Hu Wen-Yan, Yao Cui-Luan
Fisheries College, Jimei University, Xiamen, 361021, PR China.
Fisheries College, Jimei University, Xiamen, 361021, PR China.
Fish Shellfish Immunol. 2016 Sep;56:84-95. doi: 10.1016/j.fsi.2016.06.050. Epub 2016 Jun 28.
Apoptosis inducing factor (AIF) and cytochrome c (CYC) are two mitochondrial apoptogenic factors. In the present study, the cDNA sequences of AIF (LvAIF) and CYC (LvCYC) were cloned from Pacific white shrimp, Litopenaeus vannamei. The LvAIF was 1664 bp, including a 5'-terminal untranslated region (UTR) of 154 bp, an open reading frame (ORF) of 1323 bp encoding a polypeptide of 440 amino acids (aa) and a 3' UTR of 187 bp. The LvCYC was 582 bp, including a 50 bp 5' UTR, a 315 bp ORF encoding for 104 aa, and a 217 bp 3' UTR. The deduced protein of LvAIF contained a conserved Pyr_redox and AIF_C domain at the N-terminal and the predicted LvCYC included a conservative cytochrome_C domain, respectively. Phylogenetic analysis revealed that LvAIF belonged to AIF1 subfamily and showed a close relationship with AIF1 from vertebrates and LvCYC showed the closest relationship with its counterparts from shrimp Marsupenaeus japonicus. Tissue expression profiles showed that both LvAIF and LvCYC existed in most tissues, with the most predominant expression of LvAIF in intestine, then followed muscle and the weakest expression in gill. The highest expression of LvCYC was detected in muscle, and the weakest expression was in hemocytes. Additionally, after white spot syndrome virus (WSSV) infection, the significant up-regulation of LvAIF, LvCYC and caspase 3 transcripts and the increase of pro-caspase 3 and active-caspase 3 protein were detected at most time points (P < 0.05). However, all of the three genes down-regulated in hemocytes in the early stage after WSSV infection. WSSV proliferation and shrimp mortality showed a time-dependent manner and the production of ROS in hemocytes were significantly increased at 6 and 24 h after infection. Our results showed that the apoptotic genes AIF, CYC and caspase 3 might play crucial roles in hepatopancreas, however, the production of ROS in hemocytes might be important in shrimp defense against WSSV infection.
凋亡诱导因子（AIF）和细胞色素c（CYC）是两种线粒体凋亡因子。在本研究中，从凡纳滨对虾（Litopenaeus vannamei）中克隆了AIF（LvAIF）和CYC（LvCYC）的cDNA序列。LvAIF为1664 bp，包括154 bp的5′端非翻译区（UTR）、1323 bp的开放阅读框（ORF），编码一个440个氨基酸（aa）的多肽以及187 bp的3′UTR。LvCYC为582 bp，包括50 bp的5′UTR、315 bp的ORF，编码104个aa以及217 bp的3′UTR。推导的LvAIF蛋白在N端包含一个保守的Pyr_redox和AIF_C结构域，预测的LvCYC分别包含一个保守的细胞色素_C结构域。系统发育分析表明，LvAIF属于AIF1亚家族，与脊椎动物的AIF1关系密切，而LvCYC与其来自日本囊对虾（Marsupenaeus japonicus）的对应物关系最密切。组织表达谱显示，LvAIF和LvCYC在大多数组织中均有存在，其中LvAIF在肠道中表达最显著，其次是肌肉，在鳃中表达最弱。LvCYC在肌肉中表达最高，在血细胞中表达最弱。此外，在感染白斑综合征病毒（WSSV）后，在大多数时间点检测到LvAIF、LvCYC和半胱天冬酶3转录本显著上调，以及前体半胱天冬酶3和活性半胱天冬酶3蛋白增加（P < 0.05）。然而，在WSSV感染后的早期，这三个基因在血细胞中均下调。WSSV增殖和对虾死亡率呈时间依赖性，感染后6小时和24小时血细胞中活性氧的产生显著增加。我们的结果表明，凋亡基因AIF、CYC和半胱天冬酶3可能在肝胰腺中发挥关键作用，然而，血细胞中活性氧的产生可能在对虾抵御WSSV感染中起重要作用。
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