The pathogenesis of Alzheimer disease (AD) is characterized by the aggregation of amyloid-? (A?) peptides leading to deposition of senile plaques and a progressive decline of cognitive functions, which currently remains the main criterion for its diagnosis. Robust biomarkers for AD do not yet exist, although changes in the cerebrospinal fluid levels of tau and A? represent promising candidates in addition to brain imaging and genetic risk profiling. Although concentrations of soluble A?42 correlate with symptoms of AD, less is known about the biological activities of A? peptides which are generated from the amyloid-? protein precursor. An unbiased DNA microarray study showed that A?42, at sub-lethal concentrations, specifically increases expression of several genes in neuroblastoma cells, notably the insulin-like growth factor binding proteins 3 and 5 (IGFBP3/5), the transcription regulator inhibitor of DNA binding, and the transcription factor Lim only domain protein 4. Using qRT-PCR, we confirmed that mRNA levels of the identified candidate genes were exclusively increased by the potentially neurotoxic A?42 wild-type peptide, as both the less toxic A?40 and a non-toxic substitution peptide A?42 G33A did not affect mRNA levels. In vivo immunohistochemistry revealed a corresponding increase in both hippocampal and cortical IGFBP5 expression in an AD mouse model. Proteomic analyses of human AD cerebrospinal fluid displayed increased in vivo concentrations of IGFBPs. IGFBPs and transcription factors, as identified here, are modulated by soluble A?42 and may represent useful early biomarkers.
Although soluble species of the amyloid-? peptide A?42 correlate with disease symptoms in Alzheimer disease, little is known about the biological activities of amyloid-? (A?). Here, we show that A? peptides varying in lengths from 38 to 43 amino acids are internalized by cultured neuroblastoma cells and can be found in the nucleus. By three independent methods, we demonstrate direct detection of nuclear A?42 as follows: (i) biochemical analysis of nuclear fractions; (ii) detection of biotin-labeled A? in living cells by confocal laser scanning microscopy; and (iii) transmission electron microscopy of A? in cultured cells, as well as brain tissue of wild-type and transgenic APPPS1 mice (overexpression of amyloid precursor protein and presenilin 1 with Swedish and L166P mutations, respectively). Also, this study details a novel role for A?42 in nuclear signaling, distinct from the amyloid precursor protein intracellular domain. Chromatin immunoprecipitation showed that A?42 specifically interacts as a repressor of gene transcription with LRP1 and KAI1 promoters. By quantitative RT-PCR, we confirmed that mRNA levels of the examined candidate genes were exclusively decreased by the potentially neurotoxic A?42 wild-type peptide. Shorter peptides (A?38 or A?40) and other longer peptides (nontoxic A?42 G33A substitution or A?43) did not affect mRNA levels. Overall, our data indicate that the nuclear translocation of A?42 impacts gene regulation, and deleterious effects of A?42 in Alzheimer disease pathogenesis may be influenced by altering the expression profiles of disease-modifying genes.
Protein kinase B (AKT) and glycogen synthase kinase 3 beta (GSK3?) are two protein kinases involved in dopaminergic signaling. Dopamine-associated neuropsychiatric illnesses such as schizophrenia and bipolar disorder seem to be characterized by impairments in the AKT/GSK3? network. Here, we sought evidence for the presence of molecular and functional changes in the AKT/GSK3? pathway using an established infection-based mouse model of developmental neuropsychiatric disease that is based on prenatal administration of the viral mimetic poly(I:C) (=polyriboinosinic-polyribocytidilic acid). We found that adult offspring of poly(I:C)-exposed mothers displayed decreased total levels of AKT protein and reduced phosphorylation at AKT threonine residues in the medial prefrontal cortex. Prenatally immune challenged offspring also exhibited increased GSK3? protein expression and activation status, the latter of which was evidenced by a decrease in the ratio between phosphorylated and total GSK3? protein in the medial prefrontal cortex. These molecular changes were not associated with overt signs of inflammatory processes in the adult brain. We further found that acute pre-treatment with the selective GSK3? inhibitor TDZD-8 dose-dependently normalized aberrant behavior typically emerging following prenatal immune activation, including deficient spontaneous alternation in the Y-maze and increased locomotor responses to systemic amphetamine treatment. Taken together, the present mouse model demonstrates that prenatal exposure to viral-like immune activation leads to long-term alterations in GSK3? signaling, some of which are critically implicated in schizophrenia and bipolar disorder.
Amyloid plaques consisting of aggregated A? peptide are a hallmark of Alzheimers disease. Among the different forms of A?, the one of 42aa length (A?42) is most aggregation-prone and also the most neurotoxic. We find that eye-specific expression of human A?42 in Drosophila results in a degeneration of eye structures that progresses with age. Dietary supplements of zinc or copper ions exacerbate eye damage. Positive effects are seen with zinc/copper chelators, or with elevated expression of MTF-1, a transcription factor with a key role in metal homeostasis and detoxification, or with human or fly transgenes encoding metallothioneins, metal scavenger proteins. These results show that a tight control of zinc and copper availability can minimize cellular damage associated with A?42 expression.
Formation of aberrant protein conformers is a common pathological denominator of different neurodegenerative disorders, such as Alzheimers disease or prion diseases. Moreover, increasing evidence indicates that soluble oligomers are associated with early pathological alterations and that oligomeric assemblies of different disease-associated proteins may share common structural features. Previous studies revealed that toxic effects of the scrapie prion protein (PrP(Sc)), a ?-sheet-rich isoform of the cellular PrP (PrP(C)), are dependent on neuronal expression of PrP(C). In this study, we demonstrate that PrP(C) has a more general effect in mediating neurotoxic signalling by sensitizing cells to toxic effects of various ?-sheet-rich (?) conformers of completely different origins, formed by (i) heterologous PrP, (ii) amyloid ?-peptide, (iii) yeast prion proteins or (iv) designed ?-peptides. Toxic signalling via PrP(C) requires the intrinsically disordered N-terminal domain (N-PrP) and the GPI anchor of PrP. We found that the N-terminal domain is important for mediating the interaction of PrP(C) with ?-conformers. Interestingly, a secreted version of N-PrP associated with ?-conformers and antagonized their toxic signalling via PrP(C). Moreover, PrP(C)-mediated toxic signalling could be blocked by an NMDA receptor antagonist or an oligomer-specific antibody. Our study indicates that PrP(C) can mediate toxic signalling of various ?-sheet-rich conformers independent of infectious prion propagation, suggesting a pathophysiological role of the prion protein beyond of prion diseases.
N-terminally truncated A? peptides starting with pyroglutamate (A?pE3) represent a major fraction of all A? peptides in the brain of Alzheimer disease (AD) patients. A?pE3 has a higher aggregation propensity and stability and shows increased toxicity compared with full-length A?. In the present work, we generated a novel monoclonal antibody (9D5) that selectively recognizes oligomeric assemblies of A?pE3 and studied the potential involvement of oligomeric A?pE3 in vivo using transgenic mouse models as well as human brains from sporadic and familial AD cases. 9D5 showed an unusual staining pattern with almost nondetectable plaques in sporadic AD patients and non-demented controls. Interestingly, in sporadic and familial AD cases prominent intraneuronal and blood vessel staining was observed. Using a novel sandwich ELISA significantly decreased levels of oligomers in plasma samples from patients with AD compared with healthy controls were identified. Moreover, passive immunization of 5XFAD mice with 9D5 significantly reduced overall A? plaque load and A?pE3 levels, and normalized behavioral deficits. These data indicate that 9D5 is a therapeutically and diagnostically effective monoclonal antibody targeting low molecular weight A?pE3 oligomers.
The presence of Abeta(pE3) (N-terminal truncated Abeta starting with pyroglutamate) in Alzheimers disease (AD) has received considerable attention since the discovery that this peptide represents a dominant fraction of Abeta peptides in senile plaques of AD brains. This was later confirmed by other reports investigating AD and Downs syndrome postmortem brain tissue. Importantly, Abeta(pE3) has a higher aggregation propensity, and stability, and shows an increased toxicity compared to full-length Abeta. We have recently shown that intraneuronal accumulation of Abeta(pE3) peptides induces a severe neuron loss and an associated neurological phenotype in the TBA2 mouse model for AD. Given the increasing interest in Abeta(pE3), we have generated two novel monoclonal antibodies which were characterized as highly specific for Abeta(pE3) peptides and herein used to analyze plaque deposition in APP/PS1KI mice, an AD model with severe neuron loss and learning deficits. This was compared with the plaque pattern present in brain tissue from sporadic and familial AD cases. Abundant plaques positive for Abeta(pE3) were present in patients with sporadic AD and familial AD including those carrying mutations in APP (arctic and Swedish) and PS1. Interestingly, in APP/PS1KI mice we observed a continuous increase in Abeta(pE3) plaque load with increasing age, while the density for Abeta(1-x ) plaques declined with aging. We therefore assume that, in particular, the peptides starting with position 1 of Abeta are N-truncated as disease progresses, and that, Abeta(pE3) positive plaques are resistant to age-dependent degradation likely due to their high stability and propensity to aggregate.
Trace amines, compounds structurally related to classical biogenic amines, represent endogenous ligands of the trace amine-associated receptor 1 (TAAR1). Because trace amines also influence the activity of other targets, selective ligands are needed for the elucidation of TAAR1 function. Here we report on the identification and characterization of the first selective and potent TAAR1 partial agonist.
Here, we describe a novel missense mutation in the amyloid precursor protein (APP) causing a lysine-to-asparagine substitution at position 687 (APP770; herein, referred to as K16N according to amyloid-? (A?) numbering) resulting in an early onset dementia with an autosomal dominant inheritance pattern. The K16N mutation is located exactly at the ?-secretase cleavage site and influences both APP and A?. First, due to the K16N mutation APP secretion is affected and a higher amount of A? peptides is being produced. Second, A? peptides carrying the K16N mutation are unique in that the peptide itself is not harmful to neuronal cells. Severe toxicity, however, is evident upon equimolar mixture of wt and mutant peptides, mimicking the heterozygous state of the subject. Furthermore, A?42 K16N inhibits fibril formation of A?42 wild-type. Even more, A?42 K16N peptides are protected against clearance activity by the major A?-degrading enzyme neprilysin. Thus the mutation characterized here harbours a combination of risk factors that synergistically may contribute to the development of early onset Alzheimer disease.
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