Zhu Manlu, Dai Xiongfeng
Key Laboratory of Pesticide & Chemical Biology of Ministry of Education, State Key Laboratory of Green Pesticides, Hubei Key Laboratory of Genetic Regulation and Integrative Biology, School of Life Sciences, Central China Normal University, Wuhan, 430079, China.
Nucleic Acids Res. 2025 Aug 11;53(15). doi: 10.1093/nar/gkaf814.
Gene regulation and its interplay with physiological behaviors are the central topics of modern biology. Classical studies on gene regulation focus intensively on specific regulatory mechanisms of transcription. Nevertheless, the genome-wide impact of RNA polymerase (RNAP) availability on gene expression remains poorly understood. Here we developed two synthetic transcriptional switches to systematically titrate the expression of either ${\sigma ^A}$ (SigA, housekeeping sigma factor) or RpoBC (core enzyme) in Bacillus subtilis. Both systems effectively modulated cell growth, but with fundamentally distinct mechanisms. SigA limitation triggered significant resource reallocation, redirecting cellular investment from biosynthetic pathways to alternative cellular pathways, which could further facilitate the engineering of dynamic growth-bioproduction switch. In contrast, RpoBC depletion caused only weak changes of gene expression but induced ribosomal inactivation through blocking translation initiation. Notably, RpoBC depletion induced DNA damage response and increased the DNA damage sensitivity of bacteria, suggesting transcription-coupled repair as a critical survival mechanism. Our findings delineate two regulatory paradigms of resource allocation that are associated with the interplay between RNAP availability and bacterial physiological state, "abundance-based" and "activity-based" regulations. The orthogonal transcriptional switches serve as a powerful tool for dissecting the integrative role of RNAP in microbial physiology, offering meaningful implications for both fundamental studies of gene regulation and synthetic biology applications.
基因调控及其与生理行为的相互作用是现代生物学的核心主题。经典的基因调控研究主要集中在转录的特定调控机制上。然而，RNA聚合酶（RNAP）可用性对全基因组基因表达的影响仍知之甚少。在这里，我们开发了两个合成转录开关，以系统地调节枯草芽孢杆菌中${\sigma ^A}$（SigA，管家σ因子）或RpoBC（核心酶）的表达。这两个系统都有效地调节了细胞生长，但机制截然不同。SigA限制引发了显著的资源重新分配，将细胞投资从生物合成途径转移到其他细胞途径，这可能进一步促进动态生长-生物生产开关的工程设计。相比之下，RpoBC的消耗仅引起基因表达的微弱变化，但通过阻断翻译起始诱导核糖体失活。值得注意的是，RpoBC的消耗诱导了DNA损伤反应并增加了细菌对DNA损伤的敏感性，表明转录偶联修复是一种关键的生存机制。我们的研究结果描绘了两种资源分配的调控模式，它们与RNAP可用性和细菌生理状态之间的相互作用相关，即“基于丰度”和“基于活性”的调控。正交转录开关是剖析RNAP在微生物生理学中综合作用的有力工具，对基因调控的基础研究和合成生物学应用都具有重要意义。
