RNA turnover in plant stress responses

调控rna的稳定性是基因表达调控的一个关键特征。虽然管家基因通常产生高度稳定的rna，但应激反应基因通常编码低稳定性的rna。这允许快速的RNA周转；这一特征对植物来说尤为重要，因为植物必须迅速适应不断变化的环境条件和压力。在植物中执行RNA降解的途径已经被很好地理解了。然而，胁迫如何破坏特定RNA的稳定性，以及RNA降解酶的靶特异性是如何控制的，在很大程度上是未知的。为了解决这些问题，我们将把我们最近开发的eric测序技术应用于胁迫和非胁迫拟南芥植物。传统上，RNA随着时间的推移而衰变，RNA的半衰期是在高毒性转录抑制剂（如放线菌素D和虫草素）的严酷治疗后确定的。我们开发了一种新的方法（ERIC-seq），该方法使用无毒的5-乙基尿苷（5-EU）结合RNA测序进行代谢RNA标记。这使我们第一次能够非侵入性地确定植物中RNA的稳定性。我们将应用这项技术来研究核和叶绿体编码rna在多种非生物胁迫条件下的稳定性。此外，我们将研究特定的细胞质和塑性RNA降解和稳定途径在选定的胁迫条件下的作用。这种方法使我们能够识别胁迫特异性RNA序列特征，即(1)确保RNA在非生物胁迫条件下的稳定和不稳定，以及(2)确定RNA亚群的RNA降解途径的特异性。这些序列元件将进行功能研究，连接到特定的内切酶，并用于鉴定新的反调节RNA稳定因子。作为主要成果，我们的方法将在转录组范围内确定应激期间RNA翻转对基因调控的影响。

Modulating the stability of RNAs is a key feature of gene expression regulation. While housekeeping genes usually produce highly stable RNAs, stress-responsive genes often encode RNAs with low stability. This allows rapid RNA turnover; a feature particularly important for plants, which have to adapt quickly to changing environmental conditions and stresses. The pathways that execute RNA degradation in plants are well understood. However, it is largely unknown how stresses destabilize specific RNAs, and how target specificity of RNA degrading enzymes is controlled.To address these questions, we will apply our recently developed ERIC-sequencing technology to stressed and unstressed Arabidopsis thaliana plants. Traditionally, RNA decay over time and RNA half-lives have been determined after harsh treatments with highly toxic transcriptional inhibitors such as actinomycin D and cordycepin. We developed a novel method (ERIC-seq), which employs metabolic RNA labeling using the non-toxic 5-ethynyl uridine (5-EU) combined with RNA sequencing. This allowed us for the first time to determine RNA stability non-invasively in plants. We will apply this technique to study the stability of nuclear and chloroplast-encoded RNAs under a diverse set of abiotic stress conditions. In addition, we will study the role of specific cytosolic and plastidic RNA degradation and stabilization pathways under selected stress conditions. This approach enables us to identify stress-specific RNA sequence features, that (1) ensure RNA stabilization and destabilization under abiotic stress conditions, and that (2) determine the specificity of RNA degradation pathways for subpopulations of RNAs. Such sequence elements will be functionally studied, linked to specific endonucleases, and used to identify novel trans-regulatory RNA stability factors. As a major deliverable, our approach will identify the impact of RNA turn-over on gene regulation during stress on a transcriptome-wide scale.