Department of Bacteriology, University of Wisconsin-Madison, Madison, WI 53706, USA.
Department of Biochemistry, Duke University, Durham, NC 27710, USA.
Nucleic Acids Res. 2022 Jan 25;50(2):847-866. doi: 10.1093/nar/gkab1281.
The nucleotide messenger (p)ppGpp allows bacteria to adapt to fluctuating environments by reprogramming the transcriptome. Despite its well-recognized role in gene regulation, (p)ppGpp is only known to directly affect transcription in Proteobacteria by binding to the RNA polymerase. Here, we reveal a different mechanism of gene regulation by (p)ppGpp in Firmicutes: (p)ppGpp directly binds to the transcription factor PurR to downregulate purine biosynthesis gene expression upon amino acid starvation. We first identified PurR as a receptor of (p)ppGpp in Bacillus anthracis. A co-structure with Bacillus subtilis PurR reveals that (p)ppGpp binds to a PurR pocket reminiscent of the active site of phosphoribosyltransferase enzymes that has been repurposed to serve a purely regulatory role, where the effectors (p)ppGpp and PRPP compete to allosterically control transcription. PRPP inhibits PurR DNA binding to induce transcription of purine synthesis genes, whereas (p)ppGpp antagonizes PRPP to enhance PurR DNA binding and repress transcription. A (p)ppGpp-refractory purR mutant in B. subtilis fails to downregulate purine synthesis genes upon amino acid starvation. Our work establishes the precedent of (p)ppGpp as an effector of a classical transcription repressor and reveals the key function of (p)ppGpp in regulating nucleotide synthesis through gene regulation, from soil bacteria to pathogens.
核苷酸信使（p）ppGpp 通过重编程转录组使细菌能够适应不断变化的环境。尽管它在基因调控中的作用得到了广泛的认可，但（p）ppGpp 仅已知通过与 RNA 聚合酶结合直接影响变形菌中的转录。在这里，我们揭示了（p）ppGpp 在厚壁菌门中调节基因的不同机制：（p）ppGpp 直接结合转录因子 PurR，在氨基酸饥饿时下调嘌呤生物合成基因的表达。我们首先在炭疽芽孢杆菌中鉴定 PurR 为（p）ppGpp 的受体。与枯草芽孢杆菌 PurR 的共结构揭示了（p）ppGpp 结合到 PurR 口袋中，该口袋类似于磷酸核糖基转移酶酶的活性位点，该酶被重新用于纯粹的调节作用，其中效应物（p）ppGpp 和 PRPP 竞争变构控制转录。PRPP 抑制 PurR DNA 结合以诱导嘌呤合成基因的转录，而（p）ppGpp 拮抗 PRPP 以增强 PurR DNA 结合并抑制转录。枯草芽孢杆菌中（p）ppGpp 无反应性的 purR 突变体在氨基酸饥饿时不能下调嘌呤合成基因的表达。我们的工作确立了（p）ppGpp 作为经典转录抑制剂效应物的先例，并揭示了（p）ppGpp 通过基因调控在调节核苷酸合成中的关键作用，从土壤细菌到病原体。
