A Novel Biostimulant, Belonging to Protein Hydrolysates, Mitigates Abiotic Stress Effects on Maize Seedlings Grown in Hydroponics

The main challenge to agriculture worldwide is feeding a rapidly growing human population, developing more sustainable agricultural practices that do not threaten human and ecosystem health. An innovative solution relies on the use of biostimulants, as a tool to enhance nutrient use ef\ufb01ciency and crop performances under sub-optimal conditions. In this work a novel biostimulant(APR\uae,ILSAS.p.A.,ArziganoVI,Italy), belongingtothegroupofproteinhydrolysates, wassuppliedtomaizeseedlingsinhydroponicanditseffectswereassessedincontrolconditionsand in the presence of three different kinds of stresses (hypoxia, salt and nutrient de\ufb01ciency) and of their combination. OurresultsindicatethatAPR\uae issolubleandisabletoin\ufb02uencerootandshootgrowth depending on its concentration. Furthermore, its effectiveness is clearly increased in condition of single or combination of abiotic stresses, thus con\ufb01rming the previously hypothesised action of this substance as enhancer of the response to environmental adversities. Moreover, it also regulates the transcription of a set of genes involved in nitrate transport and ROS metabolism. Further work will be needed to try to transfer this basic knowledge in \ufb01eld experiments

Identification and characterization of the BZR transcription factor family and its expression in response to abiotic stresses in Zea mays L.

Brassinosteroids (BRs) are plant specific steroidal hormones that play diverse roles in regulating a broad spectrum of plant growth and developmental processes, as well as, in responding to various biotic and abiotic stresses. Extensive research over the years has established stress-impact-mitigating role of BRs and associated compounds in different plants exposed to various abiotic and biotic stresses, suggesting the idea that they may act as immunomodulators, thus opening new approaches for plant resistance against hazardous environmental conditions. In this research the characterization of the transcriptional response of 11 transcription factors (TFs) belonging to BRASSINAZOLE-RESISTANT 1 (BZR1) TF family of Zea mays L. was analyzed in seedlings subjected to different stress conditions. Being important regulators of the BR synthesis, BZR TFs might have stress resistance related activities. However, no stress resistance related functional study of BZR TFs has been reported in maize so far. In silico analyses of the selected 11 TFs validated the features of their protein domains, where a highest degree of similarity observed with recognized BZR TFs of rice and Sorghum bicolor. Additionally, we investigated the organ-specific expression of 11 ZmBZR in maize seedlings. Five of them did not show any transcript accumulation, suggesting that ZmBZR expression might be regulated in a manner dependent on plant developmental stage. For the remaining six ZmBZR, their ubiquitous expression in the whole plant indicates they could function as growth regulators during maize development. More importantly, in response to various stress conditions, the spatial transcript accumulation of all ZmBZR varies along the plant. All six ZmBZR showed up-regulation against N starvation, hypoxia and salt stress. On the contrary, heat stress clearly down-regulated gene expression of all ZmBZR analysed. Consistently with the expression results, the distribution of stress-related cis-acting elements in the promoter of these genes inferred that the maize BZR TFs might play some roles in regulating the expression of the corresponding genes in response to multifarious stresses. In conclusion, these data reveal that BZR TFs have stress signaling activity in maize, in addition to their confirmed role in regulating plant physiology and morphology

Protein Profiling of Arabidopsis Roots Treated With Humic Substances: Insights Into the Metabolic and Interactome Networks

Background and Aim: Humic substances (HSs) influence the chemical and physical properties of the soil, and are also known to affect plant physiology and nutrient uptake. This study aimed to elucidate plant metabolic pathways and physiological processes influenced by HS activity. Methods: Arabidopsis roots were treated with HS for 8 h. Quantitative mass spectrometry-based proteomics analysis of root proteins was performed using the iTRAQ (Isobaric Tag for Relative and Absolute Quantification) technique. Out of 902 protein families identified and quantified for HS treated vs. untreated roots, 92 proteins had different relative content. Bioinformatic tools such as STRING, KEGG, IIS and Cytoscape were used to interpret the biological function, pathway analysis and visualization of network amongst the identified proteins. Results: From this analysis it was possible to evaluate that all of the identified proteins were functionally classified into several categories, mainly redox homeostasis, response to inorganic substances, energy metabolism, protein synthesis, cell trafficking, and division. Conclusion: In the present study an overview of the metabolic pathways most modified by HS biological activity is provided. Activation of enzymes of the glycolytic pathway and up regulation of ribosomal protein indicated a stimulation in energy metabolism and protein synthesis. Regulation of the enzymes involved in redox homeostasis suggest a pivotal role of reactive oxygen species in the signaling and modulation of HS-induced responses

Humic substances stimulate maize nitrogen assimilation and amino acid metabolism at physiological and molecular level

The effects of a humic substance (HS) extracted from a volcanic soil on the nitrate assimilation pathway of Zea mays seedlings were thoroughly examined using physiological and molecular approaches. Plant growth, the amount of soluble proteins and amino acids, as well as the activities of the enzymes involved in nitrogen metabolism and Krebs cycle, were evaluated in response to different HS concentrations (0, 1 and 5 mg C L−1) supplied to maize seedlings for 48 h. To better understand the HS action, the transcript accumulation of selected genes encoding enzymes involved in nitrogen assimilation and Krebs cycle was additionally evaluated in seedlings grown for 2 weeks under nitrogen (N) sufficient condition and N deprivation. HS at low concentration (1 mg C L−1) positively influenced nitrate metabolism by increasing the content of soluble protein and amino acids synthesis. Furthermore, the activity and transcription of enzymes functioning in N assimilation and Krebs were significantly stimulated. HS treatment influenced the gene expression of Zea mays plants at transcriptional level and this regulation was closely dependent on the availability of nitrate in the growth medium

The Control Of Zealactone Biosynthesis And Exudation Is Involved In The Response To Nitrogen In Maize Root.

Nitrate acts as a signal in regulating plant development in response to environment. In particular nitric oxide (NO), auxin and strigolactones (SLs) were supposed to cooperate to regulate the maize root response to this anion. In this study, a combined approach based on LC-MS/MS and on physiological and molecular analyses was adopted to specify the involvement of SLs in the maize response to N. Our results showed that N deficiency strongly induces SL exudation, likely through stimulating their biosynthesis. Nitrate provision early counteracts and also ammonium lowers SL exudation, but less markedly. Exudates obtained from N-starved and ammonium-provided seedlings stimulated Phelipanche germination, whereas when seeds were treated with exudates harvested from nitrate-provided plants no germination was observed. Furthermore, our findings support the idea that the inhibition of SL production observed in response to nitrate and ammonium would contribute to the regulation of lateral root development. Moreover, the transcriptional regulation of a gene encoding a putative maize WBC transporter, in response to various nitrogen supplies, together with its mRNA tissue localization, supported its role in SL allocation. Our results highlight the dual role of SLs as molecules able to signal outwards a nutritional need and as endogenous regulators of root architecture adjustments to N, thus synchronizing plant growth with nitrogen acquisitio

Nitrate sensing by the maize root apex transition zone: A merged transcriptomic and proteomic survey

Nitrate is an essential nutrient for plants, and crops depend on its availability for growth and development, but its presence in agricultural soils is far from stable. In order to overcome nitrate fluctuations in soil, plants have developed adaptive mechanisms allowing them to grow despite changes in external nitrate availability. Nitrate can act as both nutrient and signal, regulating global gene expression in plants, and the root tip has been proposed as the sensory organ. A set of genome-wide studies has demonstrated several nitrate-regulated genes in the roots of many plants, although only a few studies have been carried out on distinct root zones. To unravel new details of the transcriptomic and proteomic responses to nitrate availability in a major food crop, a double untargeted approach was conducted on a transition zone-enriched root portion of maize seedlings subjected to differing nitrate supplies. The results highlighted a complex transcriptomic and proteomic reprogramming that occurs in response to nitrate, emphasizing the role of this root zone in sensing and transducing nitrate signal. Our findings indicated a relationship of nitrate with biosynthesis and signalling of several phytohormones, such as auxin, strigolactones, and brassinosteroids. Moreover, the already hypothesized involvement of nitric oxide in the early response to nitrate was confirmed with the use of nitric oxide inhibitors. Our results also suggested that cytoskeleton activation and cell wall modification occurred in response to nitrate provision in the transition zone

mRNA-Sequencing Analysis Reveals Transcriptional Changes in Root of Maize Seedlings Treated with Two Increasing Concentrations of a New Biostimulant

Biostimulants are a wide range of natural or synthetic products containing substances and/or microorganisms that can stimulate plant processes to improve nutrient uptake, nutrient efficiency, tolerance to abiotic stress, and crop quality ( http://www.biostimulants.eu/ , accessed September 27, 2017). The use of biostimulants is proposed as an advanced solution to face the demand for sustainable agriculture by ensuring optimal crop performances and better resilience to environment changes. The proposed approach is to predict and characterize the function of natural compounds as biostimulants. In this research, plant growth assessments and transcriptomic approaches are combined to investigate and understand the specific mode(s) of action of APR, a new product provided by the ILSA group (Arzignano, Vicenza). Maize seedlings (B73) were kept in a climatic chamber and grown in a solid medium to test the effects of two different combinations of the protein hydrolysate APR (A1 and A1/2). Data on root growth evidenced a significant enhancement of the dry weight of both roots and root/shoot ratio in response to APR. Transcriptomic profiles of lateral roots of maize seedlings treated with two increasing concentrations of APR were studied by mRNA-sequencing analysis (RNA-seq). Pairwise comparisons of the RNA-seq data identified a total of 1006 differentially expressed genes between treated and control plants. The two APR concentrations were demonstrated to affect the expression of genes involved in both common and specific pathways. On the basis of the putative function of the isolated differentially expressed genes, APR has been proposed to enhance plant response to adverse environmental conditions

Flooding Responses on Grapevine: A Physiological, Transcriptional, and Metabolic Perspective

Studies on model plants have shown that temporary soil flooding exposes roots to a significant hypoxic stress resulting in metabolic re-programming, accumulation of toxic metabolites and hormonal imbalance. To date, physiological and transcriptional responses to flooding in grapevine are poorly characterized. To fill this gap, we aimed to gain insights into the transcriptional and metabolic changes induced by flooding on grapevine roots (K5BB rootstocks), on which cv Sauvignon blanc (Vitis vinifera L.) plants were grafted. A preliminary experiment under hydroponic conditions enabled the identification of transiently and steadily regulated hypoxia-responsive marker genes and drafting a model for response to oxygen deprivation in grapevine roots. Afterward, over two consecutive vegetative seasons, flooding was imposed to potted vines during the late dormancy period, to mimick the most frequent waterlogging events occurring in the field. Untargeted transcriptomic and metabolic profiling approaches were applied to investigate early responses of grapevine roots during exposure to hypoxia and subsequent recovery after stress removal. The initial hypoxic response was marked by a significant increase of the hypoxia-inducible metabolites ethanol, GABA, succinic acid and alanine which remained high also 1 week after recovery from flooding with the exception of ethanol that leveled off. Transcriptomic data supported the metabolic changes by indicating a substantial rearrangement of primary metabolic pathways through enhancement of the glycolytic and fermentative enzymes and of a subset of enzymes involved in the TCA cycle. GO and KEGG pathway analyses of differentially expressed genes showed a general down-regulation of brassinosteroid, auxin and gibberellin biosynthesis in waterlogged plants, suggesting a general inhibition of root growth and lateral expansion. During recovery, transcriptional activation of gibberellin biosynthetic genes and down-regulation of the metabolic ones may support a role for gibberellins in signaling grapevine rootstocks waterlogging metabolic and hormonal changes to the above ground plant. The significant internode elongation measured upon budbreak during recovery in plants that had experienced flooding supported this hypothesis. Overall integration of these data enabled us to draft a first comprehensive view of the molecular and metabolic pathways involved in grapevine\u2019s root responses highlighting a deep metabolic and transcriptomic reprogramming during and after exposure to waterlogging

Metabolic and molecular rearrangements of Sauvignon Blanc (Vitis vinifera L.) berries in response to foliar applications of specific dry yeast

Dry yeast extracts (DYE) are applied to vineyards to improve aromatic and secondary metabolic compound content and wine quality; however, systematic information on the underpinning molecular mechanisms is lacking. This work aimed to unravel, through a systematic approach, the metabolic and molecular responses of Sauvignon Blanc berries to DYE treatments. To accomplish this, DYE spraying was performed in a commercial vineyard for two consecutive years. Berries were sampled at several time points after the treatment, and grapes were analyzed for sugars, acidity, free and bound aroma precursors, amino acids, and targeted and untargeted RNA-Seq transcriptional profiles. The results obtained indicated that the DYE treatment did not interfere with the technological ripening parameters of sugars and acidity. Some aroma precursors, including cys-3MH and GSH3MH, responsible for the typical aromatic nuances of Sauvignon Blanc, were stimulated by the treatment during both vintages. The levels of amino acids and the global RNA-seq transcriptional profiles indicated that DYE spraying upregulated ROS homeostatic and thermotolerance genes, as well as ethylene and jasmonic acid biosynthetic genes, and activated abiotic and biotic stress responses. Overall, the data suggested that the DYE reduced berry oxidative stress through the regulation of specific subsets of metabolic and hormonal pathway

Flooding Responses on Grapevine: A Physiological, Transcriptional, and Metabolic Perspective

Studies on model plants have shown that temporary soil flooding exposes roots to a significant hypoxic stress resulting in metabolic re-programming, accumulation of toxic metabolites and hormonal imbalance. To date, physiological and transcriptional responses to flooding in grapevine are poorly characterized. To fill this gap, we aimed to gain insights into the transcriptional and metabolic changes induced by flooding on grapevine roots (K5BB rootstocks), on which cv Sauvignon blanc (Vitis vinifera L.) plants were grafted. A preliminary experiment under hydroponic conditions enabled the identification of transiently and steadily regulated hypoxia-responsive marker genes and drafting a model for response to oxygen deprivation in grapevine roots. Afterward, over two consecutive vegetative seasons, flooding was imposed to potted vines during the late dormancy period, to mimick the most frequent waterlogging events occurring in the field. Untargeted transcriptomic and metabolic profiling approaches were applied to investigate early responses of grapevine roots during exposure to hypoxia and subsequent recovery after stress removal. The initial hypoxic response was marked by a significant increase of the hypoxia-inducible metabolites ethanol, GABA, succinic acid and alanine which remained high also 1 week after recovery from flooding with the exception of ethanol that leveled off. Transcriptomic data supported the metabolic changes by indicating a substantial rearrangement of primary metabolic pathways through enhancement of the glycolytic and fermentative enzymes and of a subset of enzymes involved in the TCA cycle. GO and KEGG pathway analyses of differentially expressed genes showed a general down-regulation of brassinosteroid, auxin and gibberellin biosynthesis in waterlogged plants, suggesting a general inhibition of root growth and lateral expansion. During recovery, transcriptional activation of gibberellin biosynthetic genes and down-regulation of the metabolic ones may support a role for gibberellins in signaling grapevine rootstocks waterlogging metabolic and hormonal changes to the above ground plant. The significant internode elongation measured upon budbreak during recovery in plants that had experienced flooding supported this hypothesis. Overall integration of these data enabled us to draft a first comprehensive view of the molecular and metabolic pathways involved in grapevine’s root responses highlighting a deep metabolic and transcriptomic reprogramming during and after exposure to waterlogging