牙鲆半乳糖凝集素6基因的克隆、表达及功能研究

Cloning, expression and function of Galectin-6 from Paralichthys olivaceus

中文关键词: 糖凝集素6  牙鲆  迟缓爱德华氏菌  免疫应答

英文关键词: Galectin-6  Paralichthys olivaceus  Edwardsiella tarda  Immune response

中文摘要:

      半乳糖凝集素6 (Galectin-6)是β-半乳糖苷结合凝集素家族的成员之一。本研究首次分离并鉴定了牙鲆(Paralichthys olivaceus) Galectin-6 (PoGalectin-6)，分析了其分子特征和表达模式，并对其免疫相关功能进行了研究。PoGalectin-6基因的开放阅读框长为1089 bp，共编码362个氨基酸，其中包含2个糖识别结构域(CRDs)。同源序列比对和系统进化树分析显示，PoGalectin-6与大菱鲆(Scophthalmus maximus) Galectin-4的相似性为80.9%。组织分布结果显示，PoGalectin-6基因主要在肠组织中特异性表达。迟缓爱德华氏菌(Edwardsiella tarda)感染后，PoGalectin-6基因在肠组织中的表达显著升高，感染后12 h表达量最高，随后逐渐降低并恢复至正常水平。细菌结合实验证实，PoGalectin-6重组蛋白(rPoGalectin-6)能够结合枯草芽孢杆菌(Bacillus subtilis)、蜡样芽孢杆菌(Bacillus cereus)、杀鲑气单胞菌(Aeromonas salmonicida)、迟缓爱德华氏菌和创伤弧菌(Vibrio vulnificus)，但并不结合短小芽孢杆菌(Bacillus pumilus)。此外，rPoGalectin-6以钙离子依赖的方式对短小芽孢杆菌、枯草芽孢杆菌、杀鲑气单胞菌和迟缓爱德华氏菌表现出明显的凝集作用。研究表明，PoGalectin-6基因参与了由迟缓爱德华氏菌感染引起的免疫应答，这一发现为探索Galectin-6在硬骨鱼类中的免疫功能奠定了基础。

英文摘要:

      Galectin-6 is a member of the β-galactoside-binding lectin family, which has been widely studied in mammals (e.g., Lgals6). Galectin-6 has been cloned from several teleost fishes, such as Maylandia zebra, Esox lucius, and Poecilia formosa, but there are few studies on its function. In this study, we analyzed the molecular characteristics of Galectin-6 extracted from Paralichthys olivaceus (PoGalectin-6) and studied its immune-related functions. The full length of the open reading frame (ORF) of PoGalectin-6 cDNA is 1089 bp, encoding 362 amino acids containing two carbohydrate recognition domains (CRDs). Multiple sequence alignment and phylogenetic tree analysis showed that PoGalectin-6 is highly similar to Scophthalmus maximus Galectin-4 (approximately 80.9%). Tissue distribution experiments demonstrated that PoGalectin-6 is specifically expressed in intestinal tissues. The expression level of PoGalectin-6 in the intestine significantly increased after Edwardsiella tarda stimulation, and the highest expression level was observed at 12 h after infection. Recombinant PoGalectin-6 (rPoGalectin-6) exhibited binding ability to gram-positive (Bacillus subtilis, and B. cereus) and gram-negative (Aeromonas salmonicida, E. tarda, and Vibrio vulnificus) bacteria. rPoGalectin-6 showed Ca2+-dependent agglutination activity against B. pumilus, B. subtilis, A. salmonicida, and E. tarda. This study suggests that PoGalectin-6 may play an important role in the immune response against E. tarda infection, laying the foundation for exploring the immune functions of Galectin-6 in teleosts.