文章基本信息

标题：Engineered temperature compensation in a synthetic genetic clock
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作者：Faiza Hussain ; Chinmaya Gupta ; Andrew J. Hirning 等
期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：2014
卷号：111
期号：3
页码：972-977
DOI：10.1073/pnas.1316298111
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：Synthetic biology promises to revolutionize biotechnology by providing the means to reengineer and reprogram cellular regulatory mechanisms. However, synthetic gene circuits are often unreliable, as changes to environmental conditions can fundamentally alter a circuit's behavior. One way to improve robustness is to use intrinsic properties of transcription factors within the circuit to buffer against intra- and extracellular variability. Here, we describe the design and construction of a synthetic gene oscillator in Escherichia coli that maintains a constant period over a range of temperatures. We started with a previously described synthetic dual-feedback oscillator with a temperature-dependent period. Computational modeling predicted and subsequent experiments confirmed that a single amino acid mutation to the core transcriptional repressor of the circuit results in temperature compensation. Specifically, we used a temperature-sensitive lactose repressor mutant that loses the ability to repress its target promoter at high temperatures. In the oscillator, this thermoinduction of the repressor leads to an increase in period at high temperatures that compensates for the decrease in period due to Arrhenius scaling of the reaction rates. The result is a transcriptional oscillator with a nearly constant period of 48 min for temperatures ranging from 30 {degrees}C to 41 {degrees}C. In contrast, in the absence of the mutation the period of the oscillator drops from 60 to 30 min over the same temperature range. This work demonstrates that synthetic gene circuits can be engineered to be robust to extracellular conditions through protein-level modifications.
关键词：LacI ; circadian oscillator ; microfluidics ; cellular dynamics ; delay