Comprehensive in silico analysis of the underutilized crop tef (Eragrostis tef (Zucc.) Trotter) genome reveals drought tolerance signatures.

BACKGROUND: Tef (Eragrostis tef) is a C4 plant known for its tiny, nutritious, and gluten-free grains. It contains higher levels of protein, vitamins, and essential minerals like calcium (Ca), iron (Fe), copper (Cu), and zinc (Zn) than common cereals. Tef is cultivated in diverse ecological zones under diverse climatic conditions. Studies have shown that tef has great diversity in withstanding environmental challenges such as drought. Drought is a major abiotic stress severely affecting crop productivity and becoming a bottleneck to global food security. Here, we used in silico-based functional genomic analysis to identify drought-responsive genes in tef and validated their expression using quantitative RT-PCR. RESULTS: We identified about 729 drought-responsive genes so far reported in six crop plants, including rice, wheat, maize, barley, sorghum, pearl millet, and the model plant Arabidopsis, and reported 20 genes having high-level of GO terms related to drought, and significantly enriched in several biological and molecular function categories. These genes were found to play diverse roles, including water and fluid transport, resistance to high salt, cold, and drought stress, abscisic acid (ABA) signaling, de novo DNA methylation, and transcriptional regulation in tef and other crops. Our analysis revealed substantial differences in the conserved domains of some tef genes from well-studied rice orthologs. We further analyzed the expression of sixteen tef orthologs using quantitative RT-PCR in response to PEG-induced osmotic stress. CONCLUSIONS: The findings showed differential regulation of some drought-responsive genes in shoots, roots, or both tissues. Hence, the genes identified in this study may be promising candidates for trait improvement in crops via transgenic or gene-editing technologies.

Integrative analysis of transcriptome and metabolism reveals potential roles of carbon fixation and photorespiratory metabolism in response to drought in Shanlan upland rice.

Shanlan upland rice is an important landrace rice resource and is characterized with high drought stress (DS) tolerance relative to cultivated rice. However, the molecular mechanism of DS response in Shanlan upland rice remains unclear. In this study, we performed an integrated analysis of transcriptome and targeted metabolism to decipher the key biological pathways that responded to drought tolerance using two Shanlan upland rice lines. Results show that SL10 possesses 64% higher photosynthetic efficiency (Pn) and 2-fold higher water use efficiency (WUE) than that in SL1 exposed to DS. The decrease in Pn by DS is not due to stomatal limitation effects for SL1. Transcriptome analysis suggests photosynthesis relevant pathways (photosynthesis-antenna proteins and carbon fixation) and photorespiration relevant pathway (glycine, serine and threonine metabolism) in SL1 under DS were significantly enriched in the down-regulated and up-regulated DEGs list, respectively. There are 412 up-regulated and 233 down-regulated drought responsive genes (DRGs) in SL10 relative to SL1 induced by DS. Targeted metabolism results suggest that the contents across five metabolites related to carbon fixation pathway were declined by 36 and 8% in SL1 and SL10 caused by DS, respectively. We finally summarized the both gene expression and metabolites involved in photorespiration and carbon fixation pathways in response to DS in both rice lines. This study provides valuable information for better understanding the molecular mechanism underlying drought tolerance in Shanlan rice.

Whole plant response of Pongamia pinnata to drought stress tolerance revealed by morpho-physiological, biochemical and transcriptome analysis.

BACKGROUND: Pongamia is considered an important biofuel species worldwide. Drought stress in the early growth stages of Pongamia influences negatively on the germination and seedling development. Due to lack of cultivar stability under drought stress conditions, establishment of successful plantation in drought hit areas becomes a major problem. To address this issue, drought stress response of four Pongamia genotypes was studied at morphological, physio-chemical and transcriptome levels. METHODS AND RESULTS: Drought stress was levied by limiting water for 15 days on three months old seedlings of four genotypes. A significant effect of water stress was observed on the traits considered. The genotype NRCP25 exhibited superior morpho-physiological, biochemical drought responses. Also, the genotype had higher root length, photosynthetic pigments, higher antioxidant enzymes and solute accumulation compared to other genotypes. In addition, transcript profiling of selected drought responsive candidate genes such as trehalose phosphate synthase 1 (TPS1), abscisic acid responsive elements-binding protein 2 (ABF2-2), heat shock protein 17 (HSP 17 kDa), tonoplast intrinsic protein 1 (TIP 1-2), zinc finger homeodomain protein 2 (ZFP 2), and xyloglucan endotransglucolase 13 (XET 13) showed only up-regulation in NRCP25. Further, the transcriptome responses are in line with key physio-chemical responses exhibited by NRCP25 for drought tolerance. CONCLUSIONS: As of now, there are no systematic studies on Pongamia drought stress tolerance; therefore this study offers a comprehensive understanding of whole plant drought stress responsiveness of Pongamia. Moreover, the results support important putative trait indices with potential candidate genes for drought tolerance improvement of Pongamia.

Genomewide identification of genes involved in the potato response to drought indicates functional evolutionary conservation with Arabidopsis plants.

Potato is one of the four most important food crop plants worldwide and is strongly affected by drought. The following two pairs of potato cultivars, which are related in ancestry but show different drought tolerances, were chosen for comparative gene expression studies: Gwiazda/Oberon and Tajfun/Owacja. Comparative RNA-seq analyses of gene expression differences in the transcriptomes obtained from drought-tolerant versus drought-sensitive plants during water shortage conditions were performed. The 23 top-ranking genes were selected, 22 of which are described here as novel potato drought-responsive genes. Moreover, all but one of the potato genes selected have homologues in the Arabidopsis genome. Of the seven tested A. thaliana mutants with altered expression of the selected homologous genes, compared to the wild-type Arabidopsis plants, six showed an improved tolerance to drought. These genes encode carbohydrate transporter, mitogen-activated protein kinase kinase kinase 15 (MAPKKK15), serine carboxypeptidase-like 19 protein (SCPL19), armadillo/beta-catenin-like repeat-containing protein, high-affinity nitrate transporter 2.7 and nonspecific lipid transfer protein type 2 (nsLPT). The evolutionary conservation of the functions of the selected genes in the plant response to drought confirms the importance of these identified potato genes in the ability of plants to cope with water shortage conditions. Knowledge regarding these gene functions can be used to generate potato cultivars that are resistant to unfavourable conditions. The approach used in this work and the obtained results allowed for the identification of new players in the plant response to drought.

Transcriptome analysis revealed the drought-responsive genes in Tibetan hulless barley.

BACKGROUND: Hulless barley, also called naked barley, is an important cereal crop worldwide, serving as a healthy food both for human consumption and animal feed. Nevertheless, it often suffered from drought stress during its growth and development, resulting in a drastic reduction in barley yields. Therefore, study on molecular mechanism of hulless barley drought-tolerance is very important for increasing barley production. To investigate molecular mechanism of barley drought-resistance, this study examined co-regulated mRNAs that show a change in expression pattern under early well water, later water deficit and finally water recovery treatments, and to identify mRNAs specific to water limiting conditions. RESULTS: Total of 853 differentially expressed genes (DEGs) were detected and categorized into nine clusters, in which VI and VIII were apparently up-regulated under low relative soil moisture content (RSMC) level. The majority of genes in these two clusters was relevant to abiotic stress responses in abscisic acid (ABA) dependent and independent signaling pathway, including NCED, PYR/PYL/RCAR, SnRK2, ABF, MYB/MYC, AP2/ERF family, LEA and DHN. In contrast, genes within clusters II and IV were generally down-regulated under water stress; cluster IX genes were up-regulated during water recovery response to both low and high RSMC levels. Genes in implicated in tetrapyrrole binding, photosystem and photosynthetic membrane were the most affected in cluster IX. CONCLUSION: Taken together, our findings indicate that the responses of hulless barley to drought stress shows differences in the pathways and genes activated. Furthermore, all these genes displayed different sensitivities to soil water deficit and might be profitable for future drought tolerance improvement in barley and other crops.

Gene expression analysis of potato drought-responsive genes under drought stress in potato (Solanum tuberosum L.) cultivars.

The potato (Solanum tuberosum L.), an important field crop consumed extensively worldwide, is adversely affected by abiotic stress factors especially drought. Therefore, it is vital to understand the genetic mechanism under drought stress to decrease loose of yield and quality . This trial aimed to screen drought-responsive gene expressions of potato and determine the drought-tolerant potato cultivar. The trial pattern is a completely randomized block design (CRBD) with four replications under greenhouse conditions. Four cultivars (Brooke, Orwell, Vr808, Shc909) were irrigated with four different water regimes (control and three stress conditions), and the gene expression levels of 10 potato genes were investigated. The stress treatments as follows: Control = 100% field capacity; slight drought = 75% field capacity; moderate drought = 50% field capacity, and severe drought 25% field capacity. To understand the gene expression under drought stress in potato genotypes, RT-qPCR analysis was performed and results showed that the genes most associated with drought tolerance were the StRD22 gene, MYB domain transcription factor, StERD7, Sucrose Synthase (SuSy), ABC Transporter, and StDHN1. The StHSP100 gene had the lowest genetic expression in all cultivars. Among the cultivars, the Orwell exhibited the highest expression of the StRD22 gene under drought stress. Overall, the cultivar with the highest gene expression was the Vr808, closely followed by the Brooke cultivar. As a result, it was determined that potato cultivars Orwell, Vr808, and Brooke could be used as parents in breeding programs to develop drought tolerant potato cultivars.

Uncovering the Genomic Regions Associated with Yield Maintenance in Rice Under Drought Stress Using an Integrated Meta-Analysis Approach.

The complex trait of yield is controlled by several quantitative trait loci (QTLs). Given the global water deficit issue, the development of rice varieties suitable for non-flooded cultivation holds significant importance in breeding programs. The powerful approach of Meta-QTL (MQTL) analysis can be used for the genetic dissection of complicated quantitative traits. In the current study, a comprehensive MQTL analysis was conducted to identify consistent QTL regions associated with drought tolerance and yield-related traits under water deficit conditions in rice. In total, 1087 QTLs from 134 rice populations, published between 2000 to 2021, were utilized in the analysis. Distinct MQTL analysis of the relevant traits resulted in the identification of 213 stable MQTLs. The confidence interval (CI) for the detected MQTLs was between 0.12 and 19.7 cM. The average CI of the identified MQTLs (4.68 cM) was 2.74 times narrower compared to the average CI of the initial QTLs. Interestingly, 63 MQTLs coincided with SNP peak positions detected by genome-wide association studies for yield and drought tolerance-associated traits under water deficit conditions in rice. Considering the genes located both in the QTL-overview peaks and the SNP peak positions, 19 novel candidate genes were introduced, which are associated with drought response index, plant height, panicle number, biomass, and grain yield. Moreover, an inclusive MQTL analysis was performed on all the traits to obtain "Breeding MQTLs". This analysis resulted in the identification of 96 MQTLs with a CI ranging from 0.01 to 9.0 cM. The mean CI of the obtained MQTLs (2.33 cM) was 4.66 times less than the mean CI of the original QTLs. Thirteen MQTLs fulfilling the criteria of having more than 10 initial QTLs, CI < 1 cM, and an average phenotypic variance explained greater than 10%, were designated as "Breeding MQTLs". These findings hold promise for assisting breeders in enhancing rice yield under drought stress conditions.

Responses to Drought Stress in Poplar: What Do We Know and What Can We Learn?

Poplar (Populus spp.) is a high-value crop for wood and biomass production and a model organism for tree physiology and genomics. The early release, in 2006, of the complete genome sequence of P. trichocarpa was followed by a wealth of studies that significantly enriched our knowledge of complex pathways inherent to woody plants, such as lignin biosynthesis and secondary cell wall deposition. Recently, in the attempt to cope with the challenges posed by ongoing climate change, fundamental studies and breeding programs with poplar have gradually shifted their focus to address the responses to abiotic stresses, particularly drought. Taking advantage from a set of modern genomic and phenotyping tools, these studies are now shedding light on important processes, including embolism formation (the entry and expansion of air bubbles in the xylem) and repair, the impact of drought stress on biomass yield and quality, and the long-term effects of drought events. In this review, we summarize the status of the research on the molecular bases of the responses to drought in poplar. We highlight how this knowledge can be exploited to select more tolerant genotypes and how it can be translated to other tree species to improve our understanding of forest dynamics under rapidly changing environmental conditions.

Foliar Application of Oil Palm Wood Vinegar Enhances Pandanus amaryllifolius Tolerance under Drought Stress.

Drought stress severely threatens plant growth, yield and survivability. Wood vinegar, formed by the condensation of smoke produced during biochar production, has been shown to promote plant growth and enhance stress tolerance. They have now been recognized as a sustainable alternative and are frequently used exogenously to support plants coping with environmental stress. This study aimed to evaluate the efficacy of oil palm wood vinegar (OPWV) in mitigating the adverse effects of drought stress on Pandanus amaryllifolius. The optimal concentrations and frequencies of OPWV application were determined before the drought treatment. The results showed that the imposed drought stress negatively affected the plant growth parameters but applying OPWV at 1:500 dilution at 3-day intervals for 12 days increased its tolerance. These include increased leaf relative water content, root-to-shoot ratio, relative stem circumference, chlorophyll pigments and antioxidant enzyme activities. In contrast, the drought-stressed plants treated with OPWV showed decreased relative electrolyte leakage, hydrogen peroxide, proline, malondialdehyde, and enhanced drought-responsive gene expressions, such as HSP70, GAPDH, and Thau, while ENO and ß-Fruc were reduced. These biostimulatory effects of OPWV might be due to several antioxidant compounds, such as anthranilic acid, tetrasiloxane, syringol, guaiacol, and catechol. Altogether, our results showed the effectiveness of OPWV in alleviating the adverse effects of drought stress, and as such, OPWV could be potentially applied in agriculture.

Unveiling resilience mechanisms of Quercus ilex seedlings to severe water stress: Changes in non-structural carbohydrates, xylem hydraulic functionality and wood anatomy.

Over the last few decades, extensive dieback and mortality episodes of Quercus ilex L. have been documented after severe drought events in many Mediterranean forests. However, the underlying physiological, anatomical, and biochemical mechanisms remain poorly understood. We investigated the physiological and biochemical processes linked to embolism formation and non-structural carbohydrates (NSCs) dynamics in Q. ilex seedlings exposed to severe water stress and rewatering. Measurements of leaf gas exchange, water relations, non-structural carbohydrates, drought-related gene expression, and anatomical changes in wood parenchyma were assessed. Under water stress, the midday stem water potential dropped below - 4.5 MPa corresponding to a ~ 50 % loss of hydraulic conductivity. A 70 % reduction in stomatal conductance led to a strong depletion of wood NSCs. Starch consumption, resulting from the upregulation of the ß-amylase gene BAM3, together with the downregulation of glucose (GPT1) and sucrose (SUC27) transport genes, suggests glucose utilization to sustain cellular metabolism in the wood parenchyma. After rewatering, the presence of residual xylem embolism led to an incomplete recovery of leaf gas exchanges. However, the partial restoration of photosynthesis allowed the accumulation of new starch reserves in the wood parenchyma and the production of new narrower vessels. In addition, changes in the cell wall composition of the wood parenchyma fibers were observed. Our findings indicate that thirty days of rewatering were sufficient to restore the NSCs reserves and growth rates of Q. ilex seedlings and that the carryover effects of water stress were primarily caused by hydraulic dysfunction.

Impact of Pre-Anthesis Drought Stress on Physiology, Yield-Related Traits, and Drought-Responsive Genes in Green Super Rice.

Optimum soil water availability is vital for maximum yield production in rice which is challenged by increasing spells of drought. The reproductive stage drought is among the main limiting factors leading to the drastic reduction in grain yield. The objective of this study was to investigate the molecular and morphophysiological responses of pre-anthesis stage drought stress in green super rice. The study assessed the performance of 26 rice lines under irrigated and drought conditions. Irrigated treatment was allowed to grow normally, while drought stress was imposed for 30 days at the pre-anthesis stage. Three important physiological traits including pollen fertility percentage (PFP), cell membrane stability (CMS), and normalized difference vegetative index (NDVI) were recorded at anthesis stage during the last week of drought stress. Agronomic traits of economic importance including grain yield were recorded at maturity stage. The analysis of variance demonstrated significant variation among the genotypes for most of the studied traits. Correlation and principal component analyses demonstrated highly significant associations of particular agronomic traits with grain yield, and genetic diversity among genotypes, respectively. Our study demonstrated a higher drought tolerance potential of GSR lines compared with local cultivars, mainly by higher pollen viability, plant biomass, CMS, and harvest index under drought. In addition, the molecular basis of drought tolerance in GSR lines was related to upregulation of certain drought-responsive genes including OsSADRI, OsDSM1, OsDT11, but not the DREB genes. Our study identified novel drought-responsive genes (LOC_Os11g36190, LOC_Os12g04500, LOC_Os12g26290, and LOC_Os02g11960) that could be further characterized using reverse genetics to be utilized in molecular breeding for drought tolerance.

Comparative analysis of drought-responsive transcriptomes of sugarcane genotypes with differential tolerance to drought.

Water stress causes considerable yield losses in sugarcane. To investigate differentially expressed genes under water stress, two sugarcane genotypes were subjected to three water-deficit levels (mild, moderate, and severe) and subsequent recovery and leaf transcriptome was generated using Illumina NextSeq sequencing. Among the differentially expressed genes, the tolerant genotype Co 06022 generated 2970 unigenes (p ≤ 0.05, functionally known, non-redundant DEGs) at 2-day stress, and there was a progressive decrease in the expressed genes as the stress period increased with 2109 unigenes at 6-day stress and 2307 unigenes at 10-day stress. There was considerable reduction at recovery with 1334 unigenes expressed at 10 days after recovery. However, in the susceptible genotype Co 8021, the number of unigenes expressed at 2 days was lower (2025) than the tolerant genotype and a further reduction was seen at 6-day stress (1552). During recovery, more differentially expressed genes were observed in the susceptible cultivar indicating that the cultivar has to activate more functions/processes to recover from the damage caused by stress. Comparison of DEGs between all stages of stress and recovery in both genotypes revealed that, the commonly up- and down-regulated genes across different stages were approximately double in the tolerant genotype. The most enriched gene ontology classes were heme binding, peroxidase activity and metal ion binding in the biological process and response to oxidative stress, hydrogen peroxide catabolic process and response to stress in the molecular function category. The cellular component was enriched with DEGs involved in extracellular region followed by integral component of membrane. The KEGG pathway analysis revealed important metabolic activities and functionally important genes involved in mitigating water-deficit stress in both the varieties. In addition, several unannotated genes in important pathways were detected and together may provide novel insights into water-deficit tolerance mechanisms in sugarcane. The reliability of the observed expression patterns was confirmed by qRT-PCR. The results of this study will help to identify useful genes for improving drought tolerance in sugarcane.

Faba bean drought responsive gene identification and validation.

This study was carried out to identify drought-responsive genes in a drought tolerant faba bean variety (Hassawi 2) using a suppressive subtraction hybridization approach (SSH). A total of 913 differentially expressed clones were sequenced from a differential cDNA library that resulted in a total of 225 differentially expressed ESTs. The genes of mitochondrial and chloroplast origin were removed, and the remaining 137 EST sequences were submitted to the gene bank EST database (LIBEST_028448). A sequence analysis identified 35 potentially drought stress-related ESTs that regulate ion channels, kinases, and energy production and utilization and transcription factors. Quantitative PCR on Hassawi 2 genotype confirmed that more than 65% of selected drought-responsive genes were drought-related. Among these induced genes, the expression levels of eight highly up-regulated unigenes were further analyzed across 38 selected faba bean genotypes that differ in their drought tolerance levels. These unigenes included ribulose 1,5-bisphosphate carboxylase (rbcL) gene, non-LTR retroelement reverse related, probable cyclic nucleotide-gated ion channel, polyubiquitin, potassium channel, calcium-dependent protein kinase and putative respiratory burst oxidase-like protein C and a novel unigene. The expression patterns of these unigenes were variable across 38 genotypes however, it was found to be very high in tolerant genotype. The up-regulation of these unigenes in majority of tolerant genotypes suggests their possible role in drought tolerance. The identification of possible drought responsive candidate genes in Vicia faba reported here is an important step toward the development of drought-tolerant genotypes that can cope with arid environments.

Dissection of early transcriptional responses to water stress in Arundo donax L. by unigene-based RNA-seq.

BACKGROUND: Arundo donax L. (Poaceae) is considered one of the most promising energy crops in the Mediterranean region because of its high biomass yield and low input requirements, but to date no information on its transcriptional responses to water stress is available. RESULTS: We obtained by Illumina-based RNA-seq the whole root and shoot transcriptomes of young A. donax plants subjected to osmotic/water stress with 10 and 20 % polyethylene glycol (PEG; 3 biological replicates/organ/condition corresponding to 18 RNA-Seq libraries), and identified a total of 3034 differentially expressed genes. Blast-based mining of stress-related genes indicated the higher responsivity of roots compared to shoots at the early stages of water stress especially under the milder PEG treatment, with a majority of genes responsive to salt, oxidative, and dehydration stress. Analysis of gene ontology terms underlined the qualitatively different responses between root and shoot tissues. Among the most significantly enriched metabolic pathways identified using a Fisher's exact test with FDR correction, a crucial role was played in both shoots and roots by genes involved in the signaling cascade of abscisic acid. We further identified relatively large organ-specific differences in the patterns of drought-related transcription factor AP2-EREBP, AUX/IAA, MYB, bZIP, C2H2, and GRAS families, which may underlie the transcriptional reprogramming differences between organs. Through comparative analyses with major Poaceae species based on Blast, we finally identified a set of 53 orthologs that can be considered as a core of evolutionary conserved genes important to mediate water stress responses in the family. CONCLUSIONS: This study provides the first characterization of A. donax transcriptome in response to water stress, thus shedding novel light at the molecular level on the mechanisms of stress response and adaptation in this emerging bioenergy species. The inventory of early-responsive genes to water stress identified could constitute useful markers of the physiological status of A. donax and be a basis for the improvement of its productivity under water limitation. The full water-stressed A. donax transcriptome is available for Blast-based homology searches through a dedicated web server (http://ecogenomics.fmach.it/arundo/).

Identification and Validation of Selected Universal Stress Protein Domain Containing Drought-Responsive Genes in Pigeonpea (Cajanus cajan L.).

Pigeonpea is a resilient crop, which is relatively more drought tolerant than many other legume crops. To understand the molecular mechanisms of this unique feature of pigeonpea, 51 genes were selected using the Hidden Markov Models (HMM) those codes for proteins having close similarity to universal stress protein domain. Validation of these genes was conducted on three pigeonpea genotypes (ICPL 151, ICPL 8755, and ICPL 227) having different levels of drought tolerance. Gene expression analysis using qRT-PCR revealed 6, 8, and 18 genes to be ≥2-fold differentially expressed in ICPL 151, ICPL 8755, and ICPL 227, respectively. A total of 10 differentially expressed genes showed ≥2-fold up-regulation in the more drought tolerant genotype, which encoded four different classes of proteins. These include plant U-box protein (four genes), universal stress protein A-like protein (four genes), cation/H(+) antiporter protein (one gene) and an uncharacterized protein (one gene). Genes C.cajan_29830 and C.cajan_33874 belonging to uspA, were found significantly expressed in all the three genotypes with ≥2-fold expression variations. Expression profiling of these two genes on the four other legume crops revealed their specific role in pigeonpea. Therefore, these genes seem to be promising candidates for conferring drought tolerance specifically to pigeonpea.