Bao Haibo, Meng Xiangkun, Liu Zewen
Key Laboratory of Integrated Management of Crop Diseases and Pests (Ministry of Education), College of Plant Protection, Nanjing Agricultural University, 1 Weigang, Nanjing 210095, China.
College of Horticulture and Plant Protection, Yangzhou University, Yangzhou 225009, China.
Insect Biochem Mol Biol. 2017 Nov;90:82-89. doi: 10.1016/j.ibmb.2017.09.014. Epub 2017 Oct 6.
Acetylcholine binding proteins (AChBPs) are homologs of extracellular domains of nicotinic acetylcholine receptors (nAChRs) and serve as models for studies on nAChRs. Particularly, studies on invertebrate nAChRs that are limited due to difficulties in their heterologous expression have benefitted from the discovery of AChBPs. Thus far, AChBPs have been characterized only in aquatic mollusks, which have shown low sensitivity to neonicotinoids, the insecticides targeting insect nAChRs. However, AChBPs were also found in spiders based on the sequence and tissue expression analysis. Here, we report five AChBP subunits in Pardosa pseudoannulata, a predator enemy against rice insect pests. Spider AChBP subunits shared higher sequence similarities with nAChR subunits of both insects and mammals compared with mollusk AChBP subunits. The AChBP1 subunit of P. pseudoannulata (Pp-AChBP) was then expressed in Sf9 cells. The Ls-AChBP from Lymnaea stagnalis was also expressed for comparison. In both AChBPs, one ligand site per subunit was present at each interface between two adjacent subunits. Neonicotinoids had higher affinities (7.9-18.4 times based on K or K values) for Pp-AChBP than for Ls-AChBP, although epibatidine and α-bungarotoxin showed higher affinities for Ls-AChBP. These results indicate that spider AChBP could be used as an alternative model to study the interaction between insect nAChRs and neonicotinoids.
乙酰胆碱结合蛋白（AChBPs）是烟碱型乙酰胆碱受体（nAChRs）胞外结构域的同源物，可作为研究nAChRs的模型。特别是，由于无脊椎动物nAChRs在异源表达方面存在困难，对其研究受到限制，而AChBPs的发现使这一情况得到改善。到目前为止，AChBPs仅在水生软体动物中得到表征，这些软体动物对新烟碱类杀虫剂（靶向昆虫nAChRs的杀虫剂）表现出低敏感性。然而，基于序列和组织表达分析，在蜘蛛中也发现了AChBPs。在此，我们报道了拟环纹豹蛛（一种水稻害虫的捕食性天敌）中的5个AChBP亚基。与软体动物AChBP亚基相比，蜘蛛AChBP亚基与昆虫和哺乳动物的nAChR亚基具有更高的序列相似性。然后，将拟环纹豹蛛的AChBP1亚基（Pp-AChBP）在Sf9细胞中表达。同时也表达了来自静水椎实螺的Ls-AChBP用于比较。在这两种AChBPs中，每个亚基在两个相邻亚基之间的每个界面处都有一个配体结合位点。新烟碱类杀虫剂对Pp-AChBP的亲和力比对Ls-AChBP的亲和力更高（基于K或K值为7.9 - 18.4倍），尽管埃博霉素和α-银环蛇毒素对Ls-AChBP的亲和力更高。这些结果表明，蜘蛛AChBP可作为研究昆虫nAChRs与新烟碱类杀虫剂之间相互作用的替代模型。
