日本囊对虾核因子κB抑制蛋白激酶ε2基因克隆及表达

Characterization and Expression of Inhibitor-κB Kinase Epsilon 2 in Marsupenaeus japonicus

【目的】了解核因子κB抑制蛋白激酶ε2(IKKε2)基因在日本囊对虾(Marsupenaeus japonicus)受细菌刺激后的应答情况,以阐释IKKε2在对虾先天免疫中所起的作用。【方法】从日本囊对虾转录组数据中获得日本囊对虾IKKε2(命名为PjIKKε2)的cDNA序列,并进行生物信息学分析,采用荧光定量PCR分析PjIKKε2的组织表达以及受哈维弧菌(Vibrio harveyi)刺激后在肝胰腺组织的表达情况。【结果】PjIKKε2全长为4009 bp,其中开放阅读框长2337 bp,编码778个氨基酸,5′UTR(Untranslated region)174 bp,3′UTR 1498 bp。PjIKKε蛋白含有一个激酶结构域、一个类泛素结构域和一个TANK结合激酶卷曲螺旋结构域。多重序列比对结果显示,PjIKKε2蛋白序列与其他甲壳类IKKε的相似性高,在激酶结构域保守性高。系统进化树结果显示,PjIKKε2与其他对虾属的IKKε聚集在一支。PjIKKε2在日本囊对虾9个组织中均有表达,在淋巴组织中表达量最高(P<0.05)。在哈维弧菌刺激6 h后,PjIKKε2在肝胰腺中表达量显著上调,达到最大值(P<0.05)。【结论】PjIKKε2参与了日本囊对虾的先天免疫。

【Objective】In order to investigate inhibitorκB kinase epsilon 2(IKKε2)response of bacterial infection and elucidate the function of IKKε2 in Marsupenaeus japonicus innate immunity.【Method】The full-length cDNA of M.japonicus IKKε2(named PjIKKε2)was identified and characterized with bioinformatics strategy.The expression of PjIKKε2 was detected in various tissues under normal condition and in hepatopancreas after the treatment with Vibrio harveyi.【Result】PjIKKε2 cDNA was 4009 bp length,including a 2337 bp open reading frame(ORF)encoding 778 amino acid,a 174 bp 5′untranslated region(5′-UTR)and a 1498 bp 3′-UTR.PjIKKε2 protien contain an N-terminal catalytic kinase domain,an ubiquitin-like domain and a TANK-binding kinase 1 coiled-coil domain.Multiple sequence alignment showed that PjIKKε2 share high identity with that of another IKKε2 of crustacean.The kinase domain was conserved in most species.The phylogenetic analysis suggested that PjIKKε2 was clustered with the closely related IKKs from Litopenaeus vannamei and Penaeus monodon.PjIKKε2 transcript was detected in all examined tissues with high mRNA expression in lymph.Following the infection of V.harveyi,the PjIKKε2 was up-regulated significantly in hepatopancreas(P<0.05)and maximize at 6 h.【Conclusion】PjIKKε2 plays an important role in innate immunity of M.japonicus.