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Authors: Vojtěch Schmidt, Roman Skokan, Thomas Depaepe, Katarina Kurtović, Samuel Haluška, Stanislav Vosolsobě, Roberta Vaculíková, Anthony Pil, Petre Ivanov Dobrev, Václav Motyka, Dominique Van Der Straeten and Jan Petrášek.
Nature Communications (2024)
Editor's view: Here, the authors show that the biosynthesis of many compounds in green algae preceded their recruitment in phytohormone signaling and metabolism in land plants.
Abstract: "The genomes of charophyte green algae, close relatives of land plants, typically do not show signs of developmental regulation by phytohormones. However, scattered reports of endogenous phytohormone production in these organisms exist. We performed a comprehensive analysis of multiple phytohormones in Viridiplantae, focusing mainly on charophytes. We show that auxin, salicylic acid, ethylene and tRNA-derived cytokinins including cis-zeatin are found ubiquitously in Viridiplantae. By contrast, land plants but not green algae contain the trans-zeatin type cytokinins as well as auxin and cytokinin conjugates. Charophytes occasionally produce jasmonates and abscisic acid, whereas the latter is detected consistently in land plants. Several phytohormones are excreted into the culture medium, including auxin by charophytes and cytokinins and salicylic acid by Viridiplantae in general. We note that the conservation of phytohormone biosynthesis and signaling pathways known from angiosperms does not match the capacity for phytohormone biosynthesis in Viridiplantae. Our phylogenetically guided analysis of established algal cultures provides an important insight into phytohormone biosynthesis and metabolism across Streptophyta. Genomic evidence dates the origins of most phytohormones to terrestrialization or later."
Authors: Jana Oklestkova, Miroslav Kvasnica and Miroslav Strnad.
Plant and Cell Physiology (2024)
Abstract: "Brassinosteroids (BRs) are plant steroidal hormones that play crucial roles in plant growth and development. Accurate quantification of BRs in plant tissues is essential for understanding their biological functions. This study presents a comprehensive overview of the latest methods used for the quantification of BRs in plants. We discuss the principles, advantages, and limitations of various analytical techniques, including immunoassays, gas chromatography-mass spectrometry (GC-MS), and liquid chromatography-tandem mass spectrometry (LC-MS/MS) that are used for the detection and quantification of BRs from complex plant matrices. We also explore the use of isotopically labeled internal standards to improve the accuracy and reliability of BR quantification."
Authors: Riya Johnson, Joy M. Joel and Jos T. Puthur.
Journal of Plant Growth Regulation (2024)
Abstract: "Anthropogenic activities have led to a surge in the use of synthetic chemical compounds in agriculture, elevating environmental toxicity levels. As a response to this concern, there is a growing demand for environmentally friendly solutions. In recent times, the focus has shifted towards the development of cost-effective and ecologically sound organic products known as biostimulants. These innovative products play a pivotal role in enhancing agricultural productivity by fostering comprehensive plant growth and development. Biostimulants encompass a diverse range of natural and synthetic substances, categorized into microbial, non-microbial, and waste-derived sources. When judiciously applied to crops, these substances exhibit the remarkable ability to enhance plant metabolism, bolster productivity, and enhance resilience to adverse environmental conditions. Through modulation of molecular mechanisms and epigenetic alterations, biostimulants achieve this by influencing critical signalling molecules, transcription factors, and hormonal levels, which collectively contribute to stress tolerance. This review paper delves into the burgeoning industrial interest surrounding biostimulants. It sheds light on their intricate modes and mechanisms of action, encompassing both physiochemical and molecular aspects. Furthermore, the paper underscores the captivating potential of biostimulants to induce trans-generational plasticity and metabolite accumulation within plants, a phenomenon warranting deeper exploration through metabolomics. This review paper focuses on valuable insights into the transformative influence of biostimulants on agricultural practices, showcasing their capacity to usher in a new era of sustainable and resilient crop cultivation."
Authors: Ivan Petřík, Aleš Pěnčík, Jakub Stýskal, Lenka Tranová, Petra Amakorová, Miroslav Strnad and Ondřej Novák.
Analitica Chimica Acta (2024)
Highlights • 37 cytokinins analysed in 9 min. • Limits of detection close to 0.01 fmol. • Accuracy and precision close to 15 %. • Low matrix effect for key metabolites. • Endogenous cytokinins determined in Arabidopsis seedlings.
Abstract: Background - The determination of plant hormones is still a very challenging analytical discipline, mainly due to their low concentration in complex plant matrices. Therefore, the involvement of very sensitive high-throughput techniques is required. Cytokinins (CKs) are semi-polar basic plant hormones regulating plant growth and development. Modern methods for CK determination are currently based on ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS), which enables the separation of CK isomeric forms occurring endogenously in plants. Here, ultra-high performance supercritical fluid chromatography coupled with tandem mass spectrometry (UHPSFC-MS/MS) was used for the simultaneous determination of 37 CK metabolites. Results - The chromatographic conditions were tested on four different columns with various retention mechanisms. Hybrid silica modified with 2-picolylamine was selected as the stationary phase. Several parameters such as column temperature, back pressure regulation, mobile phase composition and make-up solvent were investigated to achieve efficient separation of CK isomers and reasonable sensitivity. Compared to UHPLC-MS/MS, a 9-min chromatographic analysis using a mobile phase of supercritical CO2 and 5 mM ammonia in methanol represents a three-fold acceleration of total run time. The quantification limit of UHPSFC-MS/MS method was in the range of 0.03–0.19 fmol per injection and the method validation showed high accuracy and precision (below 15 % for most analytes). The method was finally applied to the complex plant matrix of the model plant Arabidopsis thaliana and the obtained profiles of CK metabolites were compared with the results from the conventional UHPLC-MS/MS method. Significance - The presented work offers a novel approach for quantification of endogenous CKs in plants. Compared to the conventional UHPLC-MS/MS, the total run time is shorter and the matrix effect lower for the key CK metabolites. This approach opens the opportunity to utilize UHPSFC-MS/MS instrumentation for targeted plant hormonomics including other plant hormone families."
Authors: Ondřej Vrobel and Petr Tarkowski.
Plant Methods (2023)
Abstract: "The field of plant hormonomics focuses on the qualitative and quantitative analysis of the hormone complement in plant samples, akin to other omics sciences. Plant hormones, alongside primary and secondary metabolites, govern vital processes throughout a plant's lifecycle. While active hormones have received significant attention, studying all related compounds provides valuable insights into internal processes. Conventional single-class plant hormone analysis employs thorough sample purification, short analysis and triple quadrupole tandem mass spectrometry. Conversely, comprehensive hormonomics analysis necessitates minimal purification, robust and efficient separation and better-performing mass spectrometry instruments. This review summarizes the current status of plant hormone analysis methods, focusing on sample preparation, advances in chromatographic separation and mass spectrometric detection, including a discussion on internal standard selection and the potential of derivatization. Moreover, current approaches for assessing the spatiotemporal distribution are evaluated. The review touches on the legitimacy of the term plant hormonomics by exploring the current status of methods and outlining possible future trends."
Authors: Václav Mik, Tomáš Pospíšil, Federica Brunoni, Jiří Grúz, Vladimíra Nožková, Claus Wasternack, Otto Miersch, Miroslav Strnad, Kristýna Floková, Ondřej Novák and Jitka Široká.
Phytochemistry (2023)
Abstract: Cis-(+)-12-oxophytodienoic acid (cis-(+)-OPDA) is a bioactive jasmonate, a precursor of jasmonic acid, which also displays signaling activity on its own. Modulation of cis-(+)-OPDA actions may be carried out via biotransformation leading to metabolites of various functions. This work introduces a methodology for the synthesis of racemic cis-OPDA conjugates with amino acids (OPDA-aa) and their deuterium-labeled analogs, which enables the unambiguous identification and accurate quantification of these compounds in plants. We have developed a highly sensitive liquid chromatography-tandem mass spectrometry-based method for the reliable determination of seven OPDA-aa (OPDA-Alanine, OPDA-Aspartate, OPDA-Glutamate, OPDA-Glycine, OPDA-Isoleucine, OPDA-Phenylalanine, and OPDA-Valine) from minute amount of plant material. The extraction from 10 mg of fresh plant tissue by 10% aqueous methanol followed by single-step sample clean-up on hydrophilic–lipophilic balanced columns prior to final analysis was optimized. The method was validated in terms of accuracy and precision, and the method parameters such as process efficiency, recovery and matrix effects were evaluated. In mechanically wounded 30-day-old Arabidopsis thaliana leaves, five endogenous (+)-OPDA-aa were identified and their endogenous levels were estimated. The time-course accumulation revealed a peak 60 min after the wounding, roughly corresponding to the accumulation of cis-(+)-OPDA. Our synthetic and analytical methodologies will support studies on cis-(+)-OPDA conjugation with amino acids and research into the biological significance of these metabolites in plants."
Authors: Moritz Lemke, Jens Reiners, Sander H. J. Smits, Nils Lakomek and Georg Groth.
Chemical Communications (2023)
Abstract: "The plant hormone receptor ETR1 regulates many highly relevant agronomic processes. Today, significant functional and structural questions remain unanswered regarding its multi-pass transmembrane sensor domain able to bind and respond to the gaseous plant hormone ethylene at femtomolar concentrations. A significant reason for this is the lack of structural data on full-length ETR1 in a lipid environment. Herein, we present the functional reconstitution of recombinant full-length ETR1 purified and solubilized from a bacterial host into lipid nanodiscs, allowing the study of the purified plant receptor for the first time in a detergent-free membrane-like environment."
Authors: Cynthia Wong, David Alabadí and Miguel A. Blázquez.
Journal of Experimental Botany (2023)
Abstract: "Although many plant cell types are capable of producing hormones and plant hormones can in most cases act in the same cells in which they are produced, they also act as signaling molecules that coordinate physiological responses between different parts of the plant, indicating that their action is subject to spatial regulation. Numerous publications have reported that all levels of plant hormonal pathways, i.e., metabolism, transport, and perception/signal transduction, can help determine the spatial ranges of hormone action. For example, polar auxin transport or localized auxin biosynthesis contribute to creating a differential hormone accumulation across tissues that is instrumental for specific growth and developmental responses. On the other hand, tissue specificity of cytokinin actions has been proposed to be regulated by mechanisms operating at the signaling stages. Here, we review and discuss current knowledge about the contribution of the three levels mentioned above in providing spatial specificity to plant hormone action. We also explore how new technological developments, such as plant hormone sensors based on FRET or single-cell RNA-seq, can provide an unprecedented level of resolution in defining the spatial domains of plant hormone action and its dynamics."
Authors: Václav Mik, Tomáš Poslíšil, Federica Brunoni, Jiří Grúz, Vladimíra Nožková, Claus Wasternack, Otto Miersch, Miroslav Strnad, Kristýna Floková, Ondřej Novák and Jitka Široká.
ChemRxiv (2023)
Highlights: Optimized methodology for synthesizing cis-OPDA conjugates with amino acids (OPDA-aa) and their deuterium-labeled analogs. Development of an analytical method based on liquid chromatography-tandem mass spectrometry for unambiguous identification and accurate quantification of seven OPDA-aa in plants. Identification of five (+)-OPDA-aa in mechanically wounded Arabidopsis thaliana leaves and their accumulation after wounding (0, 5, 15, 30, 60, and 180 min).
Abstract: "Cis-(+)-12-oxophytodienoic acid (cis-(+)-OPDA) is a bioactive jasmonate, a precursor of jasmonic acid, which also displays signaling activity on its own. Modulation of cis-(+)-OPDA actions may be carried out via biotransformation leading to metabolites of various functions, similar to other phytohormones. This work introduces a methodology for the synthesis of racemic cis-OPDA conjugates with amino acids (OPDA-aa) and their deuterium-labeled analogs, which enables the identification and accurate quantification of these compounds in plants. We have developed a highly sensitive liquid chromatography-tandem mass spectrometry-based method for the reliable determination of seven OPDA-aa (OPDA-Alanine, OPDA-Aspartate, OPDA-Glutamate, OPDA-Glycine, OPDA-Isoleucine, OPDA-Phenylalanine, and OPDA-Valine) from minute amount of plant material. The extraction from 10 mg of fresh plant tissue by 10% aqueous methanol followed by single-step sample clean-up on hydrophilic–lipophilic balanced columns prior to final analysis was optimized. The method was validated in terms of accuracy and precision, and the method parameters such as process efficiency, recovery and matrix effects were evaluated. In mechanically wounded 30-day-old Arabidopsis thaliana leaves, five endogenous (+)-OPDA-aa were identified and their endogenous levels reached a maximum of pmol/g. The time-course accumulation revealed a peak 60 min after the wounding, roughly corresponding to the accumulation of cis-(+)-OPDA. Current synthetic and analytical methodologies support studies on cis-(+)-OPDA conjugation with amino acids and research into the biological significance of these metabolites in plants."
Authors: Vojtěch Schmidt, Roman Skokan, Katarina Kurtović, Stanislav Vosolsobě, Roberta Filepová, Samuel Haluška, Petre Dobrev, Václav Motyka and Jan Petrášek.
bioRxiv (2023)
Abstract: "Multiple phytohormones act as conserved developmental regulators in land plants. Although the closely related streptophyte green algae typically lack full complements of molecular pathways underlying these responses, scattered reports of endogenous phytohormone production in these organisms exist. In this study, we performed a detailed LC/MS-based analysis of several phytohormones, their precursors and metabolites in all lineages of streptophyte algae. We also included chlorophyte algae and early-diverging land plants as outgroups. Free auxin, tRNA-derived cytokinins and certain phenolics including salicylic acid were found ubiquitously. However, land plants differed from green algae by the consistent detection of abscisic acid and the presence of auxin and cytokinin conjugates and trans-zeatin, supporting the hypotheses that these three phytohormones likely came to regulate development in the ancestral land plant. By contrast, we observed a patchy distribution of jasmonates among streptophytes. We additionally analyzed the corresponding culture and empty media to account for phytohormone excretion and environmental contamination. Extracellular auxins and cytokinins were frequently detected, while agar constituted a major external source of phenolic compounds. We provide a highly comprehensive evolution-directed screen of phytohormone compound occurrence and thoroughly discuss our data in the context of current plant hormonomics and phylogenomics."
Author: Paula Elizabeth Jameson.
Plant Physiology (2023)
Abstract: "While various labs had shown cell division-inducing activity in a variety of plant extracts for over a decade, the identification of zeatin (Z) in 1964, the first known naturally occurring cytokinin, belongs to Letham and co-workers. Using extracts from maize (Zea mays), they were the first to obtain crystals of pure Z and in sufficient quantity for structural determination by MS, NMR, chromatography, and mixed melting point analysis. This group also crystallised Z-9-riboside (ZR) from coconut (Cocos nucifera) milk. However, their chemical contributions go well beyond the identification of Z and ZR and include two unambiguous syntheses of trans-Z (to establish stereochemistry), the synthesis of 3H-cytokinins that facilitated metabolic studies, and the synthesis of deuterated internal standards for accurate mass spectral quantification. Letham and associates also unequivocally identified Z nucleotide, the 7-and 9-glucoside conjugates of Z, and the O-glucosides of Z, ZR, dihydro Z (DHZ) and DHZR as endogenous compounds and as metabolites of exogenous Z. Their contributions to the role of cytokinins in plant physiology and development were also substantial, especially the role of cytokinins moving in the xylem. These biological advances are described and briefly related to the genetic/molecular biological contributions of others that established that plants have an absolute requirement for cytokinin."
Authors: Meng Wu, Chenhui Yin, Zhixin Liu, Mengyuan Wang, Qijun Sun, Na Niu and Ligang Chen.
Analytical Chemistry (2023)
Abstract: " To fully understand the function of the phytohormone indole-3-acetic acid (IAA) in regulating plant growth and development, we need to monitor their levels and distribution with high spatial and temporal resolution. In this work, an anthracene-based fluorescent biosensor for IAA was prepared using bovine serum albumin (BSA) as a bio-template. The single linear oxygen (1O2) specifically produced by IAA catalyzed with horseradish peroxidase (HRP) turns on the fluorescence of the probe, enabling specific trace sensing of IAA in the presence of multiple structural analogues. The presence of the bio-template BSA extends the biocompatibility of the probe, enabling visual monitoring of the level and distribution signal of endogenous IAA of plants in the field of bioimaging. In addition, the strategy has shown potential for application in portable paper-based sensors and in vivo fluorescent flower culture. This work provides a technical and theoretical basis for exploring the growth regulatory mechanisms of IAA in plants at the molecular level."
Authors: Katrina H. Jensen and Brian W. Michel.
Analysis & Sensing (2023)
Abstract: "Despite its relative simplicity, ethylene is an interesting molecule with wide-ranging impact in modern chemistry and biology. Stemming from ethylene’s role as a critical plant hormone, there has been significant effort to develop selective and sensitive molecular sensors for ethylene. Late transition metal complexes have played an important role in detection strategies due to ethylene’s lack of structural complexity and limited reactivity. Two main approaches to ethylene detection are identified: (1) coordination-based sensors, wherein ethylene binds reversibly to a metal center, and (2) activity-based sensors, wherein ethylene undergoes a reaction at a metal center, resulting in the formation and destruction of covalent bonds. Herein, we describe the advantages and disadvantages of various approaches, and the challenges remaining for sensor development."
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Authors: Masato Homma, Kiyono Uchida, Takatoshi Wakabayashi, Masaharu Mizutani, Hirosato Takikawa and Yukihiro Sugimoto.
Frontiers in Plant Science (2024)
Abstract: "Strigolactones (SLs), a class of plant apocarotenoids, serve dual roles as rhizosphere-signaling molecules and plant hormones. Orobanchol, a major naturally occurring SL, along with its various derivatives, has been detected in the root exudates of plants of the Fabaceae family. Medicaol, fabacyl acetate, and orobanchyl acetate were identified in the root exudates of barrel medic (Medicago truncatula), pea (Pisum sativum), and cowpea (Vigna unguiculata), respectively. Although the biosynthetic pathway leading to orobanchol production has been elucidated, the biosynthetic pathways of the orobanchol derivatives have not yet been fully elucidated. Here, we report the identification of 2-oxoglutarate-dependent dioxygenases (DOXs) and BAHD acyltransferases responsible for converting orobanchol to these derivatives in Fabaceae plants. First, the metabolic pathways downstream of orobanchol were analyzed using substrate feeding experiments. Prohexadione, an inhibitor of DOX inhibits the conversion of orobanchol to medicaol in barrel medic. The DOX inhibitor also reduced the formation of fabacyl acetate and fabacol, a precursor of fabacyl acetate, in pea. Subsequently, we utilized a dataset based on comparative transcriptome analysis to select a candidate gene encoding DOX for medicaol synthase in barrel medic. Recombinant proteins of the gene converted orobanchol to medicaol. The candidate genes encoding DOX and BAHD acyltransferase for fabacol synthase and fabacol acetyltransferase, respectively, were selected by co-expression analysis in pea. The recombinant proteins of the candidate genes converted orobanchol to fabacol and acetylated fabacol. Furthermore, fabacol acetyltransferase and its homolog in cowpea acetylated orobanchol. The kinetics and substrate specificity analyses revealed high affinity and strict recognition of the substrates of the identified enzymes. These findings shed light on the molecular mechanisms underlying the structural diversity of SLs."
Authors: Jennifer Andres, Lisa J. Schmunk, Federico Grau-Enguix, Justine Braguy, Sophia L. Samodelov, Tim Blomeier, Rocio Ochoa-Fernandez, Wilfried Weber, Salim Al-Babili, David Alabadí, Miguel A. Blázquez and Matias D. Zurbriggen.
The Plant Journal (2024)
Significance Statement: Using the degradation-based signaling mechanism of GAs, we have built transcriptional regulator (DELLA)-based, genetically encoded ratiometric biosensors as proxies for hormone quantification at high temporal resolution and sensitivity that allow dynamic, rapid and simple analysis in a plant cell system, i.e. Arabidopsis protoplasts.
Abstract: "Gibberellins (GAs) are major regulators of developmental and growth processes in plants. Using the degradation-based signaling mechanism of GAs, we have built transcriptional regulator (DELLA)-based, genetically encoded ratiometric biosensors as proxies for hormone quantification at high temporal resolution and sensitivity that allow dynamic, rapid and simple analysis in a plant cell system, i.e. Arabidopsis protoplasts. These ratiometric biosensors incorporate a DELLA protein as a degradation target fused to a firefly luciferase connected via a 2A peptide to a renilla luciferase as a co-expressed normalization element. We have implemented these biosensors for all five Arabidopsis DELLA proteins, GA-INSENSITIVE, GAI; REPRESSOR-of-ga1-3, RGA; RGA-like1, RGL1; RGL2 and RGL3, by applying a modular design. The sensors are highly sensitive (in the low pm range), specific and dynamic. As a proof of concept, we have tested the applicability in three domains: the study of substrate specificity and activity of putative GA-oxidases, the characterization of GA transporters, and the use as a discrimination platform coupled to a GA agonists' chemical screening. This work demonstrates the development of a genetically encoded quantitative biosensor complementary to existing tools that allow the visualization of GA in planta."
Authors: Kazuo Soeno, Akiko Sato and Yukihisa Shimada.
Japan Agricultural Research Quarterly (2024)
Abstract: "Auxins are plant hormones that play key roles in almost all growth and developmental processes, such as cell division, elongation, differentiation, and environmental responses. However, biosynthetic pathways and regulatory mechanisms remain unclear. The indole-3-pyruvic acid (IPyA) pathway, from L-tryptophan (Trp) via IPyA, is the main biosynthetic pathway of the natural auxin indole-3-acetic acid (IAA). In this pathway, IAA is biosynthesized from Trp through two enzymatic reactions: aminotransferase (TRYPTOPHAN AMINOTRANSFERASE OF ARABIDOPSIS1 [TAA1]/ TRYPTOPHAN AMINOTRANSFERASE-RELATED [TARs]) and YUCCAs (YUCs), which are flavin-containing monooxygenases. We developed the inhibitors of TAA1/TARs and YUC and analyzed the physiological functions of the IPyA pathway using the biosynthesis inhibitors as chemical probes. This paper also describes the regulatory mechanism of the two-step enzymatic reactions in auxin biosynthesis, employing the novel IPyA analog compounds."
Authors: Bunta Nishikawa, Kotaro Mori, Seisuke Takimoto, Bunta Watanabe, Midori Matsuo, Takeshi Nakano, Yoshiaki Nakagawa and Hisashi Miyagawa.
Journal of Pesticide Science (2023)
Abstract: "Spiral roots are induced in germinated rice seeds through treatment with nanomolar brassinosteroids (BRs) but not with other plant hormones. Here, we determined the minimum effective concentration (MEC) of various BRs to induce spiral roots in germinated rice seeds. The reciprocal logarithm of MEC, pMEC, was used as the BL-like activity index, which was linearly correlated with the reciprocal logarithm of a 50% effective dose (pED50) as evaluated in the rice lamina inclination assay. Furthermore, a ligand-receptor docking simulation was performed against the BL receptor complex, Arabidopsis thaliana BRI1/SERK1, and the binding free energy (ΔGbind) was calculated for the tested BRs. The ΔGbind calculation was performed using the molecular mechanics/generalized Born surface area method on an ensemble of uncorrelated snapshots collected via molecular dynamics to predict biological activity."
Authors: Yiliang Chen, Bo He, Mengxu Hu, Jiawei Bao, Wei Yan, Xinya Han and Yonghao Ye.
Advanced Agrochem (2024)
Highlights: • Various types of fluorescent probes for detecting phytohormones (abscisic acid, ethylene, salicylic acid, strigolactones, jasmonic acid, auxins, and cytokinin) are reviewed. • The recognition mechanisms, sensing properties and imaging quality of fluorescent probes for phytohormones are described in detail. • The prospects of fluorescent probes for sensing phytohormones are presented, and the possible improvement directions in the future are discussed.
Abstract: "Exploring plant behavior at the cellular scale in a minimally invasive manner is critical to understanding plant adaptation to the environment. Phytohormones play vital regulatory roles in multiple aspects of plant growth and development and acclimation to environmental changes. Since the biosynthesis, modification, transportation, and degradation of plant hormones in plants change with time and space, their content level and distribution are highly dynamic. To monitor the production, transport, perception, and distribution of phytohormones within undamaged tissues, we require qualitative and quantitative tools endowed with remarkably high temporal and spatial resolution. Fluorescent probes are regarded as excellent tools for widespread plant imaging because of their high sensitivity and selectivity, reproducibility, real-time in situ detection, and uncomplicated mechanism elucidation. In this review, we provide a systematical overview of the progress in the sensing and imaging of phytohormone fluorescent probes and fluorescently labeled phytohormones to their receptors in plants. Moreover, forthcoming viewpoints and possible applications of these fluorescent probes within the realm of plants are also presented. We hold the conviction that the new perspective brought by this paper can promote the development of fluorescent probes, enabling them to have better detection performance in plant hormone imaging."
Authors: Vladimír Skalický, Ioanna Antoniadi, Aleš Pěnčík, Ivo Chamrád, René Lenobel, Martin F. Kubeš, Marek Zatloukal, Asta Žukauskaitė, Miroslav Strnad, Karin Ljung and Ondřej Novák.
The Plant Journal (2023)
Significance Statement: We introduce a breakthrough technique, so-called Fluorescence-Activated multi-Organelle Sorting (FAmOS), for simultaneous fractionation of up to four organelle populations (nuclei, chloroplasts, mitochondria and endoplasmic reticulum) from a single sample of plant cell cultures. The unique combination of our developed FAmOS tool with ultra-sensitive mass spectrometry-based analytical methods enables high-resolution mapping of plant metabolites in isolated organelles.
Abstract: "Auxins and cytokinins are two major families of phytohormones that control most aspects of plant growth, development and plasticity. Their distribution in plants has been described, but the importance of cell- and subcellular-type specific phytohormone homeostasis remains undefined. Herein, we revealed auxin and cytokinin distribution maps showing their different organelle-specific allocations within the Arabidopsis plant cell. To do so, we have developed Fluorescence-Activated multi-Organelle Sorting (FAmOS), an innovative subcellular fractionation technique based on flow cytometric principles. FAmOS allows the simultaneous sorting of four differently labelled organelles based on their individual light scatter and fluorescence parameters while ensuring hormone metabolic stability. Our data showed different subcellular distribution of auxin and cytokinins, revealing the formation of phytohormone gradients that have been suggested by the subcellular localization of auxin and cytokinin transporters, receptors and metabolic enzymes. Both hormones showed enrichment in vacuoles, while cytokinins were also accumulated in the endoplasmic reticulum."
Authors: Syed Muhammad Zaigham Abbas Naqvi, Yanyan Zhang, Muhammad Naveed Tahir, Zia Ullah, Shakeel Ahmed, Junfeng Wu, Vijaya Raghavan, Mukhtar Iderawumi Abdulraheem, Jianfeng Ping, Xinran Hu and Jiandong Hu.
Trends in Analytical Chemistry (2023)
Highlights • In vivo methods perform cellular imaging and microscopic profiling of intact organs. • Spectroscopy, biosensors, electrochemical sensors quantified trace elements precisely. • Modern methods can detect different plant hormones with accuracy and desired results. • New ultrasensitive nondestructive sensors have a higher temporal resolution.
Abstract: In vivo plant hormonal detection methods are the strategies executed on living organisms while not taking out the samples from the body. Modern methods like spectroscopy, biosensors, and electrochemical sensors have increased the quality and sensitivity of the detection and decreased the cumbersome, time, and solvent-consuming preparational efforts of traditional methods. Preparational efforts generally consisted of several samplings, extraction, purification, and enrichment steps to analyze plant hormones. Concentrations and levels of plant hormones change concerning biotic and abiotic stresses that plant faces. Spectroscopy, biosensors, and electrochemical sensors detect different plant hormones with specific accuracy and desired results, but their handling and sample preparations are challenging. At the same time, advanced ultrasensitive nondestructive methods are biocompatible and can be used for a lifetime with excellent stretchability performance. In modern science, portable, wearable, and nondestructive measurement is the trend of plant sensors that can make the “Internet of Plants” concept a reality. This review covers the significant aspects of numerous applications, advantages, and disadvantages of spectroscopy, biosensors, electrochemical sensors, and new ultrasensitive nondestructive devices to measure in vivo plant hormones. The summary of the advanced ultrasensitive plant devices, their challenges, and future prospects within the fields for in vivo plant hormone detections is presented. Furthermore, it can guide researchers to design new experiments using ultrasensitive nondestructive sensing devices for detecting in vivo plant hormones."
Authors: James Rowe, Mathieu Grangé-Guermente, Marino Exposito-Rodriguez, Rinukshi Wimalasekera, Martin O. Lenz, Kartika N. Shetty, Sean R. Cutler and Alexander M. Jones.
Nature Plants (2023)
Editor's view: ABACUS2 Förster resonance energy transfer biosensors allow an unparalleled view of abscisic acid accumulations and depletions in living plants. Well-watered roots accumulate abscisic acid in growing cells upon shoot dehydration and this is essential to maintain root growth under low humidity.
Abstract:"The plant hormone abscisic acid (ABA) accumulates under abiotic stress to recast water relations and development. To overcome a lack of high-resolution sensitive reporters, we developed ABACUS2s—next-generation Förster resonance energy transfer (FRET) biosensors for ABA with high affinity, signal-to-noise ratio and orthogonality—that reveal endogenous ABA patterns in Arabidopsis thaliana. We mapped stress-induced ABA dynamics in high resolution to reveal the cellular basis for local and systemic ABA functions. At reduced foliar humidity, root cells accumulated ABA in the elongation zone, the site of phloem-transported ABA unloading. Phloem ABA and root ABA signalling were both essential to maintain root growth at low humidity. ABA coordinates a root response to foliar stresses, enabling plants to maintain foraging of deeper soil for water uptake."
Authors: Petre I. Dobrev, Roberta Filepová, Jozef Lacek, Zuzana Vondráková, Karel Müller, Petr Maršík, Lenka Drašarová, Pavel Talacko, Petr Hošek and Jan Petrášek.
bioRxiv (2023)
Abstract: "The natural plant hormone auxin indole-3-acetic acid (IAA) influences many physiological processes in plants. Here, the metabolism of IAA was studied in detail using tobacco BY-2 cells as a model and compared with the in planta metabolism in several plant species. A combination of labeled/unlabeled substrate feeding, global untargeted mass spectrometric (MS) scanning, and selective MS filtering allowed the detection of 17 auxin metabolites, 15 of which were identified. Subsequent study of intermediate metabolism and dynamics revealed eight major pathways: three amino acid conjugation pathways with aspartate, glutamate, and glutamine, followed by their 2-oxidation with the help of the DAO enzyme; side-chain glucosyl ester formation; direct 2-oxidation; two decarboxylation pathways; and a pathway producing an unidentified metabolite. Interestingly, the first intermediates of the two decarboxylation pathways, indole-3-carbinol and oxoindole-3-carbinol, were formed outside the cells. We found that the majority of the detected auxin metabolites occur naturally in several plant species and that IAA is their precursor, indicating that the auxin metabolic pathways observed in BY-2 cells also occur in planta. Our finding that the IAA decarboxylation pathway occurs in planta, and the previous reports of auxin activity of some metabolites of this pathway, suggest that at least some of the biological effects of IAA may be explained by its conversion to decarboxylative metabolites."
Authors: Subodh, Ravina, Priyanka, Jagriti Narang and Hari Mohan.
Sensors International (2023)
Highlights: • Phytohormone ABA treats conditions like glioma, depression, glucose tolerance in rats and humans, cardiovascular risks. • LSPR (localized surface plasmon resonance system) is a colorimetric detection method based on aptamer, LSPR based biosensor is highly specific and sensitive for ABA at nanomolar concentration. • SnRK2 kinases are essential for ABA-mediated signaling and abiotic stress responses in plants by regulating stomatal closure through phosphorylation of downstream targets such as AKS1. • SNACS is a genetically encoded biosensor that can detect SnRK2 activity in live cells and is a valuable tool for studying ABA-induced changes in SnRK2 activity. • Scientists developed genetically encoded FRET sensors to detect Abscisic acid (ABA) concentration and uptake in plants. The sensors have a high signal-to-noise ratio and can provide high-resolution measurements of ABA levels in living tissues.
Abstract: "ABA (abscisic acid) is a phytohormone that is important in regulating stress and various pathological conditions in humans. ABA can be ingested exogenously or endogenously, and is naturally produced by stem cells, macrophages, and keratinocytes. ABA mediates immunological reactions such as phagocytosis, chemotaxis, and reactive oxygen species (ROS) production. In humans, LANCL2 serves as an ABA receptor. Depending on the signaling pathway involved, ABA can act as an inflammatory or anti-inflammatory agent. ABA plays a vital role in glucose tolerance, atherosclerosis treatment, prostate cancer dormancy, malaria, type 2 diabetes, and dementia. Various detection methods have been developed to detect ABA, including chromatographic, spectroscopic, and colorimetric assays, as well as high-performance liquid chromatography (HPLC) and mass spectrometry. Biosensors are commonly used to detect analytes in low concentrations. LSPR and FRET sensors are two types of biosensors that have been developed for the detection of ABA. LSPR sensors rely on light interacting with metal nanoparticles to detect analytes, while FRET sensors rely on energy transfer between a donor and acceptor molecule. LSPR sensors offer high sensitivity, label-free detection, and the ability to detect multiple targets simultaneously, while FRET sensors offer high specificity and customizability. Aptamers have recently been used as a substitute for antibodies in biosensors to provide higher sensitivity and lower cost. This review focuses on the role of ABA and the various biosensors used to detect it in the treatment of various diseases in mammals."
Authors: Kulaporn Boonyaves, Mervin Chun-Yi Ang, Minkyung Park, Jianqiao Cui, Duc Thinh Khong, Gajendra Pratap Singh, Volodymyr B. Koman, Xun Gong, Thomas Koizumi Porter, Seo Woo Choi, Kwanghun Chung, Nam-Hai Chua, Daisuke Urano and Michael S. Strano.
Nano Letters (2023)
Abstract: "Gibberellins (GAs) are a class of phytohormones, important for plant growth, and very difficult to distinguish because of their similarity in chemical structures. Herein, we develop the first nanosensors for GAs by designing and engineering polymer-wrapped single-walled carbon nanotubes (SWNTs) with unique corona phases that selectively bind to bioactive GAs, GA3 and GA4, triggering near-infrared (NIR) fluorescence intensity changes. Using a new coupled Raman/NIR fluorimeter that enables self-referencing of nanosensor NIR fluorescence with its Raman G-band, we demonstrated detection of cellular GA in Arabidopsis, lettuce, and basil roots. The nanosensors reported increased endogenous GA levels in transgenic Arabidopsis mutants that overexpress GA and in emerging lateral roots. Our approach allows rapid spatiotemporal detection of GA across species. The reversible sensor captured the decreasing GA levels in salt-treated lettuce roots, which correlated remarkably with fresh weight changes. This work demonstrates the potential for nanosensors to solve longstanding problems in plant biotechnology."
Authors: Jitka Široká, Federica Brunoni, Aleš Pěnčík, Václav Mik, Asta Žukauskaitė, Miroslav Strnad, Ondřej Novák and Kristýna Floková.
Plant Methods (2022)
Abstract: "Background - Acidic phytohormones are small molecules controlling many physiological functions in plants. A comprehensive picture of their profiles including the active forms, precursors and metabolites provides an important insight into ongoing physiological processes and is essential for many biological studies performed on plants. Results - A high-throughput sample preparation method for liquid chromatography–tandem mass spectrometry determination of 25 acidic phytohormones classed as auxins, jasmonates, abscisates and salicylic acid was optimised. The method uses a small amount of plant tissue (less than 10 mg fresh weight) and acidic extraction in 1 mol/L formic acid in 10% aqueous methanol followed by miniaturised purification on reverse phase sorbent accommodated in pipette tips organised in a 3D printed 96-place interface, capable of processing 192 samples in one run. The method was evaluated in terms of process efficiency, recovery and matrix effects as well as establishing validation parameters such as accuracy and precision. The applicability of the method in relation to the amounts of sample collected from distantly related plant species was evaluated and the results for phytohormone profiles are discussed in the context of literature reports. Conclusion - The method developed enables high-throughput profiling of acidic phytohormones with minute amounts of plant material, and it is suitable for large scale interspecies studies."
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