Regulation of Bacterial Heme Metabolism

描述（由申请人提供）：血红素及其生物合成的调节对于正常细胞功能是重要的，并且有助于细菌在不同环境中的成功，包括与高等真核生物的致病性和共生相互作用相关的环境。血红素现在被认为是一种调节分子，允许细胞感知和适应环境线索以维持稳态。拟议工作的广泛目标是了解血红素如何介导基因表达的控制，重点是铁代谢，并调和血红素的这种作用与其在细胞中的毒性。血红素生物合成途径的高潮是通过亚铁螯合酶催化的反应将铁插入原卟啉环中。我们发现了一种调节蛋白，称为Irr（铁反应调节剂），从细菌慢生根瘤菌，直接与亚铁螯合酶相互作用。这种新的机制允许Irr在血红素合成的位点局部地对血红素作出反应。Irr是一种条件稳定的蛋白质，在铁限制下发挥作用，但以血红素依赖性方式响应铁而降解。Irr最初被描述为血红素生物合成的调节因子，但新的证据表明它是铁依赖性基因的全球调节因子。因此，该提议的主要目标是解决假设B。日本血吸虫通过血红素的状态以Irr依赖的方式感受铁以调节铁稳态和代谢。此外，Irr同系物在大多数？变形菌，因此，拟议的工作作为一个模型，了解血红素和铁代谢的病原菌，实验上比B不易处理。山茱萸提出了三个具体目标。1.确定Irr识别靶基因并控制其表达的机制。Irr强烈调节编码三价铁转运蛋白的基因和许多其他铁相关基因，并且是正和负调节剂。我们特别感兴趣的激活Irr，因为细菌铁转运的阳性对照是罕见的。2.阐明Irr感知血红素状态的机制。Irr与亚铁螯合酶相互作用，后者为Irr提供调节输入。我们将描述这种相互作用的生化和遗传，并阐明随后的抑制和降解的Irr。3.阐明铁通过非Irr依赖性机制对血红素利用基因的调节。数据表明，血红素从头合成的控制不同于外源收购。我们将确定这种调节机制，并建立如何Irr依赖性和非依赖性代谢的整合。公共卫生相关性：虽然从宿主的角度来看，有益细菌和致病细菌对高等生物的入侵会导致非常不同的结果，但致病机理和共生关系在分子水平上可能非常相似。共生细菌慢生型大豆根瘤菌在实验上比相关病原体更易处理，因此是一个很好的模型系统。我们正在研究B中细菌血红素和铁代谢的新机制。japonicum的研究，以了解细菌感染和适应的分子策略。

DESCRIPTION (provided by applicant): Heme and the regulation of its biosynthesis are important for normal cell function, and contribute to the success of bacteria to diverse environments, including those associated with pathogenic and symbiotic interactions with higher eukaryotes. Heme is now known to be a regulatory molecule that allows cells to sense and adapt to environmental cues to maintain homeostasis. The broad objective of the proposed work is to understand how heme mediates the control of gene expression, with an emphasis on iron metabolism, and to reconcile this role for heme with its toxicity in cells. The heme biosynthetic pathway culminates with the insertion of iron into a protoporphyrin ring by a reaction catalyzed by ferrochelatase. We discovered a regulatory protein called Irr (iron response regulator) from the bacterium Bradyrhizobium japonicum that interacts directly with ferrochelatase. This novel mechanism allows Irr to respond to heme locally at the site of heme synthesis. Irr is a conditionally stable protein that functions under iron limitation, but degrades in response to iron in a heme-dependent manner. Irr was initially described as a regulator of heme biosynthesis, but new evidence reveals that it is a global regulator of iron-dependent genes. Thus, a major objective of this proposal is to address the hypothesis that B. japonicum senses iron through the status of heme in an Irr-dependent manner to regulate iron homeostasis and metabolism. Furthermore, Irr homologs are found in most ?-Proteobacteria, and thus the proposed work serves as a model for understanding heme and iron metabolism in pathogenic bacteria that are experimentally much less tractable than B. japonicum. Three specific aims are proposed. 1. Determine the mechanism by which Irr recognizes target genes and controls their expression. Irr strongly regulates genes encoding ferric iron transporters and many other iron-related genes, and is both a positive and negative regulator. We are particularly interested in activation by Irr since positive control of bacterial iron transport is uncommon. 2. Elucidate the mechanism by which Irr senses the status of heme. Irr interacts with ferrochelatase, which provides the regulatory input to Irr. We will characterize this interaction both biochemically and genetically, and elucidate the subsequent inhibition and degradation of Irr. 3. Elucidate the regulation of heme utilization genes by iron by an Irr-independent mechanism. Data suggest that heme synthesized de novo is controlled differently than that acquired exogenously. We will identify this regulatory mechanism, and establish how Irr-dependent and -independent metabolism is integrated. PUBLIC HEALTH RELEVANCE: Although invasion of higher organisms by beneficial and disease-causing bacteria results in very different outcomes from the host perspective, aspects of pathogenesis and symbiosis can be remarkably similar at the molecular level. The symbiotic bacterium Bradyrhizobium japonicum is experimentally more tractable than related pathogens, and is therefore a good model system. We are studying novel mechanisms of bacterial heme and iron metabolism in B. japonicum towards the end of understanding molecular strategies that bacteria employ for infection and adaptation.