Department of Bacterial Genetics, Institute of Microbiology, University of Warsaw, Miecznikowa 1, 02-096, Warsaw, Poland.
BMC Microbiol. 2011 Jul 25;11:166. doi: 10.1186/1471-2180-11-166.
Many bacterial extracytoplasmic proteins are stabilized by intramolecular disulfide bridges that are formed post-translationally between their cysteine residues. This protein modification plays an important role in bacterial pathogenesis, and is facilitated by the Dsb (disulfide bond) family of the redox proteins. These proteins function in two parallel pathways in the periplasmic space: an oxidation pathway and an isomerization pathway. The Dsb oxidative pathway in Campylobacter jejuni is more complex than the one in the laboratory E. coli K-12 strain.
In the C. jejuni 81-176 genome, the dsb genes of the oxidative pathway are arranged in three transcriptional units: dsbA2-dsbB-astA, dsbA1 and dba-dsbI. Their transcription responds to an environmental stimulus - iron availability - and is regulated in a Fur-dependent manner. Fur involvement in dsb gene regulation was proven by a reporter gene study in a C. jejuni wild type strain and its isogenic fur mutant. An electrophoretic mobility shift assay (EMSA) confirmed that analyzed genes are members of the Fur regulon but each of them is regulated by a disparate mechanism, and both the iron-free and the iron-complexed Fur are able to bind in vitro to the C. jejuni promoter regions. This study led to identification of a new iron- and Fur-regulated promoter that drives dsbA1 gene expression in an indirect way. Moreover, the present work documents that synthesis of DsbI oxidoreductase is controlled by the mechanism of translational coupling. The importance of a secondary dba-dsbI mRNA structure for dsbI mRNA translation was verified by estimating individual dsbI gene expression from its own promoter.
The present work shows that iron concentration is a significant factor in dsb gene transcription. These results support the concept that iron concentration - also through its influence on dsb gene expression - might control the abundance of extracytoplasmic proteins during different stages of infection. Our work further shows that synthesis of the DsbI membrane oxidoreductase is controlled by a translational coupling mechanism. The dba expression is not only essential for the translation of the downstream dsbI gene, but also Dba protein that is produced might regulate the activity and/or stability of DsbI.
许多细菌胞外蛋白通过分子内二硫键稳定，这些二硫键在翻译后由半胱氨酸残基之间形成。这种蛋白质修饰在细菌发病机制中起着重要作用，并且由氧化还原蛋白的 Dsb（二硫键）家族促进。这些蛋白质在周质空间中沿两条平行途径发挥作用：氧化途径和异构化途径。空肠弯曲菌的 Dsb 氧化途径比实验室大肠杆菌 K-12 菌株的途径更复杂。
在空肠弯曲菌 81-176 基因组中，氧化途径的 dsb 基因排列在三个转录单元中：dsbA2-dsbB-astA、dsbA1 和 dba-dsbI。它们的转录对环境刺激-铁的可用性-有反应，并以 Fur 依赖性方式调节。通过空肠弯曲菌野生型菌株及其同源 fur 突变体的报告基因研究证明了 Fur 参与 dsb 基因调控。电泳迁移率变动分析（EMSA）证实，分析基因是 Fur 调控子的成员，但每个基因都受到不同机制的调节，并且无铁和含铁的 Fur 都能够在体外与空肠弯曲菌启动子区域结合。这项研究导致了识别新的铁和 Fur 调节启动子的鉴定，该启动子以间接方式驱动 dsbA1 基因的表达。此外，本工作证明 DsbI 氧化还原酶的合成受翻译偶联机制的控制。通过估计从其自身启动子表达的单个 dsbI 基因，验证了 dba-dsbI mRNA 结构的次要结构对 dsbI mRNA 翻译的重要性。
本工作表明铁浓度是 dsb 基因转录的重要因素。这些结果支持这样的概念，即铁浓度-也通过其对 dsb 基因表达的影响-可能控制感染不同阶段胞外蛋白的丰度。我们的工作进一步表明，DsbI 膜氧化还原酶的合成受翻译偶联机制的控制。dba 的表达不仅对下游 dsbI 基因的翻译是必不可少的，而且产生的 Dba 蛋白也可能调节 DsbI 的活性和/或稳定性。
