Humans and other species have suffered adverse health effects thought to result from exposure to environmental chemicals that interact with the endocrine system. A significant portion of these effects have been manifested as decreased reproductive potential - often through their effects in utero leading to abnormal development of the reproductive tract. In humans and rodents these effects have been clearly demonstrated with diethylstilbestrol (DES). HOX genes are highly evolutionary conserved and impart segmental tissue identity during the development of undifferentiated body axes. We have demonstrated that HOX genes play an essential role in reproductive tract development. In the prior funding period we have shown that HOX gene expression is altered in mice after in utero exposure to DES, methoxychlor or bisphenol A. We will continue to use DES as our model endocrine disrupter due to its well characterized effects, however will also include these additional agents in selected experiments. The objective of this proposal is to test the hypothesis that the mechanism by which endocrine disrupters affect the development of the reproductive tract is by altering the epigenetic regulation of HOX gene expression. While the initial regulation is mediated through the estrogen receptor (ER a or (b), persistent defects in HOX gene expression after exposure to endocrine disrupters suggests epigenetic alteration of HOX expression. In this application we will determine the molecular mechanisms that regulate HOXA10 and HOXA11 as well as identify epigenetic modifications that regulate HOX genes in both mice and humans. First we will characterize the 5' and intronic regulatory regions of HOXA10 and HOXA11 and identify transcription factor binding sites. Based on preliminary data, we hypothesize that these regulatory regions are methylated in response to xenoestrogen exposure. We will identify the potential impact of DES induced methylation on transcriptional regulation of these genes. Second, we will define the epigenetic changes that lead to persistent Hox gene alteration in the absence of continued exposure in mice. Finally, we will determine if the molecular mechanism by which endocrine disrupters alter HOX gene expression in mice is conserved in humans. We will examine uterine tissue from women with known in utero DES exposure. We have previously shown that Hox genes are necessary for reproductive tract development and that altered Hox expression leads to developmental or functional alterations. Here we expect to demonstrate that endocrine disrupters alter HOX gene expression and will determine the molecular mechanisms that mediate this regulation. No good model exists to explain the mechanism of both the acute and long term diminished HOX gene expression following endocrine disrupter exposure in utero. We hypothesize that the effects are maintained long after exposure through epigenetic mechanisms such as methylation of HOX genes. ? ? ?
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