Department of Biochemistry and Molecular Biology, Chicago, IL, 60637, USA.
Department of Chemistry, University of Chicago, Chicago, IL, 60637, USA.
Nat Commun. 2022 May 5;13(1):2491. doi: 10.1038/s41467-022-30261-3.
Small RNAs include tRNA, snRNA, micro-RNA, tRNA fragments and others that constitute > 90% of RNA copy numbers in a human cell and perform many essential functions. Popular small RNA-seq strategies limit the insights into coordinated small RNA response to cellular stress. Small RNA-seq also lacks multiplexing capabilities. Here, we report a multiplex small RNA-seq library preparation method (MSR-seq) to investigate cellular small RNA and mRNA response to heat shock, hydrogen peroxide, and arsenite stress. Comparing stress-induced changes of total cellular RNA and polysome-associated RNA, we identify a coordinated tRNA response that involves polysome-specific tRNA abundance and synergistic N3-methylcytosine (mC) tRNA modification. Combining tRNA and mRNA response to stress we reveal a mechanism of stress-induced down-regulation in translational elongation. We also find that native tRNA molecules lacking several modifications are biased reservoirs for the biogenesis of tRNA fragments. Our results demonstrate the importance of simultaneous investigation of small RNAs and their modifications in response to varying biological conditions.
小 RNA 包括 tRNA、snRNA、microRNA、tRNA 片段等，它们在人类细胞中构成了 >90%的 RNA 拷贝数，并发挥着许多重要的功能。流行的小 RNA-seq 策略限制了对细胞应激下协调的小 RNA 反应的深入了解。小 RNA-seq 也缺乏多重检测能力。在这里，我们报告了一种多路小 RNA-seq 文库制备方法（MSR-seq），用于研究细胞小 RNA 和 mRNA 对热休克、过氧化氢和亚砷酸盐应激的反应。通过比较总细胞 RNA 和多核糖体相关 RNA 诱导的变化，我们确定了一种协调的 tRNA 反应，涉及多核糖体特异性 tRNA 丰度和协同的 N3-甲基胞嘧啶（mC）tRNA 修饰。将 tRNA 和 mRNA 对压力的反应结合起来，我们揭示了一种翻译延伸过程中应激诱导下调的机制。我们还发现，缺乏几种修饰的天然 tRNA 分子是 tRNA 片段生物发生的偏倚库。我们的研究结果表明，同时研究不同生物学条件下小 RNA 及其修饰的重要性。
