Design and characterization of a dual-mode promoter with activation and repression capability for tuning gene expression in yeast.
Design and characterization of a dual-mode promoter with activation and repression capability for tuning gene expression in yeast.
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DOI：
10.1093/nar/gku651
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发表时间：
2014-08
影响因子：
14.9
通讯作者：
McMillen DR
中科院分区：
文献类型：
--
作者：
Mazumder M;McMillen DR
人工控制基因表达的模块化正成为合成生物学的一个重要方面。基因表达的人工转录控制是设计新型合成调控网络最成熟的方法之一。这种网络旨在帮助理解自然细胞现象，并使新的生物技术应用成为可能。顺式调控元件对启动子序列的操纵是调控基因表达的关键方法。在这里，我们设计了一个既可以被激活也可以被抑制的启动子，作为合成生物学“部分”库的一部分。从酵母中最小的细胞色素C （minCYC）启动子开始，我们分别在TATA盒的上游和下游加入了5个类固醇激素响应元件（SHREs）和1个lac操作位点。这允许通过睾丸激素反应性雄激素受体激活，并通过LacI抑制因子抑制。暴露于不同浓度的睾酮（不同的激活）和IPTG（不同的抑制）证明了在大范围内调整启动子输出曲线的能力。通过整合激活和抑制信号，启动子允许一种有用的信号整合形式，我们乐观地认为，它将作为未来调节网络的一个组成部分，包括反馈控制器。
Modularity in controlling gene expression artificially is becoming an essential aspect of synthetic biology. Artificial transcriptional control of gene expression is one of the most well-developed methods for the design of novel synthetic regulatory networks. Such networks are intended to help understand natural cellular phenomena and to enable new biotechnological applications. Promoter sequence manipulation with cis-regulatory elements is a key approach to control gene expression transcriptionally. Here, we have designed a promoter that can be both activated and repressed, as a contribution to the library of synthetic biological ‘parts’. Starting with the minimal cytochrome C (minCYC) promoter in yeast, we incorporated five steroid hormone responsive elements (SHREs) and one lac operator site, respectively, upstream and downstream of the TATA box. This allows activation through the testosterone-responsive androgen receptor, and repression through the LacI repressor. Exposure to varying concentrations of testosterone (to vary activation) and IPTG (to vary repression) demonstrated the ability to tune the promoter's output curve over a wide range. By integrating activating and repressing signals, the promoter permits a useful form of signal integration, and we are optimistic that it will serve as a component in future regulatory networks, including feedback controllers.
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