Response to Cadmium Toxicity: Orchestration of Polyamines and microRNAs in Maize Plant.

Cadmium (Cd) is a heavy metal that is widely contaminating the environment due to its uses in industries as corrosive reagents, paints, batteries, etc. Cd can easily be absorbed through plant roots and may have serious negative impacts on plant growth. To investigate the mechanisms utilized by plants to cope with Cd toxicity, an experiment was conducted on maize seedlings. We observed that the plant growth and photosynthetic mechanism were negatively influenced during 20 days of Cd stress. The expression levels of ornithine decarboxylase (ORDC) increased in the six seedlings under Cd exposure compared to the control. However, Cd toxicity led to an increase in putrescine (Put) content only on day 15 when compared to the control plants. In fact, with the exception of day 15, the increases in the ORDC transcript levels did not show a direct correlation with the observed increases in Put content. Spermidine and Spermine levels were reduced on day 6 by Cd application, which was parallel with suppressed Spermidine synthase gene. However, an increase in Spermidine and Spermine levels was observed on day 12 along with a significant elevation in Spermidine synthase expression. On day 6, Cd was observed to start accumulating in the root with an increase in the expression of microRNA 528; while on day 15, Cd started to be observed in the shoot part with an increase in microRNA 390 and microRNA 168. These results imply that different miRNAs may regulate polyamines (PAs) in maize under Cd toxicity, suggesting a plant-derived strategy to commit a PAs/miRNA-regulated mechanism/s in different developmental stages (time points) in response to Cd exposure.

The regulatory role of γ-aminobutyric acid in chickpea plants depends on drought tolerance and water scarcity level.

γ-Aminobutyric acid (GABA) is a non-protein amino acid with multifunctional roles in dynamic plant responses. To determine the effects of exogenous GABA application (0, 25 and 50 µM) on drought response, two chickpea cultivars with contrasting tolerance to water deficit were examined. Plants were exposed to four irrigation levels (irrigation to 100, 60, 40 and 20% field capacity). Water deficit decreased growth, chlorophyll content, and photosynthetic efficiency. It increased electrolyte leakage and lipid peroxidation owing to both higher ROS accumulation and lower antioxidant enzyme activity. These negative effects of water deficit and the alleviating role of GABA application were more prominent in the sensitive, as compared to the tolerant cultivar. Water deficit also increased proline and GABA contents more in the tolerant cultivar, whereas their content was more enhanced by GABA application in the sensitive one. This may confer an additional level of regulation that results in better alleviation of drought damage in tolerant chickpea cultivars. In conclusion, the stimulatory effect of GABA on growth and physiological modulation depends on both the water stress severity and the cultivar sensitivity to it, implying a probable unknown GABA-related mechanism established by tolerant chickpea cultivars; a lost or not gained mechanism in susceptible ones.

Transgenic tobacco co-expressing flavodoxin and betaine aldehyde dehydrogenase confers cadmium tolerance through boosting antioxidant capacity.

Excessive heavy metal (HM) levels in soil have become a source of concern due to their adverse effects on human health and the agriculture industry. Soil contamination by HMs leads to an accumulation of reactive oxygen species (ROSs) within the plant cell and disruption of photosynthesis-related proteins. The response of tobacco lines overexpressing flavodoxin (Fld) and betaine aldehyde dehydrogenase (BADH) to cadmium (Cd) toxicity was investigated in this study. PCR results demonstrated the expected amplicon length of each gene in the transgenic lines. Absolute qRT-PCR demonstrates a single copy of T-DNA integration into each transgenic line. Relative qRT-PCR confirmed overexpression of Fld and BADH in transgenic lines. The maximum quantum yield of photosystem II (Fv/Fm) was measured under Cd toxicity stress and revealed that transgenic lines had a higher Fv/Fm than wild-type (WT) plants. Accumulation of proline, glycine betaine (GB), and higher activity of antioxidant enzymes alongside lower levels of malondialdehyde (MDA) and hydrogen peroxide (H2O2) was indicative of a robust antioxidant system in transgenic plants. Therefore, performing a loop in reducing the ROS produced in the photosynthesis electron transport chain and stimulating the ROS scavenger enzyme activity improved the plant tolerance to Cd stress.

Seed Priming with Endophytic Bacillus subtilis Modulates Physiological Responses of Two Different Triticum aestivum L. Cultivars under Drought Stress.

The protective effects against drought stress of the endophytic bacterium Bacillus subtilis 10-4 were measured by studying the priming response in two wheat (Triticum aestivum L.)-Ekada70 (E70) and Salavat Yulaev (SY)-lines, tolerant and susceptible to drought, respectively. B. subtilis 10-4 improved germination and growth parameters under normal conditions in both cultivars with the most pronounced effect observed in cv. E70. Under drought conditions, B. subtilis 10-4 significantly ameliorated the negative impact of stress on germination and growth of cv. E70, but had no protective effect on cv. SY. B. subtilis 10-4 induced an increase in the levels of photosynthetic chlorophyll (Chl) a, Chl b, and carotenoids (Car) in the leaves of cv. E70, both under normal and drought conditions. In cv. SY plants, bacterial inoculation decreased the contents of Chl a, Chl b, and Car under normal conditions, but pigment content were almost recovered under drought stress. B. subtilis 10-4 increased water holding capacity (WHC) of cv. E70 (but did not affect this parameter in cv. SY) and prevented the stress-induced decline in WHC in both cultivars. Notably, B. subtilis 10-4 increased endogenous salicylic acid (SA) concentration in both cultivars, especially in cv. E70. Moreover, B. subtilis 10-4 reduced drought-induced endogenous SA accumulation, which was correlated with the influence of endophyte on growth, indicating a possible involvement of endogenous SA in the implementation of B. subtilis-mediated effects in both cultivars. Overall, B. subtilis 10-4 inoculation was found to increase drought tolerance in seedlings of both cultivars, as evidenced by decreased lipid peroxidation, proline content, and electrolyte leakage from tissues of wheat seedlings primed with B. subtilis 10-4 under drought conditions.

γ-Aminobutyric acid confers cadmium tolerance in maize plants by concerted regulation of polyamine metabolism and antioxidant defense systems.

Gamma-Aminobutyric acid (GABA) accumulates in plants following exposure to heavy metals. To investigate the role of GABA in cadmium (Cd) tolerance and elucidate the underlying mechanisms, GABA (0, 25 and 50 µM) was applied to Cd-treated maize plants. Vegetative growth parameters were improved in both Cd-treated and control plants due to GABA application. Cd uptake and translocation were considerably inhibited by GABA. Antioxidant enzyme activity was enhanced in plants subjected to Cd. Concurrently GABA caused further increases in catalase and superoxide dismutase activities, which led to a significant reduction in hydrogen peroxide, superoxide anion and malondealdehyde contents under stress conditions. Polyamine biosynthesis-responsive genes, namely ornithine decarboxylase and spermidine synthase, were induced by GABA in plants grown under Cd shock. GABA suppressed polyamine oxidase, a gene related to polyamine catabolism, when plants were exposed to Cd. Consequently, different forms of polyamines were elevated in Cd-exposed plants following GABA application. The maximum quantum efficiency of photosystem II (Fv/Fm) was decreased by Cd-exposed plants, but was completely restored by GABA to the same value in the control. These results suggest a multifaceted contribution of GABA, through regulation of Cd uptake, production of reactive oxygen species and polyamine metabolism, in response to Cd stress.

Plastid Genotyping Reveals the Uniformity of Cytoplasmic Male Sterile-T Maize Cytoplasms.

Cytoplasmic male-sterile (CMS) lines in maize (Zea mays) have been classified by their response to specific restorer genes into three categories: cms-C, cms-S, and cms-T. A mitochondrial genome representing each of the CMS cytotypes has been sequenced, and male sterility in the cms-S and cms-T cytotypes is linked to chimeric mitochondrial genes. To identify markers for plastid genotyping, we sequenced the plastid genomes of three fertile maize lines (B37, B73, and A188) and the B37 cms-C, cms-S, and cms-T cytoplasmic substitution lines. We found that the plastid genomes of B37 and B73 lines are identical. Furthermore, the fertile and CMS plastid genomes are conserved, differing only by zero to three single-nucleotide polymorphisms (SNPs) in coding regions and by eight to 22 SNPs and 10 to 21 short insertions/deletions in noncoding regions. To gain insight into the origin and transmission of the cms-T trait, we identified three SNPs unique to the cms-T plastids and tested the three diagnostic SNPs in 27 cms-T lines, representing the HA, I, Q, RS, and T male-sterile cytoplasms. We report that each of the tested 27 cms-T group accessions have the same three diagnostic plastid SNPs, indicating a single origin and maternal cotransmission of the cms-T mitochondria and plastids to the seed progeny. Our data exclude exceptional pollen transmission of organelles or multiple horizontal gene transfer events as the source of the mitochondrial urf13-T (unidentified reading frame encoding 13-kD cms-T protein) gene in the cms-T cytoplasms. Plastid genotyping enables a reassessment of the evolutionary relationships of cytoplasms in cultivated maize.

Plastid marker-gene excision by transiently expressed CRE recombinase.

We report plastid marker-gene excision with a transiently expressed CRE, site-specific recombinase. This is a novel protocol that enables rapid removal of marker genes from the approximately 10,000 plastid genome copies without transformation of the plant nucleus. Plastid marker excision was tested in tobacco plants transformed with a prototype polycistronic plastid vector, pPRV110L, designed to express multiple genes organized in an operon. The pMHB10 and pMHB11 constructs described here are dicistronic and encode genes for herbicide (bar) and spectinomycin (aadA) resistance. In vector pMHB11, expression of herbicide resistance is dependent on conversion of an ACG codon to an AUG translation initiation codon by mRNA editing, a safety feature that prevents translation of the mRNA in prokaryotes and in the plant nucleus. In the vectors, the marker gene (aadA) is flanked by 34-bp loxP sites for excision by CRE. Marker excision by a transiently expressed CRE involves introduction of CRE in transplastomic leaves by agro-infiltration, followed by plant regeneration. In tobacco transformed with vectors pMHB10 and pMHB11, Southern analysis and PCR identified approximately 10% of the regenerated plants as marker-free.