A timeless gene-regulated pheromone binding protein governs female mating behavior in Bactrocera dorsalis

The rhythmic mating behavior of insects has been extensively documented, yet the regulation of this behavior through sex pheromone sensing olfactory genes controlled by the clock genes in the rhythm pathway remains unclear. In this study, we investigated the impact of circadian rhythm on female recognition of male rectal Bacillus -produced sex pheromone in B. dorsolis . Behavioral and electrophysiological assays revealed a peak in both mating behavior and response to sex pheromones in the evening in females. Comparative transcriptome analysis of female heads demonstrated rhythmic expression of the Timeless gene-Tim and odorant binding protein gene-Pbp5, with highest expression levels occurring in the evening. Protein structural modeling, tissue expression patterns, RNAi treatment, and physiological/behavioral studies supported Pbp5 as a sex pheromone binding protein whose expression is regulated by Tim. Furthermore, manipulation of female circadian rhythm resulted in increased morning mating activity, accompanied by consistent peak expression of Tim and Pbp5 during this time period. These findings provide evidence that insect mating behavior can be modulated by clock genes through their effects on sex pheromone sensing processes. Our results also contribute to a better understanding of the molecular mechanisms underlying rhythmic insect mating behavior.