Biochemical characterization of a unique cytokinin and nucleotide phosphoribohydrolase Lonely Guy protein from Dictyostelium discoideum.

Lonely guy (LOG) proteins are phosphoribohydrolases (PRHs) that are key cytokinin (CK)-activating enzymes in plant and non-plant CK-producing organisms. During CK biosynthesis, LOGs catalyze the conversion of precursor CK-nucleotides (CK-NTs) to biologically active free base forms. LOG/PRH activity has been detected in bacteria, archaea, algae, and fungi. However, in these organisms, the LOG/PRH activity for CK-NTs and non-CK-NTs (e.g., adenine-NTs) has not been assessed simultaneously, which leaves limited knowledge about the substrate specificity of LOGs. Thus, we performed bioinformatic analyses and a biochemical characterization of a LOG ortholog from Dictyostelium discoideum, a soil-dwelling amoeba, which produces CKs during unicellular growth and multicellular development. We show that DdLog exhibits LOG/PRH activity on two CK-NTs, N 6 -isopentenyladenosine-5'-monophosphate (iPMP) and N 6 -benzyladenosine-5'-monophosphate (BAMP), and on adenosine 5'-monophosphate (AMP) but not on 3', 5'-cyclic adenosine-monophosphate (cAMP). Additionally, there were higher turnover rates for CK-NTs over AMP. Together, these findings confirm that DdLog acts as a CK-activating enzyme; however, in contrast to plant LOGs, it maintains a wider specificity for other substrates (e.g., AMP) reflecting it has maintained its original, non-CK related role even after diversifying into a CK-activating enzyme.

Cytokinins Reduce Viral Replication and Alter Plaque Morphology of Frog Virus 3 In Vitro.

Cytokinins (CKs) are a group of N6-substituted signaling molecules whose biosynthesis and metabolism have been documented in all kingdoms of life, including vertebrates. While their biological relevance in vertebrate systems continues to be elucidated, they have broadly been documented with therapeutic effects in exogenous applications. In this study, we evaluated the virostatic potential of four types of CKs including, N6-isopentenyladenine (iP), N6-isopentenyladenosine (iPR), N6-isopentenyladenosine-5'monophosphate (iPMP), and 2-methylthiol-N6-isopentenyladenosine (2MeSiPR) against the ranavirus type species, frog virus 3 (FV3). Following concurrent treatment and infection, iP and iPR reduced viral replication by 33.8% and 59.6%, respectively, in plaque formation assays. A decrease in viral replication was also observed when CK exposure was limited to 12 h prior to infection, where iP and iPR reduced viral replication by 31% and 23.75%, respectively. Treatment with iP and iPR was also marked by 48% and 60% decreases in viral load over 72 h, respectively, as measured in single step growth curves. Plaque morphology was altered in vitro, as iP and iPR treatment increased plaque area by 83% and 112% with lytic zone formation also becoming more prevalent in corresponding treatments. Treatment with iPMP and 2MeSiPR resulted in no effect on viral kinetics in vitro. The results of this study are the first to provide evidence of CK antiviral activity against a DNA virus and highlight the importance of their structure for therapeutic investigations.

Impact of glyphosate and glyphosate-based herbicides on phyllospheric Methylobacterium.

Symbiotic Methylobacterium comprise a significant portion of the phyllospheric microbiome, and are known to benefit host plant growth, development, and confer tolerance to stress factors. The near ubiquitous use of the broad-spectrum herbicide, glyphosate, in farming operations globally has necessitated a more expansive evaluation of the impacts of the agent itself and formulations containing glyphosate on important components of the plant phyllosphere, including Methylobacterium.This study provides an investigation of the sensitivity of 18 strains of Methylobacterium to glyphosate and two commercially available glyphosate-based herbicides (GBH). Nearly all strains of Methylobacterium showed signs of sensitivity to the popular GBH formulations WeatherMax® and Transorb® in a modified Kirby Bauer experiment. However, exposure to pure forms of glyphosate did not show a significant effect on growth for any strain in both the Kirby Bauer test and in liquid broth, until polysorbate-20 (Tween20) was added as a surfactant. Artificially increasing membrane permeability through the introduction of polysorbate-20 caused a 78-84% reduction in bacterial cell biomass relative to controls containing glyphosate or high levels of surfactant only (0-9% and 6-37% reduction respectively). Concentrations of glyphosate as low as 0.05% w/v (500 µg/L) from both commercial formulations tested, inhibited the culturability of Methylobacterium on fresh nutrient-rich medium.To better understand the compatibility of important phyllospheric bacteria with commercial glyphosate-based herbicides, this study endeavours to characterize sensitivity in multiple strains of Methylobacterium, and explore possible mechanisms by which toxicity may be induced.

Euglena mutabilis exists in a FAB consortium with microbes that enhance cadmium tolerance.

BACKGROUND: Synthetic algal-fungal and algal-bacterial cultures have been investigated as a means to enhance the technological applications of the algae. This inclusion of other microbes has enhanced growth and improved stress tolerance of the algal culture. The goal of the current study was to investigate natural microbial consortia to gain an understanding of the occurrence and benefits of these associations in nature. The photosynthetic protist Euglena mutabilis is often found in association with other microbes in acidic environments with high heavy metal (HM) concentrations. This may suggest that microbial interactions are essential for the protist's ability to tolerate these extreme environments. Our study assessed the Cd tolerance of a natural fungal-algal-bacterial (FAB) association whereby the algae is E. mutabilis. RESULTS: This study provides the first assessment of antibiotic and antimycotic agents on an E. mutabilis culture. The results indicate that antibiotic and antimycotic applications significantly decreased the viability of E. mutabilis cells when they were also exposed to Cd. Similar antibiotic treatments of E. gracilis cultures had variable or non-significant impacts on Cd tolerance. E. gracilis also recovered better after pre-treatment with antibiotics and Cd than did E. mutabilis. The recoveries were assessed by heterotrophic growth without antibiotics or Cd. In contrast, both Euglena species displayed increased chlorophyll production upon Cd exposure. PacBio full-length amplicon sequencing and targeted Sanger sequencing identified the microbial species present in the E. mutabilis culture to be the fungus Talaromyces sp. and the bacterium Acidiphilium acidophilum. CONCLUSION: This study uncovers a possible fungal, algal, and bacterial relationship, what we refer to as a FAB consortium. The members of this consortium interact to enhance the response to Cd exposure. This results in a E. mutabilis culture that has a higher tolerance to Cd than the axenic E. gracilis. The description of this interaction provides a basis for explore the benefits of natural interactions. This will provide knowledge and direction for use when creating or maintaining FAB interactions for biotechnological purposes, including bioremediation.

From biosynthesis and beyond-Loss or overexpression of the cytokinin synthesis gene, iptA, alters cytokinesis and mitochondrial and amino acid metabolism in Dictyostelium discoideum.

Cytokinins (CKs) are a class of growth-promoting signaling molecules that affect multiple cellular and developmental processes. These phytohormones are well studied in plants, but their presence continues to be uncovered in organisms spanning all kingdoms, which poses new questions about their roles and functions outside of plant systems. Cytokinin production can be initiated by one of two different biosynthetic enzymes, adenylate isopentenyltransfases (IPTs) or tRNA isopentenyltransferases (tRNA-IPTs). In this study, the social amoeba, Dictyostelium discoideum, was used to study the role of CKs by generating deletion and overexpression strains of its single adenylate-IPT gene, iptA. The life cycle of D. discoideum is unique and possesses both single- and multicellular stages. Vegetative amoebae grow and divide while food resources are plentiful, and multicellular development is initiated upon starvation, which includes distinct life cycle stages. CKs are produced in D. discoideum throughout its life cycle and their functions have been well studied during the later stages of multicellular development of D. discoideum. To investigate potential expanded roles of CKs, this study focused on vegetative growth and early developmental stages. We found that iptA-deficiency results in cytokinesis defects, and both iptA-deficiency and overexpression results in dysregulated tricarboxylic acid (TCA) cycle and amino acid metabolism, as well as increased levels of adenosine monophosphate (AMP). Collectively, these findings extend our understanding of CK function in amoebae, indicating that iptA loss and overexpression alter biological processes during vegetative growth that are distinct from those reported during later development.

Profiling of adenine-derived signaling molecules, cytokinins, in myotubes reveals fluctuations in response to lipopolysaccharide-induced cell stress.

Cytokinins (CTKs) are a diverse collection of evolutionarily conserved adenine-derived signaling molecules classically studied as phytohormones; however, their roles and production have been less studied in mammalian systems. Skeletal muscles are sensitive to cellular cues such as inflammation and in response, alter their secretome to regulate the muscle stem cell and myofiber niche. Using cultured C2C12 muscle cells, we profiled CTK levels to understand (1) whether CTKs are part of the muscle secretome and (2) whether CTKs are responsive to cellular stress. To induce cellular stress, C2C12 myotubes were treated with lipopolysaccharides (LPS) for 24 h and then media and cell fractions were collected for ultra high-performance liquid chromatography tandem mass spectrometry with electrospray ionization (UHPLC-(ESI+)-HRMS/MS) for metabolomics and CTK profiling. Across LPS-treated and control cells, 11 CTKs were detected in the extracellular space while 6 were detected intracellularly. We found that muscle cells are enriched in isopentenyladenine (iP) species (from free base, riboside to nucleotide forms), and that extracellular levels are increased after LPS treatment. Our study establishes that muscle cells express various forms of CTKs, and that CTK levels are responsive to LPS-induced cell stress, suggesting a role for CTKs in intra- and extracellular signaling of mammalian cells.

Unlocking Potentially Therapeutic Phytochemicals in Capadulla (Doliocarpus dentatus) from Guyana Using Untargeted Mass Spectrometry-Based Metabolomics.

Doliocarpus dentatus is thought to have a wide variety of therapeutic phytochemicals that allegedly improve libido and cure impotence. Although a few biomarkers have been identified with potential antinociceptive and cytotoxic properties, an untargeted mass spectrometry-based metabolomics approach has never been undertaken to identify therapeutic biofingerprints for conditions, such as erectile dysfunction, in men. This study executes a preliminary phytochemical screening of the woody vine of two ecotypes of D. dentatus with renowned differences in therapeutic potential for erectile dysfunction. Liquid chromatography-mass spectrometry-based metabolomics was used to screen for flavonoids, terpenoids, and other chemical classes found to contrast between red and white ecotypes. Among the metabolite chemodiversity found in the ecotype screens, using a combination of GNPS, MS-DIAL, and SIRIUS, approximately 847 compounds were annotated at levels 2 to 4, with the majority of compounds falling under lipid and lipid-like molecules, benzenoids and phenylpropanoids, and polyketides, indicative of the contributions of the flavonoid, shikimic acid, and terpenoid biosynthesis pathways. Despite the extensive annotation, we report on 138 tentative compound identifications of potentially therapeutic compounds, with 55 selected compounds at a level-2 annotation, and 22 statistically significant therapeutic biomarkers, the majority of which were polyphenols. Epicatechin methyl gallate, catechin gallate, and proanthocyanidin A2 had the greatest significant differences and were also relatively abundant among the red and white ecotypes. These putatively identified compounds reportedly act as antioxidants, neutralizing damaging free radicals, and lowering cell oxidative stress, thus aiding in potentially preventing cellular damage and promoting overall well-being, especially for treating erectile dysfunction (ED).

Components of the Nucleotide Salvage Pathway Increase Frog Virus 3 (FV3) Replication.

Viruses are obligate intracellular parasites that alter host metabolic machinery to obtain energy and macromolecules that are pivotal for replication. Ranavirus, including the type species of the genus frog virus 3 (FV3), represent an ecologically important group of viruses that infect fish, amphibians, and reptiles. It was established that fatty acid synthesis, glucose, and glutamine metabolism exert roles during iridovirus infections; however, no information exists regarding the role of purine metabolism. In this study, we assessed the impact of exogenously applied purines adenine, adenosine, adenosine 5'-monophosphate (AMP), inosine 5'-monophosphate (IMP), inosine, S-adenosyl-L-homocysteine (SAH), and S-adenosyl-L-methionine (SAM) on FV3 replication. We found that all compounds except for SAH increased FV3 replication in a dose-dependent manner. Of the purines investigated, adenine and adenosine produced the most robust response, increasing FV3 replication by 58% and 51%, respectively. While all compounds except SAH increased FV3 replication, only adenine increased plaque area. This suggests that the stimulatory effect of adenine on FV3 replication is mediated by a mechanism that is at least in part independent from the other compounds investigated. Our results are the first to report a response to exogenously applied purines and may provide insight into the importance of purine metabolism during iridoviral infection.

Abscisic Acid: A Potential Secreted Effector Synthesized by Phytophagous Insects for Host-Plant Manipulation.

Abscisic acid (ABA) is an isoprenoid-derived plant signaling molecule involved in a wide variety of plant processes, including facets of growth and development as well as responses to abiotic and biotic stress. ABA had previously been reported in a wide variety of animals, including insects and humans. We used high-performance liquid chromatography-electrospray ionization tandem mass spectrometry (HPLC-(ESI)-MS/MS) to examine concentrations of ABA in 17 species of phytophagous insects, including gall- and non-gall-inducing species from all insect orders with species known to induce plant galls: Thysanoptera, Hemiptera, Lepidoptera, Coleoptera, Diptera, and Hymenoptera. We found ABA in insect species in all six orders, in both gall-inducing and non-gall-inducing species, with no tendency for gall-inducing insects to have higher concentrations. The concentrations of ABA in insects often markedly exceeded those typically found in plants, suggesting it is highly improbable that insects obtain all their ABA from their host plant via consumption and sequestration. As a follow-up, we used immunohistochemistry to determine that ABA localizes to the salivary glands in the larvae of the gall-inducing Eurosta solidaginis (Diptera: Tephritidae). The high concentrations of ABA, combined with its localization to salivary glands, suggest that insects are synthesizing and secreting ABA to manipulate their host plants. The pervasiveness of ABA among both gall- and non-gall-inducing insects and our current knowledge of the role of ABA in plant processes suggest that insects are using ABA to manipulate source-sink mechanisms of nutrient allocation or to suppress host-plant defenses. ABA joins the triumvirate of phytohormones, along with cytokinins (CKs) and indole-3-acetic acid (IAA), that are abundant, widespread, and localized to glandular organs in insects and used to manipulate host plants.

Silicon ameliorates clubroot responses in canola (Brassica napus): A "multi-omics"-based investigation into possible mechanisms.

Clubroot disease, caused by Plasmodiophora brassicae Woronin, results in severe yield losses in Brassica crops, including canola. Silicon (Si) mitigates several stresses and enhances plant resistance to phytopathogens. We investigated the effects of Si on clubroot disease symptoms in canola at two concentrations of Si, Si: soil in 1: 100 w/w (Si1.0) and Si: soil in 1:200 w/w (Si0.5) under greenhouse conditions. In addition, the effects of Si on P. brassicae-induced gene expression, endogenous levels of phytohormones and metabolites were studied using "omics" approaches. Si application reduced clubroot symptoms and improved plant growth parameters. Gene expression analysis revealed increased transcript-level responses in Si1.0 compared to Si0.5 plants at 7-, 14-, and 21-days post-inoculation (dpi). Pathogen-induced transcript-level changes were affected by Si treatment, with genes related to antioxidant activity (e.g., POD, CAT), phytohormone biosynthesis and signalling (e.g., PDF1.2, NPR1, JAZ, IPT, TAA), nitrogen metabolism (e.g., NRT, AAT), and secondary metabolism (e.g., PAL, BCAT4) exhibiting differential expression. Endogenous levels of phytohormones (e.g., auxin, cytokinin), a majority of the amino acids and secondary metabolites (e.g., glucosinolates) were increased at 7 dpi, followed by a decrease at 14- and 21-dpi due to Si-treatment. Stress hormones such as abscisic acid (ABA), salicylic acid (SA), and jasmonic acid (JA) also decreased at the later time points in Si0.5, and Si1.0 treated plants. Si appears to improve clubroot symptoms while enhancing plant growth and associated metabolic processes, including nitrogen metabolism and secondary metabolite biosynthesis.

Phytohormones enhance heavy metal responses in Euglena gracilis: Evidence from uptake of Ni, Pb and Cd and linkages to hormonomic and metabolomic dynamics.

Over the last decade, significant effort has been made to understand phytohormonal functions (e.g., cytokinins (CKs) and abscisic acid (ABA)) in metal stress responses of higher plants and algae. Despite the potential for these phytohormones to improve industrial remediation by Euglena gracilis (Euglenophyceae), no such roles have been elucidated for this highly adaptive species and its response to heavy metals. This study demonstrates that toxic metals (nickel, lead, cadmium) modify hormonal activity profiles (i.e., CK forms and their concentrations) in E. gracilis. Furthermore, exogenous ABA or CK (tZ) enabled higher metal uptake efficiency (i.e., 9.35% in lead and 9.2% in cadmium uptake with CK) and alleviated metal toxicity through the regulation of endogenous CKs (i.e., total CK, isoprenoid CK) and gibberellin (GAs, GA1 and GA3) levels. These responses suggest that E. gracilis regulates multiple phytohormone signals during metal stress acclimation. A deeper approach, using untargeted metabolomic analyses, gave more detailed insight into phytohormone-controlled pathways and associated modified metabolites, which were frequently related to metal accumulation and the physiological acclimation to metal presence. Significant changes in the levels of cellular metabolites, especially those involved in acclimation to metal stress, were under the influence of phytohormones in algal cells. When grown under metal stress conditions, the presence of exogenous ABA or CKs, caused changes in cellular metabolites which included those from: lipid pathways, riboflavin metabolism, the biosynthesis of cofactors/vitamins, and carbohydrate metabolism. Also, bioactive secondary metabolites (e.g., terpenoids, alkaloids, flavonoids, carotenoids) were modified in algal cells treated with phytohormones. Thus, the study gives a detailed view on the regulatory functions of ABA and CKs in algal metal bioremediation strategies, which are attributed to enhanced metal uptake and in the fine-tuning of plant hormone levels during metal stress response. The results can guide efforts to develop efficient, low-cost and environmentally friendly methods for bioremediation.

A survey of Methylobacterium species and strains reveals widespread production and varying profiles of cytokinin phytohormones.

BACKGROUND: Symbiotic Methylobacterium strains comprise a significant part of plant microbiomes. Their presence enhances plant productivity and stress resistance, prompting classification of these strains as plant growth-promoting bacteria (PGPB). Methylobacteria can synthesize unusually high levels of plant hormones, called cytokinins (CKs), including the most active form, trans-Zeatin (tZ). RESULTS: This study provides a comprehensive inventory of 46 representatives of Methylobacterium genus with respect to phytohormone production in vitro, including 16 CK forms, abscisic acid (ABA) and indole-3-acetic acid (IAA). High performance-liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) analyses revealed varying abilities of Methylobacterium strains to secrete phytohormones that ranged from 5.09 to 191.47 pmol mL-1 for total CKs, and 0.46 to 82.16 pmol mL-1 for tZ. Results indicate that reduced methanol availability, the sole carbon source for bacteria in the medium, stimulates CK secretion by Methylobacterium. Additionally, select strains were able to transform L-tryptophan into IAA while no ABA production was detected. CONCLUSIONS: To better understand features of CKs in plants, this study uncovers CK profiles of Methylobacterium that are instrumental in microbe selection for effective biofertilizer formulations.

Customizing lipids from oleaginous microbes: leveraging exogenous and endogenous approaches.

To meet the growing demands of the oleochemical industry, tailored lipid sources are expanding to oleaginous microbes. To control the fatty acid composition of microbial lipids, ground-breaking exogenous and endogenous approaches are being developed. Exogenous approaches employ extracellular tools such as product-specific feedstocks, process optimization, elicitors, and magnetic and mechanical energy, whereas endogenous approaches leverage biology through the use of product-specific microbes, adaptive laboratory evolution (ALE), and the creation of custom strains via random and targeted cellular engineering. We consolidate recent advances from both fields into a review that will serve as a resource for those striving to fulfill the vision of microbial cell factories for tailored lipid production.

Beyond transport: cytokinin ribosides are translocated and active in regulating the development and environmental responses of plants.

MAIN CONCLUSION: Riboside type cytokinins are key components in cytokinin metabolism, transport, and sensitivity, making them important functional signals in plant growth and development and environmental stress responses. Cytokinin (CKs) are phytohormones that regulate multiple processes in plants and are critical for agronomy, as they are involved in seed filling and plant responses to biotic and abiotic stress. Among the over 30 identified CKs, there is uncertainty about the roles of many of the individual CK structural forms. Cytokinin free bases (CKFBs), have been studied in great detail, but, by comparison, roles of riboside-type CKs (CKRs) in CK metabolism and associated signaling pathways and their distal impacts on plant physiology remain largely unknown. Here, recent findings on CKR abundance, transport and localization, are summarized, and their importance in planta is discussed. The history of CKR analyses is reviewed, in the context of the determination of CK metabolic pathways, and research on CKR affinity for CK receptors, all of which yield essential insights into their functions. Recent studies suggest that CKR forms are a lot more than a group of transport CKs and, beyond this, they play important roles in plant development and responses to environmental stress. In this context, this review discusses the involvement of CKRs in plant development, and highlight the less anticipated functions of CKRs in abiotic stress tolerance. Based on this, possible mechanisms for CKR modes of action are proposed and experimental approaches to further uncover their roles and future biotechnological applications are suggested.

Cytokinins are pervasive among common in vitro culture media: An analysis of their forms, concentrations and potential sources.

The number of organisms that are known to produce cytokinins (CKs) continues to increase. In fact, species from all life kingdoms have now been shown to either produce CKs or at least have the genetic components to make it possible. In vitro growth of microorganisms, plant/animal cells, and tissue cultures often requires nutrient-rich media composed of ingredients with organic origins including: yeast extract, peptone, tryptone, or various plant or animal extracts. These compounds, derived from microbial, plant and animal materials, can be the source of significant levels of exogenous CKs in the culture medium. As CK investigative work continues to expand rapidly, it is of critical importance to draw attention to this complexity; the presence of CKs in growth medium affects CK metabolism of the cultured organism and interferes with the readings of analytical instrumentation used to profile CKs in tested microorganisms or cell cultures.

Isopentenyltransferases as master regulators of crop performance: their function, manipulation, and genetic potential for stress adaptation and yield improvement.

Isopentenyltransferase (IPT) in plants regulates a rate-limiting step of cytokinin (CTK) biosynthesis. IPTs are recognized as key regulators of CTK homeostasis and phytohormone crosstalk in both biotic and abiotic stress responses. Recent research has revealed the regulatory function of IPTs in gene expression and metabolite profiles including source-sink modifications, energy metabolism, nutrient allocation and storage, stress defence and signalling pathways, protein synthesis and transport, and membrane transport. This suggests that IPTs play a crucial role in plant growth and adaptation. In planta studies of IPT-driven modifications indicate that, at a physiological level, IPTs improve stay-green characteristics, delay senescence, reduce stress-induced oxidative damage and protect photosynthetic machinery. Subsequently, these improvements often manifest as enhanced or stabilized crop yields and this is especially apparent under environmental stress. These mechanisms merit consideration of the IPTs as 'master regulators' of core cellular metabolic pathways, thus adjusting plant homeostasis/adaptive responses to altered environmental stresses, to maximize yield potential. If their expression can be adequately controlled, both spatially and temporally, IPTs can be a key driver for seed yield. In this review, we give a comprehensive overview of recent findings on how IPTs influence plant stress physiology and yield, and we highlight areas for future research.

The Hulks and the Deadpools of the Cytokinin Universe: A Dual Strategy for Cytokinin Production, Translocation, and Signal Transduction.

Cytokinins are plant hormones, derivatives of adenine with a side chain at the N6-position. They are involved in many physiological processes. While the metabolism of trans-zeatin and isopentenyladenine, which are considered to be highly active cytokinins, has been extensively studied, there are others with less obvious functions, such as cis-zeatin, dihydrozeatin, and aromatic cytokinins, which have been comparatively neglected. To help explain this duality, we present a novel hypothesis metaphorically comparing various cytokinin forms, enzymes of CK metabolism, and their signalling and transporter functions to the comics superheroes Hulk and Deadpool. Hulk is a powerful but short-lived creation, whilst Deadpool presents a more subtle and enduring force. With this dual framework in mind, this review compares different cytokinin metabolites, and their biosynthesis, translocation, and sensing to illustrate the different mechanisms behind the two CK strategies. This is put together and applied to a plant developmental scale and, beyond plants, to interactions with organisms of other kingdoms, to highlight where future study can benefit the understanding of plant fitness and productivity.

Phytohormonal Roles in Plant Responses to Heavy Metal Stress: Implications for Using Macrophytes in Phytoremediation of Aquatic Ecosystems.

Heavy metals can represent a threat to the health of aquatic ecosystems. Unlike organic chemicals, heavy metals cannot be eliminated by natural processes such as their degradation into less toxic compounds, and this creates unique challenges for their remediation from soil, water, and air. Phytoremediation, defined as the use of plants for the removal of environmental contaminants, has many benefits compared to other pollution-reducing methods. Phytoremediation is simple, efficient, cost-effective, and environmentally friendly because it can be carried out at the polluted site, which simplifies logistics and minimizes exposure to humans and wildlife. Macrophytes represent a unique tool to remediate diverse environmental media because they can accumulate heavy metals from contaminated sediment via roots, from water via submerged leaves, and from air via emergent shoots. In this review, a synopsis is presented about how plants, especially macrophytes, respond to heavy metal stress; and we propose potential roles that phytohormones can play in the alleviation of metal toxicity in the aquatic environment. We focus on the uptake, translocation, and accumulation mechanisms of heavy metals in organs of macrophytes and give examples of how phytohormones interact with plant defense systems under heavy metal exposure. We advocate for a more in-depth understanding of these processes to inform more effective metal remediation techniques from metal-polluted water bodies. Environ Toxicol Chem 2021;40:7-22. © 2020 SETAC.

The Origins and Roles of Methylthiolated Cytokinins: Evidence From Among Life Kingdoms.

Cytokinins (CKs) are a group of adenine-derived, small signaling molecules of crucial importance for growth and multiple developmental processes in plants. Biological roles of classical CKs: isopentenyladenine (iP), trans and cis isomers of zeatin (tZ, cZ), and dihydrozeatin, have been studied extensively and their functions are well defined in many aspects of plant physiology. In parallel, extensive knowledge exists for genes involved in tRNA modifications that lead to the production of tRNA-bound methylthiolated CKs, especially in bacterial and mammalian systems. However, not much is known about the origins, fates, and possible functions of the unbound methylthiolated CKs (2MeS-CKs) in biological systems. 2MeS-CKs are the free base or riboside derivatives of iP or Z-type CKs, modified by the addition of a thiol group (-SH) at position 2 of the adenine ring that is subsequently methylated. Based on the evidence to date, these distinctive CK conjugates are derived exclusively via the tRNA degradation pathway. This review summarizes the knowledge on the probable steps involved in the biosynthesis of unbound 2MeS-CKs across diverse kingdoms of life. Furthermore, it provides examples of CK profiles of organisms from which the presence of 2MeS-CKs have been detected and confirms a close association and balance between the production of classical CKs and 2MeS-CKs. Finally, it discusses available reports regarding the possible physiological functions of 2MeS-CKs in different biological systems.

Cytokinin activity during early kernel development corresponds positively with yield potential and later stage ABA accumulation in field-grown wheat (Triticum aestivum L.).

MAIN CONCLUSION: Early cytokinin activity and late abscisic acid dynamics during wheat kernel development correspond to cultivars with higher yield potential. Cytokinins represent prime targets for marker development for wheat breeding programs. Two major phytohormone groups, abscisic acid (ABA) and cytokinins (CKs), are of crucial importance for seed development. Wheat (Triticum aestivum L.) yield is, to a high degree, determined during the milk and dough stages of kernel development. Therefore, understanding the hormonal regulation of these early growth stages is fundamental for crop-improvement programs of this important cereal. Here, we profiled ABA and 25 CK metabolites (including active forms, precursors and inactive conjugates) during kernel development in five field-grown wheat cultivars. The levels of ABA and profiles of CK forms varied greatly among the tested cultivars and kernel stages suggesting that several types of CK metabolites are involved in spatiotemporal regulation of kernel development. The seed yield potential was associated with the elevated levels of active CK levels (tZ, cZ). Interestingly, the increased kernel cZ levels were followed by higher ABA production, suggesting there is an interaction between these two phytohormones. Furthermore, we analyzed the expression patterns of representatives of the four main CK metabolic gene families. The unique transcriptional patterns of the IPT (biosynthesis) and ZOG (reversible inactivation) gene family members (GFMs) in the high and low yield cultivars additionally indicate that there is a significant association between CK metabolism and yield potential in wheat. Based on these results, we suggest that both CK metabolites and their associated genes, can serve as important, early markers of yield performance in modern wheat breeding programs.