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United States Department of Agriculture

Agricultural Research Service

Title: Analysis of Cpg Methylation in the Killifish Cyp1a Promoter

Authors
item Timme-Laragy, A - DUKE UNIVERSITY
item Meyer, J - DUKE UNIVERSITY
item WATERLAND, ROBERT
item Giulio, R - DUKE UNIVERSITY

Submitted to: Comparative Biochemistry and Physiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: April 20, 2005
Publication Date: August 20, 2005
Citation: Timme-Laragy, A.R., Meyer, J.N., Waterland, R.A., Giulio, R.T. 2005. Analysis of CpG methylation in the killifish CYP1A promoter. Comparative Biochemistry And Physiology - Part C: Toxicology & Pharmacology. 141(4):406-411.

Interpretive Summary: Killifish inhabiting a creosote-contaminated site on the Elizabeth River exhibit a lack of induction of cytochrome P4501A (CYP1A). This 'refractory CYP1A phenotype' is heritable, but does not appear to be genetically based. To determine if CpG methylation at the CYP1A promoter might explain the heritability of the refractory CYP1A phenotype, we compared CYP1A promoter methylation in liver DNA from the Elizabeth River and a non-contaminated reference site. The CYP1A promoter was completely unmethylated in all the fish, indicating that DNA methylation at CYP1A does not explain the heritability of the refractory CYP1A phenotype.

Technical Abstract: Fundulus heteroclitus (Atlantic killifish or mummichog) inhabiting a creosote-contaminated Superfund site on the Elizabeth River (VA, USA), exhibit a lack of induction of cytochrome P4501A (CYP1A) mRNA, immunodetectable protein, and catalytic activity after exposure to typical inducers. This "refractory CYP1A phenotype" is not explained by alterations in mRNA expression of known CYP1A transcription factors. Furthermore, the refractory phenotype is lost progressively during development in laboratory-reared F1 generation fish. Thus, while heritable, the refractory CYP1A phenotype does not appear to be genetically based. To test the hypothesis that cytosine methylation at CpG sites in the promoter region of CYP1A underlies the refractory CYP1A phenotype, we employed bisulfite sequencing to compare the methylation status of CpG sites in the CYP1A promoter region of DNA from killifish from the Elizabeth River and a reference site. We examined genomic DNA both from livers of wild-caught adult killifish and from pools of F1 generation embryos raised in the laboratory. In fish from both the contaminated and the reference site, cytosine methylation was not detectable at any of the 34 CpG sites examined, including 3 that are part of putative xenobiotic response elements.

Last Modified: 8/19/2014
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