MethylomeDB: a database of DNA methylation profiles of the brain

MethylomeDB (http://epigenomics.columbia.edu/methylomedb/index.html) is a new database containing genome-wide brain DNA methylation profiles. DNA methylation is an important epigenetic mark in the mammalian brain. In human studies, aberrant DNA methylation alterations have been associated with various neurodevelopmental and neuropsychiatric disorders such as schizophrenia, and depression. In this database, we present methylation profiles of carefully selected non-psychiatric control, schizophrenia, and depression samples. We also include data on one mouse forebrain sample specimen to allow for cross-species comparisons. In addition to our DNA methylation data generated in-house, we have and will continue to include published DNA methylation data from other research groups with the focus on brain development and function. Users can view the methylation data at single-CpG resolution with the option of wiggle and microarray formats. They can also download methylation data for individual samples. MethylomeDB offers an important resource for research into brain function and behavior. It provides the first source of comprehensive brain methylome data, encompassing whole-genome DNA methylation profiles of human and mouse brain specimens that facilitate cross-species comparative epigenomic investigations, as well as investigations of schizophrenia and depression methylomes

Calcineurin interacts with the serotonin transporter C-terminus to modulate its plasma membrane expression and serotonin uptake

Homeostasis of serotonergic transmission critically depends on the rate of serotonin reuptake via its plasma membrane transporter (SERT). SERT activity is tightly regulated by multiple mechanisms, including physical association with intracellular proteins and post-translational modifications, such as phosphorylation, but these mechanisms remain partially understood. Here, we show that SERT C-terminal domain recruits both the catalytic and regulatory subunits of the Ca(2+)-activated protein phosphatase calcineurin (CaN) and that the physical association of SERT with CaN is promoted by CaN activity. Coexpression of constitutively active CaN with SERT increases SERT cell surface expression and 5-HT uptake in HEK-293 cells. It also prevents the reduction of 5-HT uptake induced by an acute treatment of cells with the protein kinase C activator β-PMA and concomitantly decreases PMA-elicited SERT phosphorylation. In addition, constitutive activation of CaN in vivo favors 5-HT uptake in the adult mouse brain, whereas CaN inhibition reduces cerebral 5-HT uptake. Constitutive activation of CaN also decreases immobility in the forced swim test, indicative of an antidepressant-like effect of CaN. These results identify CaN as an important regulator of SERT activity in the adult brain and provide a novel molecular substrate of clinical interest for the understanding of increased risk of mood disorders in transplanted patients treated with immunosuppressive CaN inhibitors

Genome-Wide Divergence of DNA Methylation Marks in Cerebral and Cerebellar Cortices

Emerging evidence suggests that DNA methylation plays an expansive role in the central nervous system (CNS). Large-scale whole genome DNA methylation profiling of the normal human brain offers tremendous potential in understanding the role of DNA methylation in brain development and function.Using methylation-sensitive SNP chip analysis (MSNP), we performed whole genome DNA methylation profiling of the prefrontal, occipital, and temporal regions of cerebral cortex, as well as cerebellum. These data provide an unbiased representation of CpG sites comprising 377,509 CpG dinucleotides within both the genic and intergenic euchromatic region of the genome. Our large-scale genome DNA methylation profiling reveals that the prefrontal, occipital, and temporal regions of the cerebral cortex compared to cerebellum have markedly different DNA methylation signatures, with the cerebral cortex being hypermethylated and cerebellum being hypomethylated. Such differences were observed in distinct genomic regions, including genes involved in CNS function. The MSNP data were validated for a subset of these genes, by performing bisulfite cloning and sequencing and confirming that prefrontal, occipital, and temporal cortices are significantly more methylated as compared to the cerebellum.These findings are consistent with known developmental differences in nucleosome repeat lengths in cerebral and cerebellar cortices, with cerebrum exhibiting shorter repeat lengths than cerebellum. Our observed differences in DNA methylation profiles in these regions underscores the potential role of DNA methylation in chromatin structure and organization in CNS, reflecting functional specialization within cortical regions

Minimising Immunohistochemical False Negative ER Classification Using a Complementary 23 Gene Expression Signature of ER Status

BACKGROUND: Expression of the oestrogen receptor (ER) in breast cancer predicts benefit from endocrine therapy. Minimising the frequency of false negative ER status classification is essential to identify all patients with ER positive breast cancers who should be offered endocrine therapies in order to improve clinical outcome. In routine oncological practice ER status is determined by semi-quantitative methods such as immunohistochemistry (IHC) or other immunoassays in which the ER expression level is compared to an empirical threshold. The clinical relevance of gene expression-based ER subtypes as compared to IHC-based determination has not been systematically evaluated. Here we attempt to reduce the frequency of false negative ER status classification using two gene expression approaches and compare these methods to IHC based ER status in terms of predictive and prognostic concordance with clinical outcome. METHODOLOGY/PRINCIPAL FINDINGS: Firstly, ER status was discriminated by fitting the bimodal expression of ESR1 to a mixed Gaussian model. The discriminative power of ESR1 suggested bimodal expression as an efficient way to stratify breast cancer; therefore we identified a set of genes whose expression was both strongly bimodal, mimicking ESR expression status, and highly expressed in breast epithelial cell lines, to derive a 23-gene ER expression signature-based classifier. We assessed our classifiers in seven published breast cancer cohorts by comparing the gene expression-based ER status to IHC-based ER status as a predictor of clinical outcome in both untreated and tamoxifen treated cohorts. In untreated breast cancer cohorts, the 23 gene signature-based ER status provided significantly improved prognostic power compared to IHC-based ER status (P = 0.006). In tamoxifen-treated cohorts, the 23 gene ER expression signature predicted clinical outcome (HR = 2.20, P = 0.00035). These complementary ER signature-based strategies estimated that between 15.1% and 21.8% patients of IHC-based negative ER status would be classified with ER positive breast cancer. CONCLUSION/SIGNIFICANCE: Expression-based ER status classification may complement IHC to minimise false negative ER status classification and optimise patient stratification for endocrine therapies

Modulation fonctionnelle de deux cibles des antidépresseurs, le transporteur de la sérotonine et le récepteur 5-HT2c (rôles respectifs de la NO-synthase neuronale et des agonistes inverses)

MONTPELLIER-BU Médecine UPM (341722108) / SudocMONTPELLIER-BU Médecine (341722104) / SudocPARIS-BIUP (751062107) / SudocSudocFranceF

PACAP type I receptor transactivation is essential for IGF-1 receptor signalling and antiapoptotic activity in neurons.: IGF-1 transactivates PACAP type I receptors

Insulin-like growth factor-1 (IGF-1) and pituitary adenylyl cyclase activating polypeptide (PACAP) are both potent neurotrophic and antiapoptotic factors, which exert their effects via phosphorylation cascades initiated by tyrosine kinase and G-protein-coupled receptors, respectively. Here, we have adapted a recently described phosphoproteomic approach to neuronal cultures to characterize the phosphoproteomes generated by these neurotrophic factors. Unexpectedly, IGF-1 and PACAP increased the phosphorylation state of a common set of proteins in neurons. Using PACAP type 1 receptor (PAC1R) null mice, we showed that IGF-1 transactivated PAC1Rs constitutively associated with IGF-1 receptors. This effect was mediated by Src family kinases, which induced PAC1R phosphorylation on tyrosine residues. PAC1R transactivation was responsible for a large fraction of the IGF-1-associated phosphoproteome and played a critical role in the antiapoptotic activity of IGF-1. Hence, in contrast to the general opinion that the trophic activity of IGF-1 is solely mediated by tyrosine kinase receptor-associated signalling, we show that it involves a more complex signalling network dependent on the PAC1 Gs-protein-coupled receptor in neurons

Serotonin 2B Receptor by Interacting with NMDA Receptor and CIPP Protein Complex May Control Structural Plasticity at Glutamatergic Synapses

International audienceThe serotonin 2B (5-HT2B) receptor coupled to Gq-protein contributes to the control of neuronal excitability and is implicated in various psychiatric disorders. The mechanisms underlying its brain function are not fully described. Using peptide affinity chromatography combined with mass spectrometry, we found that the PDZ binding motif of the 5-HT2B receptor located at its C-terminal end interacts with the scaffolding protein channel interacting PDZ protein (CIPP). We then showed, in COS-7 cells, that the association of the 5-HT2B receptor to CIPP enhanced receptor-operated inositol phosphate (IP) production without affecting its cell surface and intracellular levels. Co-immunoprecipitation experiments revealed that CIPP, the 5-HT2B receptor, and the NR1 subunit of the NMDA receptor form a macromolecular complex. CIPP increased 5-HT2B receptor clustering at the surface of primary cultured hippocampal neurons and prevented receptor dispersion following agonist stimulation, thus potentiating IP production and intracellular calcium mobilization in dendrites. CIPP or 5-HT2B receptor stimulation in turn dispersed NR1 clusters colocalized with 5-HT2B receptors and increased the density and maturation of dendritic spines. Collectively, our results suggest that the 5-HT2B receptor, the NMDA receptor, and CIPP may form a signaling platform by which serotonin can influence structural plasticity of excitatory glutamatergic synapses

DNA demethylation of neuronal cell death genes in depression

Background: DNA methylation may play a role in the etiology of neuropsychiatric disorders through abnormal genomic methylation patterns regulating genes involved in brain development or physiology. In this study we explored the DNA methylation profile of depression in the prefrontal cortex because converging evidence from brain imaging and postmortem studies has implicated this region in depression neuropathology. Materials and methods: In order to better understand both the wild type genomic DNA methylation patterns and aberrant methylation events occurring in disease states we profiled DNA methylation patterns in human postmortem brains of 12 depressed and non-psychiatric controls using the methylation mapping and paired-end sequencing (Methyl-MAPS) method. Methyl-MAPS is an enzymatic-base method that can delineate the methylation status of greater than 80% of CpG sites genome-wide (Xin et al., doi: 10.4161/epi.6.11.17876). Overall, these data represent relatively unbiased coverage of the genome, including CpG-rich domains such as CpG islands and repetitive elements. Results and conclusions: Comparative analysis of DNA methylation patterns among depressed cases and controls revealed little variation at the transcription start sites (TSS) across all RefSeq annotated genes, but significant variations were detected proximal to the TSS (referred to as “CpG island shores”). We observed statistically significant methylation loss in CpG island shores in depressed cases compared to controls. These findings were replicated in purified neuronal cell populations. Using an independent sample of depressed cases and matched non-psychiatric controls, we isolated neuronal nuclei from the dorsal prefrontal cortex of 11 depressed cases and 11 controls. Due to limited quantities of neuronal DNA typically obtained from isolation of nuclei using fluorescence-activated cell sorting, we used the Illumina HumanMethylation450 BeadChip. DNA methylation differences in CpG island shores revealed that, of the CpG dinucleotides with significant methylation differences, >95% showed loss of methylation in depressed brains. The underlying mechanism involved in the loss of methylation in depression psychopathology is unclear. However, the global 5-hydroxymethylcytosine levels in neuronal DNA from the same sample specimens also showed a loss of hydroxymethylation in depressed brains compared to controls. Although these data were not statistically significant, they revealed an important trend in loss of hydroxymethylation and the possible mechanism for DNA demethylation in brains of depressed patients. Gene ontology analysis of genes with significant methylation differences (primarily loss of methylation) in depressed vs. controls identified a number of cellular functions. Of note, the fourth most significant gene set identified was involved in programmed cell death and 74% of the genes in this set were associated with neuronal cell death. These changes in methylation dynamics suggest a possible mechanism linking neuronal cell death associated with oxidative stress and inflammation in the depressed brain