Codon usage optimization in pluripotent embryonic stem cells [tRNA sequencing]. Codon usage optimization in pluripotent embryonic stem cells [tRNA sequencing]

The uneven use of synonymous codons in the transcriptome regulates the efficiency and fidelity of protein translation rates. Yet, the importance of this codon bias on regulating cell state-specific expression programs is currently debated. Here, we asked whether the gene expression program in the well-defined cell states of self-renewal and differentiation in embryonic stem cells is driven by optimized codon usage. Using ribosome and transcriptome profiling, we identified distinct codon signatures for human self-renewing and differentiating embryonic stem cells. One driver for the cell state-specific codon bias was the genomic GC-content of the differentially expressed genes and thus, determined by transcription rather than translation. However, by measuring the codon frequencies at the ribosome’s active sites interacting with transfer RNAs (tRNA), we discovered that the wobble position tRNA modification inosine strongly influenced the codon optimization in self-renewing embryonic stem cells. This effect was conserved in mice and independent of the differentiation stimulus. In summary, we newly reveal how translational mechanisms based on RNA modifications can shape optimized codon usage in embryonic stem cells. Overall design: tRNA sequencing of self-renewing and differentiating human H9 cells, 4 biological replicates per condition

转录组中同义密码子的不均衡使用，可调控蛋白质翻译的效率与保真度。目前，这种密码子偏好性对细胞状态特异性表达程序的调控作用仍存在争议。本研究旨在探究：胚胎干细胞自我更新与分化这两种明确界定的细胞状态下，其基因表达程序是否由优化的密码子使用模式所驱动。通过核糖体谱分析与转录组分析，我们鉴定出了人类自我更新态与分化态胚胎干细胞的特异性密码子特征。导致细胞状态特异性密码子偏好性的一个驱动因素是差异表达基因的基因组GC含量，因此该偏好性由转录过程而非翻译过程决定。然而，通过检测核糖体与转运RNA（transfer RNA, tRNA）相互作用的活性位点处的密码子频率，我们发现，摆动位点的tRNA肌苷修饰会强烈影响自我更新态胚胎干细胞的密码子优化效果。该效应在小鼠中保守存在，且不受分化刺激的影响。综上，我们首次揭示了基于RNA修饰的翻译机制如何塑造胚胎干细胞中的优化密码子使用模式。整体实验设计：对自我更新态与分化态的人类H9细胞进行tRNA测序，每组设置4个生物学重复。