Biogenesis, Function and Turnover of Noncoding RNAs

我们实验室的长期目标是了解非编码RNA的功能，细胞如何识别和降解有缺陷和不需要的RNA，以及未能降解这些RNA如何影响细胞功能并导致人类疾病。我们工作的一个重点是一类丰富的核糖核蛋白(RNPs)，被称为Ro60RNPs，它们广泛存在于动物细胞中，存在于许多细菌中。主要的蛋白质成分是环状Ro 60 kDa自身抗原，因为它是系统性红斑狼疮和干燥综合征患者自身抗体的临床重要靶点。在所有被研究的生物中，Ro60结合了被称为Y RNA的非编码RNA。通过研究细菌中的Ro60 RNPs，我们发现了ncRNA的一个新作用，即将蛋白质辅因子与效应蛋白捆绑在一起，以改变其功能。具体地说，我们发现细菌Ro60的同源基因由Y RNA拴在环状核糖核酸酶上，形成了一种新的双环RNA降解机。我们目前正在努力确定Ro60 RNPs在哺乳动物细胞中的功能。作为这项工作的一部分，我们使用CRISPR来产生缺乏两个小鼠Y RNA中的一个或两个的小鼠胚胎干细胞。尽管有报道称Y RNAs对动物细胞中的DNA复制是必不可少的，但缺乏这两个Y RNAs的小鼠胚胎干细胞正常分裂。然而，缺乏Y RNA的细胞Ro60水平降低，我们可以通过在这些细胞中表达Y RNA来弥补这一缺陷。我们还证明了Ro60调节Ro60的亚细胞位置，并且Y RNAs将Ro60与不同的蛋白质捆绑在一起，以创建专门的RNPs。综上所述，这些数据表明Y RNAs的功能与它们的Ro60伙伴的功能密切相关。在第二个焦点中，我们正在表征RNA监视通路在哺乳动物细胞生理学中的作用。在这里，最近的一项成就是我们发现了被称为RNA外切体的多核酸酶复合体抑制人类胚胎干细胞的分化。我们发现，外切体部分通过降解编码FOXH1的前mRNAs来抑制分化，FOXH1是形成中胚层的关键转录因子，中胚层和内胚层的前体。这些研究揭示了RNA降解在维持人类胚胎干细胞多能性方面的重要性。

The long-term goals of our laboratory are to understand how noncoding RNAs function, how cells recognize and degrade defective and unneeded RNAs, and how failure to degrade these RNAs affects cell function and contributes to human disease. One focus of our work is an abundant class of ribonucleoproteins (RNPs), known as Ro60 RNPs, which are widespread in animal cells and present in many bacteria. The major protein component, the ring-shaped Ro 60 kDa autoantigen, was discovered because it is a clinically important target of autoantibodies in patients with systemic lupus erythematosus and Sjogren's syndrome. In all organisms examined, Ro60 binds noncoding RNAs called Y RNAs. By studying Ro60 RNPs in bacteria, we uncovered a novel role for ncRNA, that of tethering a protein cofactor to an effector protein to alter its function. Specifically, we discovered that a bacterial Ro60 ortholog was tethered by Y RNA to a ring-shaped ribonuclease, forming a new double-ringed RNA degradation machine. We are currently working to define the functions of Ro60 RNPs in mammalian cells. As part of this effort, we used CRISPR to generate mouse embryonic stem cells lacking one or both of the two mouse Y RNAs. Despite reports that Y RNAs are essential for DNA replication in animal cells, mouse embryonic stem cells lacking both Y RNAs divided normally. However, cells lacking Y RNAs had reduced levels of Ro60, a defect that we could complement by expressing Y RNA in these cells. We also demonstrated that Ro60 regulates the subcellular location of Ro60 and that Y RNAs tether Ro60 to diverse proteins to create specialized RNPs. Together, these data reveal that the functions of Y RNAs are closely connected to those of their Ro60 partner. In a second focus, we are characterizing the roles of RNA surveillance pathways in mammalian cell physiology. Here, a recent accomplishment was our discovery that the multinuclease complex known as the RNA exosome represses differentiation of human embryonic stem cells. We showed that the exosome restrains differentiation in part by degrading pre-mRNAs encoding FOXH1, a transcription factor crucial for formation of mesendoderm, the precursor to both mesoderm and endoderm. These studies revealed the importance of RNA degradation in maintaining human embryonic stem cell pluripotency.