【摘要】 通过生物信息学分析发现八肋游仆虫的UPF2(EoUPF2)包含三个串联的MIF4G(middle domain of translation initiation factor 4G)结构域和一个C末端UPF1的结合区域。对24种UPF2氨基酸序列的进化关系分析发现,EoUPF2与酿酒酵母的UPF2(ScUPF2)进化关系较为接近。InterPro database数据库分析结果显示,二者的核心结构域非常相似。酵母双杂交和Pull down实验证实:EoUPF2的C末端与EoUPF1的CH结构域相互作用;EoUPF2在没有UPF3存在的情况下,通过其MIF4G结构与外显子连接复合体(exon-exon junction complex,EJC)核心因子Y14a和Y14b相互作用。β-半乳糖苷酶实验证实EoUPF2与Y14a的作用更强一些。qPCR分析结果排除了EoUPF2与Y14a和Y14b相互作用与Y14a和Y14b本身的拷贝数有关。由此我们推断,在八肋游仆虫的NMD识别无义mRNA过程中,EoUPF2通过与Y14相互作用,介导了UPF1与无义mRNA上的外显子连接复合体的耦联,启动了无义mRNA的识别过程。更多还原

【Abstract】 Bioinformatics analysis in this paper showed that the EoUPF2 contained three tandem MIF4 G(middle domain of translation initiation factor 4 G)domains and a C-terminal UPF1 binding region.Analysis among 24 UPF2 amino acid sequences indicated that the evolutionary relationship between EoUPF2 and Saccharomyces cerevisiae UPF2 was very close.InterPro database analysis indicated that the core domain of EoUPF2 was similar to that of ScUPF2.Moreover,the yeast two hybrid and pull down experiments were conducted to show the role of EoUPF2 in the initiation of NMD in E.octocarinatus.We demonstrated the interaction between EoUPF1 and EoUPF2 was mediated by CH domain of EoUPF1 and C-terminal domain of EoUPF2.EoUPF2 interacted with Y14 aor Y14 b,core factor of exon-exon junction complex(EJC),by its MIF4 Gstructure in the absence of UPF3.β-galactosidase assay confirmed that Y14 aplayed a significant role.Q-PCR analysis excluded the interaction between EoUPF2 and Y14 aor Y14 bwas related to the copy number of themselves.We concluded that EoUPF2 mediated the coupling of UPF1 with EJC on nonsense mRNA and initiated the recognition of nonsense mRNA by interacting with Y14.更多还原