Narendra Tuteja

The CRISPR/Cas Genome-Editing Tool: Application in Improvement of Crops

The Clustered Regularly Interspaced Short Palindromic Repeats associated Cas9/sgRNA system is a novel targeted genome-editing technique derived from bacterial immune system. It is an inexpensive, easy, most user friendly and rapidly adopted genome editing tool transforming to revolutionary paradigm. This technique enables precise genomic modifications in many different organisms and tissues. Cas9 protein is an RNA guided endonuclease utilized for creating targeted double-stranded breaks with only a short RNA sequence to confer recognition of the target in animals and plants. Development of genetically edited (GE) crops similar to those developed by conventional or mutation breeding using this potential technique makes it a promising and extremely versatile tool for providing sustainable productive agriculture for better feeding of rapidly growing population in a changing climate. The emerging areas of research for the genome editing in plants include interrogating gene function, rewiring the regulatory signaling networks and sgRNA library for high-throughput loss-of-function screening. In this review, we have described the broad applicability of the Cas9 nuclease mediated targeted plant genome editing for development of designer crops. The regulatory uncertainty and social acceptance of plant breeding by Cas9 genome editing have also been described. With this powerful and innovative technique the designer GE non-GM plants could further advance climate resilient and sustainable agriculture in the future and maximizing yield by combating abiotic and biotic stresses.

Reactive Oxygen Species Generation-Scavenging and Signaling during Plant-Arbuscular Mycorrhizal and Piriformospora indica Interaction under Stress Condition

A defined balance between the generation and scavenging of reactive oxygen species (ROS) is essential to utilize ROS as an adaptive defense response of plants under biotic and abiotic stress conditions. Moreover, ROS are not only a major determinant of stress response but also act as signaling molecule that regulates various cellular processes including plant-microbe interaction. In particular, rhizosphere constitutes the biologically dynamic zone for plant–microbe interactions which forms a mutual link leading to reciprocal signaling in both the partners. Among plant–microbe interactions, symbiotic associations of arbuscular mycorrhizal fungi (AMF) and arbuscular mycorrhizal-like fungus especially Piriformospora indica with plants are well known to improve plant growth by alleviating the stress-impacts and consequently enhance the plant fitness. AMF and P. indica colonization mainly enhances ROS-metabolism, maintains ROS-homeostasis, and thereby averts higher ROS-level accrued inhibition in plant cellular processes and plant growth and survival under stressful environments. This article summarizes the major outcomes of the recent reports on the ROS-generation, scavenging and signaling in biotic-abiotic stressed plants with AMF and P. indica colonization. Overall, a detailed exploration of ROS-signature kinetics during plant-AMF/P. indica interaction can help in designing innovative strategies for improving plant health and productivity under stress conditions.

Fungal association and utilization of phosphate by plants: success, limitations, and future prospects

Phosphorus (P) is a major macronutrient for plant health and development. The available form of P is generally low in the rhizosphere even in fertile soils. A major proportion of applied phosphate (Pi) fertilizers in the soil become fixed into insoluble, unavailable forms, which restricts crop production throughout the world. Roots possess two distinct modes of P uptake from the soil, direct and indirect uptake. The direct uptake of P is facilitated by the plant’s own Pi transporters while indirect uptake occurs via mycorrhizal symbiosis, where the host plant obtains P primarily from the fungal partner, while the fungus benefits from plant-derived reduced carbon. So far, only one Pi transporter has been characterized from the mycorrhizal fungus Glomus versiforme. As arbuscular mycorrhizal fungi cannot be cultured axenically, their Pi transporter network is difficult to exploite for large scale sustainable agriculture. Alternatively, the root-colonizing endophytic fungus Piriformospora indica can grow axenically and provides strong growth-promoting activity during its symbiosis with a broad spectrum of plants. P. indica contains a high affinity Pi transporter (PiPT) involved in improving Pi nutrition levels in the host plant under P limiting conditions. As P. indica can be manipulated genetically, it opens new vistas to be used in P deficient fields.

Introduction to Mycorrhiza: Historical Development

Arbuscular mycorrhizal fungi (AMF) are vital component of natural ecosystem, being gifted to form symbiont with plant roots. AM fungi have mutualistic relationships with more than 80% of terrestrial plant species. Because of wide range of relationships with host plants, it becomes difficult to identify the species on the morphological bases as the spores are to be extracted from the soil. In spite of their abundance and wide range of relationship with plant species, AMF have shown low species diversity. AMF have high functional diversity because different combinations of host plants and AMF have different effects on the numerous aspects of symbiosis. Recent fossil evidence has dated the appearance of arbuscular mycorrhizae back to 460 million years, preexisting vascular plants. These studies benefit the paleoecological importance of mycorrhizae and enhancement to our understanding of the evolution of mutualisms.

PGPR-mediated expression of salt tolerance gene in soybean through volatiles under sodium nitroprusside.

Increasing evidence shows that nitric oxide (NO), a typical signaling molecule plays important role in development of plant and in bacteria‐plant interaction. In the present study, we tested the effect of sodium nitroprusside (SNP)‐a nitric oxide donor, on bacterial metabolism and its role in establishment of PGPR‐plant interaction under salinity condition. In the present study, we adopted methods namely, biofilm formation assay, GC‐MS analysis of bacterial volatiles, chemotaxis assay of root exudates (REs), measurement of electrolyte leakage and lipid peroxidation, and quantitative reverse transcription–polymerase chain reaction (qRT–PCR) for gene expression. GC‐MS analysis revealed that three new volatile organic compounds (VOCs) were expressed after treatment with SNP. Two VOCs namely, 4‐nitroguaiacol and quinoline were found to promote soybean seed germination under 100 mM NaCl stress. Chemotaxis assay revealed that SNP treatment, altered root exudates profiling (SS‐RE), found more attracted to Pseudomonas simiae bacterial cells as compared to non‐treated root exudates (S‐RE) under salt stress. Expression of Peroxidase (POX), catalase (CAT), vegetative storage protein (VSP), and nitrite reductase (NR) genes were up‐regulated in T6 treatment seedlings, whereas, high affinity K+ transporter (HKT1), lipoxygenase (LOX), polyphenol oxidase (PPO), and pyrroline‐5‐carboxylate synthase (P5CS) genes were down‐regulated under salt stress. The findings suggest that NO improves the efficiency and establishment of PGPR strain in the plant environment during salt condition. This strategy may be applied on soybean plants to increase their growth during salinity stress.

Catalase and ascorbate peroxidase—representative H2O2-detoxifying heme enzymes in plants

Plants have to counteract unavoidable stress-caused anomalies such as oxidative stress to sustain their lives and serve heterotrophic organisms including humans. Among major enzymatic antioxidants, catalase (CAT; EC 1.11.1.6) and ascorbate peroxidase (APX; EC 1.11.1.11) are representative heme enzymes meant for metabolizing stress-provoked reactive oxygen species (ROS; such as H2O2) and controlling their potential impacts on cellular metabolism and functions. CAT mainly occurs in peroxisomes and catalyzes the dismutation reaction without requiring any reductant; whereas, APX has a higher affinity for H2O2 and utilizes ascorbate (AsA) as specific electron donor for the reduction of H2O2 into H2O in organelles including chloroplasts, cytosol, mitochondria, and peroxisomes. Literature is extensive on the glutathione-associated H2O2-metabolizing systems in plants. However, discussion is meager or scattered in the literature available on the biochemical and genomic characterization as well as techniques for the assays of CAT and APX and their modulation in plants under abiotic stresses. This paper aims (a) to introduce oxidative stress-causative factors and highlights their relationship with abiotic stresses in plants; (b) to overview structure, occurrence, and significance of CAT and APX in plants; (c) to summarize the principles of current technologies used to assay CAT and APX in plants; (d) to appraise available literature on the modulation of CAT and APX in plants under major abiotic stresses; and finally, (e) to consider a brief cross-talk on the CAT and APX, and this also highlights the aspects unexplored so far.

PDH45 transgenic rice maintain cell viability through lower accumulation of Na+, ROS and calcium homeostasis in roots under salinity stress

Salinity severely affects the growth/productivity of rice, which is utilized as major staple food crop worldwide. PDH45 (pea DNA helicase 45), a member of the DEAD-box helicase family, actively provides salinity stress tolerance, but the mechanism behind this is not well known. Therefore, in order to understand the mechanism of stress tolerance, sodium ion (Na(+)), reactive oxygen species (ROS), cytosolic calcium [Ca(2+)]cyt and cell viability were analyzed in roots of PDH45 transgenic-IR64 rice lines along with wild-type (WT) IR64 rice under salinity stress (100mM and 200 mM NaCl). In addition, the roots of salinity-tolerant (FL478) and susceptible (Pusa-44) rice varieties were also analyzed under salinity stress for comparative analysis. The results reveal that, under salinity stress (100mM and 200 mM NaCl), roots of PDH45 transgenic lines accumulate lower levels of Na(+), ROS and maintain [Ca(2+)]cyt and exhibit higher cell viability as compared with roots of WT (IR64) plants. Similar results were also obtained in the salinity-tolerant FL478 rice. However, the roots of WT and salinity-susceptible Pusa-44 rice accumulated higher levels of Na(+), ROS and [Ca(2+)]cyt imbalance and lower cell viability during salinity stress, which is in contrast to the overexpressing PDH45 transgenic lines and salinity-tolerant FL478 rice. Further, to understand the mechanism of PDH45 at molecular level, comparative expression profiling of 12 cation transporters/genes was also conducted in roots of WT (IR64) and overexpressing PDH45 transgenic lines (L1 and L2) under salt stress (24h of 200 mM NaCl). The expression analysis results show altered and differential gene expression of cation transporters/genes in salt-stressed roots of WT (IR64) and overexpressing transgenic lines (L1 and L2). These observations collectively suggest that, under salinity stress conditions, PDH45 is involved in the regulation of Na(+) level, ROS production, [Ca(2+)]cyt homeostasis, cell viability and cation transporters in roots of PDH45 transgenic-IR64 rice and consequently provide salinity tolerance. Elucidating the detailed regulatory mechanism of PDH45 will provide a better understanding of salinity stress tolerance and further open new ways to manipulate genome to achieve higher agricultural production under stress.

Overexpression of Pea DNA Helicase 45 ( PDH45 ) imparts tolerance to multiple abiotic stresses in chili ( Capsicum annuum L.)

Imparting tolerance to abiotic stresses is of global importance as they inflict significant yield losses in field as well as in vegetable crops. Transcriptional activators, including helicases are identified to play a pivotal role in stress mitigation. Helicases, also known as molecular motors, are involved in myriad cellular processes that impart intrinsic tolerance to abiotic stresses in plants. Our study demonstrates the potential of a Pea DNA Helicase 45 (PDH45), in combating multiple abiotic stresses in chili. We harnessed Agrobacterium-mediated in planta transformation strategy for the generation of stable, single copy transgenic events. Precise molecular detection of the transgenes by sqRT-PCR coupled with genomic Southern analysis revealed variation in the integration of PDH45 at distinct loci in independent transgenic events. Characterization of five promising transgenic events showed both improved response to an array of simulated abiotic stresses and enhanced expression of several stress-responsive genes. While survival and recovery of transgenic events were significantly higher under gradual moisture stress conditions, under imposition of moderate stress, the transgenic events exhibited invigorated growth and productivity with concomitant improvement in water use efficiency (WUE). Thus, our study, unequivocally demonstrated the cardinal role of PDH45 in alleviating multiple abiotic stresses in chili.

Emergence of plant and rhizospheric microbiota as stable interactomes

The growing human population and depletion of resources have necessitated development of sustainable agriculture. Beneficial plant–microbe associations have been known for quite some time now. To maintain sustainability, one could show better reliance upon beneficial attributes of the rhizosphere microbiome. To harness the best agronomic traits, understanding the entire process of recruitment, establishment, and maintenance of microbiota as stable interactome within the rhizosphere is important. In this article, we highlight the process of recruitment and establishment of microbiota within rhizosphere. Further, we have discussed the interlinkages and the ability of multiple (microbial and plant) partners to interact with one another forming a stable plant holobiont system. Lastly, we address the possibility of exploring the knowledge gained from the holobiont system to tailor the rhizosphere microbiome for better productivity and maintenance of agroecosystems. The article provide new insights into the broad principles of stable plant–microbe interactions which could be useful for sustaining agriculture and food security.

Prediction and validation of cis-regulatory elements in 5' upstream regulatory regions of lectin receptor-like kinase gene family in rice.

Lectin receptor-like kinases (LecRLKs) play crucial roles in regulating plant growth and developmental processes in response to stress. In transcriptional gene regulation for normal cellular functions, cis-acting regulatory elements (CREs) direct the temporal and spatial gene expression with respect to environmental stimuli. A complete insightful of the transcriptional gene regulation system relies on effective functional analysis of CREs. Here, we analyzed the potential putative CREs present in the promoters of rice LecRLKs genes by using PlantCARE database. The CREs in LecRLKs promoters are associated with plant growth/development, light response, plant hormonal regulation processes, various stress responses, hormonal response like ABA, root-specific expression responsive, drought responsive, and cell and organ specific regulatory elements. The effect of methylation on these cis-regulatory elements was also analyzed. Real-time analysis of rice seedling under various stress conditions showed the expression levels of selected LecRLK genes superimposing the number of different CREs present in 5′ upstream region. The overall results showed that the possible CREs function in the selective expression/regulation of LecRLKs gene family and during rice plant development under stress.