美洲大蠊性信息素periplanone-A和periplanone-B诱导交配行为的分子机制

In the American cockroach, Periplaneta americana, the female's intestinal tract and feces are the main sources of sex pheromones, Periplanone-A and periplanone-B (PA/PB). PA/PB can strongly induce the mating behaviors in males; however, the detailed molecular mechanism whether and how sex pheromone receptor (PR) is involved in the regulation of males’mating behaviors is totally unknown. In a previous study, we have characterized the genomic and functional landscapes of developmental plasticity in P. americana, uncovering the mystery of the American cockroach as the “little mighty”. The applicant has recorded and analyzed the detailed aspects of males’ mating behaviors; found that RNAi knockdown of Orco significantly delayed the mating behaviors in males, suggesting that PA/PB acts through an unknown PR in the males’antenna for the induction. In this project, we will first solve the potentially positive correlation between mating behavior and antenna’s electroantennogram that are both induced by PA/PB. In order to identify the PR of PA/PB, we will carefully value the big amount of RNA-seq transcriptome data of males’antenna using bioinformatics for the screen of potential PR to PA/PB; to narrow down the potential PR gene by the combination of RNAi screen, mating behavior and antenna’s electroantennogram; to further characterize the fine distribution pattern of the potent PR using in situ hybridization or immunohistochemistry; to finally determine the actual PR by overexpressing candidate PR gene in the clawed frog oocyte in the presence of PA/PB. The molecular mechanism of PR-mediated mating behavior discovered in this study provides important cues for the development of high-efficiency, special and safe cockroach attractants.

美洲大蠊雌虫肠道和排泄物中的性信息素periplanone-A和periplanone-B(PA/PB)可诱导雄虫产生交配行为，但具体分子机制未知。实验室前期解析了美洲大蠊强大生命力的分子基础。申请人记录和解读了雄虫的详细交配行为；发现RNAi Orco显著推迟交配行为，暗示PA/PB通过触角中性信息素受体(PR)发挥作用。拟：(1)阐明PA/PB诱导交配行为的具体参数和触角电位变化的相关性。(2)分析雄虫触角转录组数据，筛选PR候选基因；RNAi PR候选基因，检测交配行为和触角电位的变化，进一步筛查PR；使用原位杂交/免疫组化检测候选PR在触角化学感受器上的精细定位；在爪蟾卵母细胞中异源表达PR候选基因，检测对PA/PB的细胞反应，最终确证PR分子。本项目中美洲大蠊性信息素诱导交配行为的分子机制研究成果可以为研发高效、专一、安全的杀蟑诱剂提供理论支持。

美洲大蠊雌虫释放的性信息素periplanone-A和periplanone-B(PA/PB)可诱导雄虫求偶和交配行为。雄虫触角的性信息素受体（pheromone receptor，PR）和共受体（Odorant receptor co-receptor，Orco）是识别PA/PB的关键。申请人分析了美洲大蠊的嗅觉受体基因集，前期研究发现，雄虫触角缺失后，对PA/PB无响应。干扰Orco显著推迟雄虫的交配行为，证明PA/PB依靠触角中特异性PR发挥作用。申请人建立了一套完善的行为测定平台，通过计算机算法优化确定了雄虫交配行为的参数，发现雄虫对雌虫性信息素粗提取物和PA/PB具有浓度依赖性，且雄虫性成熟与交配率具有显著的关联性，揭示了营养信号与发育程度对雄虫交配行为有影响。通过对雄虫不同发育时期和不同组织的转录组测序分析，发现PR1在雄虫性成熟过程中逐渐高表达，且在雄虫触角中特异性高表达，揭示PR1可能是PA/PB的潜在受体。进一步实验验证发现：（1）干扰PR1后，雄虫的求偶交配行为受到了显著抑制；（2）过表达PR1的转基因果蝇对PA/PB具有强烈的电生理响应；（3）利用原位杂交精确定位PR1于雄虫触角的锥型感器。本项目研究通过组学，分子生物学以及电生理学，结合体内和体外实验，充分证明了美洲大蠊雄虫通过特异性性信息素受体PR1响应雌虫产生性信息素PA/PB化合物，最终诱导雄虫求偶交配行为。综上所述，本研究首次揭示了嗅觉受体在美洲大蠊雄虫求偶交配行为中的关键作用，同时揭示了营养信号与发育成熟度对交配行为的影响。本研究结果为综合防控美洲大蠊等蟑螂提供新思路和新技术，也为研发高效、专一、安全的新型杀蟑诱剂奠定重要应用基础，具有重要的科学价值和现实意义。