MRI: Acquisition of Instrumentation for Spatially Relevant Plant Metabolomics and Proteomics

该项目利用资金为华盛顿州立大学的M.J.默多克代谢组学实验室(MML，http://www.murdockmetabolomics.wsu.edu，，由三个PI共同指导)增加了一个集成的基于质谱学的组织成像系统，包括一个组织切片站和新一代基于MALDI的四极杆飞行时间质谱仪(Q-TOFMS)系统，该系统结合了集成的离子迁移率光谱。将这种集成的仪器系统添加到MML中，使IBC和华盛顿州立大学园区的研究人员能够作为一个整体获得组织成像数据，不仅包括基于蛋白质和代谢物的高质量和高分辨率MS/MS鉴定，而且还包括此类化合物的离子迁移率光谱表征，增加了化合物鉴定和定量的信心，同时降低了数据采集中的噪音。过去研究中缺少的一个关键环节是在细胞或亚组织水平上识别生化过程发生在哪里或如何调节途径的能力，特别是在植物的茎、根、根茎和叶等复杂器官中。这一综合仪器的获得使几十年来由于先前分析方法的空间分辨率不足而一直没有得到回答的基本机械问题得以得到解决。集成的组织成像系统将被安置在MML内一个新装修的实验室中，旨在容纳这种仪器，并扩大MML目前在代谢组学和蛋白质组学方面的优势，推动MML成为世界一流的植物代谢组学设施的目标。植物产生各种各样的化合物和物质，用于授粉、植物防御、个体植物内部以及植物和其他生物之间的交流，以及许多其他功能。许多这些高度专门化的物质，如香料、香料、口香糖、树脂、毒素、结构成分以及营养和药用化合物，都是在极少数的细胞类型中产生的，甚至在植物的单个细胞中产生。长期以来，在植物组织的不同单细胞类型中检测和测量各种代谢物和产生这些代谢物的蛋白质的能力几乎是不可能的，尽管从长远来看，这种能力对于全面了解植物新陈代谢是至关重要的。人们普遍认为，这些知识对于今后改进植物以供人类使用的努力至关重要。在植物个体细胞水平上对这些过程的调控知之甚少，很大程度上是由于无法检测和测量特定植物细胞中的代谢物。新的组织成像技术允许检测和测量复杂生物组织中特定细胞中的代谢物和蛋白质，如茎、根、根茎、花和叶，以及植物中的其他特殊细胞类型，或其他生物(真菌、动物、昆虫等)中的其他组织。因此，这项新技术使研究人员能够测试与药用植物新陈代谢、生物燃料生产、植物-昆虫相互作用和基础植物生物化学等领域有关的重要和基本生物学过程的假说。利用该仪器系统获得的数据将成为植物生物化学、化学、跨学科组学和代谢工程领域的研究生、本科生和博士后研究员研究项目的一部分。此外，新的仪器系统将被整合到两个具体的努力中，以吸引当前和未来的学生特别是植物生物化学和新陈代谢组学领域的学生以及一般的科学领域的学生，并为西苏里大学的学生提供教育优势。首先，植物组织成像将被纳入由科比·兰格教授的高级植物生物化学课程。其次，Pi Gang将参加Copi Weller的“提升数学和科学管道：从小学到大学”项目，该项目的重点是科尔维尔保留地(太平洋西北部面积最大)以及华盛顿州奥马克周围非部落地区的原住民和拉美裔美国人，奥马克为高三学生提供暑期研究实习机会。

This project uses funds to add an integrated mass spectrometry-based tissue imaging system, including a tissue sectioning station and a new generation MALDI-based quadrupole-time of flight mass spectrometry (Q-TOFMS) system that incorporates integrated ion mobility spectrometry, to the M.J. Murdock Metabolomics Laboratory (MML, http://www.murdockmetabolomics.wsu.edu, co-directed by three of the PIs) at Washington State University. Addition of this integrated instrumentation system to the MML enables researchers in the IBC and the WSU campus as a whole to acquire tissue imaging data that incorporates not only high mass accuracy and high resolution MS and MS/MS-based identification of proteins and metabolites, but also ion mobility spectrometric characterization of such compounds, increasing confidence in compound identification and quantification, while reducing noise in data acquisition. A critical missing link in past research has been the ability to identify, at the cellular or sub-tissue level, where biochemical processes occur or how pathways are regulated, especially in complex organs such as plant stems, roots, rhizomes and leaves. Acquisition of this integrated instrumentation enables fundamental mechanistic questions to be addressed that have remained unanswered for decades due to inadequate spatial resolution of previous analytical approaches. The integrated tissue imaging system will be housed in a newly renovated lab within the MML designed to accommodate such instrumentation, and expands the MML's current strengths in metabolomics and proteomics, furthering the goal of developing the MML as a world premier plant metabolomics facility.Plants produce a fantastic diversity of compounds and substances that are used in pollination, plant defense, communication within individual plants and between plants and other organisms, among many other functions. Many of these highly specialized substances, such as flavors, fragrances, gums, resins, toxins, structural components, and nutritional and medicinal compounds, are produced in very few cell types or even in individual cells in plants. The ability to detect and measure various metabolites and the proteins that produce them in distinct single cell types in plant tissues has long been near impossible, even though this ability is vital in the long run to obtain a complete understanding of plant metabolism. It is generally considered that such knowledge is critical to future efforts to improve plants for human use. Regulation of these processes at the individual cellular level in plants is poorly understood, largely as a result of inability to detect and measure metabolites in specific plant cells. The new tissue imaging technology allows for detection and measurement of metabolites and proteins in specific cells in complex biological tissues such as stems, roots, rhizomes, flowers and leaves, and other specialized cell types in plants, or other tissues in other organisms (fungi, animals, insects, etc.). This new technology thus enables researchers to test hypotheses about important and fundamental biological processes related to medicinal plant metabolism, biofuel production, plant-insect interactions and basic plant biochemistry, among other areas. Data acquired with this instrument system will be part of the research projects of graduate students, undergraduate students and post-doctoral fellows in the fields of plant biochemistry, chemistry, interdisciplinary omics, and metabolic engineering. In addition, the new instrument system will be integrated into two specific efforts to attract current and future students to the fields of plant biochemistry and metabolomics particularly and to science in general and to provide educational advantages to the students at WSU. First, plant tissue imaging will be incorporated into the Advanced Plant Biochemistry course taught by CoPI Lange. Second, PI Gang will participate in CoPI Weller's "Pumping-Up the Math and Science Pipeline: Grade School to College" program that focuses on Native American and Hispanic populations on the Colville Reservation (largest in the Pacific Northwest) and in non-tribal regions surrounding Omak, WA, which hosts high school seniors for summer research internships.