GSE163300
Programmed suppression of oxidative phosphorylation and mitochondrial function by gestational alcohol exposure correlate with global increases in H3K9me2 that do not suppress transcription
Purpose: A critical question emerging in the field of developmental toxicology is whether alterations in chromatin structure induced by toxicant exposure control patterns of gene expression or instead are structural changes that arise due to a nuclear stress response. Previously, we used a mouse model to conduct a three-way comparison between control offspring, alcohol-exposed but phenotypically normal animals, and alcohol-exposed offspring exhibiting craniofacial and central nervous system structural defects.In the cerebral cortex of animals exhibiting alcohol-induced dysgenesis, we identified a dramatic increase in the enrichment of dimethylated histone H3, lysine 9 (H3K9me2) within the regulatory regions of key developmental factors driving histogenesis in the brain. Whether this change in chromatin structure is causally involved in the development of structural defects remains unknown. Methods: We isolated total RNA from the GD17 cortex using the RNeasy Plus Mini Kit (catalog # 74134, Qiagen) according to the manufacturer's instructions. Before RNASeq library preparation, we randomized samples and generated sequencing libraries using the TruSeq RNA Sample Preparation kit (catalog # RS-122-2001, Illumina). We sequenced pooled samples using an Illumina HiSeq 2500 at Whitehead Genomic Services (Cambridge, MA). The sequencing data were demultiplexed, aligned using STAR with default parameters (Dobin et al., 2013), and referenced against the Mus musculus genome (UCSC version mm10). Results: To identify transcriptional differences distinguishing affected fetuses from unaffected offspring, we returned to our previously published, early gestational binge model of maternal alcohol exposure (Veazey et al., 2015). Here, dams were intraperitoneally administered injections of either vehicle or alcohol on gestational-day seven. On gestational-day 17 (GD17), stage-matched control and alcohol- exposed fetuses were dissected and scored for both ocular and craniofacial congenital defects. After determining the sex of each fetus using PCR analysis targeting the Y-chromosome, we found that the majority of affected samples (13/17; 76%) were female. We then isolated RNA from the GD17 fetal cortex of three (n=3) control, EtOH-exposed unaffected, and EtOH-exposed affected offspring and conducted deep-sequencing analysis of the cortex transcriptome. Our sequencing analyses identified a small number (88) of differentially expressed transcripts between the control and unaffected samples. In contrast, comparisons between control and affected, as well as unaffected and affected, identified 876 (412 downregulated, 522 upregulated) and 2281 (1432 downregulated, 1041 upregulated) differentially expressed genes, respectively Conclusions: Alterations in the genetic pathways regulating oxidative phosphorylation and mitochondrial dysfunction distinguish alcohol-exposed affected vs. unaffected offspring. Overall design: Cortical mRNA profiles of gestation day 17 alcohol exposed offspring