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      Foliar Application of Different Vegetal-Derived Protein Hydrolysates Distinctively Modulates Tomato Root Development and Metabolism

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          Abstract

          Despite the scientific evidence supporting their biostimulant activity, the molecular mechanism(s) underlying the activity of protein hydrolysates (PHs) and the specificity among different products are still poorly explored. This work tested five different protein hydrolysates, produced from different plant sources using the same enzymatic approach, for their ability to promote rooting in tomato cuttings following quick dipping. Provided that all the different PHs increased root length (45–93%) and some of them increased root number (37–56%), untargeted metabolomics followed by multivariate statistics and pathway analysis were used to unravel the molecular processes at the basis of the biostimulant activity. Distinct metabolomic signatures could be found in roots following the PHs treatments. In general, PHs shaped the phytohormone profile, modulating the complex interaction between cytokinins and auxins, an interplay playing a pivotal role in root development, and triggered a down accumulation of brassinosteroids. Concerning secondary metabolism, PHs induced the accumulation of aliphatic glucosinolates, alkaloids, and phenylpropanoids, potentially eliciting crop resilience to stress conditions. Here, we confirm that PHs may have a hormone-like activity, and that their application can modulate plant growth, likely interfering with signaling processes. Noteworthy, the heterogenicity of the botanical origin supported the distinctive and peculiar metabolomic responses we observed across the products tested. While supporting their biostimulant activity, these findings suggest that a generalized crop response to PHs cannot be defined and that specific effects are rather to be investigated.

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          MS-DIAL: Data Independent MS/MS Deconvolution for Comprehensive Metabolome Analysis

          Data-independent acquisition (DIA) in liquid chromatography tandem mass spectrometry (LC-MS/MS) provides more comprehensive untargeted acquisition of molecular data. Here we provide an open-source software pipeline, MS-DIAL, to demonstrate how DIA improves simultaneous identification and quantification of small molecules by mass spectral deconvolution. For reversed phase LC-MS/MS, our program with an enriched LipidBlast library identified total 1,023 lipid compounds from nine algal strains to highlight their chemotaxonomic relationships.
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            Cytokinin-deficient transgenic Arabidopsis plants show multiple developmental alterations indicating opposite functions of cytokinins in the regulation of shoot and root meristem activity.

            Cytokinins are hormones that regulate cell division and development. As a result of a lack of specific mutants and biochemical tools, it has not been possible to study the consequences of cytokinin deficiency. Cytokinin-deficient plants are expected to yield information about processes in which cytokinins are limiting and that, therefore, they might regulate. We have engineered transgenic Arabidopsis plants that overexpress individually six different members of the cytokinin oxidase/dehydrogenase (AtCKX) gene family and have undertaken a detailed phenotypic analysis. Transgenic plants had increased cytokinin breakdown (30 to 45% of wild-type cytokinin content) and reduced expression of the cytokinin reporter gene ARR5:GUS (beta-glucuronidase). Cytokinin deficiency resulted in diminished activity of the vegetative and floral shoot apical meristems and leaf primordia, indicating an absolute requirement for the hormone. By contrast, cytokinins are negative regulators of root growth and lateral root formation. We show that the increased growth of the primary root is linked to an enhanced meristematic cell number, suggesting that cytokinins control the exit of cells from the root meristem. Different AtCKX-green fluorescent protein fusion proteins were localized to the vacuoles or the endoplasmic reticulum and possibly to the extracellular space, indicating that subcellular compartmentation plays an important role in cytokinin biology. Analyses of promoter:GUS fusion genes showed differential expression of AtCKX genes during plant development, the activity being confined predominantly to zones of active growth. Our results are consistent with the hypothesis that cytokinins have central, but opposite, regulatory functions in root and shoot meristems and indicate that a fine-tuned control of catabolism plays an important role in ensuring the proper regulation of cytokinin functions.
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              Hydrogen Rearrangement Rules: Computational MS/MS Fragmentation and Structure Elucidation Using MS-FINDER Software.

              Compound identification from accurate mass MS/MS spectra is a bottleneck for untargeted metabolomics. In this study, we propose nine rules of hydrogen rearrangement (HR) during bond cleavages in low-energy collision-induced dissociation (CID). These rules are based on the classic even-electron rule and cover heteroatoms and multistage fragmentation. We evaluated our HR rules by the statistics of MassBank MS/MS spectra in addition to enthalpy calculations, yielding three levels of computational MS/MS annotation: "resolved" (regular HR behavior following HR rules), "semiresolved" (irregular HR behavior), and "formula-assigned" (lacking structure assignment). With this nomenclature, 78.4% of a total of 18506 MS/MS fragment ions in the MassBank database and 84.8% of a total of 36370 MS/MS fragment ions in the GNPS database were (semi-) resolved by predicted bond cleavages. We also introduce the MS-FINDER software for structure elucidation. Molecular formulas of precursor ions are determined from accurate mass, isotope ratio, and product ion information. All isomer structures of the predicted formula are retrieved from metabolome databases, and MS/MS fragmentations are predicted in silico. The structures are ranked by a combined weighting score considering bond dissociation energies, mass accuracies, fragment linkages, and, most importantly, nine HR rules. The program was validated by its ability to correctly calculate molecular formulas with 98.0% accuracy for 5063 MassBank MS/MS records and to yield the correct structural isomer with 82.1% accuracy within the top-3 candidates. In a test with 936 manually identified spectra from an untargeted HILIC-QTOF MS data set of human plasma, formulas were correctly predicted in 90.4% of the cases, and the correct isomer structure was retrieved at 80.4% probability within the top-3 candidates, including for compounds that were absent in mass spectral libraries. The MS-FINDER software is freely available at http://prime.psc.riken.jp/ .
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                Author and article information

                Contributors
                Role: Academic Editor
                Role: Academic Editor
                Role: Academic Editor
                Journal
                Plants (Basel)
                Plants (Basel)
                plants
                Plants
                MDPI
                2223-7747
                08 February 2021
                February 2021
                : 10
                : 2
                : 326
                Affiliations
                [1 ]Department of Agriculture and Forest Sciences, University of Tuscia, 01100 Viterbo, Italy; avceccarelli@ 123456unitus.it
                [2 ]Department for Sustainable Food Process, Research Centre for Nutrigenomics and Proteomics, Università Cattolica del Sacro Cuore, 29122 Piacenza, Italy; mariabegona.mirasmoreno@ 123456unicatt.it (B.M.-M.); valentina.buffagni@ 123456unicatt.it (V.B.); biancamaria.senizza@ 123456unicatt.it (B.S.); luigi.lucini@ 123456unicatt.it (L.L.)
                [3 ]Faculty of Science and Technology, Free University of Bozen, 39100 Bolzano, Italy; youry.pii@ 123456unibz.it
                [4 ]Consiglio per la Ricerca in Agricoltura e L’analisi Dell’economia Agraria, Centro di Ricerca Orticoltura e Florovivaismo, 84098 Pontecagnano Faiano, Italy; mteresa.cardarelli@ 123456crea.gov.it
                [5 ]Department of Agricultural Sciences, University of Naples Federico II, 80055 Portici, Italy
                Author notes
                [†]

                These authors have contributed equally to this work.

                Author information
                https://orcid.org/0000-0001-5807-7113
                https://orcid.org/0000-0002-1613-851X
                https://orcid.org/0000-0002-9865-3821
                https://orcid.org/0000-0002-1002-8651
                https://orcid.org/0000-0002-3399-3622
                https://orcid.org/0000-0002-5133-9464
                Article
                plants-10-00326
                10.3390/plants10020326
                7914860
                33567668
                9d5a76d2-61d5-40e8-8af8-02756f372241
                © 2021 by the authors.

                Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license ( http://creativecommons.org/licenses/by/4.0/).

                History
                : 12 January 2021
                : 03 February 2021
                Categories
                Article

                biostimulants,metabolomics,hormone-like activity,plant bioassay,solanum lycoperscum l.

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