Modulating Pathway Performance by Perturbing Local Genetic Context.,ACS Synthetic Biology

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Modulating Pathway Performance by Perturbing Local Genetic Context.
ACS Synthetic Biology ( IF 3.7 ) Pub Date : 2020-03-24 , DOI: 10.1021/acssynbio.9b00445
Carmen Lopez ₁ , Yuxin Zhao _{2,

3} , Rick Masonbrink ₄ , Zengyi Shao _{1,

3,

5,

6}

Affiliation

Combinatorial engineering is a preferred strategy for attaining optimal pathway performance. Previous endeavors have been concentrated on regulatory elements (e.g., promoters, terminators, and ribosomal binding sites) and/or open reading frames. Accumulating evidence indicates that noncoding DNA sequences flanking a transcriptional unit on the genome strongly impact gene expression. Here, we sought to mimic the effect imposed on expression cassettes by the genome. We created variants of the model yeast Saccharomyces cerevisiae with significantly improved fluorescence or cellobiose consumption rate by randomizing the sequences adjacent to the GFP expression cassette or the cellobiose-utilization pathway, respectively. Interestingly, nucleotide specificity was observed at certain positions and showed to be essential for achieving optimal cellobiose assimilation. Further characterization suggested that the modulation effects of the short sequences flanking the expression cassettes could be potentially mediated by remodeling DNA packaging and/or recruiting transcription factors. Collectively, these results indicate that the often-overlooked contiguous DNA sequences can be exploited to rapidly achieve balanced pathway expression, and the corresponding approach could be easily stacked with other combinatorial engineering strategies.

中文翻译：

通过干扰局部遗传背景来调节途径的表现。

组合工程是获得最佳路径性能的首选策略。先前的努力集中在调节元件（例如启动子，终止子和核糖体结合位点）和/或开放阅读框上。越来越多的证据表明，位于基因组转录单元侧翼的非编码DNA序列会强烈影响基因表达。在这里，我们试图模仿基因组对表达盒的作用。通过分别随机化与GFP表达盒或纤维二糖利用途径相邻的序列，我们创建了具有明显提高的荧光或纤维二糖消耗率的模型酵母啤酒酵母变体。有趣的是在某些位置观察到核苷酸特异性，并显示对于实现最佳纤维二糖同化至关重要。进一步的特征表明，表达盒两侧的短序列的调节作用可能通过重塑DNA包装和/或募集转录因子来介导。总而言之，这些结果表明，经常被忽视的连续DNA序列可以被利用来快速实现平衡的途径表达，并且相应的方法可以很容易地与其他组合工程策略堆叠在一起。

更新日期：2020-04-23

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