title: The Methylcytosine Dioxygenase Ten-Eleven Translocase-2 (tet2) Enables Elevated GnRH Gene Expression and Maintenance of Male Reproductive Function.
abstract: Reproduction depends on the establishment and maintenance of elevated GnRH neurosecretion. The elevation of primate GnRH release is accompanied by epigenetic changes. Specifically, cytosine residues within the GnRH gene promoter are actively demethylated, whereas GnRH mRNA levels and peptide release rise. Whether active DNA demethylation has an impact on GnRH neuron development and consequently reproductive function remains unknown. In this study, we investigated whether ten-eleven translocation (tet) enzymes, which initiate the process of active DNA demethylation, influence neuronal function and reproduction. We found that tet2 expression increases with age in the developing mouse preoptic area-hypothalamus and is substantially higher in a mature (GT1-7) than an immature (GN11) GnRH cell line. GnRH mRNA levels and mean GnRH peptide release elevated after overexpression of tet2 in GN11 cells, whereas CRISPR/cas9-mediated knockdown of tet2 in GT1-7 cells led to a significant decline in GnRH expression. Manipulations of tet2 expression altered tet2 genome binding and histone 3 lysine 4 trimethylation abundance at the GnRH promoter. Mice with selective disruption of tet2 in GnRH neurons (GnRH-specific tet2 knockout mice) exhibited no sign of altered pubertal timing in either sex, although plasma LH levels were significantly lower, and fecundity was altered specifically in adult male GnRH-specific tet2 knockout animals, indicating that tet2 may participate in the maintenance GnRH neuronal function. Exposure to bisphenol A, an environmental contaminant that alters GnRH neuron activity, caused a shift in tet2 subcellular localization and a decrease in histone 3 lysine 4 trimethylation abundance at the GnRH promoter. Finally, evaluation of tet2 protein interactions in GT1-7 cells suggests that the influence of tet2 on neuronal function are not limited to nuclear mechanisms but could depend on mitochondrial function, and RNA metabolism. Together, these studies implicate tet2 in the maintenance of GnRH neuronal function and neuroendocrine control of male reproduction.
authors.affiliation: Department of Obstetrics and Gynecology (J.R.K., S.L.), Southern Illinois University School of Medicine, Springfield, Illinois 62794; St. John's Hospital Carol Jo Vecchie Women and Children's Center (J.R.K.), Springfield, Illinois 62769; Wisconsin National Primate Research Center (K.L.K., E.T., J.E.L.), Madison, Wisconsin 53705; Department of Pediatrics and Physiology (A.W.), The Johns Hopkins University School of Medicine, Baltimore, Maryland 21287; and Departments of Pediatrics (E.T.) and Neuroscience (J.E.L.), University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin 53706.
authors.forename: Joseph R
authors.initials: JR
authors.lastname: Kurian
authors.affiliation: Department of Obstetrics and Gynecology (J.R.K., S.L.), Southern Illinois University School of Medicine, Springfield, Illinois 62794; St. John's Hospital Carol Jo Vecchie Women and Children's Center (J.R.K.), Springfield, Illinois 62769; Wisconsin National Primate Research Center (K.L.K., E.T., J.E.L.), Madison, Wisconsin 53705; Department of Pediatrics and Physiology (A.W.), The Johns Hopkins University School of Medicine, Baltimore, Maryland 21287; and Departments of Pediatrics (E.T.) and Neuroscience (J.E.L.), University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin 53706.
authors.forename: Somaja
authors.initials: S
authors.lastname: Louis
authors.affiliation: Department of Obstetrics and Gynecology (J.R.K., S.L.), Southern Illinois University School of Medicine, Springfield, Illinois 62794; St. John's Hospital Carol Jo Vecchie Women and Children's Center (J.R.K.), Springfield, Illinois 62769; Wisconsin National Primate Research Center (K.L.K., E.T., J.E.L.), Madison, Wisconsin 53705; Department of Pediatrics and Physiology (A.W.), The Johns Hopkins University School of Medicine, Baltimore, Maryland 21287; and Departments of Pediatrics (E.T.) and Neuroscience (J.E.L.), University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin 53706.
authors.forename: Kim L
authors.initials: KL
authors.lastname: Keen
authors.affiliation: Department of Obstetrics and Gynecology (J.R.K., S.L.), Southern Illinois University School of Medicine, Springfield, Illinois 62794; St. John's Hospital Carol Jo Vecchie Women and Children's Center (J.R.K.), Springfield, Illinois 62769; Wisconsin National Primate Research Center (K.L.K., E.T., J.E.L.), Madison, Wisconsin 53705; Department of Pediatrics and Physiology (A.W.), The Johns Hopkins University School of Medicine, Baltimore, Maryland 21287; and Departments of Pediatrics (E.T.) and Neuroscience (J.E.L.), University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin 53706.
authors.forename: Andrew
authors.initials: A
authors.lastname: Wolfe
authors.affiliation: Department of Obstetrics and Gynecology (J.R.K., S.L.), Southern Illinois University School of Medicine, Springfield, Illinois 62794; St. John's Hospital Carol Jo Vecchie Women and Children's Center (J.R.K.), Springfield, Illinois 62769; Wisconsin National Primate Research Center (K.L.K., E.T., J.E.L.), Madison, Wisconsin 53705; Department of Pediatrics and Physiology (A.W.), The Johns Hopkins University School of Medicine, Baltimore, Maryland 21287; and Departments of Pediatrics (E.T.) and Neuroscience (J.E.L.), University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin 53706.
authors.forename: Ei
authors.initials: E
authors.lastname: Terasawa
authors.affiliation: Department of Obstetrics and Gynecology (J.R.K., S.L.), Southern Illinois University School of Medicine, Springfield, Illinois 62794; St. John's Hospital Carol Jo Vecchie Women and Children's Center (J.R.K.), Springfield, Illinois 62769; Wisconsin National Primate Research Center (K.L.K., E.T., J.E.L.), Madison, Wisconsin 53705; Department of Pediatrics and Physiology (A.W.), The Johns Hopkins University School of Medicine, Baltimore, Maryland 21287; and Departments of Pediatrics (E.T.) and Neuroscience (J.E.L.), University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin 53706.
authors.forename: Jon E
authors.initials: JE
authors.lastname: Levine
mesh_terms: D000818:Animals; D001559:Benzhydryl Compounds; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000378:metabolism*; D049308:Dioxygenases; D005260:Female; D018507:Gene Expression Regulation, Developmental; D007987:Gonadotropin-Releasing Hormone / Q000502:physiology*; D042421:Histone Code; D006801:Humans; D008297:Male; D051379:Mice; D009474:Neurons / Q000378:metabolism; D010636:Phenols; D011301:Preoptic Area / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D012098:Reproduction; D000098826:Bisphenol A Compounds
chemical_list: D001559:Benzhydryl Compounds; D004268:DNA-Binding Proteins; D049308:Dioxygenases; D007987:Gonadotropin-Releasing Hormone; D010636:Phenols; D011518:Proto-Oncogene Proteins; C551914:Tet2 protein, mouse; C006780:bisphenol A; D000098826:Bisphenol A Compounds