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

Ars2 核和细胞质异构体作为转录调控支架的表征

• 批准号: RGPIN-2020-03916
• 负责人: Howard, Perry
• 金额: $ 3.64万
• 依托单位: University of Victoria
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2021
• 资助国家: 加拿大
• 起止时间: 2021-01-01 至 2022-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=739562
• 关键词: Characterization; Ars2; nuclear; cytoplasmic; isoforms

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.

信使RNA(MRNA)的生命从转录起始到不可避免的降解都是高度协调的。这种协调需要大量的瞬时RNA-蛋白质复合体的招募、组装和分解，并在细胞对应激的反应中发挥关键作用。具体如何协调这一点还远未被理解，对这些过程的不当监管对发展产生了深远的影响。我的实验室发现了编码ARS2的基因，它是RNA Pol II RNA加工和转换的纽带。通过我的NSERC资助的项目，我们领导了ARS2的生物学特征以及它如何作为RNA和蛋白质相互作用中心的功能。最近，我们发现Ars2基因产生两种异构体：核异构体(NARS2)和新发现的细胞质异构体(Cars2)。NARS2是帽结合复合体(CBC)的关键成员，负责协调RNAPII RNA的核处理、输出和降解。我们现在发现，在细胞质CBC复合体中，Cars2取代了nARS2。这种‘ARS2转换’具有戏剧性的功能后果，将细胞质中的CBC从无意义介导的衰变(NMD)抑制因子改变为NMD启动子。具体是如何做到这一点还不清楚。了解ARS2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，除了限制带有过早终止密码子(PTCs)的mRNAs产生的异常蛋白外，NMD还通过调节内质网(ER)应激反应在所有真核生物中发挥着关键的生理作用。这是通过未折叠蛋白反应(UPR)途径发生的。有趣的是，Ars2首先被确定为参与细胞对砷引起的内质网应激反应的基因，该基因通过UPR途径。因此，我们认为我们已经发现了ARS2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。我们推测，nARS2和Cars2分别以CBC依赖的、非冗余的方式在细胞核和细胞质内引导RNAPII转录本从转录到降解的代谢。其次，我们推测ARS_2亚型在NMD中的作用可能是ARS_2的S在砷诱导的内质网应激中的重要作用。在下一个资助时期，我的研究计划的短期目标将集中在ARS2亚型生物学的两个核心方面：1)ARS2亚型如何以不同的方式协调NMD；2)ARS2亚型在NMD中的作用如何影响对内质网应激的反应？这些问题直接进入了我的研究计划的长期目标，即在分子水平上了解控制RNAPII转录本从生物发生到降解的控制过程，以及这些过程如何有助于干细胞和祖细胞的命运决定。