RESEARCH-PGR: Sieve Tube Proteomics - Unraveling the Physiology and Cell Biology of an Arcane Cell Type

人类消耗的食物是在植物中通过光合作用产生的，要么直接(想想土豆)，要么间接(吃植物的动物的肉)。光合作用发生在植物的绿色部分，但除了生菜等例外，饥饿的人类更感兴趣的是植物的非光合作用储存器官，包括谷物、块茎和水果。植物通过韧皮部组织中的微型管道网络，将光合作用的产物(化学上讲，糖)从绿色部分转移到储存器官，即所谓的筛管。由于它们的极端敏感性，活筛管的功能是出了名的难以研究，我们对筛管运输及其调节的了解仍然很差。这是不幸的，特别是因为许多吸食韧皮部的害虫，例如蚜虫，利用筛管中持续不断的富含能量的物质。更糟糕的往往是许多致病病毒和细菌搭乘这条溪流，从一个单一的入口点开始侵袭整个植物。由于这些因素每年造成大量作物损失，韧皮部在植物表现中的关键作用转化为几乎不能夸大的经济意义。该项目将对了解筛管作用的基本机理做出重要贡献，为针对害虫和病原体的治疗和预防措施的设计提供便利。可能涉及筛管生理的蛋白质将通过大规模生化方法(蛋白质组学)进行鉴定，然后通过对编码它们的基因进行定向修饰来进一步逐一分析。关于更广泛的影响，该项目将通过一个艺术展引起公众的兴趣，其中包括在普尔曼/莫斯科区域机场航站楼展示的韧皮部细胞的大幅面照片，并将继续更新“韧皮部”网页。此外，PI将与华盛顿州立大学多元文化学生服务办公室合作，招募少数族裔本科生到实验室工作。血管系统允许生物体通过大量流动在内部分配资源，从而克服扩散设置的大小限制。包括筛管在内的维管组织的发展推动了大型陆地植物(管状植物)的进化，从而大大提高了陆地生态系统的生产力。这种生产力支撑着地球上各种形式的生命。最大的光生自养海洋生物海藻汇聚地进化出运输筛管，突显了这种运输和通讯系统对大型光合作用和固着生命形式的重要性。本项目旨在阐明筛管的细胞生物学和生理学中尚未解决的问题。最先进的蛋白质组学方法将被应用于通过比较血管组织和分离的筛子元件的蛋白质组来识别潜在参与运输调节的蛋白质。候选蛋白质将通过其编码基因的修饰而被荧光标记，并将确定蛋白质在运输管道中的亚细胞定位。先进的生物成像工具将被应用于将筛管蛋白质分配到特定的细胞器或膜系统，产生筛管细胞器相互作用的3D模型，并表征蛋白质分布对压力的响应。所获得的洞察力预计将为培育更健壮和抗虫害的作物提供必要的信息。这一奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

The food consumed by humans is produced in plants by photosynthesis, either directly (think of potatoes) or indirectly (meat from plant-eating animals). Photosynthesis occurs in the green parts of plants, but, with exceptions such as lettuce, hungry humans are more interested in non-photosynthesizing storage organs of plants including grain, tubers, and fruits. Plants transfer the products of photosynthesis (sugars, chemically speaking) from their green parts to storage organs via a network of microscopic pipes, the so-called sieve tubes, in the phloem tissue. Due to their extreme sensitivity, the function of live sieve tubes is notoriously hard to study, and our understanding of sieve tube transport and its regulation has remained lamentably poor. This is unfortunate, especially since numerous phloem-sucking pests, for instance aphids, exploit the continuous stream of energy-rich substances in the sieve tubes. What often is worse are the many pathogenic viruses and bacteria that hitch rides in this stream to infest the entire plant, starting from a single point of entry. As these factors cause massive crop losses every year, the critical function of phloem in plant performance translates into an economic significance that can hardly be overstated. This project will make important contributions to comprehension of fundamental mechanisms of sieve tube function, which will facilitate the design of treatments and preventive measures targeting pests and pathogens. Proteins potentially involved in sieve tube physiology will be identified by a large-scale biochemical approach (‘proteomics’), and then further analyzed one by one through directed modification of the genes that encode them. With regard to Broader Impacts, the project will engage public interest with an artistic exhibition that incorporates large format photographs of phloem cells displayed in the terminal of the Pullman/Moscow regional airport and will also continue updating a “Phloem” webpage. In addition the PI will work with the Office of Multicultural Student Services at Washington State University to recruit minority undergraduate students to work in the lab.Vascular systems allow organisms to distribute resources internally by bulk flow, thus overcoming size limitations set by diffusion. The development of vascular tissues including sieve tubes drove the evolution of large land plants (tracheophytes) which caused a major increase in the productivity of terrestrial ecosystems. This productivity supports life on earth in its various forms. The largest photoautotroph marine organisms, kelps, have convergently evolved transporting sieve tubes, highlighting the significance of this transport and communication system for large photosynthetic and sessile life forms. This project aims at elucidating open questions in the cell biology and physiology of sieve tubes. State-of-the-art proteomics approaches will be applied to identify proteins potentially involved in transport regulation, by comparing the proteomes of vascular tissue and isolated sieve elements. Candidate proteins will be fluorescently tagged by modifications of their encoding genes, and the subcellular localization of the proteins in transporting tubes will be determined. Advanced bioimaging tools will be applied to assign sieve tube proteins to specific organelles or membrane systems, to generate 3D models of sieve tube organelle interactions, and to characterize the responses of protein distribution to stress. The insights gained are expected to provide essential information for the generation of more robust and pest-resistant crops.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.