Systematic Genome-Wide Characterization of Iron Homeostasis

 描述（由申请人提供）：作为数千种酶的辅因子，铁是一种必需的微量营养素。然而，游离铁是有毒的，因为它催化破坏性活性氧的快速形成。因此，体内平衡系统对所有生物体中的铁水平施加严格控制，并且基因表达响应于铁缺乏和铁丰度而调整：已知大肠杆菌中至少100个基因的表达是铁依赖的。在人类中，铁稳态的破坏会导致严重的疾病：大脑中的铁积累与神经退行性疾病有关，铁过载会导致肝脏疾病血色素沉着症，缺铁会导致贫血和认知发育受损。此外，入侵的细菌病原体从人类蛋白质中劫持铁以建立感染。这些考虑强调了铁稳态对人类健康的重要作用。然而，由于缺乏研究来检查对铁缺乏和铁过载的反应的总细胞框架，铁稳态如何整合到细胞生理学中仍然不清楚。在这里，为了解决这种缺陷，使用了一种双管齐下的全基因组方法，包括：（1）核糖体分析，以确定基因的表达如何随时间变化，以响应不同的铁水平;和（2）全基因组筛选，以确定在铁过载和铁缺乏下对生存重要的基因。这种方法辅之以生化和遗传研究，以机械地表征铁稳态中的新参与者。重要的是，将研究两个模型系统，革兰氏阴性细菌大肠杆菌和人类细胞，允许两个系统的信息比较，这对于开发专门针对细菌铁稳态的抗生素至关重要。这项全面的研究将提供对铁的反应的时间分辨和整体观点，并确定铁稳态的新参与者，包括转录后调节因子，从不同来源吸收铁的运输系统，以及响应铁而激活的代谢途径。从这些实验中获得的见解将有助于理解铁稳态疾病和细菌感染的治疗。

 DESCRIPTION (provided by applicant): As a cofactor for thousands of enzymes, iron is an essential micronutrient. Yet, free iron is toxic because it catalyzes rapid formation of damaging reactive oxygen species. Therefore, homeostasis systems exert tight control on iron levels in all organisms and gene expression is adjusted in response to iron deprivation and iron abundance: expression of at least 100 genes is known to be iron-dependent in Escherichia coli. In humans, disruption of iron homeostasis contributes to severe diseases: iron accumulation in the brain is linked to neurodegenerative diseases, iron overload causes the liver disease hemochromatosis, and iron deficiency leads to anemia and impaired cognitive development. Furthermore, invading bacterial pathogens hijack iron out of human proteins to establish infections. These considerations underscore the essential role of iron homeostasis to human health. Because of a lack of studies examining the total cellular framework for the response to both iron deficiency and iron overload, however, how iron homeostasis is integrated into cell physiology is still unclear. Here, to address this deficiency, a two-pronged genome-wide approach is used consisting of: (1) ribosome profiling to determine how expression of genes changes over time in response to varying iron levels; and (2) genome-wide screens to identify genes that are important for survival under iron overload and iron deficiency. This approach is complemented by biochemical and genetic studies to mechanistically characterize new players in iron homeostasis. Importantly, two model systems will be investigated, the gram-negative bacterium Escherichia coli and human cells, allowing for an informative comparison of the two systems that will be essential to develop antibiotics specifically targeted at bacterial iron homeostasis. This comprehensive study will provide a time-resolved and holistic view of the response to iron and identify new players in iron homeostasis, including post-transcriptional regulators, transport systems to take up iron from different sources, and metabolic pathways that become activated in response to iron. Insight obtained from these experiments will benefit understanding of iron homeostasis diseases and treatment of bacterial infections.