Evan Elliott

Resilience to social stress coincides with functional DNA methylation of the Crf gene in adult mice

DNA methylation regulates gene transcription and has been suggested to encode psychopathologies derived from early life stress. We found that methylation regulated the expression of the Crf (also known as Crh) gene and that chronic social stress in adult mice induced long-term demethylation of this genomic region. Demethylation was observed only in the subset of defeated mice that displayed social avoidance and site-specific knockdown of Crf attenuated the stress-induced social avoidance.

The insulin-like growth factor mRNA binding-protein IMP-1 and the Ras-regulatory protein G3BP associate with tau mRNA and HuD protein in differentiated P19 neuronal cells

Tau mRNA is axonally localized mRNA that is found in developing neurons and targeted by an axonal localization signal (ALS) that is located in the 3′UTR of the message. The tau mRNA is trafficked in an RNA–protein complex (RNP) from the neuronal cell body to the distal parts of the axon, reaching as far as the growth cone. This movement is microtubule‐dependent and is observed as granules that contain tau mRNA and additional proteins. A major protein contained in the granule is HuD, an Elav protein family member, which has an identified mRNA binding site on the tau 3′UTR and stabilizes the tau message and several axonally targeted mRNAs. Using GST‐HuD fusion protein as bait, we have identified four proteins contained within the tau RNP, in differentiated P19 neuronal cells. In this work, we studied two of the identified proteins, i.e. IGF‐II mRNA binding protein 1 (IMP‐1), the orthologue of chick β‐actin binding protein‐ZBP1, and RAS‐GAP SH3 domain binding protein (G3BP). We show that IMP‐1 associates with HuD and G3BP‐1 proteins in an RNA‐dependent manner and binds directly to tau mRNA. We also show an RNA–dependent association between G3BP‐1 and HuD proteins. These associations are investigated in relation to the neuronal differentiation of P19 cells.

Chronic stress induces sex-specific alterations in methylation and expression of corticotropin-releasing factor gene in the rat

Background Although the higher prevalence of depression in women than in men is well known, the neuronal basis of this sex difference is largely elusive. Methods Male and female rats were exposed to chronic variable mild stress (CVMS) after which immediate early gene products, corticotropin-releasing factor (CRF) mRNA and peptide, various epigenetic-associated enzymes and DNA methylation of the Crf gene were determined in the hypothalamic paraventricular nucleus (PVN), oval (BSTov) and fusiform (BSTfu) parts of the bed nucleus of the stria terminalis, and central amygdala (CeA). Results CVMS induced site-specific changes in Crf gene methylation in all brain centers studied in female rats and in the male BST and CeA, whereas the histone acetyltransferase, CREB-binding protein was increased in the female BST and the histone-deacetylase-5 decreased in the male CeA. These changes were accompanied by an increased amount of c-Fos in the PVN, BSTfu and CeA in males, and of FosB in the PVN of both sexes and in the male BSTov and BSTfu. In the PVN, CVMS increased CRF mRNA in males and CRF peptide decreased in females. Conclusions The data confirm our hypothesis that chronic stress affects gene expression and CRF transcriptional, translational and secretory activities in the PVN, BSTov, BSTfu and CeA, in a brain center-specific and sex-specific manner. Brain region-specific and sex-specific changes in epigenetic activity and neuronal activation may play, too, an important role in the sex specificity of the stress response and the susceptibility to depression.

BAG-1 Associates with Hsc70·Tau Complex and Regulates the Proteasomal Degradation of Tau Protein

Intraneuronal accumulation of phosphorylated Tau protein is a molecular pathology found in many forms of dementia, including Alzheimer disease. Research into possible mechanisms leading to the accumulation of modified Tau protein and the possibility of removing Tau protein from the system have revealed that the chaperone protein system can interact with Tau and mediate its degradation. Hsp70/Hsc70, a member of the chaperone protein family, interacts with Tau protein and mediates proper folding of Tau and can promote degradation of Tau protein under certain circumstances. However, because Hsp70/Hsc70 has many binding partners that can mediate its activity, there is still much to discover about how Hsp70 acts in vivo to regulate Tau protein. BAG-1, an Hsp70/Hsc70 binding partner, has been implicated as a mediator of neuronal function. In this work we show that BAG-1 associates with Tau protein in an Hsc70-dependent manner. Overexpression of BAG-1 induced an increase in Tau levels, which is shown to be due to an inhibition of protein degradation. We further show that BAG-1 can inhibit the degradation of Tau protein by the 20 S proteasome but does not affect the ubiquitination of Tau protein. RNA-mediated interference depletion of BAG-1 leads to a decrease in total Tau protein levels as well as promoting hyperphosphorylation of the remaining protein. Induction of Hsp70 by heat shock enhanced the increase of Tau levels in cells overexpressing BAG-1 but induced a decrease of Tau levels in cells that were depleted of BAG-1. Finally, BAG-1 is highly expressed in neurons bearing Tau tangles in a mouse model of Alzheimer disease. This data suggests a molecular mechanism through which Tau protein levels are regulated in the cell and possible consequences for the pathology and treatment of Alzheimer disease.

DNA methylation analysis of the autistic brain reveals multiple dysregulated biological pathways

Autism spectrum disorders (ASD) are a group of neurodevelopmental conditions characterized by dysfunction in social interaction, communication and stereotypic behavior. Genetic and environmental factors have been implicated in the development of ASD, but the molecular mechanisms underlying their interaction are not clear. Epigenetic modifications have been suggested as molecular mechanism that can mediate the interaction between the environment and the genome to produce adaptive or maladaptive behaviors. Here, using the Illumina 450 K methylation array we have determined the existence of many dysregulated CpGs in two cortical regions, Brodmann area 10 (BA10) and Brodmann area 24 (BA24), of individuals who had ASD. In BA10 we found a very significant enrichment for genomic areas responsible for immune functions among the hypomethylated CpGs, whereas genes related to synaptic membrane were enriched among hypermethylated CpGs. By comparing our methylome data with previously published transcriptome data, and by performing real-time PCR on selected genes that were dysregulated in our study, we show that hypomethylated genes are often overexpressed, and that there is an inverse correlation between gene expression and DNA methylation within the individuals. Among these genes there were C1Q, C3, ITGB2 (C3R), TNF-α, IRF8 and SPI1, which have recently been implicated in synaptic pruning and microglial cell specification. Finally, we determined the epigenetic dysregulation of the gene HDAC4, and we confirm that the locus encompassing C11orf21/TSPAN32 has multiple hypomethylated CpGs in the autistic brain, as previously demonstrated. Our data suggest a possible role for epigenetic processes in the etiology of ASD.

Brain‐derived neurotrophic factor induces a rapid dephosphorylation of tau protein through a PI‐3Kinase signalling mechanism

The microtubule‐associated protein tau is essential for microtubule stabilization in neuronal axons. Hyperphosphorylation and intracellular fibrillar formation of tau protein is a pathology found in Alzheimers disease (AD) brains, and in a variety of neurodegenerative disorders referred to as ‘taupathies’. In the present study, we investigated how brain‐derived neurotrophic factor (BDNF), an extracellular factor that is down‐regulated in AD brains, affects tau phosphorylation. BDNF stimulation of neuronally differentiated P19 mouse embryonic carcinoma cells resulted in a rapid decrease in tau phosphorylation, at phosphorylation sites recognized by Tau1, AT8, AT180 and p262‐Tau antibodies. K252a, a tyrosine receptor kinase (Trk) inhibitor, attenuated this dephosphorylation event, suggesting that BNDF activation of TrkB is responsible for the tau dephosphorylation. In addition, BDNF had no affect on tau phosphorylation in the presence of wortmannin, a PI‐3Kinase inhibitor, or lithium, a GSK3β inhibitor, suggesting that these two kinases are part of the signaling transduction cascade leading from TrkB receptor activation to tau dephosphorylation. These results suggest a link between a correlate of AD, decrease in BDNF levels and an AD pathology, tau hyperphosphorylation.

E3 ligase STUB1/CHIP regulates NAD(P)H:quinone oxidoreductase 1 (NQO1) accumulation in aged brain, a process impaired in certain Alzheimer disease patients.

NAD(P)H:quinone oxidoreductase 1 (NQO1) is a flavoenzyme that is important in maintaining the cellular redox state and regulating protein degradation. The NQO1 polymorphism C609T has been associated with increased susceptibility to various age-related pathologies. We show here that NQO1 protein level is regulated by the E3 ligase STUB1/CHIP (C terminus of Hsc70-interacting protein). NQO1 binds STUB1 via the Hsc70-interacting domain (tetratricopeptide repeat domain) and undergoes ubiquitination and degradation. We demonstrate here that the product of the C609T polymorphism (P187S) is a stronger STUB1 interactor with increased susceptibility to ubiquitination by the E3 ligase STUB1. Furthermore, age-dependent decrease of STUB1 correlates with increased NQO1 accumulation. Remarkably, examination of hippocampi from Alzheimer disease patients revealed that in half of the cases examined the NQO1 protein level was undetectable due to C609T polymorphism, suggesting that the age-dependent accumulation of NQO1 is impaired in certain Alzheimer disease patients.

The Interaction between the Immune System and Epigenetics in the Etiology of Autism Spectrum Disorders.

Recent studies have firmly established that the etiology of autism includes both genetic and environmental components. However, we are only just beginning to elucidate the environmental factors that might be involved in the development of autism, as well as the molecular mechanisms through which they function. Mounting epidemiological and biological evidence suggest that prenatal factors that induce a more activated immune state in the mother are involved in the development of autism. In parallel, molecular studies have highlighted the role of epigenetics in brain development as a process susceptible to environmental influences and potentially causative of autism spectrum disorders (ASD). In this review, we will discuss converging evidence for a multidirectional interaction between immune system activation in the mother during pregnancy and epigenetic regulation in the brain of the fetus that may cooperate to produce an autistic phenotype. This interaction includes immune factor-induced changes in epigenetic signatures in the brain, dysregulation of epigenetic modifications specifically in genomic regions that encode immune functions, and aberrant epigenetic regulation of microglia. Overall, the interaction between immune system activation in the mother and the subsequent epigenetic dysregulation in the developing fetal brain may be a main consideration for the environmental factors that cause autism.

Hypomethylation of miR-142 promoter and upregulation of microRNAs that target the oxytocin receptor gene in the autism prefrontal cortex

BackgroundMicroRNAs are small RNA molecules that regulate the translation of protein from gene transcripts and are a powerful mechanism to regulate gene networks. Next-generation sequencing technologies have produced important insights into gene transcription changes that occur in the brain of individuals diagnosed with autism spectrum disorder (asd). However, these technologies have not yet been employed to uncover changes in microRNAs in the brain of individuals diagnosed with asd.MethodsSmall RNA next-generation sequencing was performed on RNA extracted from 12 human autism brain samples and 12 controls. Real-time PCR was used to validate a sample of the differentially expressed microRNAs, and bioinformatic analysis determined common pathways of gene targets. MicroRNA expression data was correlated to genome-wide DNA methylation data to determine if there is epigenetic regulation of dysregulated microRNAs in the autism brain. Luciferase assays, real-time PCR, and Western blot analysis were used to determine how dysregulated microRNAs may regulate the expression and translation of an autism-related gene transcript.ResultsWe determined that miR-142-5p, miR-142-3p, miR-451a, miR-144-3p, and miR-21-5p are overexpressed in the asd brain. Furthermore, the promoter region of the miR-142 gene is hypomethylated in the same brain samples, suggesting that epigenetics plays a role in dysregulation of microRNAs in the brain. Bioinformatic analysis revealed that these microRNAs target genes that are involved in synaptic function. Further bioinformatic analysis, coupled with in vitro luciferase assays, determined that miR-451a and miR-21-5p can target the oxytocin receptor (OXTR) gene. OXTR gene expression is increased in these same brain samples, and there is a positive correlation between miR-21-5p and OXTR expression. However, miR-21-5p expression negatively correlates to production of OXTR protein from the OXTR transcript. Therefore, we suggest that miR-21-5p may attenuate OXTR expression in the human autism brain.ConclusionsOur data suggests that dysregulation of microRNAs may play a biological role in the brain of individuals of autism. In addition, we suggest an interaction between epigenetic mechanisms and microRNA dysregulation in the brain. Overall, this data adds an important link in our understanding of the molecular events that are dysregulated in the brain of individuals diagnosed with autism.

BAG-1M is up-regulated in hippocampus of Alzheimer's disease patients and associates with tau and APP proteins.

The accumulation of tau and amyloid beta proteins is the major molecular pathology of Alzheimer’s disease (AD). The mechanisms leading to the accumulation of these proteins are not completely clear. Hsc‐70/Hsp‐70, a chaperone protein, has been shown to bind both these proteins and regulate their degradation. We have previously shown that the co‐chaperone protein BAG‐1 can inhibit the degradation of tau by forming a complex with Hsc‐70 and tau. In this current work, we show that there is an increase in the BAG‐1M isoform in the hippocampus of AD patients. In addition, BAG‐1 binds to both tau and amyloid precursor protein physically, and is found highly expressed in the same neurons that contain intracellular tau or amyloid in hippocampal sections from AD patients. Over‐expression of BAG‐1M in cell culture also induced an increase in both tau and amyloid precursor protein levels. In conclusion, we report a specific increase of BAG‐1M in human AD patients, which is both physically and functionally associated to the two major molecular markers of AD.