An inquiry into the function of Ronin in embryogenesis and pluripotent stem cells

项目摘要 一个长期持有的观点是，干细胞和发育中基因调控的复杂性是通过基因调控来实现的。 数以百万计的增强子元素的明智组织。最近发现的一类新的DNA结合因子- 被称为THAP（死亡相关结构域的死亡相关蛋白）的因子可能提供新的 对干细胞基因调控的深入了解。我们发现了这个家族的一个关键成员，Ronin（Thap11） 在胚胎干细胞中。Ronin在胚胎发育的最早阶段表达， 在体外和体内维持干细胞多能性。既然罗宁缺乏可观的交易- 激活活性，但作为具有强蛋白质二聚化活性的DNA结合核蛋白发挥作用，我们 认为它可能不以转录调节因子的经典方式起作用（例如，具有酸性活性， 但相反，它可能通过促进Ronin结合蛋白之间的通讯来增强基因表达。 通过DNA循环。当我们发现大多数（83%） 由Ronin介导的成环事件涉及启动子-启动子连接，而不是增强子-启动子连接 与典型转录因子的相互作用。在这项研究中，我们将（i）建立Ro的影响， Nin介导的启动子系链对干细胞中转录和基因调控的影响;（ii）确定Nin介导的启动子系链对干细胞中转录和基因调控的影响。 启动子拴系在发育过渡上，特别是从幼稚到启动多能状态;以及 (iii)询问其他THAP是否也参与发育特异性基因聚类。

PROJECT SUMMARY A long-held view is that gene regulatory complexity in stem cells and development is achieved through the judicious organization of millions of enhancer elements. A recently identified new class of DNA binding fac- tors called THAPs (Thanatos-associated domain-containing apoptosis-associated proteins) may provide new insights into gene regulation in stem cells. One key member of this family, Ronin (Thap11), was discovered by us in embryonic stem cells. Ronin is expressed during the earliest stages of embryonic development and is essential for the maintenance of stem cell pluripotency, both in vitro and in vivo. Since Ronin lacks appreciable transac- tivation activity but functions as a DNA-binding nuclear protein with strong protein dimerization activity, we considered that it may not act in the classical manner of a transcriptional regulator (e.g., having an acidic activa- tion domain) but instead might augment gene expression by facilitating communication among Ronin binding sites through DNA looping. This working hypothesis gained traction when we found that most (83%) of the looping events mediated by Ronin involve promoter-to-promoter links rather than the enhancer-to-promoter interactions seen with typical transcription factors. In this research study we will (i) establish the effects of Ro- nin-mediated promoter tethering on transcription and gene regulation in stem cells; (ii) determine the influence of promoter tethering on developmental transitions especially from the naïve to primed pluripotent state; and (iii) ask if other THAPs participate in development specific gene clustering as well.