Environmental Toxins and Uterine Gene Expression

描述（由申请人提供）：子宫内的早期和晚期雌激素反应已被认识超过60年，但其调节机制仍存在争议。一个概念是，在前6小时内发生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准备。另一种观点认为，后期增长阶段是刺激持续存在的结果。对这两个概念的讨论通常假设所有的反应都依赖于配体与两种雌激素受体亚型（ER 1和ER 2）之一的相互作用。然而，在缺乏ER 1或所有ER活性均被雌激素拮抗剂ICI 182，780（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ER依赖性和非依赖性方式调节子宫生物学中多样化但相互依存的信号通路。虽然我们长期的假设是雌激素的某些早期反应是ER独立的，晚期反应是ER 1依赖的，并且两个阶段之间的串扰对于子宫中雌激素反应的完全补充是必要的。我们先前的和初步的观察表明，在几个这样的早期基因，Bip和Sik-SP诱导雌二醇-172（E2）和酮在小鼠子宫通过ER非依赖性的方式作为一个I相反应。此外，我们最近表明，Bip是至关重要的介导E2或酮依赖性雌激素信号，涉及ER 1功能。我们的初步研究还表明，E2诱导GPR 30和ERK 1/2磷酸化，而不涉及ER 1。此外，我们观察到在E2注射前30分钟给予ERK 1/2活化的选择性抑制剂（SL 327）消除了早期ER非依赖性基因，以及在早期和晚期反应阶段期间的子宫湿重。与此相反，注射抑制剂6小时后，E2注射并没有改变晚期效应。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，以确定阶段之间的分子关系。我们的具体目标是阐明：1。E2或kepone在子宫中涉及Sik-SP、Bip和GPR 30的早期分子信号传导。2. Sik-SP和GPR 30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产生新的分子见解，涉及Bip，Sik-SP和GPR 30在ER 1介导的雌激素信号传导。总的来说，这些结果将帮助我们确定子宫内两阶段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：1。E2或kepone在子宫中涉及Sik-SP、Bip和GPR 30的早期分子信号传导。2. Sik-SP和GPR 30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产生新的分子见解，涉及Bip，Sik-SP和GPR 30调节ER 1介导的雌激素信号。总的来说，这些结果将帮助我们确定子宫内两阶段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.