title: Quantitative methylation level of the EPHX1 promoter in peripheral blood DNA is associated with polycystic ovary syndrome.
abstract: Steroid synthesis and metabolic pathways play important roles in the pathophysiology of PCOS, but until now there have been no studies on the methylation profiles of specific genes in steroid synthesis pathways that are known to be associated with PCOS. Here we used MassARRAY quantitative methylation analysis to determine the methylation levels of each CpG site or cluster in the promoters of EPHX1, SRD5A1, and CYP11A1 in 64 peripheral blood samples. We further examined the methylation level of EPHX1 in an independent cohort consisting of 116 people. Finally, we investigated the role of EPHX1 in steroidogenesis in the KGN cell line. For SRD5A1 and CYP11A1, there was no significant difference in methylation level between patients and controls. For EPHX1, however, the methylation levels of a few consecutive CpG sites and clusters were found to be significantly associated with PCOS. The methylation levels of a number of CpG clusters or sites were significantly lower in patients than in controls in the first cohort consisting of 64 people, such as clusters 13-14 (P<0.05), 15-16 (P<0.001), and 19-24 (P<0.001) and sites CpG_53 (P<0.01) and CpG_54 (P<0.05). Among differentiated methylation sites and clusters, the methylation levels of the CpG cluster 13-14 and CpG cluster 19-24 in PCOS patients were significantly lower than in controls in the second cohort of 116 people (P<0.05 for both). In addition, knockdown and overexpression experiments in KGN cells showed that EPHX1 can regulate estradiol concentrations, and this indicates a role for EPHX1 in steroidogenesis. Our study has demonstrated that methylation of the EPHX1 promoter might be associated with PCOS. This study provides direct evidence that methylation plays an important role in PCOS and demonstrates a novel role for EPHX1 in female reproduction.
authors: State Key Laboratory of Genetic Engineering and MOE Key Laboratory of Contemporary Anthropology, School of Life Sciences, Fudan University, Shanghai, China ; Institutes of Biomedical Science, Fudan University, Shanghai, China.
authors: Qing
authors: Q
authors: Sang
authors: Department of Gynecology, Obstetrics and Gynecology Hospital of Fudan University, Shanghai, China.
authors: Xin
authors: X
authors: Li
authors: State Key Laboratory of Genetic Engineering and MOE Key Laboratory of Contemporary Anthropology, School of Life Sciences, Fudan University, Shanghai, China ; Institutes of Biomedical Science, Fudan University, Shanghai, China.
authors: Haojue
authors: H
authors: Wang
authors: State Key Laboratory of Genetic Engineering and MOE Key Laboratory of Contemporary Anthropology, School of Life Sciences, Fudan University, Shanghai, China ; Institutes of Biomedical Science, Fudan University, Shanghai, China.
authors: Huan
authors: H
authors: Wang
authors: Shanghai Ninth Hospital, Shanghai, China.
authors: Shaozhen
authors: S
authors: Zhang
authors: State Key Laboratory of Genetic Engineering and MOE Key Laboratory of Contemporary Anthropology, School of Life Sciences, Fudan University, Shanghai, China ; Institutes of Biomedical Science, Fudan University, Shanghai, China.
authors: Ruizhi
authors: R
authors: Feng
authors: State Key Laboratory of Genetic Engineering and MOE Key Laboratory of Contemporary Anthropology, School of Life Sciences, Fudan University, Shanghai, China ; Institutes of Biomedical Science, Fudan University, Shanghai, China.
authors: Yao
authors: Y
authors: Xu
authors: State Key Laboratory of Genetic Engineering and MOE Key Laboratory of Contemporary Anthropology, School of Life Sciences, Fudan University, Shanghai, China ; Institutes of Biomedical Science, Fudan University, Shanghai, China.
authors: Qiaoli
authors: Q
authors: Li
authors: State Key Laboratory of Genetic Engineering and MOE Key Laboratory of Contemporary Anthropology, School of Life Sciences, Fudan University, Shanghai, China ; Institutes of Biomedical Science, Fudan University, Shanghai, China.
authors: Xinzhi
authors: X
authors: Zhao
authors: Institutes of Biomedical Science, Fudan University, Shanghai, China ; Bio-X Center, Key Laboratory for the Genetics of Developmental and Neuropsychiatric Disorders, Ministry of Education, Shanghai Jiao Tong University, Shanghai, China.
authors: Qinghe
authors: Q
authors: Xing
authors: State Key Laboratory of Genetic Engineering and MOE Key Laboratory of Contemporary Anthropology, School of Life Sciences, Fudan University, Shanghai, China.
authors: Li
authors: L
authors: Jin
authors: Institutes of Biomedical Science, Fudan University, Shanghai, China ; Bio-X Center, Key Laboratory for the Genetics of Developmental and Neuropsychiatric Disorders, Ministry of Education, Shanghai Jiao Tong University, Shanghai, China.
authors: Lin
authors: L
authors: He
authors: State Key Laboratory of Genetic Engineering and MOE Key Laboratory of Contemporary Anthropology, School of Life Sciences, Fudan University, Shanghai, China ; Institutes of Biomedical Science, Fudan University, Shanghai, China ; Bio-X Center, Key Laboratory for the Genetics of Developmental and Neuropsychiatric Disorders, Ministry of Education, Shanghai Jiao Tong University, Shanghai, China.
authors: Lei
authors: L
authors: Wang
mesh_terms: D013741:3-Oxo-5-alpha-Steroid 4-Dehydrogenase / Q000235:genetics / Q000378:metabolism; D000328:Adult; D002786:Cholesterol Side-Chain Cleavage Enzyme / Q000235:genetics / Q000378:metabolism; D018899:CpG Islands; D019175:DNA Methylation; D004851:Epoxide Hydrolases; D005260:Female; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D011085:Polycystic Ovary Syndrome / Q000097:blood*; D011401:Promoter Regions, Genetic
chemical_list: D008565:Membrane Proteins; D002786:Cholesterol Side-Chain Cleavage Enzyme; D013741:3-Oxo-5-alpha-Steroid 4-Dehydrogenase; C548467:SRD5A1 protein, human; D004851:Epoxide Hydrolases; C522330:EPHX1 protein, human