Duan Yafei, Liu Ping, Li Jitao, Li Jian, Wang Yun, Chen Ping
Key Laboratory of Sustainable Development of Marine Fisheries, Ministry of Agriculture, Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Qingdao, 266071, People's Republic of China.
Cell Stress Chaperones. 2014 Jul;19(4):549-58. doi: 10.1007/s12192-013-0488-x. Epub 2014 Jan 10.
Chitinases are essential enzymes for crustaceans and participates in several biological processes, such as nutrient digestion, morphogenesis, pathogenesis, and pathogen defense. In the present study, the full-length cDNA of Chi (named EcChi) was cloned from the hemocytes of ridgetail white prawn Exopalaemon carinicauda by rapid amplification of cDNA ends methods. The full-length cDNA of EcChi was 1,319 bp, including contains a 5'-untranslated region (UTR) of 42 bp, 3'-UTR of 101 bp with a poly (A) tail, an open-reading frame of 1,176 bp, encoding a 391-amino acid polypeptide with the predicted molecular weight of 43.71 kDa and estimated isoelectric point of 4.78. Sequence analysis revealed that the conserved chitinases family 18 active site was predicted in the amino acid sequence of EcChi. BLAST analysis revealed that amino acids of EcChi shared high identity (61-77 %) with that of other crustaceans. Quantitative real-time PCR analysis indicated that EcChi could be detected in all the tested tissues, and strongly expressed in hepatopancreas of E. carinicauda. After challenged with Vibrio anguillarum and WSSV, EcChi transcripts both in hemocytes and hepatopancreas increased significantly in the first 3 h, respectively. These results indicated that EcChi might be involved in the innate immune responses to V. anguillarum and WSSV in E. carinicauda.
几丁质酶是甲壳类动物的必需酶，参与多种生物学过程，如营养消化、形态发生、发病机制和病原体防御。在本研究中，通过cDNA末端快速扩增方法从脊尾白虾血细胞中克隆了几丁质酶（命名为EcChi）的全长cDNA。EcChi的全长cDNA为1319bp，包括42bp的5'-非翻译区（UTR）、带有聚（A）尾的101bp的3'-UTR和1176bp的开放阅读框，编码一个391个氨基酸的多肽，预测分子量为43.71kDa，估计等电点为4.78。序列分析表明，在EcChi的氨基酸序列中预测到了保守的18家族几丁质酶活性位点。BLAST分析表明，EcChi的氨基酸与其他甲壳类动物的氨基酸具有高度同源性（61-77%）。定量实时PCR分析表明，在所有测试组织中都能检测到EcChi，并且在脊尾白虾的肝胰腺中强烈表达。在用鳗弧菌和白斑综合征病毒攻击后，血细胞和肝胰腺中的EcChi转录本在最初3小时内均显著增加。这些结果表明，EcChi可能参与了脊尾白虾对鳗弧菌和白斑综合征病毒的先天免疫反应。
