Unveiling silicon-mediated cadmium tolerance mechanisms in mungbean (Vigna radiata (L.) Wilczek): Integrative insights from gene expression, antioxidant responses, and metabolomics.

Cadmium (Cd), one of the most phytotoxic heavy metals, is a major contributor to yield losses in several crops. Silicon (Si) is recognized for its vital role in mitigating Cd toxicity, however, the specific mechanisms governing this mitigation process are still not fully understood. In the present study, the effect of Si supplementation on mungbean (Vigna radiata (L.) Wilczek) plants grown under Cd stress was investigated to unveil the intricate pathways defining Si derived stress tolerance. Non-invasive leaf imaging technique revealed improved growth, biomass, and photosynthetic efficiency in Si supplemented mungbean plants under Cd stress. Further, physiological and biochemical analysis revealed Si mediated increase in activity of glutathione reductase (GR), ascorbate peroxidase (APX), and catalase (CAT) enzymes involved in reactive oxygen species (ROS) metabolism leading to mitigation of cellular damage and oxidative stress. Untargeted metabolomic analysis using liquid chromatography coupled with mass spectrometry (LC-MS/MS) provided insights into Si mediated changes in metabolites and their respective pathways under Cd stress. Alteration in five different metabolic pathways with major changes in flavanols and flavonoids biosynthesis pathway which is essential for controlling plants antioxidant defense system and oxidative stress management were observed. The information reported here about the effects of Si on photosynthetic efficiency, antioxidant responses, and metabolic changes will be helpful in understanding the Si-mediated resistance to Cd stress in plants.

Intrinsically Disordered Kiwellin Protein-Like Effectors Target Plant Chloroplasts and are Extensively Present in Rust Fungi.

The effector proteins produced by plant pathogens are one of the essential components of host-pathogen interaction. Despite being important, most of the effector proteins remain unexplored due to the diversity in their primary sequence generated by the high selection pressure of the host immune system. However to maintain the primary function in the infection process, these effectors may tend to maintain their native protein fold to perform the corresponding biological function. In the present study, unannotated candidate secretory effector proteins of sixteen major plant fungal pathogens were analyzed to find the conserved known protein folds using homology, ab initio, and Alpha Fold/Rosetta Fold protein dimensional (3D) structure approaches. Several unannotated candidate effector proteins were found to match various known conserved protein families potentially involved in host defense manipulation in different plant pathogens. Surprisingly a large number of plant Kiwellin proteins fold like secretory proteins (> 100) were found in studied rust fungal pathogens. Many of them were predicted as potential effector proteins. Furthermore, template independent modelling using Alpha Fold/Rosetta Fold analysis and structural comparison of these candidates also predicted them to match with plant Kiwellin proteins. We also found plant Kiwellin matching proteins outside rusts including several non-pathogenic fungi suggesting the broad function of these proteins. One of the highest confidently modeled Kiwellin matching candidates effectors, Pstr_13960 (97.8%), from the Indian P. striiformis race Yr9 was characterized using overexpression, localization, and deletion studies in Nicotiana benthamiana. The Pstr_13960 suppressed the BAX-induced cell death and localized in the chloroplast. Furthermore, the expression of the Kiwellin matching region (Pst_13960_kiwi) alone suppressed the BAX-induced cell death in N. benthamiana despite the change of location to the cytoplasm and nucleus, suggesting the novel function of the Kiwellin core fold in rust fungi. Molecular docking showed that Pstr_13960 can interact with plant Chorismate mutases (CMs) using three loops conserved in plant and rust Kiwellins. Further analysis of Pstr_13960 showed to contain Intrinsically disordered regions (IDRs) in place of the N-terminal ß1/ß2 region found in plant Kiwellins suggesting the evolution of rust Kiwellins-like effectors (KLEs). Overall, this study reports the presence of a Kiwellin protein-like fold containing a novel effector protein family in rust fungi depicting a classical example of the evolution of effectors at the structure level as Kiwellin effectors show very low significant similarity to plant Kiwellin at the sequence level.

Pinpointing Genomic Regions and Candidate Genes Associated with Seed Oil and Protein Content in Soybean through an Integrative Transcriptomic and QTL Meta-Analysis.

Soybean with enriched nutrients has emerged as a prominent source of edible oil and protein. In the present study, a meta-analysis was performed by integrating quantitative trait loci (QTLs) information, region-specific association and transcriptomic analysis. Analysis of about a thousand QTLs previously identified in soybean helped to pinpoint 14 meta-QTLs for oil and 16 meta-QTLs for protein content. Similarly, region-specific association analysis using whole genome re-sequenced data was performed for the most promising meta-QTL on chromosomes 6 and 20. Only 94 out of 468 genes related to fatty acid and protein metabolic pathways identified within the meta-QTL region were found to be expressed in seeds. Allele mining and haplotyping of these selected genes were performed using whole genome resequencing data. Interestingly, a significant haplotypic association of some genes with oil and protein content was observed, for instance, in the case of FAD2-1B gene, an average seed oil content of 20.22% for haplotype 1 compared to 15.52% for haplotype 5 was observed. In addition, the mutation S86F in the FAD2-1B gene produces a destabilizing effect of (ΔΔG Stability) -0.31 kcal/mol. Transcriptomic analysis revealed the tissue-specific expression of candidate genes. Based on their higher expression in seed developmental stages, genes such as sugar transporter, fatty acid desaturase (FAD), lipid transporter, major facilitator protein and amino acid transporter can be targeted for functional validation. The approach and information generated in the present study will be helpful in the map-based cloning of regulatory genes, as well as for marker-assisted breeding in soybean.

Evolutionary Understanding of Metacaspase Genes in Cultivated and Wild Oryza Species and Its Role in Disease Resistance Mechanism in Rice.

Metacaspases (MCs), a class of cysteine-dependent proteases found in plants, fungi, and protozoa, are predominately involved in programmed cell death processes. In this study, we identified metacaspase genes in cultivated and wild rice species. Characterization of metacaspase genes identified both in cultivated subspecies of Oryza sativa, japonica, and indica and in nine wild rice species was performed. Extensive computational analysis was conducted to understand gene structures, phylogenetic relationships, cis-regulatory elements, expression patterns, and haplotypic variations. Further, the haplotyping study of metacaspase genes was conducted using the whole-genome resequencing data publicly available for 4726 diverse genotype and in-house resequencing data generated for north-east Indian rice lines. Sequence variations observed among wild and cultivated rice species for metacaspase genes were used to understand the duplication and neofunctionalization events. The expression profiles of metacaspase genes were analyzed using RNA-seq transcriptome profiling in rice during different developmental stages and stress conditions. Real-time quantitative PCR analysis of candidate metacaspase genes in rice cultivars Pusa Basmati-1 in response to Magnaporthe oryzae infection indicated a significant role in the disease resistance mechanism. The information provided here will help to understand the evolution of metacaspases and their role under stress conditions in rice.

Genome Editing in Plants: Exploration of Technological Advancements and Challenges.

Genome-editing, a recent technological advancement in the field of life sciences, is one of the great examples of techniques used to explore the understanding of the biological phenomenon. Besides having different site-directed nucleases for genome editing over a decade ago, the CRISPR/Cas (clustered regularly interspaced short palindromic repeats/CRISPR-associated protein) based genome editing approach has become a choice of technique due to its simplicity, ease of access, cost, and flexibility. In the present review, several CRISPR/Cas based approaches have been discussed, considering recent advances and challenges to implicate those in the crop improvement programs. Successful examples where CRISPR/Cas approach has been used to improve the biotic and abiotic stress tolerance, and traits related to yield and plant architecture have been discussed. The review highlights the challenges to implement the genome editing in polyploid crop plants like wheat, canola, and sugarcane. Challenges for plants difficult to transform and germline-specific gene expression have been discussed. We have also discussed the notable progress with multi-target editing approaches based on polycistronic tRNA processing, Csy4 endoribonuclease, intron processing, and Drosha ribonuclease. Potential to edit multiple targets simultaneously makes it possible to take up more challenging tasks required to engineer desired crop plants. Similarly, advances like precision gene editing, promoter bashing, and methylome-editing will also be discussed. The present review also provides a catalog of available computational tools and servers facilitating designing of guide-RNA targets, construct designs, and data analysis. The information provided here will be useful for the efficient exploration of technological advances in genome editing field for the crop improvement programs.

Si permeability of a deficient Lsi1 aquaporin in tobacco can be enhanced through a conserved residue substitution.

Silicon (Si) is a beneficial substrate for many plants, conferring heightened resilience to environmental stress. A plant's ability to absorb Si is primarily dependent on the presence of a Si-permeable Lsi1 (NIP2-1) aquaporin in its roots. Structure-function analyses of Lsi1 channels from higher plants have thus far revealed two key molecular determinants of Si permeability: (a) the amino acid motif GSGR in the aromatic/arginine selectivity filter and (b) 108 amino acids between two highly conserved NPA domains. Curiously, tobacco (Nicotiana sylvestris) stands as a rare exception as it possesses an Lsi1 (NsLsi1) with these molecular signatures but is reported as a low Si accumulator. The aim of this study was therefore to identify whether additional determinants influence Si permeability via Lsi1 channels, focusing on the role of residues that differ uniquely in NsLsi1 relative to functional Lsi1 homologs. We observed tobacco indeed absorbed Si poorly (0.1% dw), despite NsLsi1 being expressed constitutively in planta. Si influx measured in NsLsi1-expressing Xenopus oocytes was very low (<13% that of OsLsi1 from rice (Oryza sativa) over a 3-hr time course), which likely explains why tobacco is a low Si accumulator. Interestingly, NsLsi1P125F displayed a significant gain of function (threefold increase in Si influx relative to NsLsi1WT), which coincided with a threefold increase in plasma membrane localization in planta, as measured by transient expression of GFP constructs in Nicotiana benthamiana leaves. These findings thus reveal a novel molecular determinant of Si transport in plants and inform breeding, biotechnological, and agricultural practices to effectively utilize Si in the context of plant resilience to environmental stress.