Laboratory of Insect Toxicology, South China Agricultural University, Guangzhou 510642, China.
J Insect Sci. 2013;13:49. doi: 10.1673/031.013.4901.
Intensive research on the molecule structures of the gamma-nminobutyric acid (GABA) receptor in agricultural pests has great significance to the mechanism investigation, resistance prevention, and molecular design of novel pesticides. The GABA receptor a2 (SlGABARα2) subunit gene in Spodoptera litura (Fabricius) (Lepidoptera: Noctuidae) was cloned using the technologies of reverse transcription PCR and rapid amplification of cDNA ends. The gemonic DNA sequence of SlGABARα2 has 5164 bp with 8 exons and 7 introns that were in accordance with the GT-AG splicing formula. The complete mRNA sequence of SlGABARα2 was 1965 bp, with an open reading frame of 1500 bp encoding a protein of 499 amino acids. The GABA receptor is highly conserved among insects. The conserved regions include several N-glycosylation, Oglycosylation, and phosphorylation sites, as well as 4 transmembrane domains. The identities that SlGABARα2 shared with the GABA receptor a2 subunit of Spodoptera exigua, Heliothis virescens, Chilo suppressalis, Plutella xylostella, Bombyx mori ranged from 99.2% to 87.2% at the amino acid level. The comparative 3-dimensional model of SlGABARα2 showed that its tertiary structure was composed of 4 major α-helixes located at the 4 putative transmembrane domains on one side, with some β-sheets and 1 small α-helix on the other side. SlGABARα2 may be attached to the membrane by 4 α-helixes that bind ions in other conserved domains to transport them through the membrane. The results of quantitative real time PCR demonstrated that SlGABARα2 was expressed in all developmental stages of S. litura. The relative expression level of SlGABARα2 was the lowest in eggs and increased with larval growth, while it declined slightly in pupae and reached the peak in adults. The expressions of SlGABARα2 in larvae varied among different tissues; it was extremely high in the brain but was low in the midgut, epicuticle, Malpighian tube, and fat body.
对γ-氨基丁酸（GABA）受体的分子结构进行深入研究，对于农业害虫的机制研究、抗性防治和新型农药的分子设计具有重要意义。本研究采用反转录 PCR 和快速扩增 cDNA 末端技术克隆了斜纹夜蛾（Spodoptera litura）（鳞翅目：夜蛾科）GABA 受体 a2（SlGABARα2）亚基基因。该基因的基因组 DNA 序列长 5164 bp，包含 8 个外显子和 7 个内含子，符合 GT-AG 剪接规则。SlGABARα2 的完整 mRNA 序列长 1965 bp，开放阅读框长 1500 bp，编码 499 个氨基酸。昆虫的 GABA 受体具有高度保守性，SlGABARα2 与甜菜夜蛾（Spodoptera exigua）、烟青虫（Heliothis virescens）、二化螟（Chilo suppressalis）、小菜蛾（Plutella xylostella）和家蚕（Bombyx mori）的 GABA 受体 a2 亚基的氨基酸序列同源性为 99.2%~87.2%。SlGABARα2 的三维比较模型显示，其三级结构由 4 个主要的α-螺旋组成，位于一侧的 4 个假定跨膜结构域，另一侧有一些β-折叠和 1 个小的α-螺旋。SlGABARα2 可能通过与其他保守结构域中的离子结合的 4 个α-螺旋附着在膜上，以将其运输通过膜。实时荧光定量 PCR 结果表明，SlGABARα2 在斜纹夜蛾的所有发育阶段均有表达。SlGABARα2 在卵中的相对表达水平最低，随着幼虫的生长而增加，在蛹中略有下降，在成虫中达到峰值。SlGABARα2 在幼虫不同组织中的表达存在差异，在脑中的表达水平极高，而在中肠、外骨骼、马氏管和脂肪体中的表达水平较低。
