家蚕Ras3基因全长cDNA克隆及在感染BmNPV家蚕组织中的表达谱分析

Full-Length cDNA Cloning of BmRas3 Gene and Expression Profiling in Different Tissues of Bombyx mori Infected with BmNPV

Ras(rat sarcoma viral oncogene homolog)蛋白是GTPase超家族成员之一,该蛋白质可以将细胞外信号传导到不同的细胞,从而调控细胞的吞噬作用、凋亡、抗微生物侵染的酶防御系统等生理过程。运用RACE技术,从家蚕中肠中克隆出一个新的BmRas3基因,该基因cDNA全长987 bp,包含5′UTR(153 bp)、3′UTR(279 bp)和一个编码184个氨基酸残基的ORF(555 bp)。BmRas3具有小G蛋白的典型特征,其氨基酸序列中含有3个GTP/GDP结合结构域,1个效应子结合结构域以及1个烯丙基化的CAAX基序。多重比对发现BmRas3与海波斯莫科马属蛾(Hyposmocoma kahamanoa)Rap1、斜纹夜蛾(Spodoptera litura)Rap-1b、烟草天蛾(Manduca sexta)Rap1的序列相似性较高,系统进化分析也显示BmRas3与这些蛋白质的亲缘关系最近。BmRas3在食桑期的表达量高于眠期;在感染BmNPV的家蚕中肠和脂肪体中,BmRas3的表达量有不同程度提高,且与正常对照相比差异显著,暗示BmRas3可能作为免疫活化因子,在家蚕抵抗BmNPV的入侵过程中发挥着重要作用。研究结果为下一步探讨BmRas3对BmNPV感染的应答提供了理论基础。

Ras(rat sarcoma viral oncogene homolog)is a member of the GTPase superfamily,which can transmit extracellular signals to different cells,and then regulate cell phagocytosis,cell apoptosis,anti-microbial enzyme defense system and other life processes.In this study,we cloned BmRas3 gene from the midgut of Bombyx mori using RACE technology.The full-length cDNA of this gene is 987 bp including 5′UTR(153 bp),3′UTR(279 bp)and one open reading frame(555 bp)encoding 184 amino acid residues.BmRas3 has the typical characteristics of small G protein which contains three GTP/GDP binding domains,one effector binding domain and one allylated CAAX motif.Multiple sequence alignment showed that BmRas3 has high sequence identity with Hypomorpha kahamanoa Rap1,Spodoptera litura Rap-1 b and Manduca sexta Rap1.Phylogenetic analysis also showed the BmRas3 has the highest homology with these proteins.The transcription level of BmRas3 in the mulberry eating period was higher than that in the molting period.The transcription levels of BmRas3 increased in the midgut and fat body upon BmNPV challenge,suggesting that BmRas3 may serve as an immune effector and play a key role in the process of defending against BmNPV infection.This study provides a theoretical basis for further study on the function of BmRas3 in response to Bm-NPV infection.