Epigenomic convergence of genetic and immune risk factors in neurodevelopmental disorder cortex

Abstract

Neurodevelopmental disorders (NDDs) impact 7% to 14% of all children in developed countries and are one of the leading causes of lifelong disability. Epigenetic modifications are poised at the interface between genes and environment and are predicted to reveal insight into the gene networks, cell types, and developmental timing of NDD etiology. Whole-genome bisulfite sequencing was used to examine DNA methylation in 49 human cortex samples from three different NDDs (autism spectrum disorder, Rett syndrome, and Dup15q syndrome) and matched controls. Integration of methylation differences across NDDs with relevant genomic and genetic datasets revealed differentially methylated regions (DMRs) unique to each type of NDD but with shared regulatory functions in neurons and microglia. DMRs were significantly enriched for known NDD genetic risk factors, including both common inherited and rare de novo variants. Weighted region co-methylation network analysis revealed a module related to NDD diagnosis and enriched for microglial regulatory regions. Together, these results demonstrate an epigenomic signature of NDDs in human cortex shared with known genetic and immune etiological risk. Epigenomic insights into cell types and gene regulatory regions will aid in defining therapeutic targets and early biomarkers at the interface of genetic and environmental NDD risk factors. seconds on/15 seconds off on a Diagenode Bioruptor. DNA was end-repaired using 1× T4 DNA ligase buffer, 400 µM dNTPs, 15 U T4 DNA polymerase (NEB), and 50 U PNK (NEB) for 30 min at 20°C. After PCR purification (Qiagen), adenine bases were appended to the ends using 1× NEB 2 buffer, 200 µM dATP, and 15 U Klenow Fragment (3′ to 5′ exo-, NEB) for 30 min at 37°C. After another DNA purification using the PCR MinElute kit (Qiagen), 3 µL of Illumina's methylated sequencing adapters were ligated on using 1× ligase buffer and 5 µL Quick T4 DNA Ligase (NEB) for 30 min at room temperature. After a final PCR purification, 500 ng of library was bisulfite converted using Zymo's EZ DNA Methylation-Direct kit according to the manufacturer's instructions. The library was then amplified using 2.5 U PfuTurbo Cx Hotstart DNA Polymerase (Stratagene) for 12 cycles using Illumina's standard amplification protocol. The library's quality was assessed on a Bioanalyzer (Agilent) and sequenced (100 bp, singleended) on an Illumina HiSeq 2000. Each biological sample was sequenced on a single lane. Sample Acquisition, DNA Isolation, and WGBS Library Preparation Rett BA9 DNA was extracted from BA9 cortex from RTT and Control samples using the Zymo Duet Kit (Zymo). 200ng of total DNA was bisulfite converting using the Zymo Lighting kit (Zymo) and 50ng of converted DNA was used as input for library construction using the Illumina TruSeq DNA Methylation Kit (Illumina). Each sample was given a unique barcode and 14 cycles of PCR amplification. Final libraries were size selected with two rounds of KAPA Pure Beads: 0.7X Left and 0.65X Left/0.55X Right for a final library size centered around ...

Genome-wide DNA methylation profiling identifies convergent molecular signatures associated with idiopathic and syndromic autism in post-mortem human brain tissue

“…Co-methylation network analyses highlighted systems-level changes in cortical DNA methylation associated with both iASD and dup15q, with associated modules being enriched for sites annotated to genes involved in the immune system, synaptic signalling and neuronal regulation. Our results corroborate findings from other DNA methylation (26,30,31) and gene expression analyses (10,44) that have concluded that ASD-related co-methylation and co-expression modules are significantly enriched for synaptic, neuronal and immune dysfunction genes. Finally, we used existing RNA-seq data on an overlapping set of samples to show a remarkable overlap of cortical ASD-associated DMPs with differential gene expression.…”

Pharmacokinetic-Pharmacodynamic (PK-PD) Modeling of Effect of Naringenin and Its Surface Modified Nanocarriers on Associated and Core Behaviors of Autism Spectrum Disorders (ASD)

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