Post-Transcriptional Regulation of Embryo Implantation

胚胎植入的转录后调控

• 批准号: 10682386
• 负责人: Ramakrishna Kommagani
• 金额: $ 41.58万
• 依托单位: BAYLOR COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-08-11 至 2027-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10682386
• 关键词: Address; Alternative Splicing; Area; Basic Science; Biochemical; Biological Assay; Biological Factors; Biopsy; Cell Differentiation process; Cell Proliferation; Cells; Chromatin; Conceptions; Data; Decidual Cell; Decidual Cell Reactions; Diagnosis; Embryo; Endometrial; Endometrial Stromal Cell; Endometrium; Epithelium; Estrogens; Genes; Genomics; Gonadal Steroid Hormones; Homologous Gene; Hormones; Human; Impairment; Implant; In Vitro; Knockout Mice; Knowledge; Lead; LoxP-flanked allele; Measures; Mediating; Messenger RNA; Molecular; Monitor; Mus; National Institute of Child Health and Human Development; Pathway interactions; Physiology; Post-Transcriptional Regulation; Pregnancy; Pregnancy loss; Process; Progesterone; Progesterone Receptors; Proliferating; Proteins; RNA Splicing; Recurrence; Reverse Transcriptase Polymerase Chain Reaction; Role; Small Interfering RNA; Spontaneous abortion; Strategic Planning; Stromal Cells; Testing; Transcript; Ubiquitin; Uterus; Variant; Woman; Work; blastocyst; cell type; chromatin immunoprecipitation; conditional knockout; data integration; early pregnancy loss; failure Implantation; human tissue; implantation; in vivo; inhibitor; insight; knock-down; natural Blastocyst Implantation; non-genomic; novel; prevent; response; steroid hormone; transcriptome sequencing; uterine receptivity

PROJECT SUMMARY Successful establishment of pregnancy requires the uterus to undergo several well-timed cellular changes to allow the embryo to implant. Thus, even when the blastocyst develops normally, impaired uterine function can lead to implantation failure or early embryo miscarriage. The uterine endometrium prepares for implantation in two steps. First, the endometrial epithelium proliferates, loses polarity, and differentiates, allowing the embryo to attach. Second, the underlying stromal cells proliferate and differentiate into decidual cells, allowing the embryo to implant. These two processes are coordinated by cell-type specific responses to the steroid hormones estrogen and progesterone. However, we lack a complete picture of the downstream responses to these hormones, hampering our ability to develop new strategies to prevent early pregnancy loss. To address this knowledge gap, this proposal focuses on a new area in endometrial physiology, alternative mRNA splicing. Specifically, this proposal will test the central hypothesis that the splicing factor SF3B1 mediates progesterone- driven alternative splicing that is essential for uterine receptivity and decidualization. This idea is founded on the following pieces of preliminary data. First, a high-throughput siRNA screen revealed that SF3B1 was required for human endometrial stromal cell decidualization. Second, knock down of SF3B1 impaired in vitro decidualization more than knock down of eight other splicing factors. Third, treatment with the SF3B1-specific inhibitor Pladienolide B inhibited human endometrial stromal cell decidualization in vitro and murine endometrial decidualization in vivo. Fourth, treatment with Pladienolide B impaired embryo implantation and decidualization in mice. Fifth, SF3B1 protein is elevated in endometrial stromal cells during peri-implantation in mice. Finally, SF3B1 protein but not mRNA in stromal cells was elevated during artificial decidualization in mice, and progesterone stabilized SF3B1 protein but not mRNA in primary human endometrial stromal cells. The work proposed here will build on these strong preliminary data and test the hypothesis by pursuing the following specific aims: (Aim 1) Define the functions of SF3B1 in uterine receptivity and decidualization; (Aim 2) Identify progesterone-induced, SF3B1-dependent alternative splice variants in the endometrium; (Aim 3) Determine the mechanism by which progesterone regulates SF3B1. At the level of basic science, this project will identify the mechanisms that underlie SF3B1-driven mRNA splicing, which is crucial for progesterone- driven endometrial decidualization. Of translational significance, this work will identify novel transcript variants that may contribute to recurrent pregnancy loss. In the long term, such knowledge can be used to develop new strategies to diagnose or prevent early pregnancy loss. Together, this work will help advance Theme 2 of the NICHD 2020 Strategic Plan, which aims to "identify biological factors that can lead or contribute to early pregnancy loss".

项目总结 成功建立妊娠需要子宫经历几次适时的细胞变化，以 让胚胎植入。因此，即使胚泡发育正常，子宫功能受损也可能 导致着床失败或早期胚胎流产。子宫内膜为着床做准备 两个步骤。首先，子宫内膜上皮增殖，失去极性，分化，使胚胎 附在一起。其次，潜在的基质细胞增殖并分化为蜕膜细胞，使 胚胎植入。这两个过程是通过细胞类型对类固醇的特定反应来协调的 雌激素和黄体酮。然而，我们缺乏下游反应的完整图景 这些荷尔蒙阻碍了我们制定防止早孕丢失的新策略的能力。致信地址 针对这一知识空白，本提案重点研究了子宫内膜生理学的一个新领域--选择性的mRNA剪接。 具体地说，这项提议将检验剪接因子SF3B1介导孕酮的中心假设。 驱动选择性剪接，这对于子宫的接受性和蜕膜化是必不可少的。这个想法是建立在 以下是初步数据。首先，高通量siRNA筛选显示SF3B1是 人子宫内膜间质细胞蜕膜化所必需的。第二，SF3B1在体外受损的击倒 蜕膜化超过了其他八个剪接因素的影响。第三，使用针对SF3B1的治疗 普列烯内酯B抑制人子宫内膜间质细胞蜕膜化的体外和小鼠实验 活体子宫内膜蜕膜化。第四，雷公藤内酯B治疗胚胎着床障碍和 小鼠的蜕膜化。第五，子宫内膜间质细胞中SF3B1蛋白在围着床期升高。 老鼠。最后，在人工蜕膜过程中，间质细胞中SF3B1蛋白而不是mRNA表达升高。 黄体酮可以稳定人子宫内膜间质细胞中的SF3B1蛋白，但不能稳定其mRNA。 这里提出的工作将建立在这些强大的初步数据的基础上，并通过追求 下列具体目标：(目标1)确定SF3B1在子宫容受性和蜕膜形成中的功能；(目标2) 确定子宫内膜中孕酮诱导的、依赖于SF3B1的选择性剪接变异体；(目标3) 确定孕酮调节SF3B1的机制。在基础科学的层面上，这个项目 将确定SF3B1驱动的mRNA剪接的基础机制，这对孕激素至关重要- 导致子宫内膜蜕膜化。具有翻译意义，这项工作将确定新的文字记录变体 这可能会导致反复的妊娠丢失。从长远来看，这样的知识可以用来开发新的 诊断或预防早孕丢失的策略。总之，这项工作将有助于推进 《NICHD 2020年战略计划》，该计划旨在“确定可能导致或促成早期死亡的生物因素 怀孕丢失“。