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

具有铁-硫（Fe-S）簇的蛋白质实现许多基本的生物学功能，例如电子传递、氧化还原传感、催化或DNA复制和修复。氧和活性氧物质（ROS）使Fe-S簇不稳定，导致Fe 2+离子的释放，这又通过在芬顿反应中产生羟基自由基而增强氧毒性。因此，生物体必须开发促进Fe-S蛋白生物合成的系统，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，相应操纵子的调控已被深入研究。球形红细菌（Rhodobacter sphaeroides）是一种α-变形菌，它在低氧环境下形成光合复合物，增加了对Fe-S簇的需求，同时也增加了ROS形成的风险。R. sphaeroides具有排列在一起的iscRS和suf基因，并且我们有证据证明isc和suf基因的单独启动子，还有共转录。此外，差异RNAseq分析还揭示了与iscR转录物部分反义的转录物。我们的初步数据表明，铁的可用性和氧浓度有一个主要的影响isc-suf操纵子的表达和几个蛋白质调节剂参与其控制，其中一些港口铁硫，血红素或铁辅因子。在E.在大肠杆菌中，IscR本身是isc-suf表达的调节剂，并结合Fe-S中心。R.然而，sphaeroides IscR缺乏在其他细菌中参与Fe S结合的保守残基。此外，铁依赖性表达的sRNA SurS受IscR控制，并影响硫代谢和铁储存的基因。因此，这种光合细菌中Fe-S组装的调节似乎与E.我们将阐明控制isc-suf基因表达和协调铁硫组装以及光合复合物形成的机制。sphaeroides将分析不同生长条件、蛋白质和RNA调节剂对单个isc-suf启动子的影响。特别强调的问题是，是否形成光合复合物或光合作用的影响ISC-SUF表达和潜在的机制是什么，以及如何SuF机械影响光合复合物的形成。我们也将尝试定义新的Fe S结合位点类型，并测试这种连接类型是否有利于R. sphaeroides我们希望阐明在光养生物的特殊环境中调控Fe-S簇组装的新策略。

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.