Regulation of iron-sulfur cluster assemby in a facultative phototrophic alpha- proteobacterium

兼性光养α-变形菌中铁硫簇组装的调节

• 批准号: 289751504
• 负责人: Professorin Dr. Gabriele Klug
• 金额: --
• 依托单位: Institut für Mikrobiologie und Molekularbiologie
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2015
• 资助国家: 德国
• 起止时间: 2014-12-31 至 2019-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/289751504?language=en
• 关键词: Regulation; iron; sulfur; cluster; assemby

Proteins with iron-sulfur (Fe-S) clusters fulfill many essential biological functions such as electron transfer, redox sensing, catalysis, or DNA replication and repair. Oxygen and reactive oxygen species (ROS) destabilize Fe-S clusters leading to the release of Fe2+ ions that in turn potentiate oxygen toxicity by the production of hydroxyl radicals in the Fenton reaction. Thus organisms had to develop systems that promote biogenesis of Fe-S proteins while protecting the cellular surrounding from the deleterious effects of free iron. Genes for such systems are therefore regulated by iron availability and oxidative stress. Escherichia coli and related species harbor two systems for Fe-S cluster assembly, Isc and Suf and the regulation of the corresponding operons was intensively studied. However, little is known about the regulation of Fe-S assembly in other bacteria.Rhodobacter sphaeroides is an alpha-proteobacterium, which forms photosynthetic complexes at low oxygen tension increasing the demand for Fe-S clusters and at the same time the risk for the formation of ROS. R. sphaeroides has the iscRS and suf genes arranged together and we have evidence for individual promoters for isc and suf genes, yet also for co-transcription. Moreover, differential RNAseq analyses also revealed transcripts, which are partially antisense to the iscR transcript. Our preliminary data demonstrated that iron availability and oxygen concentrations have a major impact on isc-suf operon expression and that several protein regulators are involved in its control, some of which harbour Fe-S, heme or iron cofactors. As in E. coli, IscR itself is a regulator for isc-suf expression and binds an Fe-S center. R. sphaeroides IscR lacks however the conserved residues involved in Fe S binding in other bacteria. Furthermore, the iron-dependently expressed sRNA SurS is controlled by IscR and affects genes of the sulfur metabolism and for iron storage. Hence regulation of Fe-S assembly in this phototroph appears to differ significantly from that in E. coli.We will elucidate the mechanisms that control expression of the isc-suf genes and coordinate Fe-S assembly and formation of photosynthetic complexes in R. sphaeroides. The effect of different growth conditions, protein and RNA regulators on the individual isc-suf promoters will be analyzed. Special emphasis will be given to the question of whether formation of photosynthetic complexes or rather photosynthesis affects isc-suf expression and what the underlying mechanisms are and how the Suf machinery affects formation of photosynthetic complexes. We will also attempt to define the new type of Fe S binding site and test whether this ligation type is of advantage for regulation in R. sphaeroides. We expect to elucidate new strategies for the regulation of Fe-S cluster assembly in the special surrounding of a phototrophic organism.

具有铁硫（Fe-S）簇的蛋白质具有许多基本的生物学功能，如电子转移、氧化还原传感、催化或DNA复制和修复。氧和活性氧（ROS）破坏Fe-S簇的稳定性，导致Fe2+离子的释放，进而通过在芬顿反应中产生羟基自由基来增强氧毒性。因此，生物体必须发展出促进铁硫蛋白生物生成的系统，同时保护细胞周围免受游离铁的有害影响。因此，这些系统的基因受到铁的可用性和氧化应激的调节。大肠杆菌及其相关物种具有两个Fe-S簇组装系统Isc和Suf，并对其操作子的调控进行了深入研究。然而，对其他细菌中Fe-S组装的调控知之甚少。球形红杆菌是一种α -变形杆菌，在低氧张力下形成光合复合体，增加了对Fe-S簇的需求，同时也增加了形成ROS的风险。sphaeroides的iscRS和自基因排列在一起，我们有证据表明isc和自基因有单独的启动子，但也有共转录的启动子。此外，差异RNAseq分析还揭示了与iscR转录本部分反义的转录本。我们的初步数据表明，铁可用性和氧浓度对isc-suf操纵子的表达有重要影响，并且几种蛋白质调节因子参与其控制，其中一些含有Fe-S，血红素或铁辅助因子。与大肠杆菌一样，IscR本身是isc-suf表达的调节因子，并结合Fe-S中心。然而，sphaeroides在其他细菌中缺乏Fe - S结合的保守残基。此外，铁依赖性表达的sRNA SurS受IscR控制，影响硫代谢和铁储存基因。因此，Fe-S在这种光养菌中的组装调节似乎与大肠杆菌中的组装有很大不同。我们将阐明控制isc-suf基因的表达，协调铁-s组装和光合复合物形成的机制。我们将分析不同生长条件、蛋白质和RNA调节因子对单个isc-suf启动子的影响。特别强调的问题将是光合复合体的形成或光合作用是否影响isc-suf的表达，以及潜在的机制是什么以及Suf机制如何影响光合复合体的形成。我们还将尝试确定新型的Fe S结合位点，并测试这种连接类型是否有利于在圆叶藻中进行调控。我们期望阐明光养生物特殊环境中Fe-S簇组装调控的新策略。