Multi-omics reveals DEAD-box RNA helicase 20 as key protein enhancing wilt resistance in transgenic chickpea

AtNPR1 expression strengthens Fusarium wilt resistance in chickpea by activating SAR. Multi-omics analyses suggest CaDEAD-box20 as a candidate gene contributing to resistance through possible interaction with AtNPR1. Traditional breeding for broad-spectrum disease resistance in crops is often slow and resource-intensive, whereas genetic engineering provides a more precise and efficient alternative. To enhance resistance in chickpea (Cicer arietinum) against Fusarium oxysporum f. sp. ciceris, the causal agent of Fusarium wilt, we introduced the Arabidopsis NPR1 (AtNPR1) gene to activate systemic acquired resistance (SAR). We found that transgenic chickpea plants expressing AtNPR1 exhibited markedly reduced reactive oxygen species (ROS) accumulation, higher expression of defense-related genes, and up to 41% greater resistance compared with wild-type (WT) plants. qRT-PCR analysis revealed a higher fungal DNA load and increased expression of virulence genes in infected WT plants relative to transgenic lines. We also observed elevated salicylic acid (SA) levels and strong induction of pathogenesis-related (PR) genes in transgenics at 2 days post-infection (dpi). Although jasmonic acid (JA) content did not differ significantly between genotypes, methyl jasmonate (MeJA) treatment confirmed activation of JA pathway genes in both. To elucidate the molecular basis of resistance, label-free quantitative proteomics (LC-MS/MS) and metabolomics (GC-MS) analyses were performed, revealing 205 differentially expressed proteins and 38 metabolites associated with defense responses. Protein-protein interaction assays (BiFC and modeling) suggested an interaction between AtNPR1 and chickpea DEAD-box RNA helicase 20 (CaDEAD-box20). Functional validation showed that CaDEAD-box20 positively regulates resistance, as its overexpression enhanced, whereas its knockout reduced, tolerance to Fusarium wilt. Overall, we demonstrate that AtNPR1 enhances Fusarium wilt resistance in chickpea by coordinating SA- and JA-mediated defense pathways, with CaDEAD-box20 serving as a key regulatory component.