Characterization of Ars2 nuclear and cytoplasmic isoforms as a scaffold for transcriptional regulation

信使RNA（mRNA）的生命是高度协调的，从转录起始到不可避免的降解。这种协调需要大量瞬时RNA-蛋白质复合物的募集、组装和分解，并在细胞对应激的反应中起着关键作用。这一过程是如何协调的，人们还远未了解，而对这些过程的错误调节对发展有着深远的影响。我的实验室发现了编码ARS 2的基因，它是RNA Pol II RNA加工和周转的联系。通过我的NSERC资助的项目，我们领导了ARS 2生物学的表征以及它如何作为RNA和蛋白质相互作用中心的功能。 最近，我们发现Ars 2基因产生2种亚型：核亚型（nARS 2）和新鉴定的胞质亚型（cARS 2）。nARS 2是帽结合复合物（CBC）的关键成员，其协调RNAPII RNA的核加工、输出和降解。我们现在已经发现cARS 2在细胞质CBC复合物中取代nARS 2。 这种ARS 2转换具有显著的功能后果，将CBC从无义介导的衰变（NMD）抑制剂改变为细胞质中的NMD启动子。具体是如何实现的尚不清楚。 了解ARS 2亚型如何协调正确的NMD反应在生物学的不同领域具有广泛的影响。这是因为，除了限制从具有过早终止密码子（PTC）的mRNA产生异常蛋白质之外，NMD通过调节内质网（ER）应激反应在所有真核生物中具有关键的生理作用。这通过未折叠蛋白反应（UPR）途径发生。有趣的是，Ars 2首先被鉴定为通过UPR途径参与砷引起的ER应激的细胞反应的基因。因此，我们认为我们已经发现了ARS 2在砷敏感性方面的长期知识与其作为RNA伴侣的作用之间的机制联系。 我们假设nARS 2和cARS 2以CBC依赖性、非冗余的方式发挥功能，分别在细胞核和细胞质内引导RNAPII转录物代谢从转录到降解。其次，我们假设ARS 2亚型在NMD中的作用对于ARS 2在砷处理引起的ER应激中的作用是重要的。在下一个资助期内，我的研究计划的短期目标集中在ARS 2亚型生物学的两个核心方面：1）ARS 2亚型如何差异协调NMD，以及2）ARS 2亚型在NMD中的作用如何影响对ER应激的反应？ 这些问题直接进入我的研究计划的长期目标，这是在分子水平上了解控制过程RNAPII转录从他们的生物起源降解，以及这些过程如何有助于干细胞和祖细胞的命运决定。

The life of an messenger RNA (mRNA) is highly coordinated from its beginnings during transcriptional initiation to its inevitable degradation. This coordination requires the recruitment, assembly and disassembly of numerous transient RNA-protein complexes and plays a critical role in cellular responses to stress. Precisely how this is co-ordinated is far from understood and misregulation of these processes has profound effects on development. My lab discovered the gene encoding ARS2, which is a nexus for RNA Pol II RNA processing and turnover. Through my NSERC-funded program, we have led the characterization of both the biology of ARS2 and how it functions as an RNA and protein interaction hub. Recently, we discovered that the Ars2 gene generates 2 isoforms: a nuclear isoform (nARS2) and a newly identified cytoplasmic isoform (cARS2). nARS2 is a key member of the cap binding complex (CBC), which coordinates the nuclear processing, export, and degradation of RNAPII RNAs. We have now found that cARS2 replaces nARS2 in cytoplasmic CBC complexes. This ARS2 switching' has dramatic functional consequences, changing the CBC from a nonsense mediated decay (NMD) inhibitor to NMD promoter in the cytoplasm. Exactly how this is accomplished is unclear. Understanding how ARS2 isoforms orchestrate the correct NMD response has a broad impact in diverse areas of biology. This is because, in addition to limiting the production of aberrant proteins from mRNAs with premature stop codons (PTCs), NMD has a critical physiological role in all eukaryotes by regulating endoplasmic reticulum (ER) stress response. This occurs through the unfolded protein response (UPR) pathway. Intriguingly, Ars2 was first identified as a gene involved in the cellular response to ER stress caused by arsenic through the UPR pathway. Thus, we think we have discovered a mechanistic link between the longstanding knowledge of ARS2 in arsenic sensitivity and its role as a RNA chaperone. We postulate that nARS2 and cARS2 function in a CBC dependent, non-redundant manner to guide RNAPII transcript metabolism from transcription to degradation, within the nucleus and cytoplasm, respectively. Secondly, we hypothesize the roles of ARS2 isoforms in NMD are important for ARS2's role in ER stress caused by arsenic treatment. In the next funding period the short-term goals of my research program are focused on two core aspects of ARS2 isoform biology: 1) How do ARS2 isoforms differentially coordinate NMD, and 2) How do the roles of ARS2 isoforms in NMD affect response to ER stress? These questions directly feed into the long-term goal of my research program which is to understand at the molecular level the control processes governing RNAPII transcripts from their biogenesis to degradation and how these processes contribute to stem and progenitor cell fate decisions.