Genome-wide analysis of expansin superfamily in wild Arachis discloses a stress-responsive expansin-like B gene

Expansins are plant cell wall-loosening proteins involved in adaptive responses to environmental stimuli and various developmental processes. The first genome-wide analysis of the expansin superfamily in the Arachis genus identified 40 members in A. duranensis and 44 in A. ipaënsis, the wild progenitors of cultivated peanut (A. hypogaea). These expansins were further characterized regarding their subfamily classification, distribution along the genomes, duplication events, molecular structure, and phylogeny. A RNA-seq expression analysis in different Arachis species showed that the majority of these expansins are modulated in response to diverse stresses such as water deficit, rootknot nematode (RKN) infection, and UV exposure, with an expansin-like B gene (AraEXLB8) displaying a highly distinct stress-responsive expression profile. Further analysis of the AraEXLB8 coding sequences showed high conservation across the Arachis genotypes, with eight haplotypes identified. The modulation of AraEXLB8 expression in response to the aforementioned stresses was confirmed by qRT-PCR analysis in distinct Arachis genotypes, whilst in situ hybridization revealed transcripts in different root tissues according to the stress imposed. The overexpression of AraEXLB8 in soybean (Glycine max) composite plants remarkably decreased the number of galls in transformed hairy roots inoculated with RKN. This study improves the current understanding of the molecular evolution, divergence, and gene expression of expansins in Arachis, and provides molecular and functional insights into the role of expansin-like B, the less-studied plant expansin subfamily

Defining the combined stress response in wild Arachis

International audienceNematodes and drought are major constraints in tropical agriculture and often occur simultaneously. Plant responses to these stresses are complex and require crosstalk between biotic and abiotic signaling pathways. In this study, we explored the transcriptome data of wild Arachis species subjected to drought (A-metaDEG) and the root-knot nematode Meloidogyne arenaria (B-metaDEG) via meta-analysis, to identify core-stress responsive genes to each individual and concurrent stresses in these species. Transcriptome analysis of a nematode/drought bioassay (cross-stress) showed that the set of stress responsive DEGs to concurrent stress is distinct from those resulting from overlapping A- and B-metaDEGs, indicating a specialized and unique response to combined stresses in wild Arachis . Whilst individual biotic and abiotic stresses elicit hormone-responsive genes, most notably in the jasmonic and abscisic acid pathways, combined stresses seem to trigger mainly the ethylene hormone pathway. The overexpression of a cross-stress tolerance candidate gene identified here, an endochitinase-encoding gene ( AsECHI ) from Arachis stenosperma , reduced up to 30% of M. incognita infection and increased post-drought recovery in Arabidopsis plants submitted to both stresses. The elucidation of the network of cross-stress responsive genes in Arachis contributes to better understanding the complex regulation of biotic and abiotic responses in plants facilitating more adequate crop breeding for combined stress tolerance

Transcriptome Responses of Wild Arachis to UV-C Exposure Reveal Genes Involved in General Plant Defense and Priming

Stress priming is an important strategy for enhancing plant defense capacity to deal with environmental challenges and involves reprogrammed transcriptional responses. Although ultraviolet (UV) light exposure is a widely adopted approach to elicit stress memory and tolerance in plants, the molecular mechanisms underlying UV-mediated plant priming tolerance are not fully understood. Here, we investigated the changes in the global transcriptome profile of wild Arachis stenosperma leaves in response to UV-C exposure. A total of 5751 differentially expressed genes (DEGs) were identified, with the majority associated with cell signaling, protein dynamics, hormonal and transcriptional regulation, and secondary metabolic pathways. The expression profiles of DEGs known as indicators of priming state, such as transcription factors, transcriptional regulators and protein kinases, were further characterized. A meta-analysis, followed by qRT-PCR validation, identified 18 metaDEGs as being commonly regulated in response to UV and other primary stresses. These genes are involved in secondary metabolism, basal immunity, cell wall structure and integrity, and may constitute important players in the general defense processes and establishment of a priming state in A. stenosperma. Our findings contribute to a better understanding of transcriptional dynamics involved in wild Arachis adaptation to stressful conditions of their natural habitats

A novel soybean hairy root system for gene functional validation.

Agrobacterium rhizogenes-mediated transformation has long been explored as a versatile and reliable method for gene function validation in many plant species, including soybean (Glycine max). Likewise, detached-leaf assays have been widely used for rapid and mass screening of soybean genotypes for disease resistance. The present study combines these two methods to establish an efficient and practical system to generate transgenic soybean hairy roots from detached leaves and their subsequent culture under ex vitro conditions. We demonstrated that hairy roots derived from leaves of two (tropical and temperate) soybean cultivars could be successfully infected by economically important species of root-knot nematodes (Meloidogyne incognita and M. javanica). The established detached-leaf method was further explored for functional validation of two candidate genes encoding for cell wall modifying proteins (CWMPs) to promote resistance against M. incognita through distinct biotechnological strategies: the overexpression of a wild Arachis α-expansin transgene (AdEXPA24) and the dsRNA-mediated silencing of an endogenous soybean polygalacturonase gene (GmPG). AdEXPA24 overexpression in hairy roots of RKN-susceptible soybean cultivar significantly reduced nematode infection by approximately 47%, whereas GmPG downregulation caused an average decrease of 37%. This novel system of hairy root induction from detached leaves showed to be an efficient, practical, fast, and low-cost method suitable for high throughput in root analysis of candidate genes in soybean

Early responses to dehydration in contrasting wild <i>Arachis</i> species

<div><p>Wild peanut relatives (<i>Arachis</i> spp.) are genetically diverse and were selected throughout evolution to a range of environments constituting, therefore, an important source of allelic diversity for abiotic stress tolerance. In particular, <i>A</i>. <i>duranensis</i> and <i>A</i>. <i>stenosperma</i>, the parents of the reference <i>Arachis</i> A-genome genetic map, show contrasting transpiration behavior under limited water conditions. This study aimed to build a comprehensive gene expression profile of these two wild species under dehydration stress caused by the withdrawal of hydroponic nutrient solution. For this purpose, roots of both genotypes were collected at seven time-points during the early stages of dehydration and used to construct cDNA paired-end libraries. Physiological analyses indicated initial differences in gas exchange parameters between the drought-tolerant genotype of <i>A</i>. <i>duranensis</i> and the drought-sensitive genotype of <i>A</i>. <i>stenosperma</i>. High-quality Illumina reads were mapped against the <i>A</i>. <i>duranensis</i> reference genome and resulted in the identification of 1,235 and 799 Differentially Expressed Genes (DEGs) that responded to the stress treatment in roots of <i>A</i>. <i>duranensis</i> and <i>A</i>. <i>stenosperma</i>, respectively. Further analysis, including functional annotation and identification of biological pathways represented by these DEGs confirmed the distinct gene expression behavior of the two contrasting <i>Arachis</i> species genotypes under dehydration stress. Some species-exclusive and common DEGs were then selected for qRT-PCR analysis, which corroborated the <i>in silico</i> expression profiling. These included genes coding for regulators and effectors involved in drought tolerance responses, such as activation of osmosensing molecular cascades, control of hormone and osmolyte content, and protection of macromolecules. This dataset of transcripts induced during the dehydration process in two wild <i>Arachis</i> genotypes constitute new tools for the understanding of the distinct gene regulation processes in these closely related species but with contrasting drought responsiveness. In addition, our findings provide insights into the nature of drought tolerance in wild germoplasm, which might be explored as novel sources of diversity and useful wild alleles to develop climate-resilient crop varieties.</p></div

Clusters of Ortholog Groups (OGs) from four legume species.

<p>Venn diagram indicating the number of Ortholog Groups (OGs) among <i>Arachis duranensis; Arachis ipaënsis; Glycine max; and Phaseolus vulgaris</i>. The number of DEGs in the different groups is in parentheses.</p

Circos plot detailing chromosome distribution of DEGs.

<p>Distribution of the 1,602 DEGs in the ten chromosomes of <i>A</i>. <i>duranensis</i> (A01 to A10). The outer lines represent exclusive <i>A</i>. <i>duranensis</i> (red) and <i>A</i>. <i>stenosperma</i> (blue) and common (green) DEGs. The inner dots represent the distribution of Log2FC values for each up- and downregulated DEG in <i>A</i>. <i>duranensis</i> (red) and <i>A</i>. <i>stenosperma</i> (blue), with the line indicating Log2FC = 0.</p

Expression of DEGs as determined by qRT-PCR.

<p>Relative quantification of mRNA levels of 15 candidate genes in <i>A</i>. <i>duranensis</i> roots during the dehydration treatment and collected after 25 (T25); 50 (T50); 75 (T75); 100 (T100); 125 (T125) and 150 (T150) min, relative to control (T0). Bars represent the standard deviation of three biological replicates. Significantly (P < 0.05) up- or downregulated genes are indicated by asterisks.</p

Overview of DEGs expression patterns.

<p>MapMan analysis showing molecular functional categories of DEGs expression patterns in roots of dehydration-stressed plants of <i>A</i>. <i>duranensis</i> (DD) and <i>A</i>. <i>stenosperma</i> (SD) relative to control. Squares show the different genes encoding proteins related to drought perception and oxidative responses (A) and carbohydrate and amino acid metabolism steps (B). Upregulated genes are indicated by red squares and downregulated by blue squares.</p