Characterization and mitigation of gene expression burden in mammalian cells
Characterization and mitigation of gene expression burden in mammalian cells
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DOI：
10.1038/s41467-020-18392-x
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发表时间：
2020-09-15
影响因子：
16.6
通讯作者：
Siciliano, Velia
中科院分区：
文献类型：
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作者：
Frei, Timothy;Cella, Federica;Siciliano, Velia
尽管最近在电路工程方面取得了进展，但哺乳动物细胞中的基因网络设计仍然极其缓慢，并充满了令人费解的失败。在这里，我们证明了在哺乳动物细胞中瞬时表达的基因竞争有限的转录和翻译资源。这种竞争导致了原本独立的外源和内源基因的耦合，在预期功能和实际功能之间产生了分歧。在资源感知的数学模型的指导下，我们识别和设计了自然和合成的基于miRNA的非相干前馈环路(IFFL)，以减轻基因表达负担。这些电路的实现特点是使用内源性miRNAs作为设计的iFFL器件的基本组件，这是一种多功能的混合设计，允许以最小的资源需求实现跨不同细胞系的负担减轻。这项研究为在体内合成电路性能的上下文感知预测和改进奠定了基础，为在哺乳动物细胞中进行更合理的合成结构设计铺平了道路。
Despite recent advances in circuit engineering, the design of genetic networks in mammalian cells is still painstakingly slow and fraught with inexplicable failures. Here, we demonstrate that transiently expressed genes in mammalian cells compete for limited transcriptional and translational resources. This competition results in the coupling of otherwise independent exogenous and endogenous genes, creating a divergence between intended and actual function. Guided by a resource-aware mathematical model, we identify and engineer natural and synthetic miRNA-based incoherent feedforward loop (iFFL) circuits that mitigate gene expression burden. The implementation of these circuits features the use of endogenous miRNAs as elementary components of the engineered iFFL device, a versatile hybrid design that allows burden mitigation to be achieved across different cell-lines with minimal resource requirements. This study establishes the foundations for context-aware prediction and improvement of in vivo synthetic circuit performance, paving the way towards more rational synthetic construct design in mammalian cells.