Ionizing Radiation Affects Epigenetic Programming in Young Adult Mice

Humans are exposed to low and mild doses of radiation frequently, ranging from the natural environment to medical procedures like x-ray and CT scans. Ionizing radiation of various doses has been known to cause not only cellular and genomic changes, but specific neurological systems such as the limbic system have been indicated to be particularly vulnerable. Here, we demonstrated that epigenetics is also altered by radiation. Epigenetics is a subtle chemical coding above the gene, which plays a critical role in brain development, and downstream can cause the onset of cognitive aberrations and other neurological impairments. How radiation as an external environmental factor causes epigenetic changes is not clearly understood. DNA methylation, including 5-methylcytosine (5M) and 5-hydroxymethylcytosine (5-hmC) have been shown to either suppress or activate gene transcription and as such are key epigenetic players. To elucidate the role of radiation in epigenetic outcomes, we examined epigenetic, phenotypic and transcriptional markers via immunohistochemistry, in the hippocampus and cortex. In this study C57BL/6 mouse (postnatal day 21 (P21)) began a 4-week radiation treatment of various doses totaling (2Gy-4.5Gy) via global head targeting CT exposure. We found a loss of 5M and 5-hmC as well as transcriptional markers within regions of the hippocampus and cortex. There was a significant decrease in cell proliferation in the hippocampus- specifically, in the region responsible for adult neurogenesis. The cingulate cortex (a region adjacent to the hippocampus) also exhibited dramatic alterations in several epigenetic and transcriptional markers, indicating the vulnerability of the limbic system in radiation exposure. Understanding the mechanism by which ionizing radiation affects epigenetic programming will provide insight into the transmissibility of external factors to biological systems. Additionally, this work can aid the development of protective strategies against the harmful risks associated with radiation exposure

Ionizing Radiation Affects Epigenetic Programming in Adolescent Mice

poster abstractHumans are exposed to low and mild doses of radiation frequently, ranging from the natural environment to medical procedures like x-ray and CT scans. Ionizing radiation of various doses has been known to potentially cause not only cellular but also genomic changes. Here, we demonstrate that epigenetics is also altered by the radiation. Epigenetics is a chemical coding above the gene, which plays critical roles in brain development, cognitive aberrations and other neurological impairments. How radiation, as an external environmental factor, causes epigenetic change is not understood. DNA methylation, key in epigenetics, including 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) have been shown to either suppress or activate gene transcription. To aid in elucidating the role in which radiation affects epigenetic outcomes, we examined the effects of radiation on both epigenetic and phenotypic markers within the hippocampus. In this study we treated, via x-ray C57BL/6 mice, postnatal day (P) 21 with various doses (2Gy-4.5Gy) of radiation coupled with varying frequencies (0.5 Gy x 4, 1.5 Gy x 3, or 4.5Gy x 1) during a 4-week period. We used immunohistochemistry staining with cell proliferation, transcription and epigenetic markers. We found loss of 5mC in the sub-granular layer of the dentate gyrus (DG) in the upper and lower arms. Likewise a loss of 5hmC in the sub-granular layer of the DG, as well as in the cornu Ammonis (CA) layers 1 and 2. There was also loss of a transcriptional activation marker within the DG of the hippocampus. Furthermore, decreased cell proliferation in the adult neurogenesis in the hippocampus was found. Exposure to ionizing radiation altered the normal epigenetic profile of the mice. Understanding the mechanism by which ionizing radiation affects epigenetic programming will provide insight into how to develop protection against the potentially harmful risks associated with radiation exposure

Cis‐acting allele specific expression (ASE) differences induced by alcohol and impacted by sex as well as parental genotype of origin

Background Alcohol use disorders (AUDs) are influenced by complex interactions between the genetics of the individual and their environment. We have previously identified hundreds of polygenic genetic variants between the selectively bred high and low alcohol drinking (HAD and LAD) rat lines. Here we report allele specific expression (ASE) differences, between the HAD2 and LAD2 rat lines. Methods The HAD2 and LAD2 rats which have been sequenced were reciprocally crossed to generate 10 litters of F1 progeny. For 5 of these litters, the sire was HAD2; and, for the other 5 litters, the sire was a LAD2. From these 10 litters, two males and two females were picked from each F1 litter (N = 40 total). The F1‐pups were divided, with balancing for sex and direction of cross, into an alcohol (15%) vs a water control group. Alcohol‐drinking started in the middle of adolescence (~PND 35) and lasted 9 weeks. At the end of these treatments, rats were euthanized, the nucleus accumbens was dissected, and RNA was processed for RNA‐sequencing and ASE analyses. Results Analyses revealed that adolescent ethanol drinking, individual ethanol drinking levels, parentage, and sex‐of‐animal affected ASEs of about 300 genes. The identified genes included those associated with ethanol metabolism (e.g., Aldh2); neuromodulatory function [e.g., Cckbr, Slc6a7, and Slc1a1]; ion channel activity (e.g., Kcnc3); as well as other synaptic and epigenetic function. Conclusion These data indicate that ethanol drinking differentially amplified paternal vs maternal allelic contribution to the transcriptome. We hypothesize that this was due, at least in part, to ethanol‐induced changes in cis‐regulation of polymorphisms previously identified between the HAD2 and LAD2 rat lines. This report highlights the complexity of gene‐by‐environment interactions mediating a genetic predisposition for, and/or the active development of, alcohol use disorders

Localization of the dopamine transporter gene, Dat1 , on mouse Chromosome 13

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/47009/1/335_2004_Article_BF00292340.pd

Localization of the peptidylglycine α-amidating monooxygenase gene ( Pam ) introduces a region of homology between human Chromosome 5q and mouse Chromosome 1

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/47008/1/335_2004_Article_BF00426085.pd

A study of alterations in DNA epigenetic modifications (5mC and 5hmC) and gene expression influenced by simulated microgravity in human lymphoblastoid cells

Cells alter their gene expression in response to exposure to various environmental changes. Epigenetic mechanisms such as DNA methylation are believed to regulate the alterations in gene expression patterns. In vitro and in vivo studies have documented changes in cellular proliferation, cytoskeletal remodeling, signal transduction, bone mineralization and immune deficiency under the influence of microgravity conditions experienced in space. However microgravity induced changes in the epigenome have not been well characterized. In this study we have used Next-generation Sequencing (NGS) to profile ground-based “simulated” microgravity induced changes on DNA methylation (5-methylcytosine or 5mC), hydroxymethylation (5-hydroxymethylcytosine or 5hmC), and simultaneous gene expression in cultured human lymphoblastoid cells. Our results indicate that simulated microgravity induced alterations in the methylome (~60% of the differentially methylated regions or DMRs are hypomethylated and ~92% of the differentially hydroxymethylated regions or DHMRs are hyperhydroxymethylated). Simulated microgravity also induced differential expression in 370 transcripts that were associated with crucial biological processes such as oxidative stress response, carbohydrate metabolism and regulation of transcription. While we were not able to obtain any global trend correlating the changes of methylation/ hydroxylation with gene expression, we have been able to profile the simulated microgravity induced changes of 5mC over some of the differentially expressed genes that includes five genes undergoing differential methylation over their promoters and twenty five genes undergoing differential methylation over their gene-bodies. To the best of our knowledge, this is the first NGS-based study to profile epigenomic patterns induced by short time exposure of simulated microgravity and we believe that our findings can be a valuable resource for future explorations

High Resolution Genomic Scans Reveal Genetic Architecture Controlling Alcohol Preference in Bidirectionally Selected Rat Model

Investigations on the influence of nature vs. nurture on Alcoholism (Alcohol Use Disorder) in human have yet to provide a clear view on potential genomic etiologies. To address this issue, we sequenced a replicated animal model system bidirectionally-selected for alcohol preference (AP). This model is uniquely suited to map genetic effects with high reproducibility, and resolution. The origin of the rat lines (an 8-way cross) resulted in small haplotype blocks (HB) with a corresponding high level of resolution. We sequenced DNAs from 40 samples (10 per line of each replicate) to determine allele frequencies and HB. We achieved ~46X coverage per line and replicate. Excessive differentiation in the genomic architecture between lines, across replicates, termed signatures of selection (SS), were classified according to gene and region. We identified SS in 930 genes associated with AP. The majority (50%) of the SS were confined to single gene regions, the greatest numbers of which were in promoters (284) and intronic regions (169) with the least in exon\u27s (4), suggesting that differences in AP were primarily due to alterations in regulatory regions. We confirmed previously identified genes and found many new genes associated with AP. Of those newly identified genes, several demonstrated neuronal function involved in synaptic memory and reward behavior, e.g. ion channels (Kcnf1, Kcnn3, Scn5a), excitatory receptors (Grin2a, Gria3, Grip1), neurotransmitters (Pomc), and synapses (Snap29). This study not only reveals the polygenic architecture of AP, but also emphasizes the importance of regulatory elements, consistent with other complex traits

Cloning, annotation and developmental expression of the chicken intestinal MUC2 gene

Intestinal mucin 2 (MUC2) encodes a heavily glycosylated, gel-forming mucin, which creates an important protective mucosal layer along the gastrointestinal tract in humans and other species. This first line of defense guards against attacks from microorganisms and is integral to the innate immune system. As a first step towards characterizing the innate immune response of MUC2 in different species, we report the cloning of a full-length, 11,359 bp chicken MUC2cDNA, and describe the genomic organization and functional annotation of this complex, 74.5 kb locus. MUC2 contains 64 exons and demonstrates distinct spatiotemporal expression profiles throughout development in the gastrointestinal tract; expression increases with gestational age and from anterior to posterior along the gut. The chicken protein has a similar domain organization as the human orthologue, with a signal peptide and several von Willebrand domains in the N-terminus and the characteristic cystine knot at the C-terminus. The PTS domain of the chicken MUC2 protein spans ~1600 amino acids and is interspersed with four CysD motifs. However, the PTS domain in the chicken diverges significantly from the human orthologue; although the chicken domain is shorter, the repetitive unit is 69 amino acids in length, which is three times longer than the human. The amino acid composition shows very little similarity to the human motif, which potentially contributes to differences in the innate immune response between species, as glycosylation across this rapidly evolving domain provides much of the musical barrier. Future studies of the function of MUC2 in the innate immune response system in chicken could provide an important model organism to increase our understanding of the biological significance of MUC2 in host defense and highlight the potential of the chicken for creating new immune-based therapies

Evolution of Trefoil Factor(s): Genetic and Spatio- Temporal Expression of Trefoil Factor 2 in the Chicken

Trefoil factors are essential healing initiators participating in mucosal reconstitution and tissue morphogenesis, especially on the surfaces of the gastrointestinal tract. This family has been cloned and characterized predominantly from mammals and amphibians. Avian species ingest stone and grit to help digest food, which may expose their gut to severe physical conditions. To further the understanding of the function of the TFF gene family across species, we undertook this research to clone, sequence, and characterize the spatio-temporal expression patterns of chicken TFF2 (ChTFF2) cDNA. Bioinformatics analysis of the promoter region and deduced amino acid sequence demonstrated that ChTFF2 contained unique characteristics; specifically the chicken promoter has multiple start sites and the protein contains a series of Lys-Lys-Val repeats. Unlike mammals, where TFF2 is detected primarily in the stomach, and occasionally in the proximal duodenum, chicken TFF2 transcripts are found throughout the gastrointestinal tract, with major expression sites in the glandular and muscular stomach as well as evident expression in the colon, small intestine, cecal tonsil and crop. Temporal analysis of intestinal ChTFF2 transcripts by quantitative RT-PCR showed high levels in embryos and a trend of constant expression during embryonic and post-hatch development, with a reduction occurring around hatch. Phylogenetic analysis highlighted the conservation of TFF proteins and functional divergence of trefoil domains, which suggest a transitional role in the bir