Genome-Wide Identification and Characterization of HvNPR1-like Genes Reveal HvNPR1 Interacting with HvWRKY70 in Barley Resistance to Leaf Stripe
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Abstract
Background: Barley leaf stripe disease, caused by the fungal pathogen Pyrenophora graminea, significantly reduces both grain yield and quality. Previous studies have demonstrated that NPR1-like proteins, as key regulators of salicylic acid (SA) signaling, play crucial roles in plant defense against fungal pathogens. However, systematic research on NPR1-like genes involved in barley leaf stripe resistance remains limited.Results: In this study, five HvNPR1-like family members were identified at the genome-wide level in barley. These genes were unevenly distributed across three chromosomes. Phylogenetic analysis based on Arabidopsis NPR1-like proteins classified the HvNPR1-Like proteins into three major groups (I, II, and III), which exhibited conserved features in terms of intron–exon organization. Synteny analysis revealed relatively few orthologous gene pairs between barley and Arabidopsis (one pair), whereas substantially more pairs were identified between barley and rice (four pairs), between barley and maize (five pairs), and between barley and wheat (twelve pairs), consistent with the phylogenetic relationships among these species.Transcriptome sequencing (RNA-seq) of barley varieties resistant and susceptible to leaf stripe disease identified three differentially expressed HvNPR1-like genes associated with the stress response. Among these, the Group I gene HvNPR1 was significantly up-regulated upon pathogen infection. Further analysis indicated that HvNPR1 played a pivotal role in the response to leaf stripe stress. Quantitative real-time PCR (qRT-PCR) analysis validated the expression patterns observed in RNA-seq data, with significantly higher transcript levels in the resistant cultivar compared to the susceptible cultivar. Protein–protein interaction assays demonstrated that HvNPR1 specifically interacted with HvWRKY70 in the nucleus, suggesting a potential regulatory module involved in defense signaling. These findings suggest strong regulatory interactions among these genes during the barley leaf stripe response. Conclusion: This study systematically identified and characterized HvNPR1-like family members in the barley genome through comprehensive analyses of gene structures, physicochemical properties, chromosomal localization, and evolutionary relationships. A key gene associated with leaf stripe resistance, HvNPR1, was identified, and a protein-protein interaction between HvNPR1 and the transcription factor HvWRKY70 was discovered. These findings provide a foundation for elucidating the roles of HvNPR1-likes in barley resistance to leaf stripe and offer potential targets for the genetic improvement of disease-resistant barley cultivars.
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