Cells use feedback regulation to ensure robust growth despite fluctuating demands for resources and differing environmental conditions. However, the expression of foreign proteins from engineered constructs is an unnatural burden that cells are not adapted for. Here we combined RNA-seq with an in vivo assay to identify the major transcriptional changes that occur in Escherichia coli when inducible synthetic constructs are expressed. We observed that native promoters related to the heat-shock response activated expression rapidly in response to synthetic expression, regardless of the construct. Using these promoters, we built a dCas9-based feedback-regulation system that automatically adjusts the expression of a synthetic construct in response to burden. Cells equipped with this general-use controller maintained their capacity for native gene expression to ensure robust growth and thus outperformed unregulated cells in terms of protein yield in batch production. This engineered feedback is to our knowledge the first example of a universal, burden-based biomolecular control system and is modular, tunable and portable.

中文翻译：

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负担驱动的基因表达反馈控制。

尽管对资源的需求变化和环境条件的变化，电池仍使用反馈调节来确保强劲的增长。然而，来自工程构建体的外源蛋白的表达是细胞不适应的非自然负担。在这里，我们将RNA-seq与体内测定法相结合，以鉴定表达可诱导的合成构建体时在大肠杆菌中发生的主要转录变化。我们观察到，与热休克反应相关的天然启动子响应合成表达而迅速激活表达，而与构建体无关。使用这些启动子，我们构建了一个基于dCas9的反馈调节系统，该系统可根据负担自动调节合成构建体的表达。配备此通用控制器的细胞可保持其天然基因表达的能力，以确保强劲的生长，因此在批量生产中，在蛋白质产量方面胜过不受调节的细胞。就我们所知，这种工程反馈是通用的，基于负荷的生物分子控制系统的第一个示例，并且是模块化，可调式和便携式的。