anisomycin and Alzheimer-Disease

Long-term memory requires stable protein synthesis and is altered in Alzheimer's disease (AD). This study aimed to implement a method to measure the cerebral protein synthesis rate (PSR) with [

Topics: Alzheimer Disease; Animals; Anisomycin; Cross-Sectional Studies; Disease Models, Animal; Leucine; Longitudinal Studies; Positron-Emission Tomography; Rats; Rats, Inbred F344; Rats, Wistar

Alzheimer's disease (AD) is a neurodegenerative disease characterized by neurological dysfunction, including memory impairment, attributed to the accumulation of amyloid β (Aβ) in the brain. Although several studies reported possible mechanisms involved in Aβ pathology, much remains unknown. Previous findings suggested that a protein regulated in development and DNA damage response 1 (REDD1), a stress-coping regulator, is an Aβ-responsive gene involved in Aβ cytotoxicity. However, we still do not know how Aβ increases the level of REDD1 and whether REDD1 mediates Aβ-induced synaptic dysfunction. To elucidate this, we examined the effect of Aβ on REDD1-expression using acute hippocampal slices from mice, and the effect of REDD1 short hairpin RNA (shRNA) on Aβ-induced synaptic dysfunction. Lastly, we observed the effect of REDD1 shRNA on memory deficit in an AD-like mouse model. Through the experiments, we found that Aβ-incubated acute hippocampal slices showed increased REDD1 levels. Moreover, Aβ injection into the lateral ventricle increased REDD1 levels in the hippocampus. Anisomycin, but not actinomycin D, blocked Aβ-induced increase in REDD1 levels in the acute hippocampal slices, suggesting that Aβ may increase REDD1 translation rather than transcription. Aβ activated Fyn/ERK/S6 cascade, and inhibitors for Fyn/ERK/S6 or mGluR5 blocked Aβ-induced REDD1 upregulation. REDD1 inducer, a transcriptional activator, and Aβ blocked synaptic plasticity in the acute hippocampal slices. REDD1 inducer inhibited mTOR/Akt signaling. REDD1 shRNA blocked Aβ-induced synaptic deficits. REDD1 shRNA also blocked Aβ-induced memory deficits in passive-avoidance and object-recognition tests. Collectively, these results demonstrate that REDD1 participates in Aβ pathology and could be a target for AD therapy.

Topics: Alzheimer Disease; Amyloid beta-Peptides; Animals; Anisomycin; Dactinomycin; Disease Models, Animal; Extracellular Signal-Regulated MAP Kinases; Hippocampus; Male; MAP Kinase Signaling System; Memory and Learning Tests; Memory Disorders; Mice; Protein Biosynthesis; Proto-Oncogene Proteins c-akt; Proto-Oncogene Proteins c-fyn; Receptor, Metabotropic Glutamate 5; Ribosomal Protein S6 Kinases; RNA, Small Interfering; Synapses; TOR Serine-Threonine Kinases; Transcription Factors; Up-Regulation

Expression of long-lasting synaptic plasticity and long-term memory requires protein synthesis, which can be repressed by phosphorylation of eukaryotic initiation factor 2 α-subunit (eIF2α). Elevated phosphorylation of eIF2α has been observed in the brains of Alzheimer's disease patients and Alzheimer's disease model mice. Therefore, we tested whether suppressing eIF2α kinases could alleviate synaptic plasticity and memory deficits in Alzheimer's disease model mice. Genetic deletion of eIF2α kinase PERK prevented enhanced phosphorylation of eIF2α and deficits in protein synthesis, synaptic plasticity and spatial memory in mice that express familial Alzheimer's disease-related mutations in APP and PSEN1. Similarly, deletion of another eIF2α kinase, GCN2, prevented impairments of synaptic plasticity and defects in spatial memory exhibited by the Alzheimer's disease model mice. Our findings implicate aberrant eIF2α phosphorylation as a previously unidentified molecular mechanism underlying Alzheimer's disease-related synaptic pathophysioloy and memory dysfunction and suggest that PERK and GCN2 are potential therapeutic targets for treatment of individuals with Alzheimer's disease.

Topics: Aged; Aged, 80 and over; Alzheimer Disease; Amyloid beta-Protein Precursor; Animals; Anisomycin; Disease Models, Animal; eIF-2 Kinase; Female; Hippocampus; Humans; In Vitro Techniques; Male; Maze Learning; Memory Disorders; Mice; Mice, Inbred C57BL; Mice, Transgenic; Mutation; Neuronal Plasticity; Presenilin-1; Protein Synthesis Inhibitors; Recognition, Psychology

Oxidative stress and inflammation as the pathological components of Alzheimer's disease (AD) have been well understood. Among a diversity of mitogen-activated protein kinase (MAPK) family members, JNK and p38 MAPK subfamilies are relevant to the response of environmental stress, inflammatory stimuli, or other insults. Recent studies have demonstrated that epigenetic mechanisms may play a pivotal role in AD pathogenesis and development. In the present study, we have investigated epigenetic mechanisms such as DNA methylation and histone acetylation involved in the activation of stress-related signaling pathways for amyloid-β (Aβ) production. Human neuroblastoma SH-SY5Y cells were treated by anisomycin, an activator of stress-related MAPKs (JNK and p38 MAPK). A significant increase of intracellular Aβ level in anisomycin-treated SH-SY5Y cells was observed. The expression of amyloid-β precursor protein (APP), β-site APP-cleaving enzyme 1 (BACE1), and presenilin 1 (PS1) was upregulated by demethylation in three gene promoters associated with the reduction of methyltransferases (DNMTs). Meanwhile, an enhanced level of global histone H3 acetylation accompanied with upregulation of histone acetyltransferases p300/CREB-binding protein (CBP) and downregulation of histone deacetylases (HDACs) was also observed. These findings indicated that the activation of stress-related signaling pathways could result in the increased transcription of APP, BACE1, and PS1 genes through DNMT-dependent hypomethylation and histone H3 hyperacetylation, thus leading to Aβ overproduction. Moreover, our findings provided a novel insight into epigenetic mechanisms by which oxidative stress contributes to the pathogenesis of AD.

Topics: Alzheimer Disease; Amyloid beta-Peptides; Anisomycin; Cell Line, Tumor; Epigenesis, Genetic; Humans; Neuroblastoma; Oxidative Stress; Protein Synthesis Inhibitors

One of the pathological feathers of Alzheimer's disease (AD) is neurofibrillary tangles (NFTs), which consist of paired helical filaments (PHFs) formed by hyperphosphorylated microtubule-associated protein tau. To study the role of mitogen-activated protein kinase (MAPK) in tau hyperphosphorylation and the underlying mechanism, wild type mouse neuroblastoma cells (N2a) were dealt with different concentrations (0.1 microg/mL, 0.2 microg/mL and 0.4 microg/mL) of anisomycin (an activator of MAPK) for 6 h. The relationship between MAPK activity and tau phosphorylation at some Alzheimer-sites was analyzed, and the activities of protein kinase A (PKA) and glycogen synthase kinase-3 (GSK-3) were detected. The results showed that anisomycin activated MAPK in a dose-dependent manner, but tau hyperphosphorylation at Ser-198/199/202 and Ser-396/404 sites was only observed when the concentration of anisomycin was at the level of 0.4 microg/mL, and the alteration of tau phosphorylation at Ser-214 showed no significant difference in different groups. 0.2 microg/mL and 0.4 microg/mL of anisomycin led to an increase in the activity of GSK-3, respectively, but had no effect on the activity of PKA. Lithium chloride, a specific inhibitor of GSK-3, completely abolished the anisomycin-induced elevation of tau phosphorylation without any effect on the activity of MAPK. In conclusion, overactivation of MAPK up to a certain degree induces tau hyperphosphorylation at Ser-198/199/202 and Ser-396/404 sites, and this is probably related to the effect of activated GSK-3 by MAPK.

Topics: Alzheimer Disease; Animals; Anisomycin; Cell Line, Tumor; Cyclic AMP-Dependent Protein Kinases; Glycogen Synthase Kinase 3; Mice; Mitogen-Activated Protein Kinases; Neurofibrillary Tangles; Phosphorylation; tau Proteins

The c-Jun N-terminal kinase (JNK) group of mitogen-activated protein (MAP) kinases are activated by pleiotropic signals including environmental stresses, growth factors, and hormones. JNK-interacting protein 1 (JIP1) is a scaffold protein that assembles and facilitates the activation of the mixed lineage kinase-dependent JNK module and also establishes an interaction with beta-amyloid precursor protein that has been partially characterized. Here we show that, similarly to other proteins involved in various neurological diseases, JIP1 becomes hyperphosphorylated following activation of stress-activated and MAP kinases. By immobilized metal affinity chromatography and a combined microcapillary LC/MALDI-TOF/ESI-ion trap mass spectrometry approach, we identified 35 sites of mitotic phosphorylation within JIP1, among which eight were present within (Ser/Thr)-Pro sequence. This motif is modified by various kinases in aggregates of the microtubule-associated protein tau, which generates typical intraneuronal lesions occurring in Alzheimer disease. Most of the post-translational modifications found were located within the JNK, MAP kinase kinase, and RAC-alpha Ser/Thr protein kinase binding regions; no modifications occurred in protein Src homology 3 and phosphotyrosine interaction domains, which are essential for binding to kinesin, beta-amyloid precursor protein, and MAP kinase kinase kinase. Protein phosphorylation is known to affect stability and protein-protein interactions. Thus, the findings that JIP1 is extensively phosphorylated after activation of stress-activated and MAP kinases indicate that these signaling pathways might modulate JIP1 signaling by regulating its stability and association with some, but not all, interacting proteins.

Topics: Adaptor Proteins, Signal Transducing; Alzheimer Disease; Amino Acid Motifs; Amino Acid Sequence; Amyloid beta-Peptides; Anisomycin; Cells, Cultured; Chromatography, Affinity; Humans; Immunoprecipitation; MAP Kinase Kinase 4; Mitogen-Activated Protein Kinase Kinases; Molecular Sequence Data; Peptide Fragments; Phosphorylation; Protein Binding; Proto-Oncogene Proteins c-akt; Serine; Signal Transduction; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Threonine

Numerous enzymes hyperphosphorylate Tau in vivo, leading to the formation of neurofibrillary tangles (NFTs) in the neurons of Alzheimer's disease (AD). Compared with age-matched normal controls, we demonstrated here that the protein levels of WW domain-containing oxidoreductase WOX1 (also known as WWOX or FOR), its Tyr33-phosphorylated form, and WOX2 were significantly down-regulated in the neurons of AD hippocampi. Remarkably knock-down of WOX1 expression by small interfering RNA in neuroblastoma SK-N-SH cells spontaneously induced Tau phosphorylation at Thr212/Thr231 and Ser515/Ser516, enhanced phosphorylation of glycogen synthase kinase 3beta (GSK-3beta) and ERK, and enhanced NFT formation. Also an increased binding of phospho-GSK-3beta with phospho-Tau was observed in these WOX1 knock-down cells. In comparison, increased phosphorylation of Tau, GSK-3beta, and ERK, as well as NFT formation, was observed in the AD hippocampi. Activation of JNK1 by anisomycin further increased Tau phosphorylation, and SP600125 (a JNK inhibitor) and PD-98059 (an MEK1/2 inhibitor) blocked Tau phosphorylation and NFT formation in these WOX1 knock-down cells. Ectopic or endogenous WOX1 colocalized with Tau, JNK1, and GSK-3beta in neurons and cultured cells. 17Beta-estradiol, a neuronal protective hormone, increased the binding of WOX1 and GSK-3beta with Tau. Mapping analysis showed that WOX1 bound Tau via its COOH-terminal short-chain alcohol dehydrogenase/reductase domain. Together WOX1 binds Tau via its short-chain alcohol dehydrogenase/reductase domain and is likely to play a critical role in regulating Tau hyperphosphorylation and NFT formation in vivo.

Topics: Alzheimer Disease; Animals; Anisomycin; Anthracenes; Brain; Cell Line; Cell Line, Tumor; COS Cells; Down-Regulation; Enzyme Inhibitors; Estradiol; Flavonoids; Genetic Vectors; Glycogen Synthase Kinase 3; Glycogen Synthase Kinase 3 beta; Hippocampus; Humans; Immunohistochemistry; In Vitro Techniques; Mice; Microscopy, Fluorescence; Mitogen-Activated Protein Kinases; Neurons; Oxidoreductases; Phosphorylation; Precipitin Tests; Protein Binding; Protein Structure, Tertiary; Retroviridae; RNA, Small Interfering; Serine; tau Proteins; Temperature; Threonine; Time Factors; Tumor Suppressor Proteins; Two-Hybrid System Techniques; Tyrosine; WW Domain-Containing Oxidoreductase