Matrix assisted laser desorption/ionization-mass spectrometry imaging (MALDI-MSI) for direct visualization of plant metabolites in situ
Graphical abstract
Introduction
Plant based agricultural products and chemical byproducts contribute hundreds of billions dollars annually to the global economy (www.faostat.fao.org), and originate from a diversity of grain, oilseed, fiber, and numerous other crops worldwide. Optimal production of chemicals derived from these crops requires an underlying understanding of the synthesis and metabolism of these compounds, especially in support of widespread metabolic engineering efforts to generate sustainable, bio-based products. An improved understanding of plant metabolism relies on analytical capabilities for the accurate identification and quantification of metabolites, which metabolomics has largely addressed through the development of sophisticated separation techniques, mass spectrometry approaches, and computational tools [1]. Analysis of chemical/tissue extracts by these methodologies provides excellent qualitative and quantitative information about chemical composition, but provides little or no data regarding the original spatial distribution of metabolites in situ. A number of years ago, the Caprioli group pioneered the development of mass spectrometry imaging for localizing molecules of interest directly on the sections of mammalian tissues [2, 3]. This methodology, termed matrix assisted laser desorption/ionization-mass spectrometry imaging (MALDI-MSI, also referred to as imaging mass spectrometry (IMS)), has since been expanded to visualize molecules directly in plant tissues and surfaces for the localization of lipids [4, 5•, 6, 7], proteins [8], secondary metabolites [9, 10, 11•, 12••, 13], and various small molecules [14••, 15, 16, 17] at unprecedented spatial and chemical resolution.
The last several years have seen increasing applications of MALDI-MSI to plant tissues which have been enabled, in part, by novel instruments and matrices to improve spatial resolution and chemical coverage [18•, 19••], streamlined matrix and sample preparation [20], and easily accessible open-source image processing free-ware [21, 22, 23]. These and other advances have enabled scientists to not only generate high resolution chemical images of plant tissues, but also to combine spatial metabolite data with further quantitative MS studies and gene expression analysis. In addition to MALDI-MSI there are other MSI platforms being utilized for MS imaging of plant tissues in situ, including desorption electrospray ionization (DESI-MS), laser ablation electrospray ionization (LAESI-MS) and secondary ion mass spectrometry (SIMS), and several salient reviews of these platforms in the context of plant MSI are available [6, 24, 25, 26]. Here we highlight the most recent applications of MALDI-MS imaging in plant tissues (Table 1) for metabolite localization in situ. Efforts examined here have also begun to address next-level biological questions in biochemical and metabolic contexts. It is anticipated that this new dimension of spatial analysis will provide important insights into the function, regulation and manipulation of plant metabolism.
Section snippets
MALDI-MSI: a procedural overview
A typical experimental workflow for MALDI-MSI is provided in Figure 1. First, plant tissues are flash-frozen (with or without fixation) in an embedding media, often gelatin, and then cryo-sectioned and lyophilized for tissue imaging; or depending on the location of the analytes of interest and the nature of the tissue, intact plant parts can be lyophilized and imaged directly for surface metabolites. A chemical matrix, to promote desorption and ionization, is applied by either a
Lipids
The biochemical characteristics of lipids lend themselves to analysis by MALDI-MSI and not surprisingly are one of the most common types of molecules to be spatially mapped using MSI. Phospholipids, comprising the lipid-bilayer of cell membranes, and triacylglycerols (TAGs), ∼30% mass of oil seeds, have been visualized by MSI. One of the first comprehensive MALDI-MSI lipidomics studies in plant sections examined the spatial distributions and composition of the major and minor storage and
Technological advances
Several recent and notable technological advances have expanded the current capabilities of modern MALDI MSI instrumentation. First, the modification of laser optics achieved the capability of <10 μm laser spot sizes. A sampling size of this dimension can not only be used to map chemical heterogeneity by tissue type, but also has the potential to map heterogeneity on cell-to-cell, or even organelle-to-organelle basis. The Spengler group achieved spatial resolution in the range of 3 μm using a
References and recommended reading
Papers of particular interest, published within the period of review, have been highlighted as:
• of special interest
•• of outstanding interest
Acknowledgements
Support for the authors research efforts in MSI is from Cotton Incorporated (Agreement #08-395) and U.S. Department of Energy, Office of Science, Basic Energy Sciences program (DE-FG02-14ER15647) to KDC. YL was supported by the US Department of Energy, Office of Basic Energy Sciences, Division of Chemical Sciences, Geosciences, and Biosciences. The Ames Laboratory is operated by Iowa State University under Contract DE-AC02-07CH11358. We thank Maria Dueñas for obtaining MS images of Arabidopsis
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