Differential DNA methylation in somatic and sperm cells of hatchery vs wild (natural-origin) steelhead trout populations

Abstract

Environmental factors such as nutrition, stress, and toxicants can influence epigenetic programming and phenotypes of a wide variety of species from plants to humans. The current study was designed to investigate the impacts of hatchery spawning and rearing on steelhead trout (Oncorhynchus mykiss) vs the wild fish on a molecular level. Additionally, epigenetic differences between feeding practices that allow slow growth (2 years) and fast growth (1 year) hatchery trout were investigated. The sperm and red blood cells (RBC) from adult male slow growth/maturation hatchery steelhead, fast growth/maturation hatchery steelhead, and wild (natural-origin) steelhead were collected for DNA preparation to investigate potential alterations in differential DNA methylation regions (DMRs) and genetic mutations, involving copy number variations (CNVs). The sperm and RBC DNA both had a large number of DMRs when comparing the hatchery vs wild steelhead trout populations. The DMRs were cell type specific with negligible overlap. Slow growth/maturation compared to fast growth/maturation steelhead also had a larger number of DMRs in the RBC samples. A number of the DMRs had associated genes that were correlated to various biological processes and pathologies. Observations demonstrate a major epigenetic programming difference between the hatchery and wild natural-origin fish populations, but negligible genetic differences. Therefore, hatchery conditions and growth/maturation rate can alter the epigenetic developmental programming of the steelhead trout. Interestingly, epigenetic alterations in the sperm allow for potential epigenetic transgenerational inheritance of phenotypic variation to future generations. The impacts of hatchery exposures are not only important to consider on the fish exposed, but also on future generations and evolutionary trajectory of fish in the river populations.

Keywords: epigenetic inheritance; fish; generational; genomics; hatchery; phenotypic variation; rearing conditions; sperm; steelhead.

Figure 1:

Hatchery locations, rivers, adjacent dams, and steelhead trout. (a) Map of Methow River and Columbia River confluence, and Winthrop hatchery. (b) Steelhead males (hatchery-origin)

Figure 2:

sperm DMR N1 vs H chromosomal locations and analysis. (a) DMR chromosomal locations on the individual chromosomes vs size of chromosomes. All DMRs at a P-value threshold of P < 1e−05 are shown with red arrowheads and clusters DMRs with black boxes. In addition, 212 DMRs were located on the unplaced concatenated scaffolds not shown. (b) Sperm comparisons DMR overlaps, P < 1e−05. (c) Principal component analysis (PCA) of sperm DMRs N1 vs H. (d) Permutation analysis of sperm DMRs N1 vs H. (e) The number of sperm DMRs at different CpG densities. All DMRs at a P-value threshold of 1e−05 are shown. (f) The DMR lengths (kb) for sperm DMRs. All DMRs at a P-value threshold of 1e−05 are shown

Figure 3:

RBC DMR N1 vs H chromosomal locations and analysis. (a) DMR chromosomal locations on the individual chromosomes vs size of chromosomes. All DMRs at a P-value threshold of P < 1e−05 are shown with red arrowheads and DMR clusters with black boxes. In addition, 175 DMRs were located on the unplaced concatenated scaffolds not shown. (b) RBC comparisons DMR overlaps, P < 1e−05. (c) Principal component analysis (PCA) of RBC DMRs N1 vs H. (d) Permutation analysis of RBC DMRs N1 vs H. (e) The number of RBC DMRs at different CpG densities. All DMRs at a P-value threshold of 1e−05 are shown. (f) The DMR lengths (kb) for RBC DMRs. All DMRs at a P-value threshold of 1e−05 are shown

Figure 4:

DMR chromosomal locations on the individual chromosomes with red arrowheads indicating DMR and black boxes clusters of DMRs. All DMRs at a P-value threshold of P < 1e−05 are shown. (a) Sperm N2 vs S1 DMRs: 174 DMRs were located on the unplaced concatenated scaffold. (b) Sperm N2 vs S2 DMRs: 300 DMRs were located on the unplaced concatenated scaffold. (c) RBC N2 vs S1 DMRs: 110 DMRs were located on the unplaced concatenated scaffold. (d) RBC N2 vs S2 DMRs: 130 DMRs were located on the unplaced concatenated scaffold. Principal component analysis (PCA) using only DMR sites. (e) Sperm DMRs N2 vs S1 (f) Sperm DMRs N2 vs S2. (g) RBC DMRs N2 vs S1. (h) RBC DMRs N2 vs S2. Legend inserts with color N2 or S1 or S2

Figure 5:

DMR overlaps (a) DMR overlap between sperm and RBC DMR (N1 vs H and N2 vs S2). (b) Sperm and RBC overlap P < 1e−05 N2 vs S1 and N2 vs S2. (c) Extended DMR overlap P < 1e−05 vs P < 0.05. The horizontal line indicating the number of DMR overlap and percentage at P < 0.05. Gray highlight is anticipated 100% overlap and yellow highlight overlaps mentioned in the text

Figure 6:

DMR associated gene categories. The gene classification is listed and correlated to the number of DMR associated genes within the specific classification category for (a) sperm DMR associated gene categories and (b) RBC DMR associated gene categories. (c) Pathways and processes with multiple genes

Figure 7:

Sperm DMR associated gene correlations. Cellular localization of associated genes with processes and pathologies in box

Figure 8:

RBC DMR associated gene correlations. Cellular localization of associated genes with processes and pathologies in box