Tailoring crops with superior product quality through genome editing: an update.

MAIN CONCLUSION: In this review, using genome editing, the quality trait alterations in important crops have been discussed, along with the challenges encountered to maintain the crop products' quality. The delivery of economic produce with superior quality is as important as high yield since it dictates consumer's acceptance and end use. Improving product quality of various agricultural and horticultural crops is one of the important targets of plant breeders across the globe. Significant achievements have been made in various crops using conventional plant breeding approaches, albeit, at a slower rate. To keep pace with ever-changing consumer tastes and preferences and industry demands, such efforts must be supplemented with biotechnological tools. Fortunately, many of the quality attributes are resultant of well-understood biochemical pathways with characterized genes encoding enzymes at each step. Targeted mutagenesis and transgene transfer have been instrumental in bringing out desired qualitative changes in crops but have suffered from various pitfalls. Genome editing, a technique for methodical and site-specific modification of genes, has revolutionized trait manipulation. With the evolution of versatile and cost effective CRISPR/Cas9 system, genome editing has gained significant traction and is being applied in several crops. The availability of whole genome sequences with the advent of next generation sequencing (NGS) technologies further enhanced the precision of these techniques. CRISPR/Cas9 system has also been utilized for desirable modifications in quality attributes of various crops such as rice, wheat, maize, barley, potato, tomato, etc. The present review summarizes salient findings and achievements of application of genome editing for improving product quality in various crops coupled with pointers for future research endeavors.

Efficient nutrient management for enhancing crop productivity, quality and nutrient dynamics in lentil (Lens culinaris Medik.) in the semi-arid region of northern India.

Various faulty farming practices and low-performance cultivars selection are reducing crop yields, factor productivity, and soil fertility. Therefore, there is an urgent need to achieve better nutrient dynamics and sustainable production by selecting more nutrient-responsive cultivars using efficient nutrient management. The present experiment aimed to enhance crop productivity, seed quality, nutrient efficiency, and soil nutrient dynamics through efficient nutrient management under different lentil cultivars. The experiment was laid out in a split-plot design, assigning three cultivars (viz. Sapna, Garima, and HM-1) in the main plots and ten nutrient management practices in the sub-plots, replicating them thrice. Results revealed that cultivar HM-1 recorded significantly higher seed yield (1.59-1.61 Mg ha-1) and the uptake of N (67.2-67.6 kg ha-1), P (6.8-7.0 kg ha-1), K (13.8-13.9 kg ha-1), Zn (60.4-61.1 g ha-1), and Fe (162.5-165.2 g ha-1) in seed compared to Sapna and Garima. Also, the cultivar HM-1 was more efficient in terms of partial factor productivity for NPK (PFP; 24.27-24.59 kg kg-1), partial nutrient balance (PNB; 2.09-2.13 kg kg-1) and internal utilisation efficiency (IUE; 11.64-11.85 kg kg-1). The study showed that the lentil cultivar HM-1 could be successfully grown by substituting 50% RDN with organic manures, i.e., vermicompost, without compromising crop productivity and soil fertility, thereby sustaining soil-human-environment health.

Seed Longevity in Legumes: Deeper Insights Into Mechanisms and Molecular Perspectives.

Sustainable agricultural production largely depends upon the viability and longevity of high-quality seeds during storage. Legumes are considered as rich source of dietary protein that helps to ensure nutritional security, but associated with poor seed longevity that hinders their performance and productivity in farmer's fields. Seed longevity is the key determinant to assure proper seed plant value and crop yield. Thus, maintenance of seed longevity during storage is of prime concern and a pre-requisite for enhancing crop productivity of legumes. Seed longevity is significantly correlated with other seed quality parameters such as germination, vigor, viability and seed coat permeability that affect crop growth and development, consequently distressing crop yield. Therefore, information on genetic basis and regulatory networks associated with seed longevity, as well as molecular dissection of traits linked to longevity could help in developing crop varieties with good storability. Keeping this in view, the present review focuses towards highlighting the molecular basis of seed longevity, with special emphasis on candidate genes and proteins associated with seed longevity and their interplay with other quality parameters. Further, an attempt was made to provide information on 3D structures of various genetic loci (genes/proteins) associated to seed longevity that could facilitate in understanding the interactions taking place within the seed at molecular level. This review compiles and provides information on genetic and genomic approaches for the identification of molecular pathways and key players involved in the maintenance of seed longevity in legumes, in a holistic manner. Finally, a hypothetical fast-forward breeding pipeline has been provided, that could assist the breeders to successfully develop varieties with improved seed longevity in legumes.

Recent Advances for Drought Stress Tolerance in Maize (Zea mays L.): Present Status and Future Prospects.

Drought stress has severely hampered maize production, affecting the livelihood and economics of millions of people worldwide. In the future, as a result of climate change, unpredictable weather events will become more frequent hence the implementation of adaptive strategies will be inevitable. Through utilizing different genetic and breeding approaches, efforts are in progress to develop the drought tolerance in maize. The recent approaches of genomics-assisted breeding, transcriptomics, proteomics, transgenics, and genome editing have fast-tracked enhancement for drought stress tolerance under laboratory and field conditions. Drought stress tolerance in maize could be considerably improved by combining omics technologies with novel breeding methods and high-throughput phenotyping (HTP). This review focuses on maize responses against drought, as well as novel breeding and system biology approaches applied to better understand drought tolerance mechanisms and the development of drought-tolerant maize cultivars. Researchers must disentangle the molecular and physiological bases of drought tolerance features in order to increase maize yield. Therefore, the integrated investments in field-based HTP, system biology, and sophisticated breeding methodologies are expected to help increase and stabilize maize production in the face of climate change.

Salinity stress tolerance and omics approaches: revisiting the progress and achievements in major cereal crops.

Salinity stress adversely affects plant growth and causes considerable losses in cereal crops. Salinity stress tolerance is a complex phenomenon, imparted by the interaction of compounds involved in various biochemical and physiological processes. Conventional breeding for salt stress tolerance has had limited success. However, the availability of molecular marker-based high-density linkage maps in the last two decades boosted genomics-based quantitative trait loci (QTL) mapping and QTL-seq approaches for fine mapping important major QTL for salinity stress tolerance in rice, wheat, and maize. For example, in rice, 'Saltol' QTL was successfully introgressed for tolerance to salt stress, particularly at the seedling stage. Transcriptomics, proteomics and metabolomics also offer opportunities to decipher and understand the molecular basis of stress tolerance. The use of proteomics and metabolomics-based metabolite markers can serve as an efficient selection tool as a substitute for phenotype-based selection. This review covers the molecular mechanisms for salinity stress tolerance, recent progress in mapping and introgressing major gene/QTL (genomics), transcriptomics, proteomics, and metabolomics in major cereals, viz., rice, wheat and maize.

Meta-QTL analysis and candidate genes identification for various abiotic stresses in maize (Zea mays L.) and their implications in breeding programs.

Global climate change leads to the concurrence of a number of abiotic stresses including moisture stress (drought, waterlogging), temperature stress (heat, cold), and salinity stress, which are the major factors affecting maize production. To develop abiotic stress tolerance in maize, many quantitative trait loci (QTL) have been identified, but very few of them have been utilized successfully in breeding programs. In this context, the meta-QTL analysis of the reported QTL will enable the identification of stable/real QTL which will pave a reliable way to introgress these QTL into elite cultivars through marker-assisted selection. In this study, a total of 542 QTL were summarized from 33 published studies for tolerance to different abiotic stresses in maize to conduct meta-QTL analysis using BiomercatorV4.2.3. Among those, only 244 major QTL with more than 10% phenotypic variance were preferably utilised to carry out meta-QTL analysis. In total, 32 meta-QTL possessing 1907 candidate genes were detected for different abiotic stresses over diverse genetic and environmental backgrounds. The MQTL2.1, 5.1, 5.2, 5.6, 7.1, 9.1, and 9.2 control different stress-related traits for combined abiotic stress tolerance. The candidate genes for important transcription factor families such as ERF, MYB, bZIP, bHLH, NAC, LRR, ZF, MAPK, HSP, peroxidase, and WRKY have been detected for different stress tolerances. The identified meta-QTL are valuable for future climate-resilient maize breeding programs and functional validation of candidate genes studies, which will help to deepen our understanding of the complexity of these abiotic stresses. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-022-01294-9.

Pulse-based cropping systems for soil health restoration, resources conservation, and nutritional and environmental security in rainfed agroecosystems.

Pulses are an important source of energy and protein, essential amino acids, dietary fibers, minerals, and vitamins, and play a significant role in addressing global nutritional security. The global pulse area, production, and average productivity increased from 1961 to 2020 (60 years). Pulses are usually grown under rainfed, highly unstable, and complex production environments, with substantial variability in soil and environmental factors, high year-to-year output variability, and variation in soil moisture. Since the last six decades, there is not much satisfactory improvement in the yield of pulses because of their cultivation in harsh environments, coupled with their continuous ignorance of the farmers and governments in policy planning. As a result, the global food supplies through pulses remained negligible and amounted to merely ~1.0% of the total food supply and 1.2% of the vegan food system. In this situation, protein-rich food is still a question raised at the global level to make a malnutrition-free world. Pulses are a vital component of agricultural biological diversity, essential for tackling climate change, and serve as an energy diet for vegetarians. Pulses can mitigate climate change by reducing the dependence on synthetic fertilizers that artificially introduce nitrogen (N) into the soil. The high demand and manufacture of chemical fertilizers emit greenhouse gases (GHGs), and their overuse can harm the environment. In addition, the increasing demand for the vegetal protein under most global agroecosystems has to be met with under a stressed rainfed situation. The rainfed agroecosystem is a shelter for poor people from a significant part of the globe, such as Africa, South Asia, and Latin America. Nearly, 83% [over 1,260 million hectares (ha)] of cultivated land comes under rainfed agriculture, contributing significantly to global food security by supplying over 60% of the food. In rainfed areas, the limitation of natural resources with the shrinking land, continuous nutrient mining, soil fertility depletion, declining productivity factor, constantly depleting water availability, decreasing soil carbon (C) stock, augmented weed menace, ecological instability, and reduced system productivity are creating a more challenging situation. Pulses, being crops of marginal and semi-marginal soils of arid and semi-arid climates, require less input for cultivation, such as water, nutrients, tillage, labor, and energy. Furthermore, accommodation of the area for the cultivation of pulses reduces the groundwater exploitation, C and N footprints, agrochemical application in the cropping systems, and ill effects of climate change due to their inherent capacity to withstand harsh soil to exhibit phytoremediation properties and to stand well under stressed environmental condition. This article focuses on the role of pulses in ecological services, human wellbeing, soil, environmental health, and economic security for advanced sustainability. Therefore, this study will enhance the understanding of productivity improvement in a system-based approach in a rainfed agroecosystem through the involvement of pulses. Furthermore, the present study highlighted significant research findings and policy support in the direction of exploring the real yield potential of pulses. It will provide a road map to producers, researchers, policymakers, and government planners working on pulses to promote them in rainfed agroecosystems to achieve the United Nations (UN's) Sustainable Development Goals (SDGs).

A role for the Erk MAPK pathway in modulating SAX-7/L1CAM-dependent locomotion in Caenorhabditis elegans.

L1CAMs are immunoglobulin cell adhesion molecules that function in nervous system development and function. Besides being associated with autism and schizophrenia spectrum disorders, impaired L1CAM function also underlies the X-linked L1 syndrome, which encompasses a group of neurological conditions, including spastic paraplegia and congenital hydrocephalus. Studies on vertebrate and invertebrate L1CAMs established conserved roles that include axon guidance, dendrite morphogenesis, synapse development, and maintenance of neural architecture. We previously identified a genetic interaction between the Caenorhabditis elegans L1CAM encoded by the sax-7 gene and RAB-3, a GTPase that functions in synaptic neurotransmission; rab-3; sax-7 mutant animals exhibit synthetic locomotion abnormalities and neuronal dysfunction. Here, we show that this synergism also occurs when loss of SAX-7 is combined with mutants of other genes encoding key players of the synaptic vesicle (SV) cycle. In contrast, sax-7 does not interact with genes that function in synaptogenesis. These findings suggest a postdevelopmental role for sax-7 in the regulation of synaptic activity. To assess this possibility, we conducted electrophysiological recordings and ultrastructural analyses at neuromuscular junctions; these analyses did not reveal obvious synaptic abnormalities. Lastly, based on a forward genetic screen for suppressors of the rab-3; sax-7 synthetic phenotypes, we determined that mutants in the ERK Mitogen-activated Protein Kinase (MAPK) pathway can suppress the rab-3; sax-7 locomotion defects. Moreover, we established that Erk signaling acts in a subset of cholinergic neurons in the head to promote coordinated locomotion. In combination, these results suggest a modulatory role for Erk MAPK in L1CAM-dependent locomotion in C. elegans.

UNC-16 alters DLK-1 localization and negatively regulates actin and microtubule dynamics in Caenorhabditis elegans regenerating neurons.

Neuronal regeneration after injury depends on the intrinsic growth potential of neurons. Our study shows that UNC-16, a Caenorhabditis elegans JIP3 homolog, inhibits axonal regeneration by regulating initiation and rate of regrowth. This occurs through the inhibition of the regeneration-promoting activity of the long isoform of DLK-1 and independently of the inhibitory short isoform of DLK-1. We show that UNC-16 promotes DLK-1 punctate localization in a concentration-dependent manner limiting the availability of the long isoform of DLK-1 at the cut site, minutes after injury. UNC-16 negatively regulates actin dynamics through DLK-1 and microtubule dynamics partially via DLK-1. We show that post-injury cytoskeletal dynamics in unc-16 mutants are also partially dependent on CEBP-1. The faster regeneration seen in unc-16 mutants does not lead to functional recovery. Our data suggest that the inhibitory control by UNC-16 and the short isoform of DLK-1 balances the intrinsic growth-promoting function of the long isoform of DLK-1 in vivo. We propose a model where UNC-16's inhibitory role in regeneration occurs through both a tight temporal and spatial control of DLK-1 and cytoskeletal dynamics.

Protocols for electrophysiological recordings and electron microscopy at C. elegans neuromuscular junction.

Release of neurotransmitters by synaptic vesicle exocytosis at presynaptic terminals is critical for neuronal communication within the nervous system. Electrophysiology and electron microscopy are powerful and complementary approaches used to evaluate the function of synaptic proteins in synaptic transmission. Here, we provide a protocol detailing the use of these two approaches at C. elegans neuromuscular junctions, including steps for worm picking and dissection, in vivo electrophysiological recording, and sample preparation for electron microscopy, followed by imaging and analysis. For complete details on the use and execution of this protocol, please refer to Liu et al. (2021) and Li et al. (2021).

Nitrogen fixation in maize: breeding opportunities.

Maize (Zea mays L.) is a highly versatile crop with huge demand of nitrogen (N) for its growth and development. N is the most essential macronutrient for crop production. Despite being the highest abundant element in the atmosphere (~ 78%), it is scarcely available for plant growth. To fulfil the N demand, commercial agriculture is largely dependent on synthetic fertilizers. Excessive dependence on inorganic fertilizers has created extensive ecological as well as economic problems worldwide. Hence, for a sustainable solution to nitrogenous fertilizer use, development of biological nitrogen fixation (BNF) in cereals will be the best alternative. BNF is a well-known mechanism in legumes where diazotrophs convert atmospheric nitrogen (N≡N) to plant-available form, ammonium (NH4+). From many decades, researchers have dreamt to develop a similar symbiotic partnership as in legumes to the cereal crops. A large number of endophytic diazotrophs have been found associated with maize. Elucidation of the genetic and molecular aspects of their interaction will open up new avenues to introgress BNF in maize breeding. With the advanced understanding of N-fixation process, researchers are at a juncture of breeding and engineering this symbiotic relationships in cereals. Different breeding, genetic engineering, omics, gene editing, and synthetic biology approaches will be discussed in this review to make BNF a reality in cereals. It will help to provide a road map to develop/improve the BNF in maize to an advance step for the sustainable production system to achieve the food and nutritional security.

The M domain in UNC-13 regulates the probability of neurotransmitter release.

Synapses exhibit multiple forms of short-term plasticities, which have been attributed to the heterogeneity of neurotransmitter release probability. However, the molecular mechanisms that underlie the differential release states remain to be fully elucidated. The Unc-13 proteins appear to have key roles in synaptic function through multiple regulatory domains. Here, we report that deleting the M domain in Caenorhabditis elegans UNC-13MR leads to a significant increase in release probability, revealing an inhibitory function of this domain. The inhibitory effect of this domain is eliminated when the C1 and C2B domains are absent or activated, suggesting that the M domain inhibits release probability by suppressing the activity of C1 and C2B domains. When fused directly to the MUNC2C fragment of UNC-13, the M domain greatly enhances release probability. Thus, our findings reveal a mechanism by which the UNC-13 M domain regulates synaptic transmission and provides molecular insights into the regulation of release probability.

Introgression of "QTL-hotspot" region enhances drought tolerance and grain yield in three elite chickpea cultivars.

With an aim of enhancing drought tolerance using a marker-assisted backcrossing (MABC) approach, we introgressed the "QTL-hotspot" region from ICC 4958 accession that harbors quantitative trait loci (QTLs) for several drought-tolerance related traits into three elite Indian chickpea (Cicer arietinum L.) cultivars: Pusa 372, Pusa 362, and DCP 92-3. Of eight simple sequence repeat (SSR) markers in the QTL-hotspot region, two to three polymorphic markers were used for foreground selection with respective cross-combinations. A total of 47, 53, and 46 SSRs were used for background selection in case of introgression lines (ILs) developed in genetic backgrounds of Pusa 372, Pusa 362, and DCP 92-3, respectively. In total, 61 ILs (20 BC3 F3 in Pusa 372; 20 BC2 F3 in Pusa 362, and 21 BC3 F3 in DCP 92-3), with >90% recurrent parent genome recovery were developed. Six improved lines in different genetic backgrounds (e.g. BGM 10216 in Pusa 372; BG 3097 and BG 4005 in Pusa 362; IPC(L4-14), IPC(L4-16), and IPC(L19-1) in DCP 92-3) showed better performance than their respective recurrent parents. BGM 10216, with 16% yield gain over Pusa 372, has been released as Pusa Chickpea 10216 by the Central Sub-Committees on Crop Standards, Notification and Release of Varieties of Agricultural Crops, Ministry of Agriculture and Farmers Welfare, Government of India, for commercial cultivation in India. In summary, this study reports introgression of the QTL-hotspot for enhancing yield under rainfed conditions, development of several introgression lines, and release of Pusa Chickpea 10216 developed through molecular breeding in India.

Alcohol induces mitochondrial fragmentation and stress responses to maintain normal muscle function in Caenorhabditis elegans.

Chronic excessive ethanol consumption has distinct toxic and adverse effects on a variety of tissues. In skeletal muscle, ethanol causes alcoholic myopathy, which is characterized by myofiber atrophy and the loss of muscle strength. Alcoholic myopathy is more prevalent than all inherited muscle diseases combined. Current evidence indicates that ethanol directly impairs muscle organization and function. However, the underlying mechanism by which ethanol causes toxicity in muscle is poorly understood. Here, we show that the nematode Caenorhabditis elegans exhibits the key features of alcoholic myopathy when exposed to ethanol. As in mammals, ethanol exposure impairs muscle strength and induces the expression of protective genes, including oxidative stress response genes. In addition, ethanol exposure causes the fragmentation of mitochondrial networks aligned with myofibril lattices. This ethanol-induced mitochondrial fragmentation is dependent on the mitochondrial fission factor DRP-1 (dynamin-related protein 1) and its receptor proteins on the outer mitochondrial membrane. Our data indicate that this fragmentation contributes to the activation of the mitochondrial unfolded protein response (UPR). We also found that robust, perpetual mitochondrial UPR activation effectively reduces muscle weakness caused by ethanol exposure. Our results strongly suggest that the modulation of mitochondrial stress responses may provide a method to ameliorate alcohol toxicity and damage to muscle.