Seed priming to optimize germination in Arthrocnemum Moq.

BACKGROUND: Seed germination and seedling growth constitute the first stage of a plant's life cycle for crop establishment. Arthrocnemum Moq. is a halophyte of the subfamily Salicornioideae (Amaranthaceae), which could be recognized in the foreseeable future as an emerging candidate in applied biosaline agricultural programs, mainly due to the large biomass it represents in coastal and inland saltmarshes, in addition to its interesting nutritional and pharmacological properties. However, to ensure their subsequent use as a crop, it is necessary to optimize their germination through appropriate seed priming treatments. The main goal of this work was to seek the optimization of Arthrocnemum germination process using different pretreatments: exposure to sodium chloride (100 to 1200 mM) in the dark and its subsequent transferred to distilled water separately and together with the combination of pH (5, 7, 9), salinity (0, 100, 200 mM NaCl), and iron conditions (0, 200, 400 µM FeSO4). The experiments were tested on six samples of two different species: A. meridionale (from Tunisia) and A. macrostachyum (from Spain). RESULTS: Salinity priming of seeds for 15 days in darkness improved germination percentages by almost 25% at 600 mM NaCl, in both Tunisian and Spanish species. However, keeping seeds at different salt concentrations for 30 days produced higher improvement percentages at lower concentrations in A. meridionale (100-200 mM NaCl), while in A. macrostachyum the highest improvement percentages were obtained at 600 mM NaCl (percentage improvement of 47%). When the dark time period is reduced to 5 days at higher salt concentrations, the greater germination percentages were reached in all the samples at the concentration of 800 mM NaCl, increasing the improvement of germination between 17 and 50%. Finally, the conditions of pH = 7, pretreatment in darkness at 800 mM NaCl and 400 µM or iron, turned out to be an effective medium for seed germination. CONCLUSIONS: Therefore, before using Arthrocnemum seeds in applied biotechnological programs, a seed priming treatment based on prior exposure to high salt concentrations (600-1000 mM NaCl) is recommended in order to maximize germination percentages.

In-site and Ex-site Date Palm Exposure to Heavy Metals Involved Infra-Individual Biomarkers Upregulation.

As a tree of considerable importance in arid regions-date palm, Phoenix dactylifera L. survival in contaminated areas of Sfax city has drawn our attention. Leaf samples of the plants grown in the study area showed high levels of cadmium (Cd), copper (Cu), and chromium (Cr). On the basis of this finding, the cellular mechanisms that explain these metal accumulations were investigated in controlled conditions. After four months of exposure to Cd, Cr, or Cu, high bioconcentration and translocation factor (TF>1) have been shown for date palm plantlets exposed to Cd and low TF values were obtained for plantlets treated with Cr and Cu. Moreover, accumulation of oxidants and antioxidant enzyme activities occurred in exposed roots to Cu and Cd. Secondary metabolites, such as polyphenols and flavonoids, were enhanced in plants exposed at low metal concentrations and declined thereafter. Accumulation of flavonoids in cells may be correlated with the expression of the gene encoding Pdmate5, responsible for the transport of secondary metabolites, especially flavonoids. Other transporter genes responded positively to metal incorporation, especially Pdhma2, but also Pdabcc and Pdnramp6. The latter would be a new candidate gene sensitive to metallic stress in plants. Expressions of gene coding metal chelators were also investigated. Pdpcs1 and Pdmt3 exhibited a strong induction in plants exposed to Cr. These modifications of the expression of some biochemical and molecular based-markers in date palm helped to better understand the ability of the plant to tolerate metals. They could be useful in assessing heavy metal contaminations in polluted soils and may improve accumulation capacity of other plants.

Transcriptome assembly and abiotic related gene expression analysis of date palm reveal candidate genes involved in response to cadmium stress.

Date palm in Tunisia is of major economic importance but are also factors of social, environmental and economic stability. An annotated assembly of the transcriptome of cultivar Deglet Nour was reported. RNA was isolated from plant Cd-contaminated leaves, and 37,049 unique Illumina RNA-seq reads were used in a transcriptome assembly. The draft transcriptome assembly consists of 6789 contigs and 17.285 singletons with a means length of 858 bp and 1.042 bp, respectively. The final assembly was functionally annotated using Blast2GO software, allowing the identification of putative genes controlling important agronomic traits. The annotated transcriptome data sets were used to query all known Kyoto Encyclopedia of Genes and Genomes pathways. The most represented molecular functions and biological processes were nucleotide binding and transcription, transport and response to stress and abiotic and biotic stimuli. A prediction of the genes interaction network was proposed by selecting corresponding functionally similar genes from Arabidopsis datasets, downloaded by GeneMANIA version 2.1. Several Cd-responsive genes expression was monitored in in vitro isolated explant of Cd stressed Deglet Nour. Some chelators encoding genes were upregulated confirming in silico findings. Genes encoding HMs transporters in date palm showed expression enhancement more pronounced after 20 days of exposure. P. dactylifera transcriptome provides a valuable resource for future functional analysis of candidate genes involved in metal stress response.

Bioaccumulation, antioxidative response, and metallothionein expression in Lupinus luteus L. exposed to heavy metals and silver nanoparticles.

Yellow-lupin (Lupinus luteus L.) was grown on soils contaminated with heavy metals during two parallel studies. In the first one, the soil was contaminated by industrial activities whereas, in the second one, the soil was artificially contaminated with a single metal including Cd, Pb, Zn, Ni (in nitrate form), and Ag (in nitrate and nanoparticles form). The study was performed to assess a plant's response to contamination including its antioxidative response and molecular mechanisms involved in metal detoxification through the expression level of metallothioneins (MTs). Overall, the study provided insights into identification and validation of housekeeping genes (HKG) in L. luteus under exposure to metal stress and showed the effects of selected heavy metals and silver nanoparticles on the expression of metallothioneins, the activity of guaiacol peroxidase (GPX) and bioaccumulation of metals in leaves of L. luteus. As such, HKG validation using BestKeeper, NormFinder, and geNorm software allowed for the selection of four most stable reference genes in a context metal contamination for the selected plant. Moreover, a significant increase in the expression levels of MT was observed in plants grown under heavy metal stress and none on plants grown on 25 mg kg-1 of silver nanoparticles. Also, the GPX activity and MT expression showed statistically significant changes between different conditions and doses which means that they can be used as highly sensitive stress markers for planning the phytoremediation process on a large scale.

Insight into the expression variation of metal-responsive genes in the seedling of date palm (Phoenix dactylifera).

Phytochelatin synthase and metallothionein gene expressions were monitored via qPCR in order to investigate the molecular mechanisms involved in Cd and Cr detoxification in date palm (Phoenix dactylifera). A specific reference gene validation procedure using BestKeeper, NormFinder and geNorm programs allowed selection of the three most stable reference genes in a context of Cd or Cr contamination among six reference gene candidates, namely elongation factor α1, actin, aldehyde dehydrogenase, SAND family, tubulin 6 and TaTa box binding protein. Phytochelatin synthase (pcs) and metallothionein (mt) encoding gene expression were induced from the first days of exposure. At low Cd stress (0.02 mM), genes were still up-regulated until 60th day of exposure. At the highest metal concentrations, however, pcs and mt gene expressions decreased. pcs encoding gene was significantly up-regulated under Cr exposure, and was more responsive to increasing Cr concentration than mt encoding gene. Moreover, exposure to Cd or Cr influenced clearly seed germination and hypocotyls elongation. Thus, the results have proved that both analyzed genes participate in metal detoxification and their expression is regulated at transcriptional level in date palm subjected to Cr and Cd stress. Consequently, variations of expression of mt and pcs genes may serve as early-warning biomarkers of metal stress in this species.

Copper toxicity and date palm (Phoenix dactylifera) seedling tolerance: Monitoring of related biomarkers.

Date palm (Phoenix dactylifera) seeds were exposed to different copper (Cu) solutions to examine plant stress responses. Low Cu concentrations (0.02 and 0.2 mM) caused an increase of seed germination, whereas higher Cu amounts (2 mM) significantly inhibited seed germination, delayed hypocotyl elongation, increased seedling mortality, and reduced the germination index by more than 90%. Metal-related toxicity symptoms appeared after 15 d of 2 mM of Cu exposure. Biochemical activities such as amylase activity and redox balance elements were examined to study the relationship between external Cu amount and internal plant response. The present study showed that amylolytic activity was dose- and time-dependent. Likewise, H2 O2 production increased after exposure to Cu, which was correlated with thiobarbituric acid reactive substance (TBARS) accumulation. Furthermore at low Cu concentrations, superoxide dismutase (SOD) and catalase (CAT) activities increased, suggesting that date palm seed stimulated its metal homeostasis networks. However, the highest cupric ion amounts increased cell oxidant accumulation and reduced enzyme production. Gene expression level measures of P. dactylifera phytochelatin synthase (Pdpcs) and P. dactylifera metallothionein (Pdmt) encoding genes have been carried out to investigate the implication of PdPCS and PdMT proteins in Cu homeostasis and/or its sequestration. Phoenix dactylifera metallothionein induction reached a peak after 30 d of exposure to 0.2 mM of Cu. However, it was down-regulated in plants exposed to higher Cu concentrations. In the same conditions, Pdpcs was overexpressed during 1 mo of exposure before it decreased thereafter. These observations provide a new insight into date palm cell response to Cu, a metal that can be toxic but that is also an essential element. Environ Toxicol Chem 2018;37:797-806. © 2017 SETAC.

In silico characterization and Molecular modeling of double-strand break repair protein MRE11 from Phoenix dactylifera v deglet nour.

BACKGROUND: DNA double-strand breaks (DSBs) are highly cytotoxic and mutagenic. MRE11 plays an essential role in repairing DNA by cleaving broken ends through its 3' to 5' exonuclease and single-stranded DNA endonuclease activities. METHODS: The present study aimed to in silico characterization and molecular modeling of MRE11 from Phoenix dactylifera L cv deglet nour (DnMRE11) by various bioinformatic approaches. To identify DnMRE11 cDNA, assembled contigs from our cDNA libraries were analysed using the Blast2GO2.8 program. RESULTS: The DnMRE11 protein length was 726 amino acids. The results of HUMMER show that DnMRE11 is formed by three domains: the N-terminal core domain containing the nuclease and capping domains, the C-terminal half containing the DNA binding and coiled coil region. The structure of DnMRE11 is predicted using the Swiss-Model server, which contains the nuclease and capping domains. The obtained model was verified with the structure validation programs such as ProSA and QMEAN servers for reliability. Ligand binding studies using COACH indicated the interaction of DnMRE11 protein with two Mn(2+) ions and dAMP. The ConSurf server predicted that residues of the active site and Nbs binding site have high conservation scores between plant species. CONCLUSIONS: A model structure of DnMRE11 was constructed and validated with various bioinformatics programs which suggested the predicted model to be satisfactory. Further validation studies were conducted by COACH analysis for active site ligand prediction, and revealed the presence of six ligands binding sites and two ligands (2 Mn(2+) and dAMP).

Morphological and biochemical behavior of fenugreek (Trigonella foenum-graecum) under copper stress.

The effects of copper on germination and growth of fenugreek (Trigonella foenum-graecum) was investigated separately using different concentrations of CuSO4. The germination percentage and radical length had different responses to cupric ions: the root growth increased with increasing copper concentration up to 1 mM Cu²âº and was inhibited thereafter. In contrast, the germination percentage was largely unaffected by concentrations of copper below 10 mM. The reduction in root growth may have been due to inhibition of hydrolytic enzymes such as amylase. Indeed, the average total amylolytic activity decreased from the first day of treatment with [Cu²âº] greater than 1 mM. Furthermore, copper affected various plant growth parameters. Copper accumulation was markedly higher in roots as compared to shoots. While both showed a gradual decrease in growth, this was more pronounced in roots than in leaves and in stems. Excess copper induced an increase in the rate of hydrogen peroxide (H2O2) production and lipid peroxidation in all plant parts, indicating oxidative stress. This redox stress affected leaf chlorophyll and carotenoid content which decreased in response to augmented Cu levels. Additionally, the activities of proteins involved in reactive oxygen species (ROS) detoxification were affected. Cu stress elevated the ascorbate peroxidase (APX) activity more than two times at 10 mM CuSO4. In contrast, superoxide dismutase (SOD) and catalase (CAT) levels showed only minor variations, only at 1 mM Cu²âº. Likewise, total phenol and flavonoid contents were strongly induced by low concentrations of copper, consistent with the role of these potent antioxidants in scavenging ROS such as H2O2, but returned to control levels or below at high [Cu²âº]. Taken together, these results indicate a fundamental shift in the plant response to copper toxicity at low versus high concentrations.