Queuosine modification protects cognate tRNAs against ribonuclease cleavage

Eukaryotic transfer RNAs (tRNA) contain on average 13 modifications that perform a wide range of roles in translation and in the generation of tRNA fragments that regulate gene expression. Queuosine (Q) modification occurs in the wobble anticodon position of tRNAs for amino acids His, Asn, Tyr, and Asp. In eukaryotes, Q modification is fully dependent on diet or on gut microbiome in multi-cellular organisms. Despite decades of study, cellular roles of Q modification remain to be fully elucidated. Here we show that in human cells, Q modification specifically protects its cognate tRNAHis and tRNAAsn against cleavage by ribonucleases. We generated cell lines that contain completely depleted or fully Q-modified tRNAs. Using these resources, we found that Q modification significantly reduces angiogenin cleavage of its cognate tRNAs in vitro. Q modification does not change the cellular abundance of the cognate full-length tRNAs, but alters the cellular content of their fragments in vivo in the absence and presence of stress. Our results provide a new biological aspect of Q modification and a mechanism of how Q modification alters small RNA pool in human cells. Comprehensive comparative analysis of high throughput sequencing data to determine queuosine-mediated tRNA fragments

真核生物转运RNA（transfer RNAs，tRNA）平均带有13种修饰基团，这些修饰在翻译进程以及调控基因表达的tRNA片段生成过程中发挥着广泛的生物学功能。夸司核苷（queuosine，Q）修饰发生在对应组氨酸、天冬酰胺、酪氨酸以及天冬氨酸的tRNA的摆动反密码子位点处。在多细胞真核生物中，Q修饰完全依赖于膳食或肠道微生物组。尽管已有数十年的研究，但Q修饰在细胞内的具体功能仍有待完全阐明。本研究发现，在人类细胞中，Q修饰可特异性保护其同源的tRNA^His与tRNA^Asn免受核糖核酸酶（ribonucleases）的切割。我们构建了完全缺失Q修饰或完全携带Q修饰的tRNA的细胞系。利用这些细胞模型，我们发现Q修饰可在体外显著抑制血管生成素（angiogenin）对其同源tRNA的切割作用。Q修饰不会改变同源全长tRNA在细胞内的丰度，但会在应激与非应激状态下，改变细胞内其衍生片段的含量。本研究结果揭示了Q修饰的全新生物学功能，阐明了人类细胞中Q修饰调控小分子RNA库的分子机制。本研究通过对高通量测序（high throughput sequencing）数据开展全面比较分析，旨在明确夸司核苷修饰介导的tRNA片段相关特性。