Francez-Charlot Anne, Frunzke Julia, Reichen Christian, Ebneter Judith Zingg, Gourion Benjamin, Vorholt Julia A
Institute of Microbiology, Eidgenössiche Technische Hochschule Zurich, 8093 Zurich, Switzerland.
Proc Natl Acad Sci U S A. 2009 Mar 3;106(9):3467-72. doi: 10.1073/pnas.0810291106. Epub 2009 Feb 13.
Bacteria have evolved regulatory traits to rapidly adapt to changing conditions. Two principal regulatory mechanisms to modulate gene expression consist of regulation via alternative sigma factors and phosphorylation-dependent response regulators. PhyR represents a recently discovered protein family combining parts of both systems: a sigma factor-like domain of the extracytoplasmic function (ECF) subfamily linked to a receiver domain of a response regulator. Here we investigated the mode of action of this key regulator of general stress response in Methylobacterium extorquens. Our results indicate that PhyR does not act as a genuine sigma factor but instead controls gene expression indirectly through protein-protein interactions. This is evident from the analysis of additional proteins involved in PhyR-dependent gene regulation. We demonstrated that the ECF sigma factor-like domain of PhyR interacts with a protein, designated NepR, upon phosphorylation of the PhyR receiver domain. Using transcriptome analysis and phenotypic assays, we showed that NepR is a negative regulator of PhyR response. Furthermore, we provide biochemical and genetic evidence that NepR exerts this inhibitory effect through sequestration of the ECF sigma factor sigma(EcfG1). Our data support an unprecedented model according to which PhyR acts as a mimicry protein triggering a partner-switching mechanism. Such a regulation of general stress response clearly differs from the two known models operating via sigma(S) and sigma(B). Given the absence of these master regulators and the concomitant conservation of PhyR in Alphaproteobacteria, the novel mechanism presented here is most likely central to the control of general stress response in this large subclass of Proteobacteria.
细菌已经进化出调控特性以快速适应不断变化的环境。调节基因表达的两种主要调控机制包括通过替代西格玛因子和磷酸化依赖性反应调节因子进行调控。PhyR代表最近发现的一个蛋白质家族，它结合了这两种系统的部分特征：胞外功能（ECF）亚家族的一个类似西格玛因子的结构域与一个反应调节因子的接收结构域相连。在这里，我们研究了甲基营养型甲基杆菌中这种一般应激反应关键调节因子的作用模式。我们的结果表明，PhyR并不作为一个真正的西格玛因子起作用，而是通过蛋白质-蛋白质相互作用间接控制基因表达。这从对参与PhyR依赖性基因调控的其他蛋白质的分析中可以明显看出。我们证明，在PhyR接收结构域磷酸化后，PhyR的ECF西格玛因子样结构域与一种名为NepR的蛋白质相互作用。通过转录组分析和表型分析，我们表明NepR是PhyR反应的负调节因子。此外，我们提供了生化和遗传学证据，表明NepR通过隔离ECF西格玛因子sigma(EcfG1)发挥这种抑制作用。我们的数据支持了一个前所未有的模型，即PhyR作为一种模拟蛋白触发伙伴切换机制。这种一般应激反应的调节明显不同于通过sigma(S)和sigma(B)运行的两种已知模型。鉴于这些主要调节因子的缺失以及PhyR在α-变形菌中的保守性，这里提出的新机制很可能是这个大型变形菌亚类中一般应激反应控制的核心。
