Discovering the regulators of heat stress tolerance in Ziziphus nummularia (Burm.f) wight and walk.-arn.

Ziziphus nummularia an elite heat-stress tolerant shrub, grows in arid regions of desert. However, its molecular mechanism responsible for heat stress tolerance is unexplored. Therefore, we analysed whole transcriptome of Jaisalmer (heat tolerant) and Godhra (heat sensitive) genotypes of Z. nummularia to understand its molecular mechanism responsible for heat stress tolerance. De novo assembly of 16,22,25,052 clean reads yielded 276,029 transcripts. A total of 208,506 unigenes were identified which contains 4290 and 1043 differentially expressed genes (DEG) in TGO (treated Godhra at 42 °C) vs. CGO (control Godhra) and TJR (treated Jaisalmer at 42 °C) vs. CJR (control Jaisalmer), respectively. A total of 987 (67 highly enriched) and 754 (34 highly enriched) pathways were obsorved in CGO vs. TGO and CJR vs. TJR, respectively. Antioxidant pathways and TFs like Homeobox, HBP, ARR, PHD, GRAS, CPP, and E2FA were uniquely observed in Godhra genotype and SET domains were uniquely observed in Jaisalmer genotype. Further transposable elements were highly up-regulated in Godhra genotype but no activation in Jaisalmer genotype. A total of 43,093 and 39,278 simple sequence repeats were identified in the Godhra and Jaisalmer genotypes, respectively. A total of 10 DEGs linked to heat stress were validated in both genotypes for their expression under different heat stresses using quantitative real-time PCR. Comparing expression patterns of the selected DEGs identified ClpB1 as a potential candidate gene for heat tolerance in Z. nummularia. Here we present first characterized transcriptome of Z. nummularia in response to heat stress for the identification and characterization of heat stress-responsive genes. Supplementary Information: The online version contains supplementary material available at 10.1007/s12298-024-01431-y.

Plant salinity stress, sensing, and its mitigation through WRKY.

Salinity or salt stress has deleterious effects on plant growth and development. It imposes osmotic, ionic, and secondary stresses, including oxidative stress on the plants and is responsible for the reduction of overall crop productivity and therefore challenges global food security. Plants respond to salinity, by triggering homoeostatic mechanisms that counter salt-triggered disturbances in the physiology and biochemistry of plants. This involves the activation of many signaling components such as SOS pathway, ABA pathway, and ROS and osmotic stress signaling. These biochemical responses are accompanied by transcriptional modulation of stress-responsive genes, which is mostly mediated by salt-induced transcription factor (TF) activity. Among the TFs, the multifaceted significance of WRKY proteins has been realized in many diverse avenues of plants' life including regulation of plant stress response. Therefore, in this review, we aimed to highlight the significance of salinity in a global perspective, the mechanism of salt sensing in plants, and the contribution of WRKYs in the modulation of plants' response to salinity stress. This review will be a substantial tool to investigate this problem in different perspectives, targeting WRKY and offering directions to better manage salinity stress in the field to ensure food security.

Regulatory Networks of lncRNAs, miRNAs, and mRNAs in Response to Heat Stress in Wheat (Triticum Aestivum L.): An Integrated Analysis.

Climate change has become a major source of concern, particularly in agriculture, because it has a significant impact on the production of economically important crops such as wheat, rice, and maize. In the present study, an attempt has been made to identify differentially expressed heat stress-responsive long non-coding RNAs (lncRNAs) in the wheat genome using publicly available wheat transcriptome data (24 SRAs) representing two conditions, namely, control and heat-stressed. A total of 10,965 lncRNAs have been identified and, among them, 153, 143, and 211 differentially expressed transcripts have been found under 0 DAT, 1 DAT, and 4 DAT heat-stress conditions, respectively. Target prediction analysis revealed that 4098 lncRNAs were targeted by 119 different miRNA responses to a plethora of environmental stresses, including heat stress. A total of 171 hub genes had 204 SSRs (simple sequence repeats), and a set of target sequences had SNP potential as well. Furthermore, gene ontology analysis revealed that the majority of the discovered lncRNAs are engaged in a variety of cellular and biological processes related to heat stress responses. Furthermore, the modeled three-dimensional (3D) structures of hub genes encoding proteins, which had an appropriate range of similarity with solved structures, provided information on their structural roles. The current study reveals many elements of gene expression regulation in wheat under heat stress, paving the way for the development of improved climate-resilient wheat cultivars.

Morpho-biochemical characterization of a RIL population for seed parameters and identification of candidate genes regulating seed size trait in lentil (Lens culinaris Medik.).

The seed size and shape in lentil (Lens culinaris Medik.) are important quality traits as these influences the milled grain yield, cooking time, and market class of the grains. Linkage analysis was done for seed size in a RIL (F5:6) population derived by crossing L830 (20.9 g/1000 seeds) with L4602 (42.13 g/1000 seeds) which consisted of 188 lines (15.0 to 40.5 g/1000 seeds). Parental polymorphism survey using 394 SSRs identified 31 polymorphic primers, which were used for the bulked segregant analysis (BSA). Marker PBALC449 differentiated the parents and small seed size bulk only, whereas large seeded bulk or the individual plants constituting the large-seeded bulk could not be differentiated. Single plant analysis identified only six recombinant and 13 heterozygotes, of 93 small-seeded RILs (<24.0 g/1000 seed). This clearly showed that the small seed size trait is very strongly regulated by the locus near PBLAC449; whereas, large seed size trait seems governed by more than one locus. The PCR amplified products from the PBLAC449 marker (149bp from L4602 and 131bp from L830) were cloned, sequenced and BLAST searched using the lentil reference genome and was found amplified from chromosome 03. Afterward, the nearby region on chromosome 3 was searched, and a few candidate genes like ubiquitin carboxyl-terminal hydrolase, E3 ubiquitin ligase, TIFY-like protein, and hexosyltransferase having a role in seed size determination were identified. Validation study in another RIL mapping population which is differing for seed size, showed a number of SNPs and InDels among these genes when studied using whole genome resequencing (WGRS) approach. Biochemical parameters like cellulose, lignin, and xylose content showed no significant differences between parents and the extreme RILs, at maturity. Various seed morphological traits like area, length, width, compactness, volume, perimeter, etc., when measured using VideometerLab 4.0 showed significant differences for the parents and RILs. The results have ultimately helped in better understanding the region regulating the seed size trait in genomically less explored crops like lentils.

Antioxidant Potential of Glutathione and Crosstalk with Phytohormones in Enhancing Abiotic Stress Tolerance in Crop Plants.

Glutathione (GSH) is an abundant tripeptide that can enhance plant tolerance to biotic and abiotic stress. Its main role is to counter free radicals and detoxify reactive oxygen species (ROS) generated in cells under unfavorable conditions. Moreover, along with other second messengers (such as ROS, calcium, nitric oxide, cyclic nucleotides, etc.), GSH also acts as a cellular signal involved in stress signal pathways in plants, directly or along with the glutaredoxin and thioredoxin systems. While associated biochemical activities and roles in cellular stress response have been widely presented, the relationship between phytohormones and GSH has received comparatively less attention. This review, after presenting glutathione as part of plants' feedback to main abiotic stress factors, focuses on the interaction between GSH and phytohormones, and their roles in the modulation of the acclimatation and tolerance to abiotic stress in crops plants.

Effect of thermal treatments on the matrix components, inherent glycemic potential, and bioaccessibility of phenolics and micronutrients in pearl millet rotis.

Pearl millet (PM) is a nutri-cereal rich in various macro and micronutrients required for a balanced diet. Its grains have a unique phenolic and micronutrient composition; however, the lower bioaccessibility of nutrients and rancidity of flour during storage are the major constraints in its consumption and wide popularity. Here, to explore the effect of different thermal processing methods, i.e., hydrothermal (HT), microwave (MW), and infrared (IR) treatments, on the digestion of starch, phenolics, and microelements (Fe and Zn), an in vitro digestion model consisting of oral, gastric and intestinal digestion was applied to PM rotis. The hydrothermally treated PM roti was promising as it showed lower inherent glycemic potential (60.4%) than the untreated sample (72.4%) and less enzymatic activities associated with rancidity in PM flour. FTIR revealed an increased ratio of 1047/1022 cm-1 in the hydrothermally treated sample, reflecting the enhancement of the structurally ordered degree and compactness of starch compared to other thermal treatments. A tighter and more compact microstructure with an agglomeration of starch in the hydrothermally treated PM flour was observed by SEM. These structural changes could provide a better understanding of the lower starch digestion rate in the hydrothermally treated flour. However, HT treatment significantly (P < 0.05) reduced the bioaccessibility of phenolics (10.6%) compared to native PM rotis and slightly reduced the Fe (2%) and Zn (3.2%) bioaccessibility present in PM rotis.

Comparative transcriptome analysis, unfolding the pathways regulating the seed-size trait in cultivated lentil (Lens culinaris Medik.).

Market class, cooking time, quality, and milled grain yield are largely influenced by the seed size and shape of the lentil (Lens culinaris Medik.); thus, they are considered to be important quality traits. To unfold the pathways regulating seed size in lentils, a transcriptomic approach was performed using large-seeded (L4602) and small-seeded (L830) genotypes. The study has generated nearly 375 million high-quality reads, of which 98.70% were properly aligned to the reference genome. Among biological replicates, very high similarity in fragments per kilobase of exon per million mapped fragments values (R > 0.9) showed the consistency of RNA-seq results. Various differentially expressed genes associated mainly with the hormone signaling and cell division pathways, transcription factors, kinases, etc. were identified as having a role in cell expansion and seed growth. A total of 106,996 unigenes were used for differential expression (DE) analysis. String analysis identified various modules having certain key proteins like Ser/Thr protein kinase, seed storage protein, DNA-binding protein, microtubule-associated protein, etc. In addition, some growth and cell division-related micro-RNAs like miR3457 (cell wall formation), miR1440 (cell proliferation and cell cycles), and miR1533 (biosynthesis of plant hormones) were identified as having a role in seed size determination. Using RNA-seq data, 5254 EST-SSR primers were generated as a source for future studies aiming for the identification of linked markers. In silico validation using Genevestigator® was done for the Ser/Thr protein kinase, ethylene response factor, and Myb transcription factor genes. It is of interest that the xyloglucan endotransglucosylase gene was found differentially regulated, suggesting their role during seed development; however, at maturity, no significant differences were recorded for various cell wall parameters including cellulose, lignin, and xylose content. This is the first report on lentils that has unfolded the key seed size regulating pathways and unveiled a theoretical way for the development of lentil genotypes having customized seed sizes.

Morphological, Molecular, and Biochemical Characterization of a Unique Lentil (Lens culinaris Medik.) Genotype Showing Seed-Coat Color Anomalies Due to Altered Anthocyanin Pathway.

This study reports the identification of a unique lentil (Lens culinaris Medik.) genotype L4717-NM, a natural mutant (NM) derived from a variety L4717, producing brown, black, and spotted seed-coat colored seeds in a single plant, generation after generation, in different frequencies. The genetic similarity of L4717 with that of L4717-NM expressing anomalous seed-coat color was established using 54 SSR markers. In addition, various biochemical parameters such as TPC (total phenolic content), TFC (total flavonoid content), DPPH (2,2-diphenyl-1-picrylhydrazyl), FRAP (ferric reducing antioxidant power), H2O2 (peroxide quantification), TCC (total carotenoids content), TAC (total anthocyanin content), and TAA (total ascorbic acid) were also studied in the seeds, sprouts, and seedlings of L4717, brown, black, and spotted seed-coat colored seeds. Stage-specific variations for the key biochemical parameters were recorded, and seedling stage was found the best for many parameters. Moreover, seeds with black seed coat showed better nutraceutical values for most of the studied traits. A highly significant (p ≤ 0.01) and positive correlation was observed between DPPH and TPC, TAA, TFC, etc., whereas, protein content showed a negative correlation with the other studied parameters. The seed coat is maternal tissue and we expect expression of seed-coat color as per the maternal genotype. However, such an anomalous seed-coat expression, which seems to probably be governed by some transposable element in the identified genotype, warrants more detailed studies involving exploitation of the anthocyanin pathway.

Thermal treatments reduce rancidity and modulate structural and digestive properties of starch in pearl millet flour.

Pearl millet is a nutrient dense and gluten free cereal, however it's flour remains underutilized due to the onset of rancidity during its storage. To the best of our knowledge, processing methods, which could significantly reduce the rancidity of the pearl millet flour during storage, are non-existent. In this study, pearl millet grains were subjected to a preliminary hydro-treatment (HT). Subsequently, the hydrated grain-wet flour have undergone individual and combined thermal treatments viz., hydrothermal (HTh) and thermal near infrared rays (thNIR). Effects of these thermal treatments on the biochemical process of hydrolytic and oxidative rancidity were analyzed in stored flour. A significant (p < 0.05) decrease in the enzyme activities of lipase (47.8%), lipoxygenase (84.8%), peroxidase (98.1%) and polyphenol oxidase (100%) in HT-HTh-thNIR treated flour compared to the individual treatments was documented. Upon storage (90 days), decline of 67.84% and 66.4% of free fatty acid and peroxide contents were observed in flour under HT-HTh-thNIR treatment without altering starch and protein digestibility properties. HT-HTh treated flour exhibited the highest (7.6%) rapidly digestible starch, decreased viscosity and increased starch digestibility (67.17%). FTIR analysis of HT-HTh treated flour divulged destabilization of short-range ordered crystalline structure and altered protein structures with decreased in vitro digestibility of protein. Overall, these results demonstrated the effectiveness of combined thermal treatment of HT-HTh-thNIR in reducing rancidity and preserving the functional properties of the stored flour.

Meta-analysis of QTLs associated with popping traits in maize (Zea mays L.).

The rising demand for popcorn necessitates improving the popping quality with higher yield of popcorn cultivars. Towards this direction several Quantitative Traits Loci (QTLs) for popping traits have been identified. However, identification of accurate and consistent QTLs across different genetic backgrounds and environments is necessary to effectively utilize the identified QTLs in marker-assisted breeding. In the current study, 99 QTLs related to popping traits reported in 8 different studies were assembled and projected on the reference map "Genetic 2005" using BioMercator v4.2 to identify metaQTLs with consistent QTLs. Total ten metaQTLs were identified on chromosome 1 (7 metaQTLs) and 6 (3 metaQTLs) with physical distance ranging between 0.43 and 12.75 Mb, respectively. Four identified metaQTLs, viz., mQTL1_1, mQTL1_5, mQTL1_7 and mQTL6_2 harboured 5-8 QTL clusters with moderately high R2 value. The clustered QTLs were from two or more experiments. Based on the expression pattern in endosperm and pericarp tissues, a total of 229 genes were selected. Nineteen of these genes are involved in carbohydrate metabolism. Of the 19 genes specifically involved in carbohydrate metabolism, 11 of them were in these regions, implying the importance of these clustered QTLs. MetaQTL1_1 at bin location 1.01 coincided with the reported QTLs related to various agronomic traits like stalk diameter, tassel length, leaf area and plant height. The identified metaQTLs can be further explored for fine mapping and candidate gene identification, which can be validated by loss or gain of function. Identified metaQTLs can be used for introgression of popping traits towards enhancing the popping ability.

Starch molecular configuration and starch-sugar homeostasis: Key determinants of sweet sensory perception and starch hydrolysis in pearl millet (Pennisetum glaucum).

Starch-sugar homeostasis and starch molecular configuration regulates the dynamics of starch digestibility which result in sweet sensory perception and eliciting glycemic response, which has been measured in vitro as inherent glycemic potential (IGP). The objective of the research was to understand the key determinants of IGP as well as sweetness in different Pearl millet (PM) genotypes. To understand the intricate balance between starch and sugar, total starch content (TSC) and total soluble sugars (TSS) were evaluated. Higher concentrations of TSC (67.8%), TSS (2.75%), glucose (0.78%) and sucrose (1.68%) were found in Jafarabadi Bajra. Considering the role of compact molecular configuration of starch towards digestibility, X-ray powder diffraction (XRD) analysis was performed. A-type crystallinity with crystallinity degree (CD %) ranged from 53.53-62.63% among different genotypes, where the least CD% (53.53%) was found in Jafarabadi Bajra. In vitro starch hydrolyzation kinetics carried out to determine IGP, revealed a maximum of 77.05% IGP with minimum 1.42% resistant starch (RS) in Jafarabadi Bajra. Overall our results suggest higher sweet sensory perception of Jafarabadi Bajra which is contributed by the matrix composition with least molecular compactness of starch. Also, the interdependence among starch quality parameters; CD%, IGP, RS and amylose has also been discussed.

Insights into the Host-Pathogen Interaction Pathways through RNA-Seq Analysis of Lens culinaris Medik. in Response to Rhizoctonia bataticola Infection.

Dry root rot (Rhizoctonia bataticola) is an important disease of lentils (Lens culinaris Medik.).To gain an insight into the molecular aspects of host-pathogen interactions, the RNA-seq approach was used in lentils following inoculation with R.bataticola. The RNA-Seq has generated >450 million high-quality reads (HQRs) and nearly 96.97% were properly aligned to the reference genome. Very high similarity in FPKM (fragments per kilobase of exon per million mapped fragments) values (R > 0.9) among biological replicates showed the consistency of the RNA-Seq results. The study revealed various DEGs (differentially expressed genes) that were associated with changes in phenolic compounds, transcription factors (TFs), antioxidants, receptor kinases, hormone signals which corresponded to the cell wall modification enzymes, defense-related metabolites, and jasmonic acid (JA)/ethylene (ET) pathways. Gene ontology (GO) categorization also showed similar kinds of significantly enriched similar GO terms. Interestingly, of the total unigenes (42,606), 12,648 got assembled and showed significant hit with Rhizoctonia species. String analysis also revealed the role of various disease responsive proteins viz., LRR family proteins, LRR-RLKs, protein kinases, etc. in the host-pathogen interaction. Insilico validation analysis was performed using Genevestigator® and DEGs belonging to six major defense-response groups viz., defense-related enzymes, disease responsive genes, hormones, kinases, PR (pathogenesis related) proteins, and TFs were validated. For the first time some key miRNA targets viz. miR156, miR159, miR167, miR169, and miR482 were identified from the studied transcriptome, which may have some vital role in Rhizoctonia-based responses in lentils. The study has revealed the molecular mechanisms of the lentil/R.bataticola interactions and also provided a theoretical approach for the development of lentil genotypes resistant to R.bataticola.

Characterization of Atypical Protein Tyrosine Kinase (PTK) Genes and Their Role in Abiotic Stress Response in Rice.

Tyrosine phosphorylation constitutes up to 5% of the total phophoproteome. However, only limited studies are available on protein tyrosine kinases (PTKs) that catalyze protein tyrosine phosphorylation in plants. In this study, domain analysis of the 27 annotated PTK genes in rice genome led to the identification of 18 PTKs with tyrosine kinase domain. The kinase domain of rice PTKs shared high homology with that of dual specificity kinase BRASSINOSTEROID- INSENSITIVE 1 (BRI1) of Arabidopsis. In phylogenetic analysis, rice PTKs clustered with receptor-like cytoplasmic kinases-VII (RLCKs-VII) of Arabidopsis. mRNAseq analysis using Genevestigator revealed that rice PTKs except PTK9 and PTK16 express at moderate to high level in most tissues. PTK16 expression was highly abundant in panicle at flowering stage. mRNAseq data analysis led to the identification of drought, heat, salt, and submergence stress regulated PTK genes in rice. PTK14 was upregulated under all stresses. qRT-PCR analysis also showed that all PTKs except PTK10 were significantly upregulated in root under osmotic stress. Tissue specificity and abiotic stress mediated differential regulation of PTKs suggest their potential role in development and stress response of rice. The candidate dual specificity PTKs identified in this study paves way for molecular analysis of tyrosine phosphorylation in rice.

Induction of cell death by tospoviral protein NSs and the motif critical for cell death does not control RNA silencing suppression activity.

Groundnut bud necrosis virus induces necrotic symptoms in different hosts. Previous studies showed reactive oxygen species-mediated programmed cell death (PCD) resulted in necrotic symptoms. Transgenic expression of viral protein NSs mimics viral symptoms. Here, we showed a role for NSs in influencing oxidative burst in the cell, by analyzing H2O2 accumulation, activities of antioxidant enzymes and expression levels of vacuolar processing enzymes, H2O2-responsive microRNA 319a.2 plus its possible target metacaspase-8. The role of NSs in PCD, was shown using two NSs mutants: one in the Trp/GH3 motif (a homologue of pro-apototic domain) (NSsS189R) and the other in a non-Trp/GH3 motif (NSsL172R). Tobacco rattle virus (TRV) expressing NSsS189R enhanced the PCD response, but not TRV-NSsL172R, while RNA silencing suppression activity was lost in TRV-NSsL172R, but not in TRV-NSsS189R. Therefore, we propose dual roles of NSs in RNA silencing suppression and induction of cell death, controlled by different motifs.

Novel and conserved heat-responsive microRNAs in wheat (Triticum aestivum L.).

MicroRNAs (miRNAs) are small endogenous RNAs of ~22 nucleotides that have been shown to play regulatory role by negatively affecting the expression of genes at the post-transcriptional level. Information of miRNAs on some important crops like soybean, Arabidopsis, and rice, etc. are available, but no study on heat-responsive novel miRNAs has yet been reported in wheat (Triticum aestivum L.). In the present investigation, a popular wheat cultivar HD2985 was used in small RNA library construction and Illumina HiSeq 2000 was used to perform high-throughput sequencing of the library after cluster generation; 110,896,604 and 87,743,861 reads were generated in the control (22 °C) and heat-treated (42 °C for 2 h) samples, respectively. Forty-four precursor and mature miRNAs were found in T. aestivum from miRBase v 19. The frequencies of the miRNA families varied from 2 (tae-miR1117) to 60,672 (tae-miR159b). We identify 1052 and 902 mature miRNA sequences in HD2985 control and HS-treated samples by mapping on reference draft genome of T. aestivum. Maximum identified miRNAs were located on IWGSC_CSS_3B_scaff (chromosome 3B). We could identify 53 and 46 mature miRNA in the control and HS samples and more than 516 target genes by mapping on the reference genome of Oryza sativa, Zea mays, and Sorghum bicolor. Using different pipelines and plant-specific criteria, 37 novel miRNAs were identified in the control and treated samples. Six novel miRNA were validated using qRT-PCR to be heat-responsive. A negative correlation was, however, observed between the expression of novel miRNAs and their targets. Target prediction and pathway analysis revealed their involvement in the heat stress tolerance. These novel miRNAs are new additions to miRNA database of wheat, and the regulatory network will be made use of in deciphering the mechanism of thermotolerance in wheat.