Transgenerational disease specific epigenetic sperm biomarkers after ancestral exposure to dioxin

Abstract

Dioxin was historically one of the most common industrial contaminants with several major industry accidents, as well as governmental actions involving military service, having exposed large numbers of the worldwide population over the past century. Previous rat studies have demonstrated the ability of dioxin (2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)) exposure to promote the epigenetic transgenerational inheritance of disease susceptibility in subsequent generations. The types of disease previously observed include puberty abnormalities, testis, ovary, kidney, prostate and obesity pathologies. The current study was designed to use an epigenome-wide association study (EWAS) to identify potential sperm DNA methylation biomarkers for specific transgenerational diseases. Therefore, the transgenerational F3 generation dioxin lineage male rats with and without a specific disease were compared to identify differential DNA methylation regions (DMRs) as biomarkers for disease. The genomic features of the disease-specific DMRs were characterized. Observations demonstrate that disease-specific epimutation DMRs exist for the transgenerational dioxin lineage rats that can potentially be used as epigenetic biomarkers for testis, kidney, prostate and obesity diseases. These disease-specific DMRs were associated with genes that have previously been shown to be linked with the specific diseases. This EWAS for transgenerational disease identified potential epigenetic biomarkers and provides the proof of concept of the potential to develop similar biomarkers for humans to diagnose disease susceptibilities and facilitate preventative medicine.

Keywords: DNA Methylation; Dioxin; EWAS; Epigenetics; Kidney; Obesity; Pathology; Prostate; Sperm; TCDD; Testis; Transgenerational.

Figure 1.

DMR identification and numbers. The number of DMRs found using different p-value cutoff thresholds. The All Window column shows all DMRs. The Multiple Window column shows the number of DMRs containing at least two significant windows (1 kb each). The number of DMRs with the number of significant windows (1 kb per window) at a p-value threshold p<1e-04 for DMR. (A) Prostate disease DMRs; (B) Kidney disease DMRs; (C) Obesity disease DMRs; and (D) Testis disease DMRs.

Figure 2.

DMR chromosomal locations. The DMR locations on the individual chromosomes is represented with an arrowhead and a cluster of DMRs with a black box. All DMRs containing at least one significant window at the selected p-value p<1e-04 threshold are shown. The chromosome number and size of the chromosome (megabase) are presented. (A) Prostate disease DMRs; (B) Kidney disease DMRs; (C) Obesity disease DMRs; and (D) Testis disease DMRs.

Figure 3.

DMR genomic features. The number of DMRs at different CpG densities. All DMRs at a p-value threshold of p<1e-04 are shown. (A) Prostate disease DMR CpG density; (B) Prostate disease DMR length; (C) Kidney disease DMR CpG density; (D) Kidney disease DMR length; (E) Obesity disease DMR CpG density; (F) Obesity disease DMR length; (G) Testis disease DMR CpG density; (H) Testis disease DMR length.

Figure 4.

Overlap of disease DMRs. (A) Overlap of specific disease overlap epimutations p<1e-04. Venn diagram overlap analysis for specific disease states. (B) An extended overlap of disease DMRs. The p-value data set at p<1e-04 is compared to the p<0.05 data to identify potential overlap between the different pathologies with DMR number and percentage of the total presented. The gray highlight is the expected 100% overlap. (C) Overlap of the different disease DMRs at p<0.05. The Venn diagram identified 63 DMRs at p<0.05 in common between the different diseases. (D) Venn diagram overlap of the different diseases DMRs at p<1e-04 with the 63 common overlapping p<0.05 DMRs.

Figure 5.

Disease DMR associated gene categories. (A) DMR associated gene categories. The different gene categories and number of DMRs in each category is presented with a legend indicating the different disease DMR sets. (B) KEGG pathways for the different disease DMR associated genes. The top ten pathways with the number of associated genes in brackets are presented.

Figure 6.

Disease DMR associated gene correlations. The disease DMR associated genes that correlate with the specific disease tissue functions and pathologies for each individual pathology are presented. Direct gene links to pathologies and physiologic processes are shown. (A) Prostate disease and (B) kidney disease.

Figure 7.

Disease DMR associated gene correlations. The disease DMR associated genes that correlate with the specific disease tissue functions and pathologies for each individual pathology are presented. Direct gene links to pathologies and physiologic processes are shown. (A) obesity and (B) testis disease.