Selective Gene Regulation by Manganese and Iron in Bacillus subtilis

所有生物体都必须从它们的环境中获得一套特定的微量金属。因此，细胞必须能够感知细胞内单个金属离子的水平，以便适当地协调金属离子的运输、储存和细胞内运输。细胞内的金属离子水平是由金属调节蛋白控制的，金属调节蛋白可以抑制或激活金属离子稳态基因的表达。在枯草芽孢杆菌中，四种不同的金属调节因子控制锌、铁和锰的运输并调节氧化应激反应。本项目研究了两种调节因子Fur和PerR蛋白的生物学作用和机制。铁摄取抑制因子（Fur）蛋白作为铁依赖性抑制因子，协调铁摄取功能的表达。然而，最近的生化研究表明，Fur与它的靶操作位点紧密结合，独立于铁。这表明，在细胞中，Fur的活性受到其他因素的调节，可能是一种金属离子，它可以拮抗这种内在的dna结合活性。过氧化调节子（PerR）抑制因子在结构上与Fur相关。然而，PerR在生理上的功能是作为氧化应激的传感器，而不是金属离子饥饿。提出PerR与金属结合的辅因子一起与过氧化氢反应生成非活性抑制因子。导致抑郁症的PerR变化的性质目前尚不清楚。在这个项目中，将使用分子遗传学和生物化学方法的结合来研究铁传感的分子基础，通过Fur和过氧化物传感，PerR。这些研究将深入了解蛋白质如何选择性地与细胞中不同的金属离子结合，以及它们如何利用金属离子作为氧化应激的传感器。相关调控因子在多种细菌中的存在表明，通过对枯草芽孢杆菌调控因子的详细表征所建立的范式将具有深远的意义。

All organisms must obtain a specific set of trace metals from their environment. Accordingly, cells must be able to sense the intracellular level of individual metal ions in order to properly coordinate metal ion transport, storage, and intracellular trafficking. Cellular metal ion levels are monitored by metalloregulatory proteins that repress or activate the expression of metal ion homeostasis genes. In Bacillus subtilis, four distinct metalloregulators control the transport of zinc, iron, and manganese and regulate an oxidative stress response. This project addresses the biological roles and mechanisms of two regulators, the Fur and PerR proteins. The ferric uptake repressor (Fur) protein coordinates the expression of iron uptake functions by acting as an iron-dependent repressor. However, recent biochemical studies indicate that Fur binds tightly to its target operator sites independently of iron. This suggests that in the cell the activity of Fur is regulated by other factors, perhaps a metal ion, that antagonize this intrinsic DNA-binding activity. The peroxide regulon (PerR) repressor is structurally related to Fur. However, PerR functions physiologically as a sensor of oxidative stress, rather than metal ion starvation. It is proposed that PerR, together with a metal bound cofactor, reacts with hydrogen peroxide to generate an inactive repressor. The nature of the change(s) in PerR that lead to derepression are currently unknown. In this project, the molecular basis for Fe-sensing, by Fur, and peroxide-sensing, by PerR, will be investigated using a combination of molecular genetic and biochemical approaches. These studies will provide insights into how proteins bind selectively to different metal ions in the cell and how they can use metal ions to serve as sensors of oxidative stress. The presence of related regulators in diverse bacteria argues that the paradigms established by a detailed characterization of the B. subtilis regulators will have far-reaching implications.