文昌鱼VCBP免疫识别和效应功能研究：VCBP类抗体功能探索

Variable region-containing chitin binding proteins (VCBPs) are highly polymorphic, immune-type molecules that have been described in amphioxus, a cephalochordate. There is a wealth of studies about the structure, evolution, expression and function of VCBP, however many questions are still unsolved. Is VCBP capable of recognizing antigens specifically? Whether it is able to neutralize antigens and activate complement? All these questions above are the proposed research contents of this proposal..Our previous study has shown that the challenge with bacteria induced a significant increase in the degree of polymorphism of VCBP, and the mutation directions appeared to be related to the challenges with different bacteria, suggesting that the recognition of bacteria by VCBP might be specific. To test this hypothesis, we will analyze the corresponding relations between the sequence mutations and the challenges with bacteria, and on this basis, we will express these recombinant mutant sequences, and test bacteria-binding activities of these recombinant proteins. To further elucidate the function and regulation mechanism of VCBP in antibacterial immunity of amphioxus, we will conduct a series of experiments, such as bacterial adhesion experiment and complement activity experiment..In a word, we will study the antibody-like activities of VCBP via investigating the mechanism of VCBP in the recognition and elimination of antigens, and this study will provide valuable theoretical basis towards the exploration of the origins and evolution of adaptive immune system.

VCBP是最早在文昌鱼中发现的一类具有高多态性的免疫分子，对其结构、演化、表达和功能已有大量研究。然而，很多问题迄今仍不清楚，如VCBP对抗原的识别是否有特异性呢？是否具有中和抗原以及激活补体的功能呢？这些问题也构成本申请拟研究的主要内容。我们已发现文昌鱼受到细菌刺激后，其体内VCBP的多态性程度显著升高，并对于不同的细菌刺激，VCBP高变区的序列突变呈现不同的倾向性，提示VCBP可能对细菌的识别具有特异性。因此，我们拟在分析这些序列突变与细菌刺激的对应关系基础上，对重要的突变序列进行重组表达，并检测重组蛋白与细菌的结合能力，以验证VCBP对细菌的识别是否具有特异性；同时通过对VCBP抑制细菌粘附、介导补体激活等免疫功能的研究，阐明VCBP在抗原识别和抗原清除阶段的作用机制，揭示VCBP是否具有类似抗体的免疫功能。这一研究将极大丰富和提升对脊椎动物适应性免疫系统的起源的认知。

VCBP是最早在文昌鱼中发现的一类具有高多态性的免疫分子，对其结构、演化、表达和功能已有大量研究。然而，很多问题迄今仍不清楚，如VCBP能否发挥类似抗体的功能？这也是构成本研究的主要内容。我们发现文昌鱼VCBP家族基因主要在肝盲囊和后肠表达，其中VCBP5基因在文昌鱼受到同种细菌二次刺激时，表达量迅速升高，较初次刺激时反应更加强烈，该现象与脊椎动物的免疫记忆现象类似。在文昌鱼受到细菌刺激后，其体内VCBP5的多态性程度显著升高，对于不同的细菌刺激，VCBP5高变区的序列突变呈现不同的倾向性，我们进一步重组表达了这些突变蛋白，发现它们对特定细菌的识别能力显著提高，这些提示VCBP5可能对抗原的识别具有特异性。因VCBP具有多样性，并且其参与宿主的免疫防御过程呈现出记忆性和特异性，由此得出，文昌鱼VCBP能够发挥类似抗体的功能，这些研究结果将进一步拓展我们对适应性免疫系统起源的认知。