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雄性棉铃虫利用气味结合蛋白(OBP)感受雌性性信息素(Z11-16Ald).md

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雄性棉铃虫利用气味结合蛋白(OBP)感受雌性性信息素(Z11-16:Ald)

大家好，本来想着最近能回京的，没想到北京的疫情突然开始出现反复，希望在京的小伙伴保护好自己啊。

今天分享的这篇文章是我们组郭浩师兄，郭平平师兄和孙亚兰师姐共同完成的，目前文章在bioRxiv上peer review。

Abstract

​ 棉铃虫利用(Z)-11-hexadecenal (Z11-16:Ald)作为主要的性信息素组分，在雄性棉铃虫触角中高表达pheromone binding proteins (PBPs) and two general odorant binding proteins (GOBPs)，而这些气味结合蛋白在感受Z11-16:Ald的具体作用却不清楚。

​ 首先我们先合成HarmOR13(感受Z11-16:Ald的一种性信息素受体)抗体来检测PBPs / GOBPs与HarmOR13的相互作用。免疫染色实验表明HarmPBPs和HarmGOBPs各自定位于雄性触角的毛形和锥形感器中的支持细胞中。具体来说，HarmPBP1和HarmPBP2共定位于表达HarmOR13的olfactory receptor neurons (ORNs)中。接下来我们利用果蝇空神经元系统来验证PBP2与HarmOR13的相互作用，我们发现HarmPBP1和HarmPBP2可以显著增加HarmOR13对Z11-16:Ald的敏感性，然而HarmPBP1和HarmPBP2不能拓宽HarmOR13的反应谱。综上，PBP可以增加外周嗅觉敏感性但对嗅觉选择性没有作用。最后，我们发现棉铃虫HarmOR13与果蝇OR67d利用相似的嗅觉编码机制来感受性信息素，这可能表明性信息素的感受机制在不同昆虫目中保守型较高。

Introduction

一般来说，蛾子的毛形感器感受性信息素，而锥形和栓锥形感器感受普通气味化合物。昆虫外周气味感受主要通过odorant binding proteins (OBPs), olfactory receptors (ORs) and odorant degrading enzymes (ODEs)来感受。

Insect ORs are predicted as heterotetramers consisting of two subunits of narrowly tuned ORs and two subunits of highly conserved odorant receptor co receptor (ORco).

OBP是一类可溶性，高浓度的小分子蛋白，主要存在于嗅觉受体神经元感器的淋巴液中。OBPs可以通过结合气味化合物并把它们转运到树突膜ORNs上的ORs，进行识别。

Pheromone binding proteins (PBPs)是一种特异表达在毛形感器中感受性信息素的一类OBPs，PBPs具有物种特异性的结合亲和性，但不同系统下，PBP-PR的相互作用似乎有所差异。结合实验表明GOBPs也会优先结合性信息素。

棉铃虫触角上的毛形感器可以分为type A, B and C (ratio: 84:5:11)，type A responds specifically to Z11-16:Ald, type B to (Z)-9-tetradecenal 104 (Z9-14:Ald), and type C to (Z)-9-hexadecenal (Z9-16:Ald), Z9-14:Ald, (Z)-11-105 hexadecenyl acetate (Z11-16:OAc) and (Z)-11-hexadecenol (Z11-16:OH)。

在这里，我们采用果蝇的两个互不干扰的GAL4-UAS and LexA-LexAOP系统，表达HarmPBPs在果蝇T1感器，HarmOR13表达在果蝇OR67d神经元中，再结合免疫染色实验来鉴定HarmPBPs和HarmGOBP的分布以及与HarmOR13的相互作用。

Results

Spatial distribution of HarmPBPs and HarmGOBPs on male antennae of H.armigera

OBPs are synthesized in the tormogen cells and thecogen cells and then 643 secreted into the sensillum lymph bathing ORNs.

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  • HarmPBPs and HarmGOBPs were mutually exclusive and co-localization was never found in the longitudinal sections.

  • HarmPBPs were only detected in the auxiliary cells underneath the sensilla trichodea, while the expression of HarmPBPs were stricktly restricted to the cells associated with the sensilla basiconica.

这种分布表明HarmPBPs特定参与性信息素感受，而HarmGOBPs感受普通气味化合物。

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  • The immunostaining results showed that HarmPBP1and HarmPBP2 were always co-localized.
  • these results identify two categories of PBP positive cells: category A expressing HarmPBP1 & HarmPBP2 and category B harboring all three PBPs.
  • HarmPBP3所占比例为18.7%，这反应了雄性触角上B和C型感器的分布情况。

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  • HarmPBP1 and HarmPBP2 expressing cells were closely located to HarmOR13 positive ORNs, whereas the cells additionally expressing HarmPBP3 were not. HarmPBP1 and HarmPBP2也表达在非A型感器中。
  • Neither GOBP1 nor GOBP2 expressing cells showed spatial relationships with HarmOR13 ORNs.这表明GOBP1和GOBP2主要表达在栓锥形感器中感受普通气味化合物。

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下图表示PBP1和PBP2和OR13表达在同一个感器中，而PBP3表达在其它感器中。

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RNA in situ hybridization results confirmed the immunofluorescence results

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Heterologous expression of HarmPBP1, HarmPBP2 and HarmOR13 to Drosophila T1 sensilla

  • Using LUSH-GAL4, we successfully expressed HarmPBP1 and HarmPBP2 in the lymph of Drosophila T1 sensilla through supporting cells secretory pathway.

  • In parallel, HarmOR13 was expressed in the ORNs by the driver line ORco-LexA and the receptor was fully functional as revealed by its response to Z11-16:Ald sensitivity in OR67d ORN.

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Functional interactions between HarmPBP1 / HarmPBP2 and HarmOR13

  • HarmPBP1 and HarmPBP2 have comparable binding affinities to Z11-16:Ald and are functioning in a similar manner.

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The molecular principle of ligand-receptor coupling

HarmOR13可以对Z11-14:Ald有反应，这表明HarmOR13对更长碳链的性信息素可以产生反应，但对18个碳的Z11-18:Ald没反应。而且对16:Ald, Z9-16:Ald and Z7-16:Ald没反应，表明双键的位置也很重要，此外对几何异构体E11-16:Ald也没有反应。对这些Z11-16:OAc, Z11-16:OH and Z11-16:COOH这些不同功能团的性信息素也没有反应。

我们发现果蝇OR67d对Z11-18:OAc (cVA), Z11-16:OAc and Z11-14:OAc有反应，但对16:OAc，异构体(E11-16:OAc), Z9-16:OA and Z7-16:OAc, 以及aldehyde(Z11-16:Ald)，alcohol (Z11-16:OH) and acid(Z11-16:COOH)没有反应。

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Discussion

  • We found that the presence of HarmPBP1 and HarmPBP2 in Drosophila T1 sensilla enhanced the sensitivity of HarmOR13 to Z11-16:Ald.
  • We observed that HarmPBP1 and HarmPBP2 are colocalized and expressed in the type A sensilla trichodea which house HarmOR13 expressing ORNs.
  • We also noticed that a small fraction of HarmPBP1 and HarmPBP2 positive cells are not physically associated with HarmOR13, hinting at their presences in type B and C sensilla trichodea and their roles in detection of other pheromone components, such as Z9-16:Ald, Z9-14:Ald, Z11-254 16:OAc.
  • Our results favor a model in which PBPs do not have a significant effect on the selectivity of ORNs and PRs determine the specificity of pheromones detection.
  • Our work shows the importance of different structural parameters, such as double bonds, functional groups and isomerism for the ligand-receptor affinity.

值得注意的是，最近的实验表明，敲除掉PBP对蛾子感受性信息素的作用不是非常明显，这表明PBPs可能会有其它的作用。

在不同物种PBP研究中出现的矛盾现象可能反映了，这种物种特异性的PBPs在不同物种可能有不同的行使功能的方式。=

参考文献

https://doi.org/10.1101/2020.06.10.145235