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Developmental profiles and expression of the DNA methyltransferase genes in the fathead minnow (Pimephales promelas) following exposure to di-2-ethylhexyl phthalate

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Abstract

DNA methylation is an epigenetic regulator of gene expression, and this process has been shown to be disrupted by environmental contaminants. Di-2-(ethylhexyl) phthalate (DEHP) and related phthalate esters have been shown to affect development in early life stages of fish and can alter genomic methylation patterns in vertebrates. The objectives of this study were the following: (1) Describe the expression patterns of the DNA methyltransferase (dnmt) genes during early fathead minnow (FHM) development. These genes are critical for methylation and imprinting during development. (2) Determine the effects of DEHP on the development of FHM larvae [1 and 14 days post-hatch (dph)]. (3) Determine the effect of DEHP on dnmt expression and global methylation status in larval FHM. FHMs were first collected over a developmental time course [1, 3, 5, 6, and 14 days post-fertilization (dpf)] to investigate the expression patterns of five dnmt isoforms. The expression of dnmt1 and dnmt7 was relatively high in embryos at 1 dpf but was variable in expression, and these transcripts were later expressed at a lower level (>3 dpf); dnmt3 was significantly higher in embryos at 1 dpf compared to those at 3 dpf. Dnmt6 showed more of a constitutive pattern of expression during the first 2 weeks of development, and the mRNA levels of dnmt8 were higher in embryos at 5 and 6 dpf compared to those at 1 and 3 dpf, corresponding to the hatching period of the embryos. A waterborne exposure to three concentrations of DEHP (1, 10 and 100 µg/L) was conducted on 1-day FHM embryos for 24 h and on larval fish for 2 weeks, ending at 14 dpf. DEHP did not negatively affect survival, hatch rate, or the expression of dnmt isoforms in FHMs. There were no differences in global cytosine methylation following DEHP treatments in 14 dpf larvae, suggesting that environmentally relevant levels of DEHP may not affect global methylation at this stage of FHM development. However, additional targeted methylome studies are required to determine whether specific gene promoters are differently methylated following exposure to DEHP. This study offers new insight into the roles of the dnmt enzymes during FHM development.

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Acknowledgments

The authors have no conflict of interest to declare. This research was funded by a Natural Science and Engineering Research Council (NSERC) Discovery Grant to CJM (386275-2010) and an NSERC USRA to RKW. We thank Rosalinda Knight, Jennifer Loughery, and Kathleena Sarty for their assistance with FHM rearing.

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  1. Christopher J. Martyniuk

    Present address: Department of Physiological Sciences, Center for Environmental and Human Toxicology, University of Florida Genetics Institute, University of Florida, Gainesville, FL, 32611, USA

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  1. Department of Biology, Canadian Rivers Institute, University of New Brunswick, Saint John, NB, E2L 4L5, Canada

    Richard K. Wood, Emma Crowley & Christopher J. Martyniuk

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Wood, R.K., Crowley, E. & Martyniuk, C.J. Developmental profiles and expression of the DNA methyltransferase genes in the fathead minnow (Pimephales promelas) following exposure to di-2-ethylhexyl phthalate. Fish Physiol Biochem 42, 7–18 (2016). https://doi.org/10.1007/s10695-015-0112-3

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