Aberrant Amyloid Precursor Protein Processing Research Articles

Dysregulated Wnt Signalling in the Alzheimer's Brain.

The Wnt signalling system is essential for both the developing and adult central nervous system. It regulates numerous cellular functions ranging from neurogenesis to blood brain barrier biology. Dysregulated Wnt signalling can thus have significant consequences for normal brain function, which is becoming increasingly clear in Alzheimer’s disease (AD), an age-related neurodegenerative disorder that is the most prevalent form of dementia. AD exhibits a range of pathophysiological manifestations including aberrant amyloid precursor protein processing, tau pathology, synapse loss, neuroinflammation and blood brain barrier breakdown, which have been associated to a greater or lesser degree with abnormal Wnt signalling. Here we provide a comprehensive overview of the role of Wnt signalling in the CNS, and the research that implicates dysregulated Wnt signalling in the ageing brain and in AD pathogenesis. We also discuss the opportunities for therapeutic intervention in AD via modulation of the Wnt signalling pathway, and highlight some of the challenges and the gaps in our current understanding that need to be met to enable that goal.

Upregulated function of mitochondria-associated ER membranes in Alzheimer disease

Alzheimer disease (AD) is associated with aberrant processing of the amyloid precursor protein (APP) by γ-secretase, via an unknown mechanism. We recently showed that presenilin-1 and -2, the catalytic components of γ-secretase, and γ-secretase activity itself, are highly enriched in a subcompartment of the endoplasmic reticulum (ER) that is physically and biochemically connected to mitochondria, called mitochondria-associated ER membranes (MAMs). We now show that MAM function and ER-mitochondrial communication-as measured by cholesteryl ester and phospholipid synthesis, respectively-are increased significantly in presenilin-mutant cells and in fibroblasts from patients with both the familial and sporadic forms of AD. We also show that MAM is an intracellular detergent-resistant lipid raft (LR)-like domain, consistent with the known presence of presenilins and γ-secretase activity in rafts. These findings may help explain not only the aberrant APP processing but also a number of other biochemical features of AD, including altered lipid metabolism and calcium homeostasis. We propose that upregulated MAM function at the ER-mitochondrial interface, and increased cross-talk between these two organelles, may play a hitherto unrecognized role in the pathogenesis of AD.

Cdk5/p25-Induced Cytosolic PLA2-Mediated Lysophosphatidylcholine Production Regulates Neuroinflammation and Triggers Neurodegeneration

The deregulation of cyclin-dependent kinase 5 (Cdk5) by p25 has been shown to contribute to the pathogenesis in a number of neurodegenerative diseases such as amyotrophic lateral sclerosis (ALS), Parkinson's disease (PD) and Alzheimer's disease (AD). In particular, p25/Cdk5 has been shown to produce hyperphosphorylated tau, neurofibrillary tangles as well as aberrant amyloid precursor protein processing found in AD. Neuroinflammation has been observed alongside the pathogenic process in these neurodegenerative diseases, however the precise mechanism behind the induction of neuroinflammation and the significance in the AD pathogenesis has not been fully elucidated. In this report, we uncover a novel pathway for p25-induced neuroinflammation where p25 expression induces an early trigger of neuroinflammation in vivo in mice. Lipidomic mass spectrometry, in vitro coculture and conditioned media transfer experiments show that the soluble lipid mediator lysophosphatidylcholine (LPC) is released by p25 overexpressing neurons to initiate astrogliosis, neuroinflammation and subsequent neurodegeneration. Reverse transcriptase PCR and gene silencing experiments show that cytosolic phospholipase 2 (cPLA2) is the key enzyme mediating the p25-induced LPC production and cPLA2 upregulation is critical in triggering the p25-mediated inflammatory and neurodegenerative process. Together, our findings delineate a potential therapeutic target for the reduction of neuroinflammation in neurodegenerative diseases including AD.

Lactic acid induces aberrant amyloid precursor protein processing by promoting its interaction with endoplasmic reticulum chaperone proteins

Lactic acid induces aberrant amyloid precursor protein processing by promoting its interaction with endoplasmic reticulum chaperone proteins

Expression of aquaporins in a transgenic mouse model of Alzheimer's disease

Background Several studies reported altered expression of the water channels, aquaporins (AQPs), in brains of Alzheimer’s disease (AD) but the relevance of the observed changes with respect to the neuropathology of AD is poorly understood. Because aberrant processing of the amyloid precursor protein (APP) to amyloid beta-peptide (A-beta), the principle component of amyloid plaques, is central to the pathogenesis of AD, we sought to determine the temporal and spatial expression of AQPs in the cortex and hippocampus of the triple transgenic mouse model of Alzheimer’s disease (3xTg-AD) and to elucidate whether AQPs contribute directly to the aberrant processing of APP into A-beta.

Lactic Acid Induces Aberrant Amyloid Precursor Protein Processing by Promoting Its Interaction with Endoplasmic Reticulum Chaperone Proteins

BackgroundLactic acid, a natural by-product of glycolysis, is produced at excess levels in response to impaired mitochondrial function, high-energy demand, and low oxygen availability. The enzyme involved in the production of β-amyloid peptide (Aβ) of Alzheimer's disease, BACE1, functions optimally at lower pH, which led us to investigate a potential role of lactic acid in the processing of amyloid precursor protein (APP).Methodology/Principal FindingsLactic acid increased levels of Aβ40 and 42, as measured by ELISA, in culture medium of human neuroblastoma cells (SH-SY5Y), whereas it decreased APP metabolites, such as sAPPα. In cell lysates, APP levels were increased and APP was found to interact with ER-chaperones in a perinuclear region, as determined by co-immunoprecipitation and fluorescence microscopy studies. Lactic acid had only a very modest effect on cellular pH, did increase the levels of ER chaperones Grp78 and Grp94 and led to APP aggregate formation reminiscent of aggresomes.Conclusions/SignificanceThese findings suggest that sustained elevations in lactic acid levels could be a risk factor in amyloidogenesis related to Alzheimer's disease through enhanced APP interaction with ER chaperone proteins and aberrant APP processing leading to increased generation of amyloid peptides and APP aggregates.

The role of retromer in sorla-dependent APP transport and processing

The retromer complex has been implicated in the intracellular transport of the amyloid precursor protein (APP). Retromer-deficiency leads to aberrant APP processing and the level of retromer proteins are altered in Alzheimer's disease (AD). SorLA is a neuronal sorting receptor genetically linked to late-onset AD with important functions in neuronal sorting of APP. The current study describes the interaction between sorLA and retromer and its role in APP transport and processing. Protein interactions were determined by fluorescence life-time imaging microscopy (FLIM) and co-immunoprecipitation analysis. APP processing products were determined by WB and ELISA. The subcellular localizations of sorLA and APP were characterized by confocal microscopy and electron microscopy analysis. Knockdown of expression of the VPS26 retromer subunit was undertaken by siRNA and shRNA technologies. We find that sorLA and retromer co-localize in neurons within Golgi and endosomal compartments, and that knockdown of retromer leads to a mislocalization of sorLA. Mutations of key amino acids responsible for retromer binding in the cytoplasmic domain of sorLA results in a redistribution of the receptors that is indistinguishable from that observed in cells following knockdown of VPS26. In retromer expressing cells, wild-type sorLA binds APP in the trans-Golgi network (TGN), diminishing processing of APP by both the amyloidogeninc and non-amyloidogenic pathway. The sorLA mutant that does not associate with retromer binds APP, as demonstrated by FLIM studies, but within distinct cellular compartments as compared to wildtype sorLA. Interestingly, the mutant receptor exhibits no protection against amyloid production. We have mapped the binding site for retromer in sorLA and provide evidence for a regulatory role of this receptor in retromer-dependent APP sorting and processing. We propose a model whereby retromer functions in the retrieval of sorLA from endosomes to the TGN.

Effect of creatine on the processing of amyloid precursor protein isoforms and mitochondrial function.

Glucose hypo‐metabolism is an early feature in Alzheimer's disease (AD), preceding the onset of the clinical symptoms of dementia and the appearance of amyloidbeta (Aβ) peptide, a proteolytic product of amyloid precursor protein (APP). Studies showed that this metabolic deficit is associated with altered expression and activity of enzymes involved in mitochondrial energy production. Mitochondrial dysfunction can lead to ATP depletion and trigger apoptosis. Inhibition of energy metabolism can also alter APP processing. Hence, buffering intracellular energy levels could exert neuroprotective effects. One such buffering system is creatine kinase and its substrate creatine. However, it is unclear if energy buffering can modulate aberrant APP processing. In human brain, APP695 is the predominant APP isoforms and APP751 and APP770 are two other major APP transcripts. APP751 and APP770 isoforms contain a Kunitz protease inhibitory (KPI) motif that APP695 lacks, and APP770 possesses an additional OX2 domain that is absent in both APP695 and APP751. In AD‐afflicted human brain tissues, APP751 and APP770 expression were elevated. In this study using an immortalized APP‐null neuronal cell line expressing APP695, APP751 and APP77, we observed creatine is able to reduce the production of amyloidogenic derivatives and lessen the impairment of mitochondrial function in cells expressing the KPI‐containing APP isoforms. This neuroprotective effect is due in part to the activation of the insulin signalling pathway. Taken together, we speculate that creatine may have an important role in preventing energy dysfunction and possibly modulating the neurodegenerative cascade in AD.

Impaired "episodic-like" object memory in adult APPswe transgenic mice.

An early clinical symptom of Alzheimer's disease is impaired episodic memory. However, the precise pathological event(s) that underpins this deficit remains unclear. In the present study, the authors examined whether wild-type mice and Tg2576 mice expressing an amyloid precursor protein (APP) mutation are able to form an integrated memory of the spatio-temporal context in which objects are presented. In Experiment 1, wild-type mice, but not Tg2576 mice that were 10-12 months old, explored objects presented in a novel location. In Experiment 2, wild-type mice explored an object that was presented both earlier in a sequence and in a different location relative to other objects that possessed only one of these properties (i.e., memory for "what," "where," and "when" items were presented). In contrast, the behavior of adult Tg2576 mice was influenced only by the temporal order in which objects were presented. These results demonstrate that wild-type, but not APP-mutant, mice are able to form an "episodic-like" memory of the spatio-temporal properties of objects and support the hypothesis that aberrant APP processing contributes to impairments in event memory.

P25/Cyclin-Dependent Kinase 5 Induces Production and Intraneuronal Accumulation of Amyloid βIn Vivo

Aberrant processing of the amyloid precursor protein (APP) and the subsequent accumulation of amyloid beta (Abeta) peptide has been widely established as a central event in Alzheimer's disease (AD) pathogenesis. The sequential cleavage steps required for the generation of Abeta are well outlined; however, there is a relative dearth of knowledge pertaining to signaling pathways and molecular mechanisms that can modulate this process. Here, we demonstrate a novel role for p25/cyclin-dependent kinase 5 (Cdk5) in regulating APP processing, Abeta peptide generation, and intraneuronal Abeta accumulation in inducible p25 transgenic and compound PD-APP transgenic mouse models that demonstrate deregulated Cdk5 activity and a neurodegenerative phenotype. Induction of p25 resulted in enhanced forebrain Abeta levels before any evidence of neuropathology in these mice. Intracellular Abeta accumulated in perinuclear regions and distended axons within the forebrains of these mice. Evidence for modulations in axonal transport or beta-site APP cleaving enzyme 1 protein levels and activity are presented as mechanisms that may account for the Abeta accumulation caused by p25/Cdk5 deregulation. Collectively, these findings delineate a novel pathological mechanism involving aberrant APP processing by p25/Cdk5 and have important implications in AD pathogenesis.

Coated-platelets retain amyloid precursor protein on their surface

Coated-Platelets are a subset of platelets produced by dual-agonist activation with collagen plus thrombin and are characterized by strong retention of several procoagulant, α-granule proteins on the cell surface. In this report we demonstrate that coated-platelets also retain full-length amyloid precursor protein (APP) on their surface in contrast to the cleavage of APP in platelets activated with a single agonist. In addition, western blot analysis indicated that APP is derivatized during coated-platelet synthesis. We subsequently measured coated-platelet production in patients with Alzheimer's disease (AD). Twenty-two AD patients showed a wide distribution of coated-platelet values; however the least impaired AD patients produced coated-platelets at a level significantly above that of aged controls (41.0 ± 9.9 vs. 28.7 ± 11.4%; mean ± 1SD; p = 0.017). These findings suggest that coated-platelets may be a model of aberrant APP processing in early AD patients.

ε-Glycation, APP and Aβ in ageing and Alzheimer disease: A hypothesis

The post-translational modifications of protein molecules include glycation, which may not only occur enzymatically controlled in N and O position, but also wherever proteins meet reducing sugars non-enzymatically in epsilon position at lysines (non-enzymatic (epsilon) glycation (NEG)). The formation of keto-amines from the amine-sugar compounds (Amadori re-arrangement) and further processing of the largely undigestible Amadori compounds eventually results in insoluble advanced glycation end products (AGEs). The latter can induce or favour disease including mental disorders. Preferential targets of NEG include large cell surface proteins. Ample evidence has been provided that NEG also occurs in the brain where cross-linking of epsilon-glycated proteins, induction of oxidative stress and signalling of AGEs through their specific receptor (RAGE) likely play a role in (brain) ageing and Alzheimer disease (AD). This is underscored by the demonstration of particular interactions between AGE/RAGE and amyloid-beta (Abeta) that favour the aggregation and deposition of Abeta and, perhaps, the formation of Abeta itself. The close relationship between NEG and Abeta, as well as other facts foster the hypothesis that NEG of the large trans-membrane amyloid precursor protein (APP) might be a significant factor in the induction of aberrant APP cleavage with production of Abeta, not only in normal ageing, but also in AD. Blockade of lysine cleavage sites on APP by sugar chains or marker effects induced by NEG akin to ubiquitination of proteins for degradation at lysines could be expected to contribute to altered processing of APP. The hypothesis of epsilon-glycation in APP proposed here and the review of evidences for the significance of NEG in brain ageing and AD are aimed at the stimulation of investigations into the still open question which role NEG plays with respect to APP and its abnormal processing in AD. It can be rendered likely that such research might open new avenues towards decreasing the risk of AD and/or slowing its progression through the prevention of NEG in APP with aberrant APP processing, increased generation of Abeta and the formation of AGEs from epsilon-glycated APP.

Accelerated amyloid deposition, neurofibrillary degeneration and neuronal loss in double mutant APP/tau transgenic mice

Even though the idea that amyloid beta peptide accumulation is the primary event in the pathogenesis of Alzheimer's disease has become the leading hypothesis, the causal link between aberrant amyloid precursor protein processing and tau alterations in this type of dementia remains controversial. We further investigated the role of beta-amyloid production/deposition in tau pathology and neuronal cell death in the mouse brain by crossing Tg2576 and VLW lines expressing human mutant amyloid precursor protein and human mutant tau, respectively. The resulting double transgenic mice showed enhanced amyloid deposition accompanied by neurofibrillary degeneration and overt neuronal loss in selectively vulnerable brain limbic areas. These findings challenge the idea that tau pathology in Alzheimer's disease is merely a downstream effect of amyloid production/deposition and suggest that reciprocal interactions between beta-amyloid and tau alterations may take place in vivo.

Increased β-secretase activity and expression in rats following transient cerebral ischemia

The aberrant proteolytic processing of the amyloid precursor protein (APP) by β- and γ-secretases is key to amyloid plaque formation in Alzheimer's disease (AD). Identification of an aspartyl protease as the β-secretase (β-site APP cleaving enzyme, BACE) involved in APP processing provides a pharmaceutical target for potential AD treatment. In the present studies, we demonstrate that transient cerebral ischemia in female rats caused a 30% increase in β-secretase activity. α-Secretase activity did not increase significantly. We examined protein levels of BACE1, and its analogue BACE2, in ischemic brain extracts. BACE1 protein levels increased 67%, while BACE2 protein level did not change after such a transient ischemia. Immunohistochemical studies demonstrated that BACE1 protein was increased in the ischemic neocortex, when compared with its contralateral cortex. Further, colocalization assessment indicated that BACE1 strongly associated with staining for the apoptotic marker, TUNEL. These results may partially explain epidemiological study, which demonstrate a higher incidence of dementia after stroke. Further, our results support the hypothesis that apoptosis and aberrant APP processing are correlated events in AD brain, and suggest that inhibition of BACE may have a therapeutic effect in the prevention of dementia after stroke recovery.

β -amyloid: friend or foe

The function of the amyloid precursor protein (APP) and its product, β-amyloid, (Aβ) is at present unknown. The deposition of Aβ in senile plaques as well as meningeal and cerebral vessels has led many researchers to discount the possibility of a beneficial protective function for the protein. Thus it is generally believed that the aberrant processing of APP leads to increased β-amyloid secretion that in turn leads to subsequent plaque formation and Alzheimer’s disease. Here, a hypothesis is presented that the protein may indeed be protective and that a potential role for beta amyloid in innate immunity may exist.

Neuronal overexpression of APPL, the Drosophila homologue of the amyloid precursor protein (APP), disrupts axonal transport

The two pathological hallmarks of Alzheimer's disease, amyloid plaques and neurofibrillary tangles, involve two apparently unrelated proteins, the amyloid precursor protein (APP) and Tau. Although it is known that aberrant processing of APP is associated with Alzheimer's disease [1], the definitive role of APP in neurons is not yet clear [2]. Tau regulates microtubule stabilization and assembly in axons [3] and is, thus, an essential component of the microtubule-associated organelle transport machinery [4,5]. Although several groups have reported physical interaction between APP and Tau [6,7], and induction of Tau phosphorylation by APP [8] and β-amyloid peptide [9], the functional connection between APP and Tau is unclear. To explore the possibility that the functions of these two proteins may somehow converge on the same cellular process, we overexpressed APPL, the Drosophila homologue of APP, along with Tau in Drosophila neurons. Panneural coexpression of APPL and Tau resulted in adults that, upon eclosion, failed to expand wings and harden the cuticle, which is suggestive of neuroendocrine dysfunction. We analyzed axonal transport when Tau and APPL were coexpressed and found that transport of axonal cargo was disrupted, as evidenced by increased retention of synaptic proteins in axons and scarcity of neuropeptide-containing vesicles in the distal processes of peptidergic neurons. In an independent approach, we demonstrated genetic interaction and phenotypic similarity between APPL overexpression and mutations in the Kinesin heavy chain (Khc) gene, the product of which is a motor for anterograde vesicle trafficking [10].

Identification of Amyloid Precursor Protein in Synaptic Plasma Membrane

Although the etiology of Alzheimer′s disease has not been elucidated yet, dysfunction and loss of synapses are believed to cause dementia. Recent studies suggest that the primary cause of the disease is closely related to the aberrant processing of the amyloid precursor protein (APP). To investigate the localization of APP at synaptic sites, we obtained synaptic plasma membrane and synaptic vesicles from rat brain. Enhanced chemiluminescence (ECL) western blot analysis using two specific polyclonal antibodies against APP revealed strong APP immunoreactivity in the synaptic plasama membrane, but not in the synaptic vesicle fraction. These data indicate that APP is localized at the synaptic plasma membrane and may play a role in physiological synaptic activity. Alternative localization or aberrant processing of APP at the synaptic site may cause impairment of synaptic function in Alzheimer′s disease.

Mutation of the gene for the human lysosomal serine protease cathepsin G is not the cause of aberrant APP processing in familial Alzheimer disease

Recent genetic linkage studies have implicated a gene on chromosome 14 in the pathogenesis of FAD. The identify of this gene remains unknown but it has been speculated that it may be involved in the cellular processing of the amyloid precursor protein ( APP). We have analyzed the nucleotide sequence of the entire open reading frame of the cathepsin G gene located on chromosome 14q. No mutations were observed, suggesting that defects in this lysosomal protease are not responsible for aberrant accumulation of proteolytic products of APP in FAD brain tissue.