EIN2 and COI1 control the antagonism between ethylene and jasmonate in adventitious rooting of Arabidopsis thaliana thin cell layers

Auxins induce adventitious roots (ARs) in numerous culture-systems, and indole-3-butyric acid (IBA) is frequently the best AR-inducer. Vitamin requirements vary according to species, explant, and culture-conditions. Arabidopsis thaliana thin cell layers (AtTCLs) are uncapable of AR-formation on hormone-free medium containing thiamine and myo-inositol, whereas ARs are induced when IBA (10 μM), with/without kinetin (Kin, 0.1 μM), is added. The research frst aim was to determine whether a synergism between IBA and myo-inositol and thiamine was necessary for AR-formation. Results showed that IBA induced AR-formation without myo-inositol and thiamine, but better when both vitamins were also present. Deciphering hormonal action on AR formation under optimal vitamin content would be essential for improving the AR process. Ethylene (ET)/jasmonic acid (JA) signaling cross-talk has been demonstrated as being involved in AR-formation in IBA+Kincultured AtTCLs, by using ein3eil1 and coi1-16 mutants. ETHYLENE INSENSITIVE3 (EIN3)/EIN3-LIKE1 (EIL1) are positive regulators of ethylene (ET)-signaling, whereas CORONATINE INSENSITIVE1 (COI1) is involved in JA-signaling. The ETHYLENE INSENSITIVE2 (EIN2) protein activates EIN3/EIL1 in ET-presence. To understand whether EIN2 was also involved, the AR-response of ein2-1 and coi1-16 TCLs was evaluated adding the ET-precursor 1-aminocyclopropane1-carboxylic acid (ACC, 0.1 μM) and/or the JA-donor methyl jasmonate (JAMe, 0.01 μM) to IBA+vitamins-containing medium. AR-formation was enhanced by JAMe, reduced by ACC, but unchanged by JAMe+ACC in the wild type TCLs, whereas remained similarly low in ein2-1 and coi1-16 under all treatments. Collectively, these results demonstrate that the antagonism between JA and ET in AR-formation from AtTCLs involves a cross-talk by EIN2 and COI1

Indole-3-butyric acid induces ectopic formation of metaxylem in the hypocotyl of Arabidopsis thaliana without conversion into indole-3-acetic acid and with a positive interaction with ethylene

The role of the auxins indole-3-acetic acid (IAA) and indole-3-butyric acid (IBA) and of the auxin-interacting phytohormone ethylene, on the ectopic formation of primary xylem (xylogenesis in planta) is still little known. In particular, auxin/ethylene-target tissue(s), modality of the xylary process (trans-differentiation vs. de novo formation), and the kind of ectopic elements formed (metaxylem vs. protoxylem) are currently unknown. It is also unclear whether IBA may act on the process independently of conversion into IAA. To investigate these topics, histological analyses were carried out in the hypocotyls of Arabidopsis wild type seedlings and ech2ibr10 and ein3eil1 mutants, which are blocked in IBA-to-IAA conversion and ethylene signalling, respectively. The seedlings were grown under darkness with either IAA or IBA, combined or not with the ethylene precursor 1-aminocyclopropane-1-carboxylic acid. Adventitious root formation was also investigated because this process may compete with xylogenesis. Our results show that ectopic formation of protoxylem and metaxylem occurred as an indirect process starting from the pericycle periclinal derivatives of the hypocotyl basal part. IAA favoured protoxylem formation, whereas IBA induced ectopic metaxylem with ethylene cooperation through the EIN3EIL1 network. Ectopic metaxylem differentiation occurred independently of IBA-to-IAA conversion as mediated by ECH2 and IBR10, and in the place of IBA-induced adventitious root formation

Jasmonate and nitric oxide roles in the control of xylary cell formation and identity in Arabidopsis seedlings

In basal hypocotyls of dark-grown Arabidopsis seedlings, xylary cells may form from the pericycle as an alternative to another developmental program, i.e. adventitious roots. It is known that several hormones may induce xylogenesis, as jasmonic acid (JA), indole-3-acetic acid (IAA) and indole-3-butyric acid (IBA), which also affect xylary cell identity. Recent studies with the ethylene (ET)-perception mutant ein3eil1 and the ET-precursor 1-aminocyclopropane-1-carboxylic acid (ACC) have shown ET involvement in IBA induced ectopic metaxylem. Nitric oxide (NO) is a reactive free radical molecule, which acts as a messenger in several cell differentiation events, including programmed cell death, moreover it can be produced after IBA/IAA-treatments influencing JA signalling and interacting positively/negatively with ET. To date, NO involvement in ET/JA-mediated xylogenesis has never been investigated.The aim of the present research was to determine the involvement of JA, ET and NO in the control of endogenous/exogenous auxin-induced xylogenesis through a possible crosstalk mediated by EIN3/EIL1. To this aim, ectopic xylem formation was investigated in the hypocotyl of dark-grown Arabidopsis seedlings exposed to various concentrations of JA methyl-ester (JAMe) with/without ACC, IBA or IAA. The xylogenic response in the wild-type (wt) was compared with that of the ein3eil1 mutant, the NO signal was quantified and the its role evaluated by measuring the effects of treatments with a NO donor/scavenger (SNP/cPTIO). Results show that the ectopic formation of protoxylem was enhanced in the wt by JAMe when applied alone at a specific concentration (i.e. 10μM), whereas in ein3eil1 mutant it occurred with any JAMe concentration (i.e. 0.01, 1 and 10 μM). This stimulation of xylary elements mediated by JAMe suggests that a negative interaction between JA and ET-signalling is involved in this developmental program. The negative interaction was confirmed by the reduction in xylogenesis observed in the wt after the combined application of JAMe with ACC, in comparison with JAMe alone. Nitric oxide was detected at early stages of both xylogenesis and adventitious rooting in the hypocotyl pericycle cells and its production was highly enhanced by JAMe at the highest concentration, combined or not with IBA (10 μM). Histological analyses showed that the xylary identity changed when JAMe was applied with each auxin in comparison with treatments with auxin alone. In addition, the IBA/IAA-induced adventitious rooting was increased by the same JAMe concentration enhancing xylogenesis when applied alone. This suggests a role for JA in modulating both developmental programs (adventitious rooting and xylogenesis) in the same target cells (hypocotyl pericycle cells), through an interaction with NO, as summarized in the model proposed (Fig. 1)

Sulphur fertilization influences the sulphur species composition in Allium sativum : sulphomics using HPLC-ICPMS/MS-ESI-MS/MS

We thank Agilent, UK for access to the Agilent 6200 series TOF/6500 series Q-TOF. M.R. especially thanks the ERASMUS programme and G.Falasca for support.Peer reviewedPostprin

A printed luminescent flier inspired by plant seeds for eco-friendly physical sensing

Continuous and distributed monitoring of environmental parameters may pave the way for developing sustainable strategies to tackle climate challenges. State-of-the-art technologies, made with electronic systems, are often costly, heavy, and generate e-waste. Here, we propose a new generation of self-deployable, biocompatible, and luminescent artificial flying seeds for wireless, optical, and eco-friendly monitoring of environmental parameters (i.e., temperature). Inspired by natural Acer campestre plant seeds, we developed three-dimensional functional printed luminescent seed–like fliers, selecting polylactic acid as a biocompatible matrix and temperature as a physical parameter to be monitored. The artificial seeds mimic the aerodynamic and wind dispersal performance of the natural ones. The sensing properties are given by the integration of fluorescent lanthanide–doped particles, whose photoluminescence properties depend on temperature. The luminescent artificial flying seeds can be optically read from a distance using eye-safe near-infrared wavelengths, thus acting as a deployable sensor for distributed monitoring of topsoil environmental temperatures

Influence of sulphur fertilization on speciation of small water-soluble sulphur compounds and selenium uptake in garlic

Garlic is known for its high sulphur content and for its ability to metabolize selenium. We studied the influence of sulphur fertilisation (0.1, 0.5, 2mM SO4 2 ) in hydroponically grown garlic on Se uptake (7.8 mg Se/L for 24 h) and the influence on small water-soluble S and Se species. Beside total S and Se in root and bulb, acid soluble S and Se species were identified and quantified. Roots of control plants contained more sulphur than Se treated plants, showing that short-term exposure to Se has generally a negative impact on S-content in roots. Se uptake by roots was highest in plants grown at low S-fertilisation. Bulbs on the other hand showed the opposite trend, with high S-fertilisation plants having higher Se accumulation. The main Se-species in roots were N-acetyl-selenocystathionine and Se-Met. Bulbs contained generally less acid-extractable Se than roots and this was present mostly in form of Se-Met.This experiment shows that the metabolism of small acid-extractable Se and S compounds is interlinked. Considering garlic as nutritional supplement to increase Se-uptake it seems that the application of increased S-fertilisation in addition to the exposure to inorganic Se would improve the transport of Se into the edible bulb and therefore produce a more valuable crop

The quiescent center and the stem cell niche in the adventitious roots of Arabidopsis thaliana

Adventitious rooting is essential for the survival of numerous species from vascular cryptogams to monocots, and is required for successful micropropagation. The tissues involved in AR initiation may differ in planta and in in vitro systems. For example, in Arabidopsis thaliana, ARs originate from the hypocotyl pericycle in planta and the stem endodermis in in vitro cultured thin cell layers. The formation of adventitious roots (ARs) depends on numerous factors, among which the hormones, auxin, in particular. In both primary and lateral roots, growth depends on a functional stem cell niche in the apex, maintained by an active quiescent center (QC), and involving the expression of genes controlled by auxin and cytokinin. This review summarizes current knowledge about auxin and cytokinin control on genes involved in the definition and maintenance of QC, and stem cell niche, in the apex of Arabidopsis ARs in planta and in longitudinal thin cell layers

Cadmium and Arsenic alter auxin homeostasis during adventitious root formation in Arabidopsis thaliana L. (Heynh)

The metalloid arsenic (As) and the heavy metal cadmium (Cd) are environmental pollutants with a wide-spread and persistent presence in all ecosystems. Cadmium toxicity affects plant development, in particular altering root growth and differentiation (Brunetti et al., 2011, J Experiment Bot 62:5509; Zanella et al., 2016, Planta 243:605). Arsenic is a metalloid present in the environment in inorganic and organic forms. The inorganic ones are more toxic. It was reported that Cd and As mainly localize in the root meristems (Feng et al, 2013, Environ Sci Pollut Res 20:5449; Ko-pittke et al, 2012, Plant Physiol 159:1149). Arabidopsis thaliana (L.) Heynh exhibits a root system composed of primary (PR), lateral (LRs) and adventitious roots (ARs). Indole-3-acetic acid (IAA) is the main auxin in plants, and its homeostasis is regulated by an integrated and coordinated action between synthesis and polar transport, both essential for a proper root formation and development (Blilou et al. 2005, Nature 433:39; Della Rovere et al.,2013, Ann Bot 112:1395). YUCCA6 gene is involved in the tryptophan-dependent IAA biosynthesis, with an important role in root formation and development (Kim et al., 2007, Plant Physiol 145:722). Influx and efflux membrane proteins regulate IAA shoot-to-root polar transport. Among them, key roles in root development have been described for PIN1, member of the PIN-FORMED fami-ly of auxin efflux carriers (Adamowski and Friml 2015, Plant Cell 27:20) and for LAX3, member of the AUXIN1/LIKE-AUX1 (AUX/LAX) family of auxin influx carriers (Swarup et al., 2008, Nat Cell Bio 10:946). The coordinated auxin efflux/influx activities, generating the IAA gradients and maxima, are required for LR and AR initiation and development. Stress caused by toxic met-als alters growth and development by interfering with auxin levels and homeostasis (Potters et al., 2009 Plant Cell Environ 32:158; Sofo et al., 2013, Physiol Plant 149:487). However, less is known about the combined effect of Cd and As on IAA biosynthesis, levels and transport in ARs and LRs. The study’s aim was to determine if Cd and/or As affected root formation/development by alter-ing IAA biosynthesis and/or transport. To the aim, the expression pattern of the quiescent centre (QC) marker QC25::GUS, YUCCA6 transcript levels, the expression patterns of PIN1 and LAX3, detection of IAA levels, and auxin localization monitored by the DR5::GUS system, were investi-gated in Arabidopsis seedlings exposed to Cd and/or As. The results show that Cd and/or As significantly inhibited PR and hypocotyl growth. The mean density of LRs and ARs were significantly increased in the seedlings exposed to either Cd alone or to both pollutants, while root density was reduced by As alone. Cd and As disrupted QC formation, and stem cell niche maintenance over time in both LRs and ARs. Cadmium in-creased total auxin levels through an overexpression of YUCCA6. The toxic elements altered the expression pattern of PIN1 and LAX3, negatively affecting IAA accumulation in ARs and LRs. In conclusion, our results show that the most severe damages that Cd and/or As cause in Arabidopsis root system are due to a strong alteration of auxin biosynthesis, transport and accumulation in LRs and ARs

Exogenous nitric oxide enhances Cd tolerance in the rice root system by interacting with auxin

Oryza sativa L. is a worldwide food-crop frequently growing in cadmium (Cd) polluted soils. High Cd concentrations alter plant development and, in particular, the root-system, both by affecting auxin metabolism and by triggering reactive oxygen/nitrogen species (ROS/RNS), thereby affecting rice yield. In addition, Cd2+ easily enters in the rice root cells through passive transport, reaching the grains after xylem-tophloem transfer, thus becoming a threat to food security. Nitrogen monoxide (nitric oxide – NO) is a ubiquitous gaseous molecule involved in numerous animal and plant physiological processes, and it is also a mediator of plant development and of abiotic/biotic stresses response. Various reports highlight that NO has an important role in alleviating heavy metal toxicity and reducing the oxidative damages in plant organs either by enhancing the activity of antioxidant enzymes or by directly scavenging ROS. On the other hand, heavy metal-induced accumulation of NO was reported to be responsible for heavy metal toxicity. Indeed, NO can act either as a stress-inducing agent or as a protective molecule depending on its concentration, the plant tissue or age, and the type/severity of stress. At optimal levels, NO interacts with auxins [both indole-3 butyric acid (IBA) and indole-3 acetic acid (IAA)] during root growth and development. An auxin-induced NO production during many plant root responses has been suggested trough the modulation of the activity of enzymes involved in NO biosynthesis, while studies carried out with exogenous application of NO-specific donor compounds (i.e. sodium-nitroprusside, SNP) have demonstrated the involvement of the signal molecule in auxin metabolism, transport and signalling. However, the complex mechanisms underlying the interaction between NO and auxin during the metal stress is still poorly understood and need to be better investigated, together with further elucidations about the multifaceted role of NO (i.e. as a mitigating or a stressor agent) during Cd toxicity. To this aim, the effects of Cd toxicity on rice root anatomy/morphology and on H2O2 and O2●ˉ production, and the possible recovery by NO, was evaluated after 100μM Cd exposure, combined or not with SNP at 50μM. Moreover, endogenous IAA/IBA contents, transcription-levels of OsYUCCA1 and OsASA2 IAA-biosynthetic-genes, and expression of the IAA-responsive DR5::GUS construct were analysed, and the NO-epifluorescence levels measured. Our results show that exogenous treatments with the NO-donor SNP increase intracellular root NO levels in in vitro grown rice seedlings not exposed to Cd and restore the NO-levels reduced by the heavy metal. In addition, SNP treatments mitigate both the increase in the HPLC-measured root IAA levels and the alteration of its distribution monitored by the DR5::GUS system due to the toxic metal exposure. Notably, treatments with Cd alone or combined with SNP reduced YUCCA1 expression compared to the Control, while no effects were detected on ASA2, suggesting no involvement of the two IAA biosynthetic genes in the Cd-related increase of the IAA levels detected. Finally, the enhanced cellular NO-content alleviates the Cd-induced root morphological and histological damages and the root H2O2 and O2●ˉ overproduction. Moreover, exogenous NO decreases the heavy-metal uptake. All together our data highlight the beneficial effects of the NO in alleviating Cd toxicity in rice

Arabidopsis root formation is altered by cadmium and arsenic

The semimetal arsenic (As) and the heavy metal cadmium (Cd) are highly toxic for plants and animals, evoking enormous concern due to their widespread and persistent presence in polluted ecosystems. Both elements are not essential for plants but easily absorbed by their roots using the same membrane transporters of essential nutrients. The exposure to Cd or As causes inhibition of plant growth, especially in sensitive plants as Arabidopsis thaliana, the model species used in this research. It was reported that Cd and As mainly localize in root meristems. The correct organization and functionality of primary (PR), lateral (LR) and adventitious roots (AR) depends on the integrity of their apical meristem, on the correct activity and maintenance over time of a small group of cells which rarely divide, i.e. the quiescent centre (QC) cells. The QC inhibits the differentiation of the surrounding stem cells, allowing the apical root growth and the correct root differentiation. In A. thaliana LR and AR originate from pericycle founder cells in the PR and hypocotyl, respectively, their QC is established in a precise stage of primordium development. It was demonstrated that the positioning and maintenance of the QC in these roots is strictly related to a correct transport and biosynthesis of indole-3-acetic acid (IAA), the main plant auxin. To the aim to investigate the effect of Cd and As on auxin-mediated LR and AR development and QC maintenance, the expression of the IAA-sensitive DR5: GUS, of QC25: GUS (QC-marker), of the auxin biosynthetic gene YUCCA6, of the IAA carriers GUS-lines PIN1: GUS and LAX3: GUS and IAA levels in seedlings exposed to Na2HAsO47H2O and/or CdSO4 were evaluated. Results indicate that Cd and As alter auxin biosynthesis and transport during root formation, with consequent negative effects on their growth