques and by the occurrence of neurofibrillary tangles (NFTs) composed primarily of hyperphosphorylated tau protein. Activation of cyclin-dependent kinase 5 (Cdk5) via its potent activator p25 has recently been shown to promote phosphorylation of tau at AD-specific phosphoepitopes, and increased cleavage of p35 to p25 has been demonstrated in AD patients, suggesting that Cdk5 may represent a pathogenic tau protein kinase. We were interested in the potential effect of soluble forms of Abeta on Cdk5-mediated AD-like tau phosphorylation, insofar as previous studies of human biopsies and aged canine and primate brains have shown that dystrophic neurites appear before the formation of neuritic plaques. We transfected N2a cells with a p35 vector (N2a/p35 cells) and, after differentiation, challenged these cells with Abeta(1-42) peptide in soluble form (sAbeta(1-42)). Results show that sAbeta(1-42) at relatively low levels (1-5 microM) dose-dependently increases tau phosphorylation at AD-specific phosphoepitopes in differentiated N2a/p35 cells compared with controls, an effect that is blocked by antisense oligonucleotides against p35. sAbeta(1-42)-induced tau phosphorylation is concomitant with an increase in both p25 to p35 ratio and Cdk5 activity (but not protein levels). Additionally, blockade of L-type calcium channels or inhibition of calpain completely abolishes this effect. Taken together, these data indicate that sAbeta is a potent activator of the p25/Cdk5 pathway, resulting in promotion of AD-like tau phosphorylation in vitro.     

The role of Cdk5 in cognition and neuropsychiatric and neurological pathology

Cyclin-dependent kinase 5 (Cdk5) is a proline-directed serine/threonine kinase that is ubiquitous in the nervous system and interacts with a myriad of substrates. Its modulation of synaptic plasticity and associated mechanisms of learning and memory as well as neurodegeneration and cognitive disease highlights its importance in the human brain. Cdk5 is active throughout the neuron via its kinase activity, protein-protein interactions, and nuclear associations. It regulates functions thought vital to memory and plasticity, including synaptic vesicle recycling, dendritic spine formation, neurotransmitter receptor density, and neuronal excitability. Although conditional knockout of Cdk5 improves learning and plasticity, the associated deleterious effects of increased excitability cast doubts on the therapeutic efficacy of systemic inhibitors. However, through further work on the regulation of Cdk5 and its effectors, this important molecule promises to aid in elucidating key pathways involved in learning and memory and uncover innovative therapeutic targets to treat neurodegenerative and neuropsychiatric diseases.