转录因子-CR46018-靶基因轴在果蝇Imd通路免疫应答中的功能研究

目前关于长链非编码RNA（lncRNA）自身表达调控及其在动物先天免疫调控中的作用仍少有研究。申请人前期通过遗传学筛选发现lncRNA（CR46018）能通过果蝇Imd通路调控抗菌肽基因的表达，并预测Dfd、hb和gt等转录因子能结合CR46018启动子区参与Imd免疫应答。为揭示其机制，本项目拟利用组学及生物信息学技术筛选调控CR46018的转录因子和CR46018调控的靶基因；通过ChIP技术和双荧光素酶报告系统鉴定转录因子对CR46018的调控；ChIRP技术验证CR46018与靶基因的互作；并构建转录因子（如Dfd）、CR46018和靶基因不同（过表达或敲低）基因型组合的挽救型果蝇品系，利用qRT-PCR和Western blot等技术阐明转录因子-CR46018-靶基因轴在果蝇Imd免疫通路中的作用机制。项目的完成对进一步完善动物先天免疫应答的复杂调控机制具有重要的理论意义。

At present, the self-expression regulation of long non-coding RNA (lncRNA) and its role in the regulation of animal innate immunity are still few studied. Through genetic screening, the applicant previously has found that lncRNA (CR46018) could regulate the expression of antimicrobial peptide genes through Drosophila Imd pathway, and has predicted that transcription factors such as Dfd, hb and gt could combine with the promoter region of CR46018 to participate in the Imd immune response. In order to reveal its mechanism, this project will intend to screen transcription factors that regulate CR46018 and target genes regulated by CR46018 by using omics analyses and bioinformatics techniques. The regulation of transcription factors on CR46018 will be identified by ChIP technique and Dual-luciferase reporter system. The interaction between CR46018 and target genes will be verified by ChIRP technology. In addition, we will construct the Drosophila rescued strains with different genotype combinations of transcription factors (such as Dfd), CR46018 and target genes (overexpression or knockdown). And the regulation mechanism of transcription factor-CR46018-target genes axis in Drosophila Imd immune pathway will be elucidated by qRT-PCR and Western blot techniques. Taken together，the completion of the project is of great theoretical significance for further improving the complex regulatory mechanism of animal innate immune response.