Cyanobacteria are photosynthetic prokaryotes being developed as sustainable platforms that use renewable resources (light, water, and air) for diverse applications in energy, food, environment, and medicine. Despite the attractive promise that cyanobacteria offer to industrial biotechnology, slow growth rates pose a major challenge in processes which typically require large amounts of biomass and are often toxic to the cells. Two-stage cultivation strategies are an attractive solution to prevent any undesired growth inhibition by de-coupling biomass accumulation (stage I) and the industrial process (stage II). In cyanobacteria, two-stage strategies involve costly transfer methods between stages I and II, and little work has been focussed on using the distinct growth and stationary phases of batch cultures to autoregulate stage transition. In the present study, we identified and characterised a growth phase-specific promoter, which can serve as an auto-inducible switch to regulate two-stage bioprocesses in cyanobacteria. First, growth phase-specific genes were identified from a new RNAseq dataset comparing two growth phases and six nutrient conditions in Synechocystis sp. PCC 6803, including two new transcriptomes for low Mg and low K. A type II NADH dehydrogenase (ndbA) showed robust induction when the cultures transitioned from exponential to stationary phase growth. Behaviour of a 600-bp promoter sequence (PndbA600) was then characterised in detail following the expression of PndbA600:GFP in Synechococcus sp. PCC 7002. Culture density and growth media analyses showed that PndbA600 activation was not dependent on increases in culture density per se but on N availability and on another activating factor present in the spent media of stationary phase cultures (Factor X). PndbA600 deactivation was dependent on the changes in culture density and in either N availability or Factor X. Electron transport inhibition studies revealed a photosynthesis-specific enhancement of active PndbA600 levels. Our findings are summarised in a model describing the environmental regulation of PndbA600, which can now inform the rational design of two-stage industrial processes in cyanobacteria.

中文翻译：

---

PndbA600 的环境调节，一种用于蓝藻两阶段工业生物技术的自诱导启动子

蓝藻是一种光合原核生物，正被开发为可持续平台，利用可再生资源（光、水和空气）在能源、食品、环境和医学方面的各种应用。尽管蓝藻为工业生物技术提供了诱人的前景，但缓慢的生长速度对通常需要大量生物质且通常对细胞有毒的过程构成了重大挑战。两阶段培养策略是一种有吸引力的解决方案，可通过将生物质积累（阶段 I）和工业过程（阶段 II）解耦来防止任何不希望的生长抑制。在蓝藻中，两阶段策略涉及在阶段 I 和 II 之间昂贵的转移方法，并且很少有工作关注使用分批培养的不同生长和静止阶段来自动调节阶段过渡。在本研究中，我们鉴定并表征了一种生长期特异性启动子，它可以作为自诱导开关来调节蓝藻中的两阶段生物过程。首先，从新的 RNAseq 数据集中鉴定了生长阶段特异性基因，该数据集比较了集胞藻属的两个生长阶段和六个营养条件。PCC 6803，包括低镁和低钾的两个新转录组。当培养物从指数生长过渡到稳定期生长时，II 型 NADH 脱氢酶 (ndbA) 表现出强烈的诱导作用。600 bp 启动子序列 (PndbA600) 的行为然后在 PndbA600: GFP 在聚球藻中表达后被详细表征。PCC 7002。培养密度和生长培养基分析表明，PndbA600 的激活不依赖于培养密度本身的增加，而是依赖于 N 可用性和固定相培养物用过的培养基中存在的另一种激活因子（因子 X）。PndbA600 失活取决于培养密度和 N 可用性或因子 X 的变化。电子传输抑制研究揭示了活性 PndbA600 水平的光合作用特异性增强。我们的研究结果总结在一个描述 PndbA600 环境调节的模型中，该模型现在可以为蓝藻中两阶段工业过程的合理设计提供信息。PndbA600 失活取决于培养密度和 N 可用性或因子 X 的变化。电子传输抑制研究揭示了活性 PndbA600 水平的光合作用特异性增强。我们的研究结果总结在一个描述 PndbA600 环境调节的模型中，该模型现在可以为蓝藻中两阶段工业过程的合理设计提供信息。PndbA600 失活取决于培养密度和 N 可用性或因子 X 的变化。电子传输抑制研究揭示了活性 PndbA600 水平的光合作用特异性增强。我们的研究结果总结在一个描述 PndbA600 环境调节的模型中，该模型现在可以为蓝藻中两阶段工业过程的合理设计提供信息。