Spatial Mapping and Profiling of Metabolite Distributions during Germination

Abstract

Germination is a highly complex process by which seeds begin to develop and establish themselves as viable organisms. In this study, we utilize a combination of gas chromatography-mass spectrometry, liquid chromatography-fluorescence, and mass spectrometry imaging approaches to profile and visualize the metabolic distributions of germinating seeds from two different inbreds of maize (Zea mays) seeds, B73 and Mo17. Gas chromatography and liquid chromatography analyses demonstrate that the two inbreds are highly differentiated in their metabolite profiles throughout the course of germination, especially with regard to amino acids, sugar alcohols, and small organic acids. Crude dissection of the seed followed by gas chromatography-mass spectrometry analysis of polar metabolites also revealed that many compounds were highly sequestered among the various seed tissue types. To further localize compounds, matrix-assisted laser desorption/ionization mass spectrometry imaging was utilized to visualize compounds in fine detail in their native environments over the course of germination. Most notably, the fatty acyl chain-dependent differential localization of phospholipids and triacylglycerols was observed within the embryo and radicle, showing correlation with the heterogeneous distribution of fatty acids. Other interesting observations include unusual localization of ceramides on the endosperm/scutellum boundary and subcellular localization of ferulate in the aleurone.Plants use seeds as the propagule to ensure reproduction to the next generation, and over the past 10,000 years, human civilizations have established agricultural practices to ensure a seed-based food supply (Larson et al., 2014). Therefore, deciphering the processes that enable seeds to perform their biological functions is of importance in understanding how plants are propagated and also is of practical importance to improve agriculture. Seeds are designed to survive long periods of dormancy in a relatively dry state, and the process of germination is initiated by the imbibition of water. During this germination process, many metabolic changes occur, most of which are associated with the catabolism of seed storage products (proteins, polysaccharides, and lipids) into metabolically usable, simpler chemical forms that are either used as precursors to assemble the growing seedling or are oxidized via energy-producing biochemical pathways to thermodynamically support growth (Bewley, 1997(Bewley, , 2001.The specific metabolic processes that support seed germination are somewhat dependent on the taxonomic clade of the plant, which determines seed tissue/ organ organization and the nature of the seed storage compounds. Specifically, the seeds of the Poaceae family of monocots are characterized by a starch-and protein-filled endosperm, and their catabolism by digestive enzymes produced in the outermost layer of the endosperm (i.e. the aleurone) provides the carbonbased and nitrogenous precursors for the growth of the embryo, which is composed of the embryonic ax...

Metabolic and gene expression hallmarks of seed germination uncovered by sodium butyrate in Medicago truncatula

“…truncatula (Balestrazzi et al, ), metabolomics provides the opportunity to identify novel markers that might speed up the early prediction of seed vigour. Metabolomics of seed germination is expanding (Das, Kim, Khadka, Rakwal, & Rohifa, ; Feenstra et al, ; Gorzolka, Kolling, Nattkemper, & Niehaus, ); however, the metabolome of the seed response to HDAC inhibitors is still untargeted. The aim of this work was to investigate novel signatures of the seed response in M .…”

“…Whereas basic knowledge on the expression profiles of DDR genes during early germination has already proved to be a useful source of seed quality indicators in M. truncatula , metabolomics provides the opportunity to identify novel markers that might speed up the early prediction of seed vigour. Metabolomics of seed germination is expanding (Das, Kim, Khadka, Rakwal, & Rohifa, 2017;Feenstra et al, 2017;Gorzolka, Kolling, Nattkemper, & Niehaus, 2016); however, the metabolome of the seed response to HDAC inhibitors is still untargeted. The aim of this work was to investigate novel signatures of the seed response in M. truncatula, using the HDAC inhibitor sodium butyrate (NaB) as a stress agent, by conducting multiple molecular phenotyping analyses ranging from gene expression to metabolite profiling.…”

Interrogation of spatial metabolome of Ginkgo biloba with high‐resolution matrix‐assisted laser desorption/ionization and laser desorption/ionization mass spectrometry imaging

“…MS imaging (MSI) has become an attractive molecular imaging tool for capturing the spatial distribution of plant metabolites (Boughton, Thinagaran, Sarabia, Bacic, & Roessner, ; Feenstra et al, ; Kaspar, Peukert, Svatos, Matros, & Mock, ; Lee, Perdian, Song, Yeung, & Nikolau, ; Li, Bohn, & Sweedler, ). It is label‐free, untargeted, multiplexed, and capable of de novo chemical discovery.…”

‘In plaque-mass spectrometry imaging’ reveals a major metabolic shift towards odd-chain fatty acid lipids induced by host-virus interactions

“…Recent advances in mass spectrometry imaging (MSI) technologies allow to localize specific metabolites of various dynamic processes and biotic interactions at the microscale level, and consequently to monitor metabolic changes in high spatiotemporal resolution 16,17 . MSI-based approaches were used to study seed germination 18 , metabolic footprints of host-associated bacteria 19,20 , embryonic development of zebrafish 21 , bacterial population dynamics [22][23][24][25] , to detect human pathogens 26 and to visualize the spatial distribution of anti-viral drugs 27 . An attractive host-virus model system is the cosmopolitan alga Emiliania huxleyi and its specific virus, E. huxleyi virus (EhV), which plays a key role in regulating the fate of carbon and sulfur cycles in contemporary oceans 28,29 .…”