The bacterium Francisella tularensis (Ft) is one of the most infectious agents known. Ft virulence is controlled by a unique combination of transcription regulators: the MglA-SspA heterodimer, PigR, and the stress signal, ppGpp. MglA-SspA assembles with the σ⁷⁰-associated RNAP holoenzyme (RNAPσ⁷⁰), forming a virulence-specialized polymerase. These factors activate Francisella pathogenicity island (FPI) gene expression, which is required for virulence, but the mechanism is unknown. Here we report FtRNAPσ⁷⁰-promoter-DNA, FtRNAPσ⁷⁰-(MglA-SspA)-promoter DNA, and FtRNAPσ⁷⁰-(MglA-SspA)-ppGpp-PigR-promoter DNA cryo-EM structures. Structural and genetic analyses show MglA-SspA facilitates σ⁷⁰ binding to DNA to regulate virulence and virulence-enhancing genes. Our Escherichia coli RNAPσ^70-homodimeric EcSspA structure suggests this is a general SspA-transcription regulation mechanism. Strikingly, our FtRNAPσ⁷⁰-(MglA-SspA)-ppGpp-PigR-DNA structure reveals ppGpp binding to MglA-SspA tethers PigR to promoters. PigR in turn recruits FtRNAP αCTDs to DNA UP elements. Thus, these studies unveil a unique mechanism for Ft pathogenesis involving a virulence-specialized RNAP that employs two (MglA-SspA)-based strategies to activate virulence genes.

土拉弗朗西斯菌 ( Ft ) 是已知最具传染性的病原体之一。 Ft毒力由转录调节因子的独特组合控制：MglA-SspA 异二聚体、PigR 和应激信号 ppGpp。 MglA-SspA 与 σ ⁷⁰相关 RNAP 全酶 (RNAPσ ⁷⁰ ) 组装，形成毒力特异性聚合酶。这些因子激活弗朗西斯菌致病岛（FPI）基因表达，这是毒力所必需的，但其机制尚不清楚。在这里，我们报告了Ft RNAPσ ^{70 -}启动子-DNA、 Ft RNAPσ ⁷⁰ -(MglA-SspA)-启动子 DNA 和Ft RNAPσ ⁷⁰ -(MglA-SspA)-ppGpp-PigR-启动子 DNA 冷冻电镜结构。结构和遗传分析表明 MglA-SspA 促进 σ ⁷⁰与 DNA 结合，从而调节毒力和毒力增强基因。我们的大肠杆菌RNAPσ ^70-同源二聚体Ec SspA 结构表明这是一种通用的 SspA 转录调控机制。引人注目的是，我们的Ft RNAPσ ⁷⁰ -(MglA-SspA)-ppGpp-PigR-DNA 结构揭示了 ppGpp 与 MglA-SspA 的结合将 PigR 束缚到启动子上。 PigR 反过来将Ft RNAP αCTD 招募到 DNA UP 元件上。因此，这些研究揭示了Ft发病机制的独特机制，涉及毒力特异性 RNAP，该 RNAP 采用两种基于 (MglA-SspA) 的策略来激活毒力基因。