siRNAs were resuspended at 20 µM and complexed with in vivo-jetPEI. First, equal volumes of 20 µM siRNAs and 10% glucose solution were mixed to 20 µl total. Next, 1.6 ml of in vivo-jetPEI reagent was diluted to 20 µl of glucose solution (final concentration of 5%). The two solutions were combined, resulting in 5 µM siRNA and incubated for 15 min at room temperature before injections. Each ant was injected with a calibrated glass capillary directly in the head with 0.5 µl of siRNA-PEI complexes. For molecular analyses (RT-qPCR, RNA-seq), ant brains were collected 24 hr post injections. Hunting assays were conducted as described above 24 and 48 hr after injections.

Social insects are emerging models to study how gene regulation affects behavior because their colonies comprise individuals with the same genomes but greatly different behavioral repertoires. To investigate the molecular mechanisms that activate distinct behaviors in different castes, we exploit a natural behavioral plasticity in Harpegnathos saltator, where adult workers can transition to a reproductive, queen-like state called gamergate. Analysis of brain transcriptomes during the transition reveals that corazonin, a neuropeptide homologous to the vertebrate gonadotropin-releasing hormone, is downregulated as workers become gamergates. Corazonin is also preferentially expressed in workers and/or foragers from other social insect species. Injection of corazonin in transitioning Harpegnathos individuals suppresses expression of vitellogenin in the brain and stimulates worker-like hunting behaviors, while inhibiting gamergate behaviors, such as dueling and egg deposition. We propose that corazonin is a central regulator of caste identity and behavior in social insects.