家蚕谷氨酸转运蛋白基因克隆及序列分析

Molecular Cloning and Sequence Analysis of Glutamate Transporter Genes in Bombyx mori

谷氨酸转运蛋白(EAAT)主要存在于神经、神经原及神经胶质细胞浆膜上,对胞外谷氨酸浓度进行调节,在维持细胞正常的生理状态、避免神经元过度兴奋方面起着重要的作用.利用鳞翅目粉纹夜蛾(Trichoplusia ni)和双翅目拟暗果蝇(Drosophila pseudoobscura)的谷氨酸转运蛋白序列为问询序列,在家蚕基因组数据库中进行同源检索,找到2个EAAT基因的同源基因(BmEAAT),并进行了克隆测序.两个BmEAAT基因均含8个外显子,编码蛋白均为膜蛋白,各有8个跨膜结构域.多物种的同源基因序列的系统发生分析结果表明:家蚕的两个EAAT拷贝至少在鳞翅目与双翅目分化前就已存在.基因芯片数据显示BmEAAT的两个拷贝在家蚕5龄3天的组织表达模式有明显差异,表明这两个基因的功能可能已经分化.

Glutamate transporters are mainly distributed in the nerve,neuron and glial cells on the serosal.They regulate extracellular glutamate concentrations and play an important role in maintaining the normal physiological state of cells and in avoiding over-excitation of neurons.Blasting the silkworm genome with the glutamate transporter protein sequences in Trichoplusia ni and Drosophila melanogaster as querying probes,we obtained two homologous sequences（BmEAAT1 and BmEAAT2） in Bombyx mori,which were then cloned and sequenced.Both BmEAAT genes were shown to contain 8 exons and the deduced proteins were glycoproteins with 8 transmembrane domains each.Phylogenetic analysis showed that the two copies of BmEAAT genes in B.mori duplicated at least before divergence between Lepidoptera and Diptera.The information from microarray indicated that the expression profiles considerably differ between BmEAAT1 and BmEAAT2 in 10 tissues on the third day of the 5th instar silkworm,suggesting that function differentiation may have occurred between the two gene copies.