GmIQD63 functions as a novel GmCDPK38-interacting protein in soybean defense against the common cutworm.

Main conclusion: GmIQD63 interacts with GmCDPK38 to mediate soybean defense against the common cutworm, and its elite haplotype enhances resistance without a significant yield penalty, providing a potential breeding resource. Soybean (Glycine max) production is severely threatened by the common cutworm (Spodoptera litura Fabricius), necessitating the identification of resistance genes for molecular breeding. Calcium-dependent protein kinases (CDPKs) are critical regulators of plant defense responses; however, the substrate network of GmCDPK38, a negative regulator of soybean resistance to this pest, remains elusive. In this study, we combined phosphoproteomic analysis of the gmcdpk38 mutants with bimolecular fluorescence complementation assays to identify 12 potential GmCDPK38-interacting partners. Among these, GmIQD63, an IQ67 domain protein homologous to Arabidopsis AtIQD1 (a known insect resistance regulator), emerged as a key candidate. The physical interaction between GmCDPK38 and GmIQD63 was further validated through yeast two-hybrid and luciferase complementation imaging assays. Sequence analysis revealed that GmIQD63 encodes a 477-amino acid polypeptide containing multiple stress- and hormone-responsive cis-acting regulatory elements in its promoter region, along with a conserved IQ67 domain characteristic of plant IQD IIIb subfamily members. Subcellular localization experiments confirmed its plasma membrane targeting. Functional characterization via RNA interference-mediated silencing of GmIQD63 in soybean hairy roots significantly enhanced susceptibility to the common cutworm. Population genetic analysis of 341 soybean accessions uncovered natural variation in GmIQD63, with haplotype H5 in wild soybeans conferring enhanced resistance to the common cutworm without compromising agronomic traits. Notably, H5 is rare in cultivated soybeans, suggesting its untapped potential for breeding insect-resistant varieties. This study identifies GmIQD63 as a novel GmCDPK38-interacting partner essential for soybean defense and provides a molecular foundation for developing insect-resistant cultivars.