七鳃鳗含硒谷胱甘肽过氧化物酶2的分子克隆、生物学特性及表达分析

摘要

谷胱甘肽过氧化酶2(GPx2)，是生物体内重要的一类抗氧化酶，能够通过酶解过氧化氢阻断活性氧(ROS)的有害作用。本研究从七鳃鳗的血液 cDNA 文库中克隆了 GPx2 cDNA 全长和基因组 DNA 序列。序列分析表明，其 cDNA全长为868 bp(GenBank 序列号 KM211710)，包括53bp 5′非编码区、251bp 3′非编码区和564 bp 开放阅读框，编码由187个氨基酸组成的多肽。基因组 DNA 序列(GenBank 序列号 KM211711)揭示了基因组特征含有2个外显子和1个内含子结构。系统发育分析表明，七鳃鳗 GPx2(LjGPx2)与其它的 GPx2具有很高的同源性。利用实时荧光定量 PCR检测到 GPx2基因在七鳃鳗各组织中广泛表达，其中在口腔腺、心脏、卵、生殖腺中表达量较高。此外，用脂多糖(LPS)体内刺激七鳃鳗后发现，LjGPx2 mRNA 在生殖腺中表达量显著升高。本研究为探讨 GPx2在七鳃鳗的免疫应答中的作用提供了理论依据。%Glutathione peroxidase 2 (GPx2) is an important antioxidant enzyme in vivo that enzymatically reduces hydrogen peroxide to limit harmful effects of reactive oxygen species (ROS) in living organisms. In this study, the full-length GPx2 gene and the genomic DNA sequences were cloned from the blood cDNA library of Lampetra japonica (L. japonica). The sequence analysis showed that the full-length cDNA of 868 bp (GenBank accession no. KM211710) includes an open reading frame (ORF) of 564 bp plus 53 bp of the 5'-untranslated region (5'UTR) and 251 bp of the 3'-untranslated region (3'UTR). Genomic DNA sequences (GenBank accession no. KM211711) included a two exon-one intron structure. Phylogenetic analysis revealed that GPx2 (LjGPx2) is closely related to other GPx2. The real-time PCR results demonstrated that the GPx2 gene was widely expressed in adult lamprey in which higher levels in buccal glands, heart, ovum, and gonad. Furthermore, upon in vivo stimulation of lampreys with lipopolysaccharide (LPS), LjGPx2 mRNA was upregulated in gonad. The results obtained in the present study will help us to understand the roles of GPx2 in immune responses in jawless vertebrates.