Functional analysis of the classical complement pathway in bony fish

哺乳动物的经典补体途径是由抗原结合抗体激活的，在清除入侵病原体方面起着至关重要的作用。尽管经典途径在硬骨鱼中的存在是显而易见的，但其反应机制仍有待阐明。本研究通过分子克隆和功能分析，对鲤鱼补体经典激活途径中的成分进行了鉴定。从鲤鱼肝胰腺cDNA文库中分离到编码甘露糖结合蛋白相关丝氨酸蛋白酶的两个C1r/S亚型、两个C4亚型和一个新的截短亚型的互补克隆。这些序列在几个功能位点上显示出相当大的序列多样性，这表明它们的功能差异。此外，还从同一文库中克隆了三个B/C2-A亚型。它们在不同的组织中不同水平地合成。特别是，在注射β-葡聚糖时，一个新的亚型B/C2-A3仅在肾和脾等淋巴器官中表达，并且其表达上调，表明B/C2-A3亚型是一个急性时相反应物。补体成分同种类型的功能差异表明，这种多样性是硬骨鱼为扩大其固有免疫识别而采取的一种独特的进化策略。

The classical complement pathway of mammals is activated by antigen-bound antibody and plays a crucial role in elimination of invading pathgens. Although the presence of the classical pathway is evident in bony fish, its reaction mechanism remains to be elucidated. In the present study, components involved in the classical pathway activation of the common carp complement was identified by molecular cloning and functional analysis. Complementary DNA clones encoding two C1r/s isotypes, two C4 isotypes and a novel truncated isoform of the mannose-binding protein-associated serine protease (MASP) have been isolated from carp hepatopancreas CDNA library. These show a substantial sequence diversity in several functional sites, suggesting their functional difference. Furthermore, three B/C2-A isotypes were also cloned from the same library. They are synthesized in different tissues at different levels. In particular, a novel isotype, designated B/C2-A3, is expressed exclusively in the lymphoid organs such as kidney and spleen, and its expression is up-regulated, on beta-glucan-injection, suggesting that B/C2-A3 isotype is an acute-phase reactant. The functional divergence of the complement component isotypes implicates that the diversity is a unique evolutioary strategy taken by bony fish to expand its innate immune-recognition.