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Abstract：
Differences in codon frequency between genomes, genes, or positions along a gene, modulate transcription and translation efficiency, leading to phenotypic and functional differences. Here, we present a multiscale analysis of the effects of synonymous codon recoding during heterologous gene expression in human cells, quantifying the phenotypic consequences of codon usage bias at different molecular and cellular levels, with an emphasis on translation elongation. Six synonymous versions of an antibiotic resistance gene were generated, fused to a fluorescent reporter, and independently expressed in HEK293 cells. Multiscale phenotype was analyzed by means of quantitative transcriptome and proteome assessment, as proxies for gene expression; cellular fluorescence, as a proxy for single-cell level expression; and real-time cell proliferation in absence or presence of antibiotic, as a proxy for the cell fitness. We show that differences in codon usage bias strongly impact the molecular and cellular phenotype: (i) they result in large differences in mRNA levels and protein levels, leading to differences of over 15 times in translation efficiency; (ii) they introduce unpredicted splicing events; (iii) they lead to reproducible phenotypic heterogeneity; and (iv) they lead to a trade-off between the benefit of antibiotic resistance and the burden of heterologous expression. In human cells in culture, codon usage bias modulates gene expression by modifying mRNA availability and suitability for translation, leading to differences in protein levels and eventually eliciting functional phenotypic changes.
摘要：
基因组之间密码子频率的差异，基因，或者沿着基因的位置，调节转录和翻译效率，导致表型和功能差异。这里,我们提出了一个多尺度分析的影响，同义密码子重新编码在异源基因表达在人类细胞中，量化不同分子和细胞水平的密码子使用偏倚的表型后果，强调翻译的延伸。产生了六个同义版本的抗生素抗性基因，融合到一个荧光报告，并且在HEK293细胞中独立表达。通过定量转录组和蛋白质组评估分析了多尺度表型，作为基因表达的代理；细胞荧光，作为单细胞水平表达的代表;和在不存在或存在抗生素的情况下的实时细胞增殖，作为细胞适应性的代理。我们表明，密码子使用偏差的差异强烈影响分子和细胞表型：（i）它们导致mRNA水平和蛋白质水平的巨大差异，导致翻译效率差异超过15倍;(ii)它们引入未预测的剪接事件;(iii)它们导致可重复的表型异质性;和(iv)它们导致抗生素抗性的益处和异源表达的负担之间的权衡。在培养的人类细胞中，密码子使用偏倚通过改变mRNA的可用性和翻译的适用性来调节基因表达，导致蛋白质水平的差异，并最终引发功能表型的变化。
