Genome-wide identification and functional characterisation of the EDS1 gene family reveals evolutionary conservation and stress-responsive regulatory roles in barley.

Enhanced Disease Susceptibility 1 (EDS1) genes are central regulators of plant innate immunity and have emerging roles in biotic stress adaptation. Although extensively characterized in dicots, a genome-wide investigation of the EDS1 gene family in barley (Hordeum vulgare L.) remains unavailable. Thirteen HvEDS1 genes were systematically identified by integrated HMM and BLASTP approaches. These genes encode structurally distinct proteins that differ in length, charge, stability and subcellular localization, with several of them reported to occur in the nucleus, chloroplasts, mitochondria and cytoskeleton. Phylogenetic analyses subdivided the HvEDS1 proteins into three major clades, revealing both a diversification of monocots and conserved ancestors. Analyses of gene structure and conserved motifs revealed clade-specific exon–intron architectures and domain configurations, suggesting functional specialization. Analysis of the genome distribution showed that the spread of HvEDS1 is primarily due to scattered duplications, with purifying selection acting on the duplicated pairs. Analysis of the promoters identified numerous cis-regulatory elements that respond to hormones (ABA, JA), light (G-box) and abiotic influences (MBS), suggesting multi-layered transcriptional regulation. The predicted miRNA–mRNA interactions revealed that hvu-miR6192, a stress-responsive microRNA, targets receptor-like kinases involved in abiotic stress signaling, suggesting a post-transcriptional regulatory role in HvEDS1-mediated stress responses. KEGG enrichment linked HvEDS1 genes to lipid signaling pathways, including glycerophospholipid and α-linolenic acid metabolism, as well as ubiquitin-mediated proteolysis. Structural modelling indicated conservation of key EDS1 domains, but showed protein-specific variations in loop flexibility and secondary structure content, suggesting different functional dynamics. The overall transition from a specialized, low-redundancy network under normal conditions to a highly coordinated and overlapping network under fungal stress condition reflects the versatility of the HvEDS1 family in rewiring of signaling cascades stress responsive pathways during different physiological conditions. This first genome-wide characterization of the HvEDS1 gene family reveals its structural diversity, evolutionary development and potential role in immunity and abiotic stress signaling. The identified candidates provide valuable targets for functional validation and molecular breeding of stress-resistant barley cultivars.