Three stress-responsive peptides modulate immunity and development in the fall armyworm Spodoptera frugiperda

Insect stress-responsive peptides (SRPs) orchestrate development and immunity, yet their roles in the globally invasive pest Spodoptera frugiperda remain poorly understood. Here, we identify three SRP genes, SfSRP1, SfSRP2, and SfSRP3, that are strongly induced after Beauveria bassiana infection. Each encodes a 115-119 aa peptide with a conserved five-β-strand scaffold and divergent loop regions. RT-qPCR analysis reveals that all three are most abundant in hemocytes and head tissues, with stage-specific peaks at 2nd to 4th instars. Upon challenge with fungal or bacterial pathogens, SfSRP1 and SfSRP3 display strong, tissue- and time-dependent induction, whereas SfSRP2 only shows moderate upregulation. All three are also upregulated by heat, cold, starvation, and injury. Inhibitor assays implicate both NF-κB and JAK-STAT pathways in SfSRP regulation. Functional assays demonstrate that peptide injection into larvae suppresses feeding, stunts growth, accelerates pupation, and reduces pupal weight in a dose-dependent manner, and enhances spreading behavior of Sf9 cells. Exogenous SfSRP3 triggers broad induction of antimicrobial peptides, lectins, lysozymes, PGRPs, and prophenoloxidase, while RNAi of SfSRP3 compromises survival after B. bassiana infection. Transcriptomic profiling of SfSRP3-treated larvae uncovers 212 upregulated and 28 downregulated genes enriched in metabolic, peroxisomal, and xenobiotic pathways. Mechanistically, SfSRP3 evokes inositol 1,4,5-trisphosphate (IP₃)-mediated Ca²⁺ release in Sf9 cells, which is required for upregulating the key immune effector S. frugiperda immulectin-1 (SfIML-1). Together, these data establish SfSRP1-3 as integrators of developmental and immune signaling and highlight their potential as novel targets for pest management strategies against S. frugiperda.