Drosophila odorant receptors are novel seven transmembrane domain proteins that can signal independently of heterotrimeric G proteins
Drosophila odorant receptors are novel seven transmembrane domain proteins that can signal independently of heterotrimeric G proteins
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DOI：
10.1016/j.ibmb.2008.05.002
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发表时间：
2008-08-01
影响因子：
3.8
通讯作者：
Warr, Coral G.
中科院分区：
文献类型：
--
作者：
Smart, Renee;Kiely, Aidan;Warr, Coral G.
果蝇的嗅觉是由一大家族膜结合气味受体蛋白(ORS)介导的。在异种细胞中，我们研究了配体结合的果蝇ORs的结构特征和信号机制是否与它们是G蛋白偶联受体(GPCRs)一致。通过在末端和预测的环区插入表位标签来确定Or22a的详细膜拓扑结构。在果蝇S2细胞中的免疫细胞化学实验表明，Or22a具有7个跨膜区，但其膜拓扑结构与GPCRs相反，具有胞内N端和胞外C端。为了研究Or43b的信号转导机制，我们在Sf9和HEK293细胞中表达了Or43b，发现异源三聚体G蛋白(GDp-β-S)、腺苷环化酶(SQ22536)、鸟苷酸环化酶(ODQ)、环核苷酸磷酸二酯酶(IBMX)和磷脂酶C(U73122)的抑制剂对Or43b的反应几乎没有影响。全细胞贴片显示，加入配体后，细胞质膜阳离子通道开放。该反应可被镧和2-APB阻断，但不能被Ru红或SKF96365阻断。基于这些数据，我们得出结论，果蝇Ors包括一个新的家族，由七个跨膜受体组成，在HEK293细胞中，这些受体通过开放阳离子通道来传递信号，这种机制在很大程度上不依赖于G蛋白。(C)2008爱思唯尔有限公司。保留所有权利。
Olfaction in Drosophila is mediated by a large family of membrane-bound odorant receptor proteins (Ors). In heterologous cells, we investigated whether the structural features and signalling mechanisms of ligand-binding Drosophila Ors are consistent with them being G protein-coupled receptors (GPCRs). The detailed membrane topology of Or22a was determined by inserting epitope tags into the termini and predicted loop regions. Immunocytochemistry experiments in Drosophila S2 cells imply that Or22a has seven transmembrane domains but that its membrane topology is opposite to that of GPCRs, with a cytoplasmic N-terminus and extracellular C-terminus. To investigate Or signalling mechanisms, we expressed Or43b in Sf9 and HEK293 cells, and show that inhibitors of heterotrimeric G proteins (GDP-beta-S), adenylate cyclase (SQ22536), guanylyl cyclase (ODQ), cyclic nucleotide phosphodiesterases (IBMX) and phospholipase C (U73122) have negligible impact on Or43b responses. Whole cell patching Or43b/Or83b-transfected HEK293 cells revealed the opening of plasma membrane cation channels on addition of ligand. The response was blocked by lanthanum and by 2-APB, but not by Ruthenium red or SKF96365. Based on these data, we conclude that Drosophila Ors comprise a novel family of seven transmembrane receptors that in HEK293 cells signal by opening cation channels, through a mechanism that is largely independent of G proteins. (c) 2008 Elsevier Ltd. All rights reserved.