Epigenetic Regulation of Myometrial Contractility in Pregnancy and Labor

项目总结/摘要 早产是全世界婴儿出生后前四周死亡的主要原因。的 这项研究的首要目标是提高我们对遗传和表观遗传机制的理解 在妊娠和分娩期间介导子宫肌层静止和收缩，以及它们的失调 导致早产。我们认为，孕酮（P4）/孕酮受体（PR）维持子宫肌层 通过抑制炎性因子（如IL-1b、IL-8、考克斯-2）的表达，在妊娠期大部分时间内保持静止 和收缩（CAP）（例如OXTR、CX 43）基因。我们认为，这是通过两个基本机制发生的：（1） PR可能与转录因子（例如NF-κ B、AP-1）相连，转录因子与炎性基因的启动子结合， 招募辅阻遏物以抑制基因表达;（2）PR也可直接结合编码以下基因的启动子： 转录抑制因子（例如ZEB 1）激活其表达。ZEB 1反过来与CAP启动子结合， 并招募一种抑制复合物来抑制它们的表达。相反，在学期开始时， 和早产时，子宫肌层中的PR功能因其与NF-κ B的直接相互作用而受损， 通过减少PR辅激活因子的表达，将P4代谢为无活性产物的酶的表达 以及通过上调截短的PR同种型（例如PR-A）。截短的PR亚型可能具有减少的 转录和反式阻遏活性。我们推测这些炎症和CAP基因的变化可能与 表达由染色质修饰和结构的改变介导。本提案的目标是 确定P4/PR抑制作用和PR差异作用的基因和机制， A和PR-B对炎症和CAP基因表达的影响，并表征 介导子宫肌层静止，并伴随CAP和炎症基因表达增强， 足月和早产。为了实现这些目标，我们将使用小鼠模型和人类子宫肌层细胞， （1）定义和表征转录因子/辅调节因子复合物的组分， 与妊娠子宫肌层中的PR相互作用以介导其抗炎作用， ZEB 1抑制CAP基因表达;（2）分析这些PR相互作用的 怀孕期间的因素和足月和早产，以及激素和microRNA在其 （3）使用RNA-seq和ChIP-seq来发现对维持子宫肌层细胞增殖至关重要的基因。 妊娠和分娩的启动及其调节的潜在转录机制。RNA-seq 这将有助于发现与妊娠维持和分娩启动有关的基因。将合并的 ChIP-seq的使用将提供对转录和表观遗传机制的全面了解， 这些基因表达的变化。使用PR-B-KO小鼠的研究将阐明PR-A与PR-B-KO小鼠的不同作用。 PR-B在子宫肌层转录组调节中的作用总的来说，我们的发现将揭示新的基因， 这些途径可以作为预防早产及其后果的治疗靶点。

PROJECT SUMMARY/ABSTRACT Preterm birth is the leading cause of infant mortality during the first four weeks of life world-wide. The overarching goal of this research is to enhance our understanding of the genetic and epigenetic mechanisms that mediate myometrial quiescence and contractility during pregnancy and labor, and their dysregulation leading to preterm birth. We suggest that progesterone (P4)/progesterone receptor (PR) maintains myometrial quiescence throughout most of pregnancy by inhibiting expression of inflammatory (e.g. IL-1b, IL-8, COX-2) and contractile (CAP) (e.g. OXTR, CX43) genes. We propose that this occurs via two basic mechanisms: (1) PR may tether to transcription factors (e.g. NF-kB, AP-1) bound to promoters of inflammatory genes and recruit corepressors to inhibit gene expression; (2) PR also may bind directly to promoters of genes encoding transcriptional repressors (e.g. ZEB1) to activate their expression. ZEB1, in turn, binds to promoters of CAP genes and recruits a repressive complex to inhibit their expression. By contrast, during the initiation of term and preterm labor, PR function in myometrium is impaired by its direct interaction with NF-kB, by increased expression of enzymes that metabolize P4 to inactive products, by decreased expression of PR coactivators and by upregulation of truncated PR isoforms (e.g. PR-A). The truncated PR isoforms may have reduced transcriptional and transrepressive activity. We postulate that these changes in inflammatory and CAP gene expression are mediated by alterations in chromatin modifications and structure. The goals of this proposal are to define the genes and mechanisms that underlie the inhibitory actions of P4/PR and differential actions of PR- A and PR-B on inflammatory and CAP gene expression, and to characterize the chromatin modifications that mediate myometrial quiescence and accompany enhanced CAP and inflammatory gene expression leading to term and preterm labor. To achieve these goals, we will use mouse models and human myometrial cells and tissues to: (1) define and characterize components of the complex of transcription factors/coregulators that interact with PR in the pregnant myometrium to mediate its anti-inflammatory actions and that interact with ZEB1 to inhibit CAP gene expression; (2) analyze expression and promoter-binding of these PR-interacting factors during pregnancy and with term and preterm labor and the effects of hormones and microRNAs in their regulation; (3) use RNA-seq and ChIP-seq to discover myometrial genes that are critical for the maintenance of pregnancy and initiation of labor and the underlying transcriptional mechanisms for their regulation. RNA-seq will enable discovery of genes involved in maintenance of pregnancy and the initiation of labor. The combined use of ChIP-seq will provide global insight into the transcriptional and epigenetic mechanisms that underlie these gene expression changes. Studies using PR-B-KO mice will elucidate the differential roles of PR-A vs. PR-B in regulation of the myometrial transcriptome. Collectively, our findings will reveal novel genes and pathways that can serve as therapeutic targets to prevent preterm birth and its consequences.