Li Yang, Wu Yaokang, Xu Xianhao, Liu Yanfeng, Li Jianghua, Du Guocheng, Lv Xueqin, Li Yangyang, Liu Long
Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, Jiangnan University, NO.1800, Lihu avenue, Wuxi 214122, China.
Science Center for Future Foods, Jiangnan University, NO.1800, Lihu avenue, Wuxi 214122, China.
Nucleic Acids Res. 2025 Jan 11;53(2). doi: 10.1093/nar/gkae1315.
Inducible systems are crucial to metabolic engineering and synthetic biology, enabling organisms that function as biosensors and produce valuable compounds. However, almost all inducible systems are strain-specific, limiting comparative analyses and applications across strains rapidly. This study designed and presented a robust workflow for developing the cross-species inducible system. By applying this approach, two reconstructed inducible systems (a 2,4-diacetylphloroglucinol-inducible system PphlF3R1 and an anhydrotetracycline-inducible system Ptet2R2*) were successfully developed and demonstrated to function in three model microorganisms, including Escherichia coli, Bacillus subtilis and Corynebacterium glutamicum. To enhance their practicality, both inducible systems were subsequently placed on the plasmid and genome for detailed characterization to determine the optimal expression conditions. Furthermore, the more efficient inducible system Ptet2R2* was employed to express various reporter proteins and gene clusters in these three strains. Moreover, the aTc-inducible system Ptet2R2*, combined with T7 RNA polymerase and dCas12a, was utilized to develop a single-input genetic circuit that enables the simultaneous activation and repression of gene expression. Overall, the cross-species inducible system serves as a stringent, controllable and effective tool for protein expression and metabolic pathway control in different bacteria.
诱导系统对于代谢工程和合成生物学至关重要，它能使生物体充当生物传感器并产生有价值的化合物。然而，几乎所有诱导系统都具有菌株特异性，这限制了跨菌株的快速比较分析和应用。本研究设计并展示了一种用于开发跨物种诱导系统的强大工作流程。通过应用这种方法，成功开发了两个重构的诱导系统（一个2,4 - 二乙酰基间苯三酚诱导系统PphlF3R1和一个脱水四环素诱导系统Ptet2R2*），并证明它们在三种模式微生物中发挥作用，包括大肠杆菌、枯草芽孢杆菌和谷氨酸棒杆菌。为提高其实用性，随后将这两个诱导系统分别置于质粒和基因组上进行详细表征，以确定最佳表达条件。此外，更高效的诱导系统Ptet2R2被用于在这三种菌株中表达各种报告蛋白和基因簇。此外，aTc诱导系统Ptet2R2与T7 RNA聚合酶和dCas12a相结合，被用于开发一种单输入遗传电路，该电路能够同时激活和抑制基因表达。总体而言，跨物种诱导系统是一种用于不同细菌中蛋白质表达和代谢途径控制的严格、可控且有效的工具。
