Molecular cloning, characterization and expression analysis of heat shock protein 90 from Pacific abalone, Haliotis discus hannai Ino in response to dietary selenium.
Molecular cloning, characterization and expression analysis of heat shock protein 90 from Pacific abalone, Haliotis discus hannai Ino in response to dietary selenium.
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DOI：
10.1016/j.fsi.2010.10.019
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发表时间：
2011
影响因子：
4.7
通讯作者：
Wenbing Zhang;Chenglong Wu;K. Mai;Qiyong Chen;Wei Xu
中科院分区：
文献类型：
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作者：
Wenbing Zhang;Chenglong Wu;K. Mai;Qiyong Chen;Wei Xu
本研究采用同源克隆和cDNA末端快速扩增（RACE）相结合的方法，克隆了太平洋鲍鱼热休克蛋白90 （HdhHSP90）的cDNA。HdhHSP90全长2660 bp，其中开放阅读框（ORF）长度为2187 bp，编码728个氨基酸的多肽，预测分子量为84.134 kDa，理论等电点为4.619。BLAST分析显示，HdhHSP90与其他已知的HSP90具有很高的相似性，并且在HdhHSP90中还鉴定出5个被定义为HSP90蛋白家族特征的保守氨基酸块，这表明HdhHSP90应该是HSP90家族的细胞质成员。用含硒水平分别为0.15、1.32和48.70 mg Kg−1的半纯化饲料饲喂20周的太平洋鲍鱼，用实时荧光定量PCR检测其血细胞和肝胰腺中HdhHSP90的表达水平。结果表明，饲粮硒水平为1.32 mg Kg−1时，HdhHSP90转录本的表达量显著上调，表达量最高（0.47倍）（p < 0.05）。然而，在饲粮硒过量（48.70 mg Kg−1）时，肝胰腺中这些水平显著降低（p < 0.01）。在血细胞中，HdhHSP90 mRNA的表达量在饲粮硒过量（48.70 mg Kg−1）时升高，达到最大值（0.96倍）（p < 0.01）。提示HdhHSP90在肝胰腺和血细胞中的表达水平可能受硒的影响，HdhHSP90可能参与了鲍鱼的抗氧化反应。
In the present study, the cDNA of heat shock protein 90 from Pacific abalone Haliotis discus hannai Ino (HdhHSP90) was cloned by the combination of homology cloning and rapid amplification of cDNA ends (RACE) approaches. The full-length cDNA of HdhHSP90 was of 2660 bp, including an open reading frame (ORF) of 2187 bp encoding a polypeptide of 728 amino acids with predicted molecular weight of 84.134 kDa and theoretical isoelectric point of 4.619. BLAST analysis revealed that HdhHSP90 shared high similarity with other known HSP90s, and the five conserved amino acid blocks defined as HSP90 protein family signatures were also identified in HdhHSP90, which indicated that HdhHSP90 should be a cytosolic member of the HSP90 family. The expression levels of HdhHSP90 in haemocytes and hepatopancreas were measured by real-time PCR after Pacific abalone were fed with semi-purified diets containing graded levels of selenium (0.15, 1.32 and 48.70 mg Kg−1) for 20 weeks, respectively. The results showed that the expression levels of HdhHSP90 transcript were significantly up-regulated and reached the maximum (0.47-fold) in hepatopancreas of Pacific abalone fed with optimal dietary Se (1.32 mg Kg−1) (p < 0.05). However, these levels significantly decreased in hepatopancreas at the excessive dietary Se (48.70 mg Kg−1) (p < 0.01). In haemocytes, the expression of HdhHSP90 mRNA increased and reached the maximum (0.96-fold) at the excessive dietary Se (48.70 mg Kg−1) (p < 0.01). It is implied that the expression levels of HdhHSP90 could be affected by dietary Se in hepatopancreas and haemocytes, and HdhHSP90 was potentially involved in the anti-oxidation responses in Pacific abalone H. discus hannai.