title: Laser-ablation electrospray ionization mass spectrometry with ion mobility separation reveals metabolites in the symbiotic interactions of soybean roots and rhizobia.
abstract: Technologies enabling in situ metabolic profiling of living plant systems are invaluable for understanding physiological processes and could be used for rapid phenotypic screening (e.g., to produce plants with superior biological nitrogen-fixing ability). The symbiotic interaction between legumes and nitrogen-fixing soil bacteria results in a specialized plant organ (i.e., root nodule) where the exchange of nutrients between host and endosymbiont occurs. Laser-ablation electrospray ionization mass spectrometry (LAESI-MS) is a method that can be performed under ambient conditions requiring minimal sample preparation. Here, we employed LAESI-MS to explore the well characterized symbiosis between soybean (Glycine max L. Merr.) and its compatible symbiont, Bradyrhizobium japonicum. The utilization of ion mobility separation (IMS) improved the molecular coverage, selectivity, and identification of the detected biomolecules. Specifically, incorporation of IMS resulted in an increase of 153 differentially abundant spectral features in the nodule samples. The data presented demonstrate the advantages of using LAESI-IMS-MS for the rapid analysis of intact root nodules, uninfected root segments, and free-living rhizobia. Untargeted pathway analysis revealed several metabolic processes within the nodule (e.g., zeatin, riboflavin, and purine synthesis). Compounds specific to the uninfected root and bacteria were also detected. Lastly, we performed depth profiling of intact nodules to reveal the location of metabolites to the cortex and inside the infected region, and lateral profiling of sectioned nodules confirmed these molecular distributions. Our results established the feasibility of LAESI-IMS-MS for the analysis and spatial mapping of plant tissues, with its specific demonstration to improve our understanding of the soybean-rhizobial symbiosis.
authors.affiliation: Department of Chemistry, W. M. Keck Institute for Proteomics Technology and Applications, The George Washington University, Washington, DC, 20052, USA.
authors.forename: Sylwia A
authors.initials: SA
authors.lastname: Stopka
authors.affiliation: Divisions of Plant Sciences and Biochemistry, C. S. Bond Life Sciences Center, University of Missouri, Columbia, MO, 65211, USA.
authors.forename: Beverly J
authors.initials: BJ
authors.lastname: Agtuca
authors.affiliation: Environmental Molecular Sciences Laboratory, Earth and Biological Sciences Directorate, Pacific Northwest National Laboratory, 902 Battelle Boulevard, Richland, WA, 99354, USA.
authors.forename: David W
authors.initials: DW
authors.lastname: Koppenaal
authors.affiliation: Environmental Molecular Sciences Laboratory, Earth and Biological Sciences Directorate, Pacific Northwest National Laboratory, 902 Battelle Boulevard, Richland, WA, 99354, USA.
authors.forename: Ljiljana
authors.initials: L
authors.lastname: Paša-Tolić
authors.affiliation: Divisions of Plant Sciences and Biochemistry, C. S. Bond Life Sciences Center, University of Missouri, Columbia, MO, 65211, USA.
authors.forename: Gary
authors.initials: G
authors.lastname: Stacey
authors.affiliation: Department of Chemistry, W. M. Keck Institute for Proteomics Technology and Applications, The George Washington University, Washington, DC, 20052, USA.
authors.forename: Akos
authors.initials: A
authors.lastname: Vertes
authors.affiliation: Environmental Molecular Sciences Laboratory, Earth and Biological Sciences Directorate, Pacific Northwest National Laboratory, 902 Battelle Boulevard, Richland, WA, 99354, USA.
authors.forename: Christopher R
authors.identifier: http://orcid.org/0000-0002-6170-1033
authors.initials: CR
authors.lastname: Anderton
mesh_terms: D020369:Bradyrhizobium / Q000502:physiology*; D004867:Equipment Design; D007834:Lasers; D018517:Plant Roots / Q000378:metabolism / Q000382:microbiology*; D053204:Root Nodules, Plant / Q000382:microbiology; D013025:Glycine max / Q000378:metabolism* / Q000382:microbiology*; D021241:Spectrometry, Mass, Electrospray Ionization / Q000295:instrumentation / Q000379:methods; D013559:Symbiosis
keywords: Bradyrhizobium japonicum; Glycine max; LAESI; ion mobility separation; metabolites; nitrogen fixation; root nodules; technical advance