Natural diversity uncovers P5CS1 regulation and its role in drought stress tolerance and yield sustainability in barley.

Proline accumulation is one of the major responses of plants to many abiotic stresses. However, the significance of proline accumulation for drought stress tolerance remains enigmatic in crop plants. First, we examined the natural variation of the pyrolline-5-carboxylate synthase (P5CS1) among 49 barley genotypes. Allele mining identified a previously unknown allelic series that showed polymorphisms at 42 cis-elements across the P5CS1 promoter. Selected haplotypes had quantitative variation in P5CS1 gene expression and proline accumulation, putatively influenced by both  abscisic acid-dependent and independent pathways under drought stress. Next, we introgressed the P5CS1 allele from a high proline accumulating wild barley accession ISR42-8 into cultivar Scarlett developing a near-isogenic line (NIL-143). NIL-143 accumulated higher proline concentrations than Scarlett under drought stress at seedling and reproductive stages. Under drought stress, NIL-143 showed less pigment damage, sustained photosynthetic health, and higher drought stress recovery compared to Scarlett. Further, the drought-induced damage to yield-related traits, mainly thousand-grain weight, was lower in NIL-143 compared with Scarlett in field conditions. Our data uncovered new variants of the P5CS1 promoter and the significance of the increased proline accumulation regulated by the P5CS1 allele of ISR42-8 in drought stress tolerance and yield stability in barley.

Proline-Mediated Drought Tolerance in the Barley (Hordeum vulgare L.) Isogenic Line Is Associated with Lateral Root Growth at the Early Seedling Stage.

A vigorous root system in barley promotes water uptake from the soil under water-limited conditions. We investigated three spring barley genotypes with varying water stress responses using rhizoboxes at the seedling stage. The genotypes comprised two elite German cultivars, Barke and Scarlett, and a near-isogenic line, NIL 143. The isogenic line harbors the wild allele pyrroline-5-carboxylate synthase1-P5cs1. Root growth in rhizoboxes under reduced water availability conditions caused a significant reduction in total root length, rooting depth, root maximum width, and root length density. On average, root growth was reduced by more than 20% due to water stress. Differences in organ proline concentrations were observed for all genotypes, with shoots grown under water stress exhibiting at least a 30% higher concentration than the roots. Drought induced higher leaf and root proline concentrations in NIL 143 compared with any of the other genotypes. Under reduced water availability conditions, NIL 143 showed less severe symptoms of drought, higher lateral root length, rooting depth, maximum root width, root length density, and convex hull area compared with Barke and Scarlett. Within the same comparison, under water stress, NIL 143 had a higher proportion of lateral roots (+30%), which were also placed at deeper substrate horizons. NIL 143 had a less negative plant water potential and higher relative leaf water content and stomatal conductance compared with the other genotypes under water stress. Under these conditions, this genotype also maintained an enhanced net photosynthetic rate and exhibited considerable fine root growth (diameter class 0.05-0.35 mm). These results show that water stress induces increased shoot and root proline accumulation in the NIL 143 barley genotype at the seedling stage and that this effect is associated with increased lateral root growth.

Lr21 diversity unveils footprints of wheat evolution and its new role in broad-spectrum leaf rust resistance.

Aegilops tauschii, the progenitor of the wheat D genome, contains extensive diversity for biotic and abiotic resistance. Lr21 is a leaf rust resistance gene, which did not enter the initial gene flow from Ae. tauschii into hexaploid wheat due to restrictive hybridization events. Here, we used population genetics and high-resolution comparative genomics to study evolutionary and functional divergence of Lr21 in diploid and hexaploid wheats. Population genetics identified the original Lr21, lr21-1 and lr21-2 alleles and their evolutionary history among Ae. tauschii accessions. Comparative genetics of Lr21 variants between Ae. tauschii and cultivated genotypes suggested at least two independent polyploidization events in bread wheat evolution. Further, a recent re-birth of a unique Lr21-tbk allele and its neofunctionalization was discovered in the hexaploid wheat cv. Tobak. Altogether, four independent alleles were investigated and validated for leaf rust resistance in diploid, synthetic hexaploid and cultivated wheat backgrounds. Besides seedling resistance, we uncover a new role of the Lr21 gene in conferring an adult plant field resistance. Seedling and adult plant resistance turned out to be correlated with developmentally dependent variation in Lr21 expression. Our results contribute to understand Lr21 evolution and its role in establishing a broad-spectrum leaf rust resistance in wheat.

A Wild Allele of Pyrroline-5-Carboxylate Synthase1 Leads to Proline Accumulation in Spikes and Leaves of Barley Contributing to Improved Performance Under Reduced Water Availability.

Water stress (WS) during spike development strongly affects final grain yield and grain quality in cereals. Proline, an osmoprotectant amino-acid, may contribute to alleviating the effects of cell and tissue dehydration. We studied five spring barley genotypes contrasting in their drought response, including two introgression lines, S42IL-143 and S42IL-141, harboring a Pyrroline-5-carboxylate synthase1- P5cs1 allele originating from the wild barley accession ISR42-8. We tested the hypothesis that barley genotypes harboring a wild allele at P5cs1 locus are comparatively more drought-tolerant at the reproductive stage by inducing proline accumulation in their immature spikes. At the booting stage, we subjected plants to well-watered and WS treatments until physiological maturity. Several morpho-physiological traits had significant genotype by treatment interaction and reduction under WS. Varying levels of genotypic proline accumulation and differences in WS tolerance were observed. Spike proline accumulation was higher than leaf proline accumulation for all genotypes under WS. Also, introgression lines carrying a wild allele at P5cs1 locus had a markedly higher spike and leaf proline content compared with the other genotypes. These introgression lines showed milder drought symptoms compared with elite genotypes, remained photosynthetically active under WS, and maintained their intrinsic water use efficiency. These combined responses contributed to the achievement of higher final seed productivity. Magnetic resonance imaging (MRI) of whole spikes at the soft dough stage showed an increase in seed abortion among the elite genotypes compared with the introgression lines 15 days after WS treatment. Our results suggest that proline accumulation at the reproductive stage contributes to the maintenance of grain formation under water shortage.

Genetics and genomics of root system variation in adaptation to drought stress in cereal crops.

Cereals are important crops worldwide that help meet food demands and nutritional needs. In recent years, cereal production has been challenged globally by frequent droughts and hot spells. A plant's root is the most relevant organ for the plant adaptation to stress conditions, playing pivotal roles in anchorage and the acquisition of soil-based resources. Thus, dissecting root system variations and trait selection for enhancing yield and sustainability under drought stress conditions should aid in future global food security. This review highlights the variations in root system attributes and their interplay with shoot architecture features to face water scarcity and maintain thus yield of major cereal crops. Further, we compile the root-related drought responsive quantitative trait loci/genes in cereal crops including their interspecies relationships using microsynteny to facilitate comparative genomic analyses. We then discuss the potential of an integrated strategy combining genomics and phenomics at genetic and epigenetic levels to explore natural genetic diversity as a basis for knowledge-based genome editing. Finally, we present an outline to establish innovative breeding leads for the rapid and optimized selection of root traits necessary to develop resilient crop varieties.

Genetic components of root architecture and anatomy adjustments to water-deficit stress in spring barley.

Roots perform vital roles for adaptation and productivity under water-deficit stress, even though their specific functions are poorly understood. In this study, the genetic control of the nodal-root architectural and anatomical response to water deficit were investigated among diverse spring barley accessions. Water deficit induced substantial variations in the nodal root traits. The cortical, stele, and total root cross-sectional areas of the main-shoot nodal roots decreased under water deficit, but increased in the tiller nodal roots. Root xylem density and arrested nodal roots increased under water deficit, with the formation of root suberization/lignification and large cortical aerenchyma. Genome-wide association study implicated 11 QTL intervals in the architectural and anatomical nodal root response to water deficit. Among them, three and four QTL intervals had strong effects across seasons and on both root architectural and anatomical traits, respectively. Genome-wide epistasis analysis revealed 44 epistatically interacting SNP loci. Further analyses showed that these QTL intervals contain important candidate genes, including ZIFL2, MATE, and PPIB, whose functions are shown to be related to the root adaptive response to water deprivation in plants. These results give novel insight into the genetic architectures of barley nodal root response to soil water deficit stress in the fields, and thus offer useful resources for root-targeted marker-assisted selection.

Genome-wide association study uncovers a novel QTL allele of AtS40-3 that affects the sex ratio of cyst nematodes in Arabidopsis.

Plant-parasitic cyst nematodes are obligate sedentary parasites that infect the roots of a broad range of host plants. Cyst nematodes are sexually dimorphic, but differentiation into male or female is strongly influenced by interactions with the host environment. Female populations typically predominate under favorable conditions, whereas male populations predominate under adverse conditions. Here, we performed a genome-wide association study (GWAS) in an Arabidopsis diversity panel to identify host loci underlying variation in susceptibility to cyst nematode infection. Three different susceptibility parameters were examined, with the aim of providing insights into the infection process, the number of females and males present in the infected plant, and the female-to-male sex ratio. GWAS results suggested that variation in sex ratio is associated with a novel quantitative trait locus allele on chromosome 4. Subsequent candidate genes and functional analyses revealed that a senescence-associated transcription factor, AtS40-3, and PPR may act in combination to influence nematode sex ratio. A detailed molecular characterization revealed that variation in nematode sex ratio was due to the disturbed common promoter of AtS40-3 and PPR genes. Additionally, single nucleotide polymorphisms in the coding sequence of AtS40-3 might contribute to the natural variation in nematode sex ratio.

Genome-Wide Association Mapping in the Global Diversity Set Reveals New QTL Controlling Root System and Related Shoot Variation in Barley.

The fibrous root system is a visible sign of ecological adaptation among barley natural populations. In the present study, we utilized rich barley diversity to dissect the genetic basis of root system variation and its link with shoot attributes under well-water and drought conditions. Genome-wide association mapping of phenotype data using a dense genetic map (5892 SNP markers) revealed 17 putative QTL for root and shoot traits. Among these, at 14 loci the preeminence of exotic QTL alleles resulted in trait improvements. The most promising QTL were quantified using haplotype analysis at local and global genome levels. The strongest QTL was found on chromosome 1H which accounted for root dry weight and tiller number simultaneously. Candidate gene analysis across the targeted region detected a crucial amino acid substitution mutation in the conserved domain of a WRKY29 transcription factor among genotypes bearing major and minor QTL alleles. Similarly, the drought inducible QTL QRdw.5H (5H, 95.0 cM) seems to underlie 37 amino acid deletion and substitution mutations in the conserved domain of two related genes CBF10B and CBF10A, respectively. The identification and further characterization of these candidate genes will be essential to decipher genetics behind developmental and natural adaptation mechanisms of barley.

Landscape genomics reveal signatures of local adaptation in barley (Hordeum vulgare L.).

Land plants are sessile organisms that cannot escape the adverse climatic conditions of a given environment. Hence, adaptation is one of the solutions to surviving in a challenging environment. This study was aimed at detecting adaptive loci in barley landraces that are affected by selection. To that end, a diverse population of barley landraces was analyzed using the genotyping by sequencing approach. Climatic data for altitude, rainfall and temperature were collected from 61 weather sites near the origin of selected landraces across Ethiopia. Population structure analysis revealed three groups whereas spatial analysis accounted significant similarities at shorter geographic distances (< 40 Km) among barley landraces. Partitioning the variance between climate variables and geographic distances indicated that climate variables accounted for most of the explainable genetic variation. Markers by climatic variables association analysis resulted in altogether 18 and 62 putative adaptive loci using Bayenv and latent factor mixed model (LFMM), respectively. Subsequent analysis of the associated SNPs revealed putative candidate genes for plant adaptation. This study highlights the presence of putative adaptive loci among barley landraces representing original gene pool of the farming communities.

AB-QTL analysis reveals new alleles associated to proline accumulation and leaf wilting under drought stress conditions in barley (Hordeum vulgare L.).

BACKGROUND: Land plants have evolved several measures to maintain their life against abiotic stresses. The accumulation of proline is the most generalized response of plants under drought, heat or salt stress conditions. It is known as an osmoprotectant which also acts as an instant source of energy during drought recovery process. But, both its role and genetic inheritance are poorly understood in agriculture crops. In the present work, advanced backcross quantitative trait locus (AB-QTL) analysis was performed to elucidate genetic mechanisms controlling proline accumulation and leaf wilting in barley under drought stress conditions. RESULTS: The analysis revealed eight QTL associated to proline content (PC) and leaf wilting (WS). QTL for PC were localized on chromosome 3H, 4H, 5H and 6H. The strongest QTL effect QPC.S42.5H was detected on chromosome 5H where drought inducible exotic allele was associated to increase PC by 54%. QTL effects QPC.S42.3H, QPC.S42.4H and QPC.S42.6H were responsible to heighten PC due to the preeminence of elite alleles over the exotic alleles which ranged from 26% to 43%. For WS, QTL have been localized on chromosome 1H, 2H, 3H and 4H. Among these, QWS.S42.1H and QWS.S42.4H were associated to decrease in WS due to the introgression of exotic alleles. In addition, two digenic epistatic interaction effects were detected for WS where the additive effect of exotic alleles imparted a favorable increase in the trait value. CONCLUSIONS: The present data represents a first report on whole-genome mapping of proline accumulation and leaf wilting in barley. The detected QTL are linked to new alleles from both cultivated and wild accessions which bring out an initial insight on the genetic inheritance of PC and WS. These QTL alleles are fixed in the isogenic background of Scarlett, which will allow for positional cloning of underlying genes and to develop drought resilient barley cultivars.