Advances in synthetic biology have allowed photosynthetic cyanobacteria as promising “green cell factories” for sustainable production of biofuels and biochemicals. However, a limited of genetic switches developed in cyanobacteria restrict the complex and orthogonal metabolic regulation. In addition, suitable and controllable switches sensing and responding to specific inducers would allow for the separation of cellular growth and expression of exogenous genes or pathways that cause metabolic burden or toxicity. Here in this study, we developed a genetic switch repressed by NanR and induced by N-acetylneuraminic acid (Neu5Ac) in a fast-growing cyanobacterium Synechococcus elongatus UTEX 2973 along with its highly homologous strain S. elongatus PCC 7942. First, nanR from Escherichia coli and a previously optimized cognate promoter P_J23119H10 were introduced into Syn2973 to control the expression of the reporter gene lacZ encoding β-galactosidase, achieving induction with negligible leakage. Second, the switch was systemically optimized to reach ∼738-fold induction by fine-tuning the expression level of NanR and introducing additional transporter of Neu5Ac. Finally, the orthogonality between the NanR/Neu5Ac switch and theophylline-responsive riboregulator was investigated, achieving a coordinated regulation or binary regulation toward the target gene. Our work here provided a new switch for transcriptional control and orthogonal regulation strategies in cyanobacteria, which would promote the metabolic regulation for the cyanobacterial chassis in the future.

中文翻译：

---

开发基于 N-乙酰神经氨酸的传感和响应开关，用于蓝藻聚球藻菌株的正交基因调控

合成生物学的进步使光合蓝藻成为有前途的“绿色细胞工厂”，用于可持续生产生物燃料和生物化学品。然而，蓝藻中开发的有限的遗传开关限制了复杂和正交的代谢调节。此外，感应和响应特定诱导物的合适且可控的开关将允许分离细胞生长和外源基因或引起代谢负担或毒性的途径的表达。在这项研究中，我们在快速生长的蓝藻细长聚球藻UTEX 2973 及其高度同源的菌株S. elongatus中开发了由 NanR 抑制并由N-乙酰神经氨酸 (Neu5Ac) 诱导的遗传开关PCC 7942. 首先，将来自大肠杆菌的nanR和先前优化的同源启动子 P _J23119H10引入 Syn2973 以控制报告基因lacZ的表达编码β-半乳糖苷酶，实现诱导而泄漏可忽略不计。其次，通过微调 NanR 的表达水平和引入额外的 Neu5Ac 转运蛋白，对开关进行了系统优化，以达到 738 倍的诱导。最后，研究了NanR/Neu5Ac开关与茶碱反应性核糖调节剂之间的正交性，实现了对靶基因的协同调控或二元调控。我们在这里的工作为蓝藻的转录控制和正交调控策略提供了一个新的开关，这将促进未来蓝藻底盘的代谢调控。