The Role of Membrane Transporters in Plant Growth and Development, and Abiotic Stress Tolerance

The proteins of membrane transporters (MTs) are embedded within membrane-bounded organelles and are the prime targets for improvements in the efficiency of water and nutrient transportation. Their function is to maintain cellular homeostasis by controlling ionic movements across cellular channels from roots to upper plant parts, xylem loading and remobilization of sugar molecules from photosynthesis tissues in the leaf (source) to roots, stem and seeds (sink) via phloem loading. The plant’s entire source-to-sink relationship is regulated by multiple transporting proteins in a highly sophisticated manner and driven based on different stages of plant growth and development (PG&D) and environmental changes. The MTs play a pivotal role in PG&D in terms of increased plant height, branches/tiller numbers, enhanced numbers, length and filled panicles per plant, seed yield and grain quality. Dynamic climatic changes disturbed ionic balance (salt, drought and heavy metals) and sugar supply (cold and heat stress) in plants. Due to poor selectivity, some of the MTs also uptake toxic elements in roots negatively impact PG&D and are later on also exported to upper parts where they deteriorate grain quality. As an adaptive strategy, in response to salt and heavy metals, plants activate plasma membranes and vacuolar membrane-localized MTs that export toxic elements into vacuole and also translocate in the root’s tips and shoot. However, in case of drought, cold and heat stresses, MTs increased water and sugar supplies to all organs. In this review, we mainly review recent literature from Arabidopsis, halophytes and major field crops such as rice, wheat, maize and oilseed rape in order to argue the global role of MTs in PG&D, and abiotic stress tolerance. We also discussed gene expression level changes and genomic variations within a species as well as within a family in response to developmental and environmental cues

Reduced Glutathione Mediates Pheno-Ultrastructure, Kinome and Transportome in Chromium-Induced Brassica napus L.

Chromium (Cr) as a toxic metal is widely used for commercial purposes and its residues have become a potential environmental threat to both human and plant health. Oilseed rape (Brassica napus L.) is one of the candidate plants that can absorb the considerable quantity of toxic metals from the soil. Here, we used two cultivars of B. napus cvs. ZS 758 (metal-tolerant) and Zheda 622 (metal-susceptible) to investigate the phenological attributes, cell ultrastructure, protein kinases (PKs) and molecular transporters (MTs) under the combined treatments of Cr stress and reduced glutathione (GSH). Seeds of these cultivars were grown in vitro at different treatments i.e., 0, 400 μM Cr, and 400 μM Cr + 1 mM GSH in control growth chamber for 6 days. Results had confirmed that Cr significantly reduced the plant length, stem and root, and fresh biomass such as leaf, stem and root. Cr noticeably caused the damages in leaf mesophyll cells. Exogenous application of GSH significantly recovered both phenological and cell structural damages in two cultivars under Cr stress. For the PKs, transcriptomic data advocated that Cr stress alone significantly increased the gene expressions of BnaA08g16610D, BnaCnng19320D, and BnaA08g00390D over that seen in controls (Ck). These genes encoded both nucleic acid and transition metal ion binding proteins, and protein kinase activity (PKA) and phosphotransferase activities in both cultivars. Similarly, the presence of Cr revealed elite MT genes [BnaA04g26560D, BnaA02g28130D, and BnaA02g01980D (novel)] that were responsible for water transmembrane transporter activity. However, GSH in combination with Cr stress significantly up-regulated the genes for PKs [such as BnaCnng69940D (novel) and BnaC08g49360D] that were related to PKA, signal transduction, and oxidoreductase activities. For MTs, BnaC01g29930D and BnaA07g14320D were responsible for secondary active transmembrane transporter and protein transporter activities that were expressed more in GSH treatment than either Ck or Cr-treated cells. In general, it can be concluded that cultivar ZS 758 is more tolerant toward Cr-induced stress than Zheda 622

Alleviation of Lead Toxicity by 5-Aminolevulinic Acid Is Related to Elevated Growth, Photosynthesis, and Suppressed Ultrastructural Damages in Oilseed Rape

Lead (Pb) is a widely spread pollutant and leads to diverse morphological and structural changes in the plants. In this study, alleviating role of 5-aminolevulinic acid (ALA) in oilseed rape (Brassica napus L.) was investigated with or without foliar application of ALA (25 mg L−1) in hydroponic environment under different Pb levels (0, 100, and 400 µM). Outcomes stated that plant morphology and photosynthetic attributes were reduced under the application of Pb alone. However, ALA application significantly increased the plant growth and photosynthetic parameters under Pb toxicity. Moreover, ALA also lowered the Pb concentration in shoots and roots under Pb toxicity. The microscopic studies depicted that exogenously applied ALA ameliorated the Pb stress and significantly improved the cell ultrastructures. After application of ALA under Pb stress, mesophyll cell had well-developed nucleus and chloroplast having a number of starch granules. Moreover, micrographs illustrated that root tip cell contained well-developed nucleus, a number of mitochondria, and golgi bodies. These results proposed that under 15-day Pb-induced stress, ALA improved the plant growth, chlorophyll content, photosynthetic parameters, and ultrastructural modifications in leaf mesophyll and root tip cells of the B. napus plants

The Role of Membrane Transporters in Plant Growth and Development, and Abiotic Stress Tolerance

The proteins of membrane transporters (MTs) are embedded within membrane-bounded organelles and are the prime targets for improvements in the efficiency of water and nutrient transportation. Their function is to maintain cellular homeostasis by controlling ionic movements across cellular channels from roots to upper plant parts, xylem loading and remobilization of sugar molecules from photosynthesis tissues in the leaf (source) to roots, stem and seeds (sink) via phloem loading. The plant’s entire source-to-sink relationship is regulated by multiple transporting proteins in a highly sophisticated manner and driven based on different stages of plant growth and development (PG&D) and environmental changes. The MTs play a pivotal role in PG&D in terms of increased plant height, branches/tiller numbers, enhanced numbers, length and filled panicles per plant, seed yield and grain quality. Dynamic climatic changes disturbed ionic balance (salt, drought and heavy metals) and sugar supply (cold and heat stress) in plants. Due to poor selectivity, some of the MTs also uptake toxic elements in roots negatively impact PG&D and are later on also exported to upper parts where they deteriorate grain quality. As an adaptive strategy, in response to salt and heavy metals, plants activate plasma membranes and vacuolar membrane-localized MTs that export toxic elements into vacuole and also translocate in the root’s tips and shoot. However, in case of drought, cold and heat stresses, MTs increased water and sugar supplies to all organs. In this review, we mainly review recent literature from Arabidopsis, halophytes and major field crops such as rice, wheat, maize and oilseed rape in order to argue the global role of MTs in PG&D, and abiotic stress tolerance. We also discussed gene expression level changes and genomic variations within a species as well as within a family in response to developmental and environmental cues

Regulation of Cadmium-Induced Proteomic and Metabolic Changes by 5-Aminolevulinic Acid in Leaves of Brassica napus L.

It is evident from previous reports that 5-aminolevulinic acid (ALA), like other known plant growth regulators, is effective in countering the injurious effects of heavy metal-stress in oilseed rape (Brassica napus L.). The present study was carried out to explore the capability of ALA to improve cadmium (Cd2+) tolerance in B. napus through physiological, molecular, and proteomic analytical approaches. Results showed that application of ALA helped the plants to adjust Cd2+-induced metabolic and photosynthetic fluorescence changes in the leaves of B. napus under Cd2+ stress. The data revealed that ALA treatment enhanced the gene expressions of antioxidant enzyme activities substantially and could increase the expression to a certain degree under Cd2+ stress conditions. In the present study, 34 protein spots were identified that differentially regulated due to Cd2+ and/or ALA treatments. Among them, 18 proteins were significantly regulated by ALA, including the proteins associated with stress related, carbohydrate metabolism, catalysis, dehydration of damaged protein, CO2 assimilation/photosynthesis and protein synthesis/regulation. From these 18 ALA-regulated proteins, 12 proteins were significantly down-regulated and 6 proteins were up-regulated. Interestingly, it was observed that ALA-induced the up-regulation of dihydrolipoyl dehydrogenase, light harvesting complex photo-system II subunit 6 and 30S ribosomal proteins in the presence of Cd2+ stress. In addition, it was also observed that ALA-induced the down-regulation in thioredoxin-like protein, 2, 3-bisphosphoglycerate, proteasome and thiamine thiazole synthase proteins under Cd2+ stress. Taken together, the present study sheds light on molecular mechanisms involved in ALA-induced Cd2+ tolerance in B. napus leaves and suggests a more active involvement of ALA in plant physiological processes than previously proposed

Insights on the responses of Brassica napus cultivars against the cobalt-stress as revealed by carbon assimilation, anatomical changes and secondary metabolites

Cobalt (Co) is a toxin environmental pollutant, and its elevated concentrations in agricultural soils can negatively impact the crop productivity. In the current study, we evaluated the toxic effects of Co levels (0, 100, 400 μM) on the hydroponically grown seedlings of four different cultivars of Brassica napus L. cvs. ZS 758, ZY 50, Zheda 619 and Zheda 622. Results showed that lower Co dose (100 μM) executed significant but less toxic effects as compared to 400 μM Co among all studied cultivars. Excessive Co (400 μM) level had significantly declined the plant growth, biomass, chlorophyll contents, nutrients uptake, activities and transcript levels of antioxidant enzymes in leaf and root tissues of all tested cultivars.The decline in physiochemical traits was more prominent in Zheda 622, and ZS 758 performed better under Co toxicity as compared with other cultivars. Co-induced root impairment led to greater ROS and MDA accumulation and inhibit nutrients uptake. Ultimately, the lower translocation to leaves causes damages in carbon assimilation process.These results were confirmed by root staining with nitro-blue tetrazolium (NBT) and 3,3-diaminobenzidine (DAB) and damages in leaf mesophyll and root tip cells by electron microscopic analysis. More anatomical damages were noted in Zheda 622 and least in ZS 758. A significant induction of stress-related proteins (HSP90-1 and MT-1) and secondary metabolites (PAL, PPO, and CAD) under higher Co level (400 μM) indicated the greater plant-defense system against Co-induced oxidative stress and Co-homeostasis that helps in toleraance to Co-phytotoxicity. The comparison of sensitive (Zheda 622) and tolerant (ZS 758) genotypes showed that root tissues were the major target of Co toxicity. Overall, higher Co-accumulation in leaf and root tissues cause cellular toxicity and overall carbon assimilation process. This phytotoxicity was more pronounced in Zheda 622 and least in ZS 758 as compared with other cultivars

Conventional and Molecular Techniques from Simple Breeding to Speed Breeding in Crop Plants: Recent Advances and Future Outlook

In most crop breeding programs, the rate of yield increment is insufficient to cope with the increased food demand caused by a rapidly expanding global population. In plant breeding, the development of improved crop varieties is limited by the very long crop duration. Given the many phases of crossing, selection, and testing involved in the production of new plant varieties, it can take one or two decades to create a new cultivar. One possible way of alleviating food scarcity problems and increasing food security is to develop improved plant varieties rapidly. Traditional farming methods practiced since quite some time have decreased the genetic variability of crops. To improve agronomic traits associated with yield, quality, and resistance to biotic and abiotic stresses in crop plants, several conventional and molecular approaches have been used, including genetic selection, mutagenic breeding, somaclonal variations, whole-genome sequence-based approaches, physical maps, and functional genomic tools. However, recent advances in genome editing technology using programmable nucleases, clustered regularly interspaced short palindromic repeats (CRISPR), and CRISPR-associated (Cas) proteins have opened the door to a new plant breeding era. Therefore, to increase the efficiency of crop breeding, plant breeders and researchers around the world are using novel strategies such as speed breeding, genome editing tools, and high-throughput phenotyping. In this review, we summarize recent findings on several aspects of crop breeding to describe the evolution of plant breeding practices, from traditional to modern speed breeding combined with genome editing tools, which aim to produce crop generations with desired traits annually

Beryllium Stress-Induced Modifications in Antioxidant Machinery and Plant Ultrastructure in the Seedlings of Black and Yellow Seeded Oilseed Rape

Beryllium (Be) could be a threatening heavy metal pollutant in the agroecosystem that may severely affect the performance of crops. The present study was conducted to evaluate the toxic effects of Be (0, 100, 200, and 400 μM) on physiological, ultrastructure, and biochemical attributes in hydroponically grown six-day-old seedlings of two cultivars of Brassica napus L., one tolerant (ZS 758, black seeded) and one sensitive (Zheda 622, yellow seeded). Higher Be concentrations reduced the plant growth, biomass production, chlorophyll contents, and the total soluble protein contents. A significant accumulation of ROS (H2O2, OH−) and MDA contents was observed in a dose-dependent manner. Antioxidant enzymatic activities including SOD, POD, GR, APX, and GSH (except CAT) were enhanced with the increase in Be concentrations in both cultivars. Relative transcript gene expression of above-mentioned antioxidant enzymes further confirmed the alterations induced by Be as depicted from higher involvement in the least susceptible cultivar ZS 758 as compared to Zheda 622. The electron microscopic study showed that higher level of Be (400 μM) greatly damaged the leaf mesophyll and root tip cells. More damage was observed in cultivar Zheda 622 as compared to ZS 758. The damage in leaf mesophyll cells was highlighted as the disruption in cell wall, immature nucleus, damaged mitochondria, and chloroplast structures. In root tip cells, disruption in Golgi bodies and damage in cell wall were clearly noticed. As a whole, the present study confirmed that more inhibitory effects were recorded in yellow seeded Zheda 622 as compared to black seeded ZS 758 cultivar, which is regarded as more sensitive cultivar

Computational Tools and Resources for CRISPR/Cas Genome Editing

The past decade has witnessed a rapid evolution in identifying more versatile clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein (Cas) nucleases and their functional variants, as well as in developing precise CRISPR/Cas-derived genome editors. The programmable and robust features of the genome editors provide an effective RNA-guided platform for fundamental life science research and subsequent applications in diverse scenarios, including biomedical innovation and targeted crop improvement. One of the most essential principles is to guide alterations in genomic sequences or genes in the intended manner without undesired off-target impacts, which strongly depends on the efficiency and specificity of single guide RNA (sgRNA)-directed recognition of targeted DNA sequences. Recent advances in empirical scoring algorithms and machine learning models have facilitated sgRNA design and off-target prediction. In this review, we first briefly introduce the different features of CRISPR/Cas tools that should be taken into consideration to achieve specific purposes. Secondly, we focus on the computer-assisted tools and resources that are widely used in designing sgRNAs and analyzing CRISPR/Cas-induced on- and off-target mutations. Thirdly, we provide insights into the limitations of available computational tools that would help researchers of this field for further optimization. Lastly, we suggest a simple but effective workflow for choosing and applying web-based resources and tools for CRISPR/Cas genome editing