Environmental Toxins and Uterine Gene Expression

描述（由申请人提供）：子宫的早期和晚期雌激素反应已经被认识了60多年，但其调节机制仍然存在争议。一种观点认为，发生在前6小时内的早期事件为子宫后期（18-30小时）DNA合成、细胞增殖和蛋白质合成的增加做好了准备。另一种观点认为，增长后期是刺激持续存在的结果。讨论这两个概念时，通常假设所有的反应都依赖于配体与两种雌激素受体同种异构体（ER1和ER2）之一的相互作用。然而，在缺乏ER1或所有er活性被雌激素拮抗剂ICI 182780 （ICI）抑制的小鼠中，注射雌激素或异种雌激素后基因表达增加，表明这是一种过度简化。在这方面，越来越多的证据表明，雌激素通过雌激素依赖和不依赖的方式调节子宫生物学中多种相互依赖的信号通路。虽然我们长期以来的假设是雌激素的某些早期反应是er独立的，但晚期反应是ER1依赖的，两个阶段之间的相互作用对于子宫内雌激素反应的充分补充是必要的。我们之前和初步的观察表明，在几个这样的早期基因中，Bip和Sik-SP是由雌二醇- 172 （E2）和kepone在小鼠子宫中通过er不依赖的方式诱导的，作为i期反应。此外，我们最近发现Bip在介导涉及ER1功能的E2或酮依赖的雌激素信号传导中是至关重要的。我们的初步研究也表明E2诱导GPR30和ERK1/2磷酸化而不涉及ER1。此外，我们观察到，在E2注射前30分钟给予选择性ERK1/2激活抑制剂（SL327），可以消除早期ER独立基因，以及早期和晚期反应期的子宫湿重。相比之下，E2注射后6小时注射抑制剂没有改变晚期效应。总的来说，这些结果使我们能够研究早期和晚期雌激素反应，以确定这些阶段之间的分子关系。我们的具体目的是阐明：1。E2或kepone参与子宫中Sik-SP、Bip和GPR30的早期分子信号传导。2. Sik-SP和GPR30在E2或酮介导的子宫晚期雌激素反应中的功能意义。研究应该会在er1介导的雌激素信号传导过程中产生涉及Bip、Sik-SP和GPR30的新的分子见解。总的来说，这些结果将有助于我们确定子宫内两期雌激素反应之间的关系和分子串扰。公共卫生相关性：我们的具体目标是阐明：E2或kepone参与子宫中Sik-SP、Bip和GPR30的早期分子信号传导。2. Sik-SP和GPR30在E2或酮介导的子宫晚期雌激素反应中的功能意义。研究应该会产生涉及Bip、Sik-SP和GPR30调控er1介导的雌激素信号的新的分子见解。总的来说，这些结果将有助于我们确定子宫内两期雌激素反应之间的关系和分子串扰。

DESCRIPTION (provided by applicant): Early- and late-phase estrogenic responses in the uterus have been recognized for more than 60 years, yet mechanisms involved in their regulation remain controversial. One concept is that an early events(s), occurring within the first 6 h, prepares the uterus for later (18-30 h) increase in DNA synthesis, cell proliferation and protein synthesis. An alternate view is that the late growth phase is a result of the continuous presence of a stimulus. Discussion of either concept usually makes the assumption that all of the responses are dependent upon ligand interaction with one of the two estrogen receptor isoforms (ER1 and ER2). However, increased gene expression following injection of estrogen or xenoestrogen, in mice lacking ER1, or in which all ER-activity has been suppressed by an estrogen-antagonist, ICI 182,780 (ICI), has shown this to be an oversimplification. In this regard, accumulating evidence suggests that estrogen regulates diverse but interdependent signaling pathways in uterine biology via ER- dependent and -independent manners. While our long standing hypothesis is that estrogenic certain early responses are ER-independent, late responses are ER1- dependent, and a cross-talk between the two phases is necessary for a full complement of estrogenic responses in the uterus. Our previous and preliminary observations suggest that among several such early genes, Bip and Sik-SP are induced by estradiol- 172 (E2) and kepone in the mouse uterus via ER-independent manner as a phase-I response. Moreover, we recently showed that Bip is critically necessary in mediating E2- or kepone-dependent estrogen signaling that involves ER1 functions. Our preliminary studies also indicated that E2 induces GPR30 and ERK1/2 phoshorylation without involving ER1. Furthermore, we observed that the administration of a selective inhibitor of ERK1/2 activation (SL327), 30 min prior to E2 injection abrogates early ER- independent genes, as well as uterine wet weights during the early and late responsive phases. In contrast, injection of the inhibitor 6 h after E2 injection did not alter late effects. Collectively, these results positioned us to study early and late estrogenic responses to determine molecular relationship between the phases. Our specific aims are to elucidate: 1. Early molecular signaling by E2 or kepone involving Sik-SP, Bip and GPR30 in the uterus. 2. Functional significance of Sik-SP and GPR30 in E2- or kepone- mediated late estrogenic responses in the uterus. Studies should yield new molecular insights involving Bip, Sik-SP and GPR30 during ER1-mediated estrogen signaling. Overall, the results will help us to define relationship and a molecular cross-talk between the two-phase estrogenic responses in the uterus. PUBLIC HEALTH RELEVANCE: Our specific aims are to elucidate: 1. Early molecular signaling by E2 or kepone involving Sik-SP, Bip and GPR30 in the uterus. 2. Functional significance of Sik-SP and GPR30 in E2 or kepone mediated late estrogenic responses in the uterus. Studies should yield new molecular insights involving Bip, Sik-SP and GPR30 regulation of ER1-mediated estrogen signaling. Overall, the results will help us to define the relationship and a molecular cross-talk between the two-phase estrogenic responses in the uterus.