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Practical Guidelines for High-Resolution Epigenomic Profiling of Nucleosomal Histones in Postmortem Human Brain Tissue.

ABSTRACT: BACKGROUND:The nervous system may include more than 100 residue-specific posttranslational modifications of histones forming the nucleosome core that are often regulated in cell-type-specific manner. On a genome-wide scale, some of the histone posttranslational modification landscapes show significant overlap with the genetic risk architecture for several psychiatric disorders, fueling PsychENCODE and other large-scale efforts to comprehensively map neuronal and nonneuronal epigenomes in hundreds of specimens. However, practical guidelines for efficient generation of histone chromatin immunoprecipitation followed by deep sequencing (ChIP-seq) datasets from postmortem brains are needed. METHODS:Protocols and quality controls are given for the following: 1) extraction, purification, and NeuN neuronal marker immunotagging of nuclei from adult human cerebral cortex; 2) fluorescence-activated nuclei sorting; 3) preparation of chromatin by micrococcal nuclease digest; 4) ChIP for open chromatin-associated histone methylation and acetylation; and 5) generation and sequencing of ChIP-seq libraries. RESULTS:We present a ChIP-seq pipeline for epigenome mapping in the neuronal and nonneuronal nuclei from the postmortem brain. This includes a stepwise system of quality controls and user-friendly data presentation platforms. CONCLUSIONS:Our practical guidelines will be useful for projects aimed at histone posttranslational modification mapping in chromatin extracted from hundreds of postmortem brain samples in cell-type-specific manner.

SUBMITTER: Kundakovic M

PROVIDER: S-EPMC5017897 | biostudies-literature | 2017 Jan

REPOSITORIES: biostudies-literature

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Practical Guidelines for High-Resolution Epigenomic Profiling of Nucleosomal Histones in Postmortem Human Brain Tissue.

Kundakovic Marija M Jiang Yan Y Kavanagh David H DH Dincer Aslihan A Brown Leanne L Pothula Venu V Zharovsky Elizabeth E Park Royce R Jacobov Rivka R Magro Isabelle I Kassim Bibi B Wiseman Jennifer J Dang Kristen K Sieberts Solveig K SK Roussos Panos P Fromer Menachem M Harris Brent B Lipska Barbara K BK Peters Mette A MA Sklar Pamela P Akbarian Schahram S

Biological psychiatry 20160309 2

<h4>Background</h4>The nervous system may include more than 100 residue-specific posttranslational modifications of histones forming the nucleosome core that are often regulated in cell-type-specific manner. On a genome-wide scale, some of the histone posttranslational modification landscapes show significant overlap with the genetic risk architecture for several psychiatric disorders, fueling PsychENCODE and other large-scale efforts to comprehensively map neuronal and nonneuronal epigenomes in ...[more]

PMID: 27113501

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Project description:The MAPT gene, encoding the microtubule-associated protein tau on chromosome 17q21.31, is result of an inversion polymorphism, leading to two allelic variants (H1 and H2). Homozygosity for the more common haplotype H1 is associated with an increased risk for several tauopathies, but also for the synucleinopathy Parkinson's disease (PD). In the present study, we aimed to clarify whether the MAPT haplotype influences expression of MAPT and SNCA, encoding the protein α-synuclein (α-syn), on mRNA and protein levels in postmortem brains of PD patients and controls. We also investigated mRNA expression of several other MAPT haplotype-encoded genes. Postmortem tissues from cortex of fusiform gyrus (ctx-fg) and of the cerebellar hemisphere (ctx-cbl) of neuropathologically confirmed PD patients (n = 95) and age- and sex-matched controls (n = 81) were MAPT haplotype genotyped to identify cases homozygous for either H1 or H2. Relative expression of genes was quantified using real-time qPCR; soluble and insoluble protein levels of tau and α-syn were determined by Western blotting. Homozygosity for H1 versus H2 was associated with increased total MAPT mRNA expression in ctx-fg regardless of disease state. Inversely, H2 homozygosity was associated with markedly increased expression of the corresponding antisense MAPT-AS1 in ctx-cbl. PD patients had higher levels of insoluble 0N3R and 1N4R tau isoforms regardless of the MAPT genotype. The increased presence of insoluble α-syn in PD patients in ctx-fg validated the selected postmortem brain tissue. Our findings in this small, but well controlled cohort of PD and controls support a putative biological relevance of tau in PD. However, we did not identify any link between the disease-predisposing H1/H1 associated overexpression of MAPT with PD status. Further studies are required to gain a deeper understanding of the potential regulatory role of MAPT-AS1 and its association to the disease-protective H2/H2 condition in the context of PD.

Dataset Information

Practical Guidelines for High-Resolution Epigenomic Profiling of Nucleosomal Histones in Postmortem Human Brain Tissue.

Publications

Practical Guidelines for High-Resolution Epigenomic Profiling of Nucleosomal Histones in Postmortem Human Brain Tissue.

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