Design of Adjacent Transcriptional Regions to Tune Gene Expression and Facilitate Circuit Construction.
Design of Adjacent Transcriptional Regions to Tune Gene Expression and Facilitate Circuit Construction.
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DOI：
10.1016/j.cels.2018.01.010
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发表时间：
2018-02-28
期刊：
影响因子：
9.3
通讯作者：
Wang X
中科院分区：
文献类型：
--
作者：
Wu F;Zhang Q;Wang X
多顺反子结构对于合成基因回路是常见的，然而，仍然不知道一个基因的表达如何受到操纵子中其他基因/非编码区（称为相邻转录区（ATR））的影响。在这里，我们构建了合成操纵子与报告基因侧翼不同的ATR，我们发现与GC含量高，小尺寸，低折叠能量的ATR导致高基因表达。基于这些结果，我们建立了一个基因表达模型，并生成了一个考虑ATR的度量。利用该度量设计和构造了具有低基表达式、高灵敏度和高非线性的逻辑门。此外，我们合理设计了具有不同GC含量和大小的合成5 'ATR，以将蛋白质表达水平调节到300倍范围内，并使用这些来构建具有不同基础表达和双稳态程度的合成切换开关。我们的综合模型和基因表达度量可以促进未来更复杂的合成基因电路的工程。Wu等人量化了相邻转录区（ATR）对约120个合成多顺反子基因回路中蛋白质表达的影响。数据驱动的分析产生了一种新的蛋白质表达度量，其与ATR特征（包括GC含量、大小和核糖体结合位点附近mRNA折叠的稳定性）密切相关。该度量的效用在预测每个基因在合成逻辑门和电路输出中的相对表达水平以及调谐基因表达和非线性动力学中被证明。
Polycistronic architecture is common for synthetic gene circuits, however, it remains unknown how expression of one gene is affected by the presence of other genes/non-coding regions in the operon, termed adjacent transcriptional regions (ATR). Here, we constructed synthetic operons with a reporter gene flanked by different ATRs, and we found ATRs with high GC content, small size, and low folding energy lead to high gene expression. Based on these results we built a model of gene expression and generated a metric that takes into account ATRs. We used the metric to design and construct logic gates with low basal expression and high sensitivity and nonlinearity. Furthermore, we rationally designed synthetic 5′ATRs with different GC content and sizes to tune protein expression levels over a 300-fold range and use these to build synthetic toggle switches with varying basal expression and degrees of bistability. Our comprehensive model and gene expression metric could facilitate the future engineering of more complex synthetic gene circuits. Wu et al. quantify the adjacent transcriptional regions’ (ATR) effect on protein expression in ~120 synthetic polycistronic gene circuits. Data-driven analysis yields a new protein expression metric that strongly correlates with ATRs features including GC content, size, and stability of mRNA folding near ribosomal binding site. This metric’s utility is demonstrated in predicting each gene’s relative expression levels in synthetic logic gates and circuit outputs and tuning gene expression and nonlinear dynamics of bistable gene networks.
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