Walter Tatjana, Veldmann Kareen H, Götker Susanne, Busche Tobias, Rückert Christian, Kashkooli Arman Beyraghdar, Paulus Jannik, Cankar Katarina, Wendisch Volker F
Genetics of Prokaryotes, Faculty of Biology and CeBiTec, Bielefeld University, 33615 Bielefeld, Germany.
Center for Biotechnology, Bielefeld University, 33615 Bielefeld, Germany.
Microorganisms. 2020 Dec 8;8(12):1945. doi: 10.3390/microorganisms8121945.
The aromatic heterocyclic compound indole is widely spread in nature. Due to its floral odor indole finds application in dairy, flavor, and fragrance products. Indole is an inter- and intracellular signaling molecule influencing cell division, sporulation, or virulence in some bacteria that synthesize it from tryptophan by tryptophanase. that is used for the industrial production of amino acids including tryptophan lacks tryptophanase. To test if indole is metabolized by or has a regulatory role, the physiological response to indole by this bacterium was studied. As shown by RNAseq analysis, indole, which inhibited growth at low concentrations, increased expression of genes involved in the metabolism of iron, copper, and aromatic compounds. In part, this may be due to iron reduction as indole was shown to reduce Fe to Fe in the culture medium. Mutants with improved tolerance to indole were selected by adaptive laboratory evolution. Among the mutations identified by genome sequencing, mutations in three transcriptional regulator genes were demonstrated to be causal for increased indole tolerance. These code for the regulator of iron homeostasis DtxR, the regulator of oxidative stress response RosR, and the hitherto uncharacterized Cg3388. Gel mobility shift analysis revealed that Cg3388 binds to the intergenic region between its own gene and the operon encoding inositol uptake system IolT2, maleylacetate reductase, and catechol 1,2-dioxygenase. Increased RNA levels of in a deletion strain indicated that Cg3388 acts as repressor. Indole, hydroquinone, and 1,2,4-trihydroxybenzene may function as inducers of the operon in vivo as they interfered with DNA binding of Cg3388 at physiological concentrations in vitro. Cg3388 was named IhtR.
芳香杂环化合物吲哚在自然界中广泛分布。由于其花香气味，吲哚被应用于乳制品、香料和香精产品中。吲哚是一种细胞间和细胞内信号分子，在一些能通过色氨酸酶将色氨酸合成吲哚的细菌中，它会影响细胞分裂、孢子形成或毒力。用于包括色氨酸在内的氨基酸工业生产的[细菌名称未给出]缺乏色氨酸酶。为了测试吲哚是否会被[细菌名称未给出]代谢或具有调节作用，研究了该细菌对吲哚的生理反应。如RNA测序分析所示，低浓度时抑制生长的吲哚会增加参与铁、铜和芳香族化合物代谢的基因的表达。部分原因可能是铁还原，因为吲哚在培养基中能将Fe³⁺还原为Fe²⁺。通过适应性实验室进化筛选出了对吲哚耐受性增强的突变体。在通过基因组测序鉴定出的突变中，三个转录调节基因的突变被证明是吲哚耐受性增强的原因。它们分别编码铁稳态调节因子DtxR、氧化应激反应调节因子RosR以及此前未被表征的Cg3388。凝胶迁移率变动分析表明，Cg3388与它自身基因和编码肌醇摄取系统IolT2、马来酰乙酸还原酶以及儿茶酚1,2 - 双加氧酶的操纵子之间的基因间区域结合。在[基因名称未给出]缺失菌株中[基因名称未给出]的RNA水平升高表明Cg3388作为阻遏物起作用。吲哚、对苯二酚和1,2,4 - 三羟基苯在体内可能作为该操纵子的诱导物，因为它们在体外生理浓度下会干扰Cg3388与DNA的结合。Cg3388被命名为IhtR。
