Queuosine-tRNA promotes sex-dependent learning and memory formation by maintaining codon-biased translation elongation speed.

Queuosine (Q) is a modified nucleoside at the wobble position of specific tRNAs. In mammals, queuosinylation is facilitated by queuine uptake from the gut microbiota and is introduced into tRNA by the QTRT1-QTRT2 enzyme complex. By establishing a Qtrt1 knockout mouse model, we discovered that the loss of QtRNA leads to learning and memory deficits. Ribo-Seq analysis in the hippocampus of Qtrt1-deficient mice revealed not only stalling of ribosomes on Q-decoded codons but also a global imbalance in translation elongation speed between codons that engage in weak and strong interactions with their cognate anticodons. While Qdependent molecular and behavioral phenotypes were identified in both sexes, female mice were affected more severely than males. Proteomics analysis confirmed deregulation of synaptogenesis and neuronal morphology. Together, our findings provide a link between tRNA modification and brain functions and reveal an unexpected role of protein synthesis in sex-dependent cognitive performance. Cytoplasmic RNA and Cytoplasmic RNA protected by ribosomes from RNase I (Ribonuclease I) cleavage, were sequenced. Small RNA libraries were performed according to the protocol using NEB NEXT Small RNA library Prep Set for Illumina (Multiplex Compatible) E7330, and size-selected (120-170 base pairs) before sequencing.Q核苷（Queuosine, Q）是一类定位于特定tRNA反密码子摆动位点的修饰核苷。在哺乳动物体内，Q核苷化过程需通过肠道菌群摄取q碱（Queuine）完成，并由QTRT1-QTRT2酶复合物将其修饰至tRNA上。本研究通过构建Qtrt1基因敲除小鼠模型，发现tRNA的Q修饰缺失会引发学习与记忆功能缺陷。对Qtrt1缺陷小鼠海马体的核糖体印记测序（Ribo-Seq）分析显示，不仅核糖体在Q解码密码子处出现停滞，而且与对应反密码子存在强弱相互作用的密码子之间，翻译延伸速度呈现全局失衡。尽管在雌雄小鼠中均观察到依赖Q修饰的分子与行为表型，但雌性小鼠受影响的程度显著高于雄性。蛋白质组学分析证实，突触发生与神经元形态的调控出现异常。综上，本研究结果揭示了tRNA修饰与脑功能之间的关联，并阐明了蛋白质合成在性别依赖型认知表现中此前未被发现的作用。 本研究对两类RNA进行了测序：细胞质RNA，以及被核糖体保护、免受RNase I（核糖核酸酶I）降解的细胞质RNA。小型RNA文库构建严格遵循实验方案，采用适配Illumina平台的NEB NEXT小型RNA文库制备试剂盒（多重兼容型）E7330，并在测序前对文库进行120-170碱基对的片段大小筛选。