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  1. Abstract

    Mass spectrometry imaging (MSI) enables simultaneous spatial mapping for diverse molecules in biological tissues. Matrix‐assisted laser desorption ionization (MALDI) mass spectrometry (MS) has been a mainstream MSI method for a wide range of biomolecules. However, MALDI‐MSI of biological homopolymers used for energy storage and molecular feedstock is limited by, e.g., preferential ionization for certain molecular classes. Matrix‐free nanophotonic ionization from silicon nanopost arrays (NAPAs) is an emerging laser desorption ionization (LDI) platform with ultra‐trace sensitivity and molecular imaging capabilities. Here, we show complementary analysis and MSI of polyhydroxybutyric acid (PHB), polyglutamic acid (PGA), and polysaccharide oligomers in soybean root nodule sections by NAPA‐LDI and MALDI. For PHB, number and weight average molar mass, polydispersity, and oligomer size distributions across the tissue section and in regions of interest were characterized by NAPA‐LDI‐MSI.

     
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  2. Abstract

    Mass spectrometry imaging (MSI) has become an important analytical tool for the label‐free chemical imaging of diverse molecules in biological specimens. This minireview surveys some emerging methods in the context of factors that can lead to inaccurate information in MSI, chemical and spatial aberrations, along with their common sources. Matrix‐assisted laser desorption ionization, based on organic matrices, has become the most widely used MSI technique for biomolecules. However, due to inherent limitations associated with the use of organic matrices, for example, heterogeneous matrix‐analyte cocrystallization, and spectral interferences due to the matrix, laser desorption ionization (LDI) from inorganic and nanophotonic platforms has emerged as an alternative MSI modality with complementary advantages. In this review, inorganic and nanophotonic platforms for LDI‐MSI, their applications in imaging, notable merits, and limitations are described.

     
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  3. SUMMARY

    The establishment of the nitrogen‐fixing symbiosis between soybean andBradyrhizobium japonicumis a complex process. To document the changes in plant metabolism as a result of symbiosis, we utilized laser ablation electrospray ionization‐mass spectrometry (LAESI‐MS) forin situmetabolic profiling of wild‐type nodules, nodules infected with aB. japonicum nifHmutant unable to fix nitrogen, nodules doubly infected by both strains, and nodules formed on plants mutated in thestearoyl‐acyl carrier protein desaturase(sacpd‐c) gene, which were previously shown to have an altered nodule ultrastructure. The results showed that the relative abundance of fatty acids, purines, and lipids was significantly changed in response to the symbiosis. ThenifHmutant nodules had elevated levels of jasmonic acid, correlating with signs of nitrogen deprivation. Nodules resulting from the mixed inoculant displayed similar, overlapping metabolic distributions within the sectors of effective (fix+) and ineffective (nifHmutant, fix) endosymbionts. These data are inconsistent with the notion that plant sanctioning is cell autonomous. Nodules lackingsacpd‐cdisplayed an elevation of soyasaponins and organic acids in the central necrotic regions. The present study demonstrates the utility of LAESI‐MS for high‐throughput screening of plant phenotypes. Overall, nodules disrupted in the symbiosis were elevated in metabolites related to plant defense.

     
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  4. Abstract

    In this study, the three-dimensional spatial distributions of a number of metabolites involved in regulating symbiosis and biological nitrogen fixation (BNF) within soybean root nodules were revealed using matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI). While many metabolites exhibited distinct spatial compartmentalization, some metabolites were asymmetrically distributed throughout the nodule (e.g., S-adenosylmethionine). These results establish a more complex metabolic view of plant–bacteria symbiosis (and BNF) within soybean nodules than previously hypothesized. Collectively these findings suggest that spatial perspectives in metabolic regulation should be considered to unravel the overall complexity of interacting organisms, like those relating to associations of nitrogen-fixing bacteria with host plants.

     
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  5. Abstract

    The Soybean Gene Atlas project provides a comprehensive map for understanding gene expression patterns in major soybean tissues from flower, root, leaf, nodule, seed, and shoot and stem. The RNA‐Seq data generated in the project serve as a valuable resource for discovering tissue‐specific transcriptome behavior of soybean genes in different tissues. We developed a computational pipeline for Soybean context‐specific network (SoyCSN) inference with a suite of prediction tools to analyze, annotate, retrieve, and visualize soybean context‐specific networks at both transcriptome and interactome levels. BicMix and Cross‐Conditions Cluster Detection algorithms were applied to detect modules based on co‐expression relationships across all the tissues. Soybean context‐specific interactomes were predicted by combining soybean tissue gene expression and protein–protein interaction data. Functional analyses of these predicted networks provide insights into soybean tissue specificities. For example, under symbiotic, nitrogen‐fixing conditions, the constructed soybean leaf network highlights the connection between the photosynthesis function and rhizobium–legume symbiosis. SoyCSN data and all its results are publicly available via an interactive web service within the Soybean Knowledge Base (SoyKB) athttp://soykb.org/SoyCSN. SoyCSN provides a useful web‐based access for exploring context specificities systematically in gene regulatory mechanisms and gene relationships for soybean researchers and molecular breeders.

     
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  6. Nodule organogenesis in legumes is regulated temporally and spatially through gene networks. Genome-wide transcriptome, proteomic, and metabolomic analyses have been used previously to define the functional role of various plant genes in the nodulation process. However, while significant progress has been made, most of these studies have suffered from tissue dilution since only a few cells/root regions respond to rhizobial infection, with much of the root non-responsive. To partially overcome this issue, we adopted translating ribosome affinity purification (TRAP) to specifically monitor the response of the root cortex to rhizobial inoculation using a cortex-specific promoter. While previous studies have largely focused on the plant response within the root epidermis (e.g., root hairs) or within developing nodules, much less is known about the early responses within the root cortex, such as in relation to the development of the nodule primordium or growth of the infection thread. We focused on identifying genes specifically regulated during early nodule organogenesis using roots inoculated with Bradyrhizobium japonicum . A number of novel nodulation gene candidates were discovered, as well as soybean orthologs of nodulation genes previously reported in other legumes. The differential cortex expression of several genes was confirmed using a promoter-GUS analysis, and RNAi was used to investigate gene function. Notably, a number of differentially regulated genes involved in phytohormone signaling, including auxin, cytokinin, and gibberellic acid (GA), were also discovered, providing deep insight into phytohormone signaling during early nodule development. 
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  7. null (Ed.)
    Laser ablation electrospray ionization (LAESI) driven by mid-infrared laser pulses allows the direct analysis of biological tissues with minimal sample preparation. Dedicated remote ablation chambers have been developed to eliminate the need for close proximity between the sample and the mass spectrometer inlet. This also allows for the analysis of large or irregularly shaped objects, and incorporation of additional optics for microscopic imaging. Here we report on the characterization of a newly designed conical inner volume ablation chamber working in transmission geometry, where a reduced zone of stagnation was achieved by tapering the sample platform and the chamber outlet. As a result, the transmission efficiency of both large (>7.5 μm) and smaller particulates (<6.5 μm) has increased significantly. Improved analytical figures of merit, including 300 fmol limit of detection, and three orders of magnitude in dynamic range, were established. Particle residence time, measured by the FWHM of the analyte signal, was reduced from 2.0 s to 0.5 s enabling higher ablation rates and shorter analysis time. A total of six glucosinolates (sinigrin, gluconapin, progoitrin, glucoiberin, glucoraphanin, and glucohirsutin) were detected in plant samples with ion abundances higher by a factor of 2 to 8 for the redesigned ablation chamber. 
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