Gustatory receptors interact with the serotonin pathway to regulate silkmoth stereotyped courtship behavior

Gustatory receptors (GRs) are known to mediate responses to various chemical stimuli including sugars and bitter compounds and sex pheromones. Although numerous types of GRs have been identified across insect species, the physiological roles of most GRs remain largely unknown. The silkworm, Bombyx mori, is representative of the order Lepidoptera, which includes over 70% of agriculturally important pest species. A notable feature of GRs in B. mori is that most GRs are organized in clusters, making them highly suitable for large-scale genetic studies.

Here we developed a transcription activator-like effector nuclease-mediated gene cluster replacement system, enabling functional studies beyond single-gene resolution. Using this system, we deleted the GR27-31 cluster in B. mori, which includes 12 GRs spanning approximately 82 kb of genomic sequence. Loss of GR27-31 function results in significant time retardation of courtship by impairing the ability of male silkmoths to seek female silkmoths. Electroantennogram analysis revealed that GR27-31 mutant male silkmoths did not change their responses to volatile sex pheromones of bombykol and bombykal compared with wild-type silkmoths, indicating that GR27-31 mutation did not affect male-female sex pheromone responses at the peripheral level. Subsequent molecular analysis revealed that mRNA relative expression of serotonin receptors was significantly affected in mutant silkmoths.

The current study provides the first genetic and phenotypic evidence that GRs modulate male-female courtship in B. mori. Furthermore, the results revealed for the first time that the serotonin pathway is associated with GR regulation of courtship behavior in insects.