EAGER: Iron sensing and signaling in Arabidopsis thaliana

EAGER：拟南芥中的铁感应和信号传导

• 批准号: 1441450
• 负责人: Elsbeth Walker
• 金额: $ 21.75万
• 依托单位: University of Massachusetts Amherst
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-06-15 至 2018-05-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1441450&HistoricalAwards=false
• 关键词: EAGER; Iron; sensing; signaling; Arabidopsis

Living organisms keep the level of the highly reactive metal, iron, carefully controlled to avoid cellular damage. In plants, part of this control is a mechanism to allow the shoot, where iron is most heavily needed, to signal iron status to roots where primary iron uptake occurs. The molecular details of this shoot to root signaling mechanism are unknown, but are likely to be essential for successful engineering of plants to accumulate additional bioavailable iron in edible parts. This project explores the hypothesis that Yellow Stripe1-Like proteins are responsible for signaling the iron status between the shoot and the root of plants. Our current state of ignorance about many of the mechanisms involved in plant iron homeostasis is a major obstacle to devising approaches for biofortification of staple foods with iron. Biofortification refers to the genetic engineering of staple crops to accumulate additional bioavailable iron in edible parts; it is widely regarded as a sustainable means of improving the iron nutrition of the 2-3 billion people worldwide whose inadequate diet causes iron deficiency anemia. Improving our understanding of plant iron homeostatic mechanisms is also critical if we wish to improve growth of crops in marginal soils, where iron deficiency frequently limits crop growth.A double mutant with null mutations in the Arabidopsis Yellow Stripe-Like1 and Yellow Stripe-Like3 (ysl1ysl3) has severe pleiotropic growth defects related to iron. YSLs are transporters of iron and other transition metals (Zn, Cu, Mn, Ni) complexed with the plant-specific metal chelating compound nicotianamine (NA). It is important to note that both tissue localization (mainly in the leaf veins) and regulation (down-regulated by iron deficiency) argue against the idea that these proteins participate in primary metal uptake from the soil. The phenotype of ysl1ysl3 plants, which includes iron deficiency chlorosis, low Fe levels in tissues, failure to re-translocate Cu and Zn from leaves during senescence, pollen failure, and incomplete embryo development, were at first all thought to be consequences of failed metal-NA transport by AtYSL1 and AtYSL3. But ysl1ysl3 plants also exhibit defects in Fe signaling, which are difficult to explain as a simple consequence of metal transport defects in leaf veins. New evidence has indicated that ysl1ysl3 plants have a pervasive inability to respond vigorously to iron deficiency, and a new hypothesis is put forward that AtYSL1 and AtYSL3 function as "transceptors": transporters that also function as receptors for the iron status of the plant. In this project, we will investigate the hypothesis that, in addition to their roles as metal-NA transporters, AtYSL1 and AtYSL3 function in signaling the iron status of shoots. A key test of this idea is to make mutations in YSL1 and YSL3 that affect receptor function without affecting transporter function, and vice versa. In this project, transport defective versions of YSL1 and YSL3 will be made using site directed mutagenesis and will be tested for functionality in yeast. Three properly localized transport defective versions will then be introduced into the ysl1ysl3 double mutant under their own promoters, and the resulting plants will be tested for ysl1ysl3 phenotypes. In a second, more limited experiment, the acidic N-terminal domain(s) of YSL1 and YSL3 will be tested. These domains may be iron binding, and thus could be involved in iron sensing. We will test whether removing these domains affects transport function in yeast, and if it does not, the proteins will be tested in planta for complementation of ysl1ysl3 phenotypes. These experiments will elucidate whether YSL1 and YSL3 are directly involved in iron signaling. This project will be integrated into a laboratory course for undergraduates and provide training in bioinformatics and genomics methodologies, as well as the process of hypothesis-driven research.

生物体保持高度反应性的金属铁的水平，仔细控制以避免细胞损伤。在植物中，这种控制的一部分是一种机制，允许最需要铁的芽向发生初级铁吸收的根发出铁状态的信号。这种茎到根信号传导机制的分子细节尚不清楚，但可能是成功地工程化植物在可食用部分积累额外的生物可利用铁所必需的。该项目探讨了黄条1样蛋白负责在植物的茎和根之间传递铁状态的假设。我们目前对植物铁稳态所涉及的许多机制的无知是设计铁主食生物强化方法的主要障碍。生物强化是指对主要作物进行基因工程，以在可食用部分积累额外的生物可利用铁;它被广泛认为是改善全球20亿至30亿人铁营养的可持续手段，这些人的饮食不足导致缺铁性贫血。提高我们对植物铁稳态机制的理解也是至关重要的，如果我们希望改善作物在边缘土壤中的生长，缺铁经常限制作物的生长。拟南芥黄条样1和黄条样3（ysl 1 ysl 3）的无效突变的双突变体具有严重的多效性生长缺陷与铁有关。YSL是铁和其他过渡金属（Zn、Cu、Mn、Ni）与植物特异性金属螯合化合物烟酰胺（NA）络合的转运蛋白。值得注意的是，组织定位（主要在叶脉中）和调节（缺铁下调）都反对这些蛋白质参与土壤中初级金属吸收的观点。ysl 1 ysl 3植物的表型，其中包括缺铁失绿，组织中的低铁水平，未能重新转运铜和锌从叶片衰老过程中，花粉失败，胚胎发育不完全，首先被认为是失败的金属-NA运输AtYSL 1和AtYSL 3的后果。但ysl 1 ysl 3植物也表现出缺陷的Fe信号，这是很难解释为一个简单的结果，金属运输缺陷的叶脉。新的证据表明，ysl 1 ysl 3植物普遍不能积极响应缺铁，并提出了一个新的假设，AtYSL 1和AtYSL 3的功能作为“transeptor”：转运蛋白，也作为受体的铁状态的植物。在这个项目中，我们将调查的假设，除了作为金属-NA转运蛋白的作用，AtYSL 1和AtYSL 3的功能，在信号的铁状态的芽。这个想法的一个关键测试是在YSL 1和YSL 3中进行突变，影响受体功能而不影响转运蛋白功能，反之亦然。 在本项目中，将使用定点诱变制备YSL 1和YSL 3的转运缺陷型，并将在酵母中检测其功能。 然后将三个适当定位的转运缺陷型在它们自己的启动子下引入ysl 1 ysl 3双突变体中，并测试所得植物的ysl 1 ysl 3表型。在第二个更有限的实验中，将测试YSL 1和YSL 3的酸性N-末端结构域。 这些结构域可能是铁结合的，因此可能参与铁传感。 我们将测试去除这些结构域是否会影响酵母的运输功能，如果不会，将在植物中测试这些蛋白质的ysl 1 ysl 3表型互补。 这些实验将阐明YSL 1和YSL 3是否直接参与铁信号传导。该项目将纳入本科生实验室课程，并提供生物信息学和基因组学方法以及假设驱动研究过程方面的培训。