Doublesex-mediated regulation of insulin signaling drives sex-specific body growth

Most animals develop sex-biased body sizes driven by sexually divergent plasticity in nutrient-dependent growth. Prior work in Drosophila, the sex-determination gene transformer (tra) has been shown to regulate sex differences in body size. However, tra does not widely mediate sex determination across insects, the unifying molecular pathway governing female-biased sexual size dimorphism (SSD) remains poorly defined in non-drosophilid insects. The rice stem borer Chilo suppressalis, a devastating lepidopteran crop pest, exhibits robust female-enlarged SSD, offering an ideal system to dissect underlying mechanisms. We report that female-specific splice isoforms of the sex-determining gene Csdsx are the master regulator of female-biased growth. Disruption of the female-specific Csdsx exon 3 via CRISPR knockout or RNA interference drastically reduces female body size and completely erases SSD. Csilp2, encoding a key insulin-like peptide, is selectively upregulated in late-instar female larvae, and female CsDsx proteins directly bind and activate the Csilp2 promoter to boost transcription. Loss-of-function of Csilp2 also eliminates SSD by suppressing female somatic overgrowth. Our results define a conserved regulatory cascade: female-specific DSX directly stimulates insulin signaling via Csilp2, bridging core sex-determination circuitry and nutrient-responsive growth control. Beyond fundamental advances in developmental and evolutionary biology, Csdsx and Csilp2 provide tractable molecular targets for targeted lepidopteran pest management. SignificanceSexual size dimorphism (SSD), widespread across insects with larger females, strongly shapes reproductive fitness. Yet molecular reports on the connection between sex determination and dimorphic growth have long been scarce. Using the destructive rice stem borer with prominent female-biased SSD, we show female-specific Chilo suppressalis Doublesex (CsDsx) isoforms directly activate insulin effector Chilo suppressalis Insulin-like peptide 2 (Csilp2) in late larvae. Knocking out either gene fully abolishes female size advantage and eliminates SSD. Our findings resolve a decades-long developmental biology puzzle by establishing dsx as the missing bridge connecting sex-determination and nutrient-dependent insulin signaling. These two genes also serve as actionable molecular targets for sustainable lepidopteran pest control.