ß-Amyloid Precursor Protein Intracellular Domain Controls Mitochondrial Function by Modulating Phosphatase and Tensin Homolog-Induced Kinase 1 Transcription in Cells and in Alzheimer Mice Models.

BACKGROUND: Mitophagy and mitochondrial dynamics alterations are two major hallmarks of neurodegenerative diseases. Dysfunctional mitochondria accumulate in Alzheimer's disease-affected brains by yet unexplained mechanisms. METHODS: We combined cell biology, molecular biology, and pharmacological approaches to unravel a novel molecular pathway by which presenilins control phosphatase and tensin homolog-induced kinase 1 (Pink-1) expression and transcription. In vivo approaches were carried out on various transgenic and knockout animals as well as in adeno-associated virus-infected mice. Functional readout and mitochondrial physiology (mitochondrial potential) were assessed by combined procedures including flow cytometry, live imaging analysis, and immunohistochemistry. RESULTS: We show that presenilins 1 and 2 trigger opposite effects on promoter transactivation, messenger RNA, and protein expression of Pink-1. This control is linked to γ-secretase activity and ß-amyloid precursor protein but is independent of phosphatase and tensin homolog. We show that amyloid precursor protein intracellular domain (AICD) accounts for presenilin-dependent phenotype and upregulates Pink-1 transactivation in cells as well as in vivo in a Forkhead box O3a-dependent manner. Interestingly, the modulation of γ-secretase activity or AICD expression affects Pink-1-related control of mitophagy and mitochondrial dynamics. Finally, we show that parkin acts upstream of presenilins to control Pink-1 promoter transactivation and protein expression. CONCLUSIONS: Overall, we delineate a molecular cascade presenilins-AICD-Forkhead box O3a linking parkin to Pink-1. Our study demonstrates AICD-mediated Pink-1-dependent control of mitochondrial physiology by presenilins. Furthermore, it unravels a parkin-Pink-1 feedback loop controlling mitochondrial physiology that could be disrupted in neurodegenerative conditions.

Tau Protein Mediates APP Intracellular Domain (AICD)-Induced Alzheimer's-Like Pathological Features in Mice.

Amyloid precursor protein (APP) is cleaved by gamma-secretase to simultaneously generate amyloid beta (Aß) and APP Intracellular Domain (AICD) peptides. Aß plays a pivotal role in Alzheimer's disease (AD) pathogenesis but recent studies suggest that amyloid-independent mechanisms also contribute to the disease. We previously showed that AICD transgenic mice (AICD-Tg) exhibit AD-like features such as tau pathology, aberrant neuronal activity, memory deficits and neurodegeneration in an age-dependent manner. Since AD is a tauopathy and tau has been shown to mediate Aß-induced toxicity, we examined the role of tau in AICD-induced pathological features. We report that ablating endogenous tau protects AICD-Tg mice from deficits in adult neurogenesis, seizure severity, short-term memory deficits and neurodegeneration. Deletion of tau restored abnormal phosphorylation of NMDA receptors, which is likely to underlie hyperexcitability and associated excitotoxicity in AICD-Tg mice. Conversely, overexpression of wild-type human tau aggravated receptor phosphorylation, impaired adult neurogenesis, memory deficits and neurodegeneration. Our findings show that tau is essential for mediating the deleterious effects of AICD. Since tau also mediates Aß-induced toxic effects, our findings suggest that tau is a common downstream factor in both amyloid-dependent and-independent pathogenic mechanisms and therefore could be a more effective drug target for therapeutic intervention in AD.

Decrease in levels of the evolutionarily conserved microRNA miR-124 affects oligodendrocyte numbers in Zebrafish, Danio rerio.

Oligodendrocytes produce multi-lamellar myelin membranes that surround axons in the central nervous system (CNS). Preservation and generation of myelin are potential therapeutic targets for dysmyelinating and demyelinating diseases. MicroRNAs (miRNAs) play a vital role in oligodendrocyte differentiation and overall CNS development. miR-124 is a well-conserved neuronal miRNA with important roles in neuronal differentiation and function. miR-124 levels increase following loss of myelin in both human and rodent brains. While the role of neuronal miR-124 in neurogenesis has been established, its effects on axonal outgrowth and oligodendrocytes are not currently known. We therefore explored the possible effect of selective knockdown of miR-124 in Danio rerio using a morpholino-based knockdown approach. No morphological abnormalities or loss of motor neurons were detected despite loss of axonal outgrowth. Morpholino-based knockdown of miR-124 led to reciprocal increases in mRNA levels of target genes that inhibit axonal and dendritic projections. Importantly, loss of miR-124 led to decreased oligodendrocyte cell numbers and myelination of axonal projections in the ventral hindbrain. Taken together, our results add a new dimension to the existing complexity of neuron-glial relationships and highlight the utility of Danio rerio as a model system to investigate such interactions.

JNK-interacting protein 1 mediates Alzheimer's-like pathological features in AICD-transgenic mice.

Amyloid precursor protein, which generates amyloid beta peptides, is intimately associated with Alzheimer's disease (AD) pathogenesis. We previously showed that transgenic mice overexpressing amyloid precursor protein intracellular domain (AICD), a peptide generated simultaneously with amyloid beta, develop AD-like pathologies, including hyperphosphorylated tau, loss of synapses, and memory impairments. AICD is known to bind c-Jun N-terminal kinase (JNK)-interacting protein 1 (JIP1), a scaffold protein that associates with and activates JNK. The aim of this study was to examine the role of JIP1 in AICD-induced AD-like pathologies in vivo, since the JNK pathway is aberrantly activated in AD brains and contributes to AD pathologies. We generated AICD-Tg mice lacking the JIP1 gene (AICD; JIP1(-/-)) and found that although AICD; JIP1(-/-) mice exhibit increased AICD, the absence of JIP1 results in decreased levels of hyperphosphorylated tau and activated JNK. AICD; JIP1(-/-) mice are also protected from synaptic loss and show improved performance in behavioral tests. These results suggest that JIP1 mediates AD-like pathologies in AICD-Tg mice and that JNK signaling may contribute to amyloid-independent mechanisms of AD pathogenesis.

TREM2 deficiency eliminates TREM2+ inflammatory macrophages and ameliorates pathology in Alzheimer's disease mouse models.

Variants in triggering receptor expressed on myeloid cells 2 (TREM2) confer high risk for Alzheimer's disease (AD) and other neurodegenerative diseases. However, the cell types and mechanisms underlying TREM2's involvement in neurodegeneration remain to be established. Here, we report that TREM2 is up-regulated on myeloid cells surrounding amyloid deposits in AD mouse models and human AD tissue. TREM2 was detected on CD45(hi)Ly6C(+) myeloid cells, but not on P2RY12(+) parenchymal microglia. In AD mice deficient for TREM2, the CD45(hi)Ly6C(+) macrophages are virtually eliminated, resulting in reduced inflammation and ameliorated amyloid and tau pathologies. These data suggest a functionally important role for TREM2(+) macrophages in AD pathogenesis and an unexpected, detrimental role of TREM2 in AD pathology. These findings have direct implications for future development of TREM2-targeted therapeutics.

Neuroinflammation in Alzheimer's disease: from pathogenesis to a therapeutic target.

The top-down, reductionist approach of the past three decades has resulted in remarkable progress in identifying genes and proteins involved in Alzheimer's disease (AD), including ß-amyloid (Aß) peptides and tau protein. Recently, a number of genes of the innate immune pathway have been identified as AD risk factors and several microglial proteins have been shown to be chronically activated in AD brains. Together, these observations suggest a crucial role for neuroinflammation in AD pathogenesis and emerging evidence suggests that neuroinflammation is both a cause and a consequence of AD. Epidemiological studies show that long-term users of anti-inflammatory drugs are protected from AD but anti-inflammatory treatment in mild AD patients has not been successful. These observations suggest that anti-inflammatory treatment is likely to be successful if initiated prior to the onset of neurological symptoms. Finally, after the remarkable success of the reductionist approach, a complimentary bottom-up systems approach is necessary to gain a better understanding of the highly complex, multifactorial nature of AD pathogenesis.

The apolipoprotein-E-mimetic COG112 protects amyloid precursor protein intracellular domain-overexpressing animals from Alzheimer's disease-like pathological features.

BACKGROUND: Amyloid-ß (Aß) peptides derive from the amyloid precursor protein (APP) and play a pivotal role in Alzheimer's disease (AD) pathogenesis. Our previous work showed that the APP intracellular domain (AICD), which is produced simultaneously with Aß, also contributes to the development of AD-like features. Studies show that administration of apolipoprotein E (apoE) and apoE-derived small peptide mimetics protect AD mouse models against these AD-like features. However, the effects of apoE-mimetic treatment on AICD-mediated AD-like pathologies remain to be elucidated. OBJECTIVE: To study the effects of an apoE mimetic (COG112) on neuroinflammation, hyperphosphorylation of tau and defects in adult neurogenesis in AICD- overexpressing transgenic mice (FeCγ25 line). METHODS: Beginning at 1 month of age, animals were administered subcutaneous COG112 3 times per week for 3 months, followed by immunohistochemical analysis for neuroinflammation, neurogenesis and phosphorylated tau. RESULTS: Treatment with COG112 significantly reduced neuroinflammation in AICD mice and protected against impaired adult hippocampal neurogenesis. We also found that COG112 treatment reduced hyperphosphorylation and somatodendritic accumulation of tau in the hippocampus and cerebral cortex of AICD mice. CONCLUSIONS: Reduction of neuroinflammation by the apoE-mimetic COG112 protects against impaired neurogenesis and tau pathology in AICD transgenic mice. These data suggest that neuroinflammation plays an important role in AICD-induced AD-like pathologies.

Knockdown of amyloid precursor protein in zebrafish causes defects in motor axon outgrowth.

Amyloid precursor protein (APP) plays a pivotal role in Alzheimer's disease (AD) pathogenesis, but its normal physiological functions are less clear. Combined deletion of the APP and APP-like protein 2 (APLP2) genes in mice results in post-natal lethality, suggesting that APP performs an essential, if redundant, function during embryogenesis. We previously showed that injection of antisense morpholino to reduce APP levels in zebrafish embryos caused convergent-extension defects. Here we report that a reduction in APP levels causes defective axonal outgrowth of facial branchiomotor and spinal motor neurons, which involves disorganized axonal cytoskeletal elements. The defective outgrowth is caused in a cell-autonomous manner and both extracellular and intracellular domains of human APP are required to rescue the defective phenotype. Interestingly, wild-type human APP rescues the defective phenotype but APPswe mutation, which causes familial AD, does not. Our results show that the zebrafish model provides a powerful system to delineate APP functions in vivo and to study the biological effects of APP mutations.

Amyloid precursor protein: more than just neurodegeneration.

Amyloid precursor protein (APP) fascinates cell biologists because it is proteolytically processed to generate multiple peptides, including amyloid-ß, which is implicated in Alzheimer's disease. However, a large body of data also shows that the extracellular soluble fragment of APP produced by α-secretase (sAPPα) is neuroprotective and promotes neuronal outgrowth. A study by Demars and colleagues appearing in the previous issue provides data showing that sAPPα is a general growth factor for stem cells of multiple lineages. Thus, APP seems to play complex and disparate roles in neurodegeneration and neuroprotection.

Detection of APP intracellular domain in brain tissue.

The cleavage of amyloid precursor protein (APP) by γ-secretase produces Aß peptides, which are prominent features in Alzheimer's disease and have been extensively studied. By contrast, APP intracellular domain (AICD), also a product of this cleavage event, has received little or no investigative attention. A major reason for this is that AICD is generally not detected in tissue lysates and, therefore, is neglected as a non-relevant product of APP metabolism. However, recent studies have shown that AICD regulates a number of important cellular events. Furthermore, we found that contrary to previous assertions, AICD can be detected in brain lysates using Western blotting if an antigen retrieval protocol is employed. Here we describe the protocol for AICD detection and note the biological relevance of AICD in physiological and pathological conditions.

PAT1 induces cell death signal and SET mislocalization into the cytoplasm by increasing APP/APLP2 at the cell surface.

The cleavage of amyloid precursor protein (APP) by caspases unmasks a domain extending from membrane to caspase cleavage site. This domain induces apoptosis in vitro and in vivo when overexpressed in neurons through the help of an internalization vector. In this model, we previously showed that SET rapidly binds to the internalized domain and is involved in downstream deleterious effects. Under these conditions SET mislocalizes from the nucleus to the cytoplasm, as in Alzheimer's disease (AD). In this report using the same model, we show that PAT1 attaches to the internalized domain earlier than SET and that this binding causes an increase in the levels of APP and APLP2 at the cell surface. Down regulation experiments of PAT1 and of APP and APLP2 show that the increase of the levels of APP and APLP2 at the cell surface triggers the cell death signal and SET mislocalization into the cytoplasm. In the context of AD these data suggest that mislocalization of SET into the cytoplasm may occur downstream of first cell death signal events involving PAT1 protein.

Aging and excitotoxic stress exacerbate neural circuit reorganization in amyloid precursor protein intracellular domain transgenic mice.

The cleavage of amyloid precursor protein (APP) by presenilins simultaneously generates amyloid-ß (Aß) and APP intracellular Domain (AICD) peptides. Aß plays a pivotal role in Alzheimer's disease (AD) pathology and recently AICD was also shown to contribute to AD. Transgenic mice overexpressing AICD show age-dependent tau phosphorylation and aggregation, memory deficits, and neurodegeneration. Moreover, these mice show aberrant electrical activity and silent seizures beginning at 3-4 months of age. Here we show that AICD mice also displayed abnormal mossy fiber sprouting beginning about the same time and that this sprouting intensified as the animals aged. Expression of neuropeptide Y was increased in mossy fiber terminals in aged but not young AICD mice. Importantly, young AICD mice injected with kainic acid showed similar pathology to that observed in aged AICD mice. These data show that elevated levels of AICD render neurons hypersensitive to stress and induce hippocampal circuit reorganization, which can further exacerbate hyperexcitability. These results further demonstrate that AICD, in addition to Aß, can play a significant role in AD pathogenesis.

Amyloid-independent mechanisms in Alzheimer's disease pathogenesis.

Despite the progress of the past two decades, the cause of Alzheimer's disease (AD) and effective treatments against it remain elusive. The hypothesis that amyloid-ß (Aß) peptides are the primary causative agents of AD retains significant support among researchers. Nonetheless, a growing body of evidence shows that Aß peptides are unlikely to be the sole factor in AD etiology. Evidence that Aß/amyloid-independent factors, including the actions of AD-related genes, also contribute significantly to AD pathogenesis was presented in a symposium at the 2010 Annual Meeting of the Society for Neuroscience. Here we summarize the studies showing how amyloid-independent mechanisms cause defective endo-lysosomal trafficking, altered intracellular signaling cascades, or impaired neurotransmitter release and contribute to synaptic dysfunction and/or neurodegeneration, leading to dementia in AD. A view of AD pathogenesis that encompasses both the amyloid-dependent and -independent mechanisms will help fill the gaps in our knowledge and reconcile the findings that cannot be explained solely by the amyloid hypothesis.

When neurogenesis encounters aging and disease.

In this review, we consider the evidence that a reduction in neurogenesis underlies aging-related cognitive deficits and impairments in disorders such as Alzheimer's disease (AD). The molecular and cellular alterations associated with impaired neurogenesis in the aging brain are discussed. Dysfunction of presenilin-1, misprocessing of amyloid precursor protein and toxic effects of hyperphosphorylated tau and ß-amyloid probably contribute to impaired neurogenesis in AD. Because factors such as exercise, environmental enrichment and dietary energy restriction enhance neurogenesis, and protect against age-related cognitive decline and AD, knowledge of the underlying neurogenic signaling pathways could lead to novel therapeutic strategies for preserving brain function. In addition, manipulation of endogenous neural stem cells and stem cell transplantation, as stand-alone or adjunct treatments, seems promising.

APP intracellular domain impairs adult neurogenesis in transgenic mice by inducing neuroinflammation.

BACKGROUND: A devastating aspect of Alzheimer's disease (AD) is the progressive deterioration of memory due to neuronal loss. Amyloid precursor protein (APP) occupies a central position in AD and APP-derived amyloid-beta (Abeta) peptides are thought to play a pivotal role in disease pathogenesis. Nonetheless, it is becoming clear that AD etiology is highly complex and that factors other than Abeta also contribute to AD pathogenesis. APP intracellular domain (AICD) is generated together with Abeta and we recently showed that AICD transgenic mice recapitulate pathological features of AD such as tau hyperphosphorylation, memory deficits and neurodegeneration without increasing the Abeta levels. Since impaired adult neurogenesis is shown to augment memory deficits in AD mouse models, here we examined the status of adult neurogenesis in AICD transgenic mice. METHODOLOGY/PRINCIPAL FINDING: We previously generated transgenic mice co-expressing 59-residue long AICD fragment and its binding partner Fe65. Hippocampal progenitor cell proliferation was determined by BrdU incorporation at 1.5, 3 and 12 months of age. Only male transgenic and their respective wilt type littermate control mice were used. We find age-dependent decrease in BrdU incorporation and doublecortin-positive cells in the dentate gyrus of AICD transgenic mice suggesting impaired adult neurogenesis. This deficit resulted from decreased proliferation and survival, whereas neuronal differentiation remained unaffected. Importantly, this impairment was independent of Abeta since APP-KO mice expressing AICD also exhibit reduced neurogenesis. The defects in adult neurogenesis are prevented by long-term treatment with the non-steroidal anti-inflammatory agents ibuprofen or naproxen suggesting that neuroinflammation is critically involved in impaired adult neurogenesis in AICD transgenic mice. CONCLUSION/SIGNIFICANCE: Since adult neurogenesis is crucial for spatial memory, which is particularly vulnerable in AD, these findings suggest that AICD can exacerbate memory defects in AD by impairing adult neurogenesis. Our findings further establish that AICD, in addition to Abeta, contributes to AD pathology and that neuroinflammation plays a much broader role in AD pathogenesis than previously thought.

Alzheimer's disease-like pathological features in transgenic mice expressing the APP intracellular domain.

The hypothesis that amyloid-beta (Abeta) peptides are the primary cause of Alzheimer's disease (AD) remains the best supported theory of AD pathogenesis. Yet, many observations are inconsistent with the hypothesis. Abeta peptides are generated when amyloid precursor protein (APP) is cleaved by presenilins, a process that also produces APP intracellular domain (AICD). We previously generated AICD-overexpressing transgenic mice that showed abnormal activation of GSK-3beta, a pathological feature of AD. We now report that these mice exhibit additional AD-like characteristics, including hyperphosphorylation and aggregation of tau, neurodegeneration and working memory deficits that are prevented by treatment with lithium, a GSK-3beta inhibitor. Consistent with its potential role in AD pathogenesis, we find AICD levels to be elevated in brains from AD patients. The in vivo findings that AICD can contribute to AD pathology independently of Abeta have important therapeutic implications and may explain some observations that are discordant with the amyloid hypothesis.

Amyloid precursor protein is required for convergent-extension movements during Zebrafish development.

Amyloid precursor protein (APP) has been a focus of intense investigation because of its role in Alzheimer's disease (AD), however, its biological function remains uncertain. Loss of APP and APP-like proteins results in postnatal lethality in mice, suggesting a role during embryogenesis. Here we show that in a zebrafish model system, knock down of APP results in the generation of fish with dramatically reduced body length and a short, curly tail. In situ examination of gene expression suggests that the APP morphant embryos have defective convergent-extension movements. We also show that wild-type human APP rescues the morphant phenotype, but the Swedish mutant APP, which causes familial AD (fAD), does not rescue the developmental defects. Collectively, this work demonstrates that the zebrafish model is a powerful system to define the role of APP during embryonic development and to evaluate the functional activity of fAD mutant APP.

Reassessing the amyloid cascade hypothesis of Alzheimer's disease.

Since its inception, the amyloid cascade hypothesis has dominated the field of Alzheimer's disease (AD) research and has provided the intellectual framework for therapeutic intervention. Although the details of the hypothesis continue to evolve, its core principle has remained essentially unaltered. It posits that the amyloid-beta peptides, derived from amyloid precursor protein (APP), are the root cause of AD. Substantial genetic and biochemical data support this view, and yet a number of findings also run contrary to its tenets. The presence of familial AD mutations in APP and presenilins, demonstration of Abeta toxicity, and studies in mouse models of AD all support the hypothesis, whereas the presence of Abeta plaques in normal individuals, the uncertain nature of the pathogenic Abeta species, and repeated disappointments with Abeta-centered therapeutic trials are inconsistent with the hypothesis. The current state of knowledge does not prove nor disprove the amyloid hypothesis, but rather points to the need for its reassessment. A view that Abeta is one of the factors, as opposed to the factor, that causes AD is more consistent with the present knowledge, and is more likely to promote comprehensive and effective therapeutic strategies.