Dorsal是Toll通路的核转录因子。昆虫Dorsal或/和Dif组成二聚体入核激活抗菌肽的表达，但在甲壳动物中只报道了Dorsal基因。申请人在克氏原螯虾中鉴定到两种含有形成二聚体所需RHD结构域的Dorsal异构体，并发现Dorsal-a应对WSSV刺激转录水平明显增加，Dorsal-b应对弧菌刺激转录水平增加。基于此，拟开展：1）Dorsal异构体可变剪接方式（预测是内含子保留型）与时空表达分析，2）利用Native-PAGE、Pull-down、co-IP验证Dorsal-a和Dorsal-b能否形成不同二聚体组合（同源或/和异源），3）利用免疫荧光明确不同病原感染是否导致不同组合的异构体入核，干扰、过表达和EMSA进一步探讨不同二聚体入核是否调控不同抗菌肽的表达。最终阐明可变剪接形成的Dorsal异构体在免疫中的功能和机制，拓展先天免疫知识体系，为螯虾抗病免疫防御提供新思路。

Dorsal is a nuclear transcription factor of the Toll pathway. Insects Dorsal or/and Dif form dimers into the nucleus to activate the expression of antimicrobial peptides (AMPs), but only the Dorsal gene has been reported in crustaceans. We identified two Dorsal isoforms containing the Rel homology domains (RHD) required for dimer formation in Procambarus clarkii and found that transcription level of Dorsal-a was significantly increased after WSSV stimulation, and Dorsal-b expression was remarkably increased after Vibrio challenge. Based on this, it is planned to carry out: 1) analysis of alternative splicing method (predicted to be intron-retained) and spatiotemporal expression of two Dorsal isoforms; 2) use Native-PAGE, Pull-down and co-IP to verify whether Dorsal-a and Dorsal-b can form different dimer combinations (homologous or/and heterologous); 3) use immunofluorescence to clarify whether different pathogen infections cause different combinations of isoforms to enter the nucleus. RNA interference in vivo, overexpression in Drosophila S2 cells and EMSA in vitro to further explore whether the entry of different dimers into the nucleus regulate the expression of different AMPs. Finally, it is expected to clarify the function and mechanism of two Dorsal isoforms formed by alternative splicing in immunity, expand the knowledge system of innate immunity and provide fresh ideas for disease prevention and immune defense of crayfish.