刺参呼吸树抗氧化防御酶类基因在夏眠期的表达特征

Transcriptional regulation of antioxidant enzymes in respiratory tree of sea cucumber Apostichopus japonicus(Selenka)during aestivation

以夏眠刺参呼吸树为研究材料,研究了刺参抗氧化防御酶类的基因表达特征,共获得夏眠刺参的5个基因片段序列:铜锌超氧化物歧化酶(Cu/Zn-SOD)、过氧化氢酶(CAT)、磷脂氢谷胱甘肽过氧化物酶(PHGPx)、过氧化物还原酶-5(PRDX5)及过氧化物还原酶-6(PRDX6),完成基因序列初步分析;利用荧光定量PCR技术,以NDUFA13(NADH dehydrogenase[ubiquinone]1 alpha subcomplex subunit 13)和ACTB(beta actin)基因为多内参,定量分析实验各阶段刺参抗氧化防御酶类基因表达量。结果表明,刺参抗氧化酶类基因与近、远源物种同源性均较高;现场调查实验结果显示,相对5月非夏眠刺参样品,8月夏眠刺参呼吸树组织Cu/Zn-SOD及PRDX5基因表达上调,CAT及PHGPx基因表达量均出现显著下降。温度诱导夏眠实验证实Cu/Zn-SOD基因在实验第0天表达量显著上升至2.01倍,并一直保持较高水平;CAT基因表达在实验第0天下调,第5天时出现上调,而在第10天和40天表达量均出现显著下降;PHGPX基因表达实验期间均出现显著下调,在第40天时降到最低,为0.34倍;PRDX5基因表达在实验第0天出现显著上调至1.55倍,第5天回落后呈现上升,于第40天达到高值1.91倍;PRDX6基因表达在实验第10、20天出现显著下调。

Aestivation is a kind of dormancy that is probably defined as a survival strategy for dealing with arid conditions,food shortage and high environmental temperature.As an adaptation for life in stressful environments,multiple adjustments occurred in aestivated organism including the regulation of defense system.Antioxidant enzymes are regulated in aestivating animals usually.To evaluate the transcriptional regulations associated with antioxidant defense in sea cucumber during aestivation,gene expression of antioxidant enzymes in respiratory tree was analyzed in this study.Five partial sequences of antioxidant enzyme genes were obtained from Apostichopus japonicus transcriptome database：Cu/Zn-SOD（copper-zinc superoxide dismutase）,CAT（catalase）,PHGPx（Phospholipid hydroperoxide glutathione peroxidas）,PRDX5（Peroxiredoxin-5）and PRDX6（Peroxiredoxin-6）.The multiple sequence alignment（MSA）was conducted and special primers were designed for Real-time PCR assay.These genes expressions were normalized by multiple internal control genes of NDUFA13（NADH dehydrogenase[ubiquinone]1 alpha subcomplex subunit 13）and ACTB（beta actin）,and their variations during aestivation of A.japonicus were analyzed.Compared with the active sea cucumbers（sampled in May）,the antioxidant enzymes got transcriptional change in respiratory tree of aestivated sea cucumbers（sampled in Aug）：the expression of Cu/Zn-SOD genes got increase to 1.66 fold（P0.05）;the expression of PRDX5 genes got significant increase to 1.69 fold（P0.05）;the expression of CAT and PHGPx got significant decrease to 0.60 and 0.38 fold respectively（P0.05）;no notable expression variation of PRDX6 was found.It was mainly consistent with the variation of these genes expressions in later phase of thermal induced aestivation of A.japonicus（aestivated sea cucumbers sampled on 40 d）：the expressions of Cu/Zn-SOD and PRDX5 genes got significant increase to 1.85 and 1.91 fold respectively（P0.05）;the expressions of CAT and PHGPx genes got significant decrease to 0.60 and 0.34 fold respectively（P0.05）;no notable expression variation of PRDX6 was found.The result indicated that the gene expressions of different antioxidant enzymes were regulated differently during aestivation of sea cucumber.