Fine-Tuned Immune Antagonism and Nodule-Specific Cysteine-Rich Peptides Govern the Symbiotic Specificity Between Alfalfa Cultivars and Ensifer meliloti

Abstract Alfalfa expresses significantly distinct sets of genes in response to infection with different rhizobial strains at the below species level (i.e., biotype or strain). However, differences transcriptomic profiles two alfalfa cultivars nodulated by a single rhizobium strain have been largely unexamined. In this study, comparative RNA-seq analysis cultivars, Medicago sativa cv. Gannong No. 3 (G3) and 9 (G9) inoculated one Ensifer meliloti LL2, varying symbiotic performance, was conducted, followed hub gene interaction network construction based on weighted co-expression (WGCNA). The G9-LL2 system showed better nodule formation, nitrogen fixation, growth characteristics than G3-LL2 system. Compared non-inoculated control, LL2-inoculated G9 plants (10,053) produced more differentially expressed (DEGs) G3 (7112). A group 227 displayed completely distinguished expression (6.63 < log 2 (FC) 15.45) (‒ 3.05 12.05), which are primarily involved encoding nodule-specific cysteine-rich peptides (NCRs), nodulin, leghemoglobin. Although predicted roles metabolism were upregulated almost all those ubiquitin-mediated proteolysis plant–pathogen suppressed, interestingly, consistently higher measured observed plants. Hub networks that NCRs, late related plant immunity (TIR-NBS-LRR, defensin, thioredoxin, thionine, polygalacturonase) regulate other source node positions. After successful initiation nodulation both E. achieved preferable outcomes rhizobia–alfalfa symbiosis equilibrating antagonism compatibility immunity. It elevated PTI, suppressed defense ETI, enhanced fixation utilization efficiency inducing NIN, NFH1, LysM-RLK, LRP, predominantly associated highly specific positively governed NCRs fine-tuned immune antagonism, comprising TIR-NBS-LRR. These findings provide insights into genetic mechanisms underlying modification efficient semi-compatible incompatible resources.