天然免疫信号调控肠道上皮细胞脱落的分子机制研究

The intestinal epithelium is not only the primary organ of digestion, but also constitutes a crucial barrier between the host and the external environment. It undergoes constant epithelial renewal through cell shedding and stem cell activation. Despite the dynamic nature of the intestinal epithelium, intestinal barrier integrity has to be tightly maintained in order to prevent penetration of harmful intraluminal entities including foreign antigens, microorganisms and their toxins. Conversely, pathological epithelial shedding compromises gut barrier function and acts as a fundamental cause of inflammatory bowel diseases. Despite extensive focus on gut regenerative growth and general epithelial biology over the past years, the molecular and cellular mechanisms underlying epithelial shedding have never been systematically investigated using a genetic approach, and consequently our knowledge on epithelial shedding remains rudimentary. We have recently revealed that the epithelium-intrinsic Imd/NFκB immune pathway triggered by enteric bacterial infection is required for intestinal epithelial shedding in Drosophila, pointing to an evolutionarily conserved crosstalk between immunity and epithelial turnover (Zhai et al., 2018 Immunity). Built on this work and taking advantage of the versatile Drosophila genetic tools, the present project is designed to obtain an in-depth understanding of immunity-induced epithelial shedding at the mechanistic level, by using a combination of the state-of-the-art approaches ranging from comparative genomics, large-scale genetic screening to in vivo live imaging. Its execution is expected to identify novel genes and pathways that contribute to epithelial shedding, elucidate the role of genes responsible for the epithelial cell polarity program in cell shedding, and further uncover regulatory mechanisms linking the immune pathway to epithelial shedding. This project is significant and timely, as it will promote a mechanistic understanding of both intestinal epithelial renewal and host-microbe interactions, two current topics with extensive focus. Because epithelial cell biology and architecture are broadly conserved, understanding the basic mechanisms that are essential for epithelial shedding will ultimately open new therapeutic avenues to combat intestinal inflammation and improve intestinal health of animals, including human.

肠道上皮是动物生理和防御的结构基础。上皮细胞脱落启动肠道更新，作为肠道最根本的细胞过程与干细胞分裂分化一同构筑起动态性的肠道上皮。非生理性的细胞脱落破坏上皮屏障功能，是肠道炎症发生发展最根本的原因。以往工作多侧重研究肠道干细胞维持和上皮细胞生物学方面的机制，尚缺少对肠上皮细胞脱落分子细胞机制的系统探索。我们的最新研究成果揭示，细菌感染激活肠道上皮Imd/NFκB天然免疫通路是导致上皮细胞脱落的重要原因。在此基础上，本项目拟继续发挥果蝇遗传模式的优势，通过比较基因组学、大规模遗传筛选等手段，系统发现并功能鉴定参与肠上皮细胞脱落的新基因，检测维持上皮细胞特性的蛋白在细胞脱落中的作用，并深入解析免疫信号调控上皮细胞脱落的分子机制。相关结果将加深对肠道上皮更新机制及宿主与微生物互作机理的认识，并为干预非生理性上皮细胞脱落提供理论依据和有效靶点，为改善动物肠道健康指明新方向。

微生物感染导致肠道上皮免疫激活和肠道上皮细胞脱落是从果蝇到哺乳动物都保守的细胞过程。我们之前报道了感染导致的IMD/NFkB/Relish信号激活导致细胞特异性的免疫功能输出：上皮细胞脱落或抗菌肽的表达，建立起免疫信号除调控抗细菌免疫之外在调节肠道上皮更新方面的作用。感谢自然基金的支持，本项目继续深入揭示免疫信号调控肠道上皮细胞脱落的分子机制。通过构建并使用Relish的GFP敲入品系，我们发现Relish被激活而入核的强度可能决定了免疫通路的细胞特异性功能输出，并进一步提示JNK信号通路可能通过增强Relish入核而实现调控抗菌肽表达和上皮细胞脱落之间的转换。对维持上皮细胞特性相关蛋白的系统筛选实验揭示整合素（integrin）在上皮细胞脱落中发挥重要作用，免疫信号可能作用于整合素来调控细胞脱落。在生理意义方面，本项目揭示抑制肠道上皮细胞脱落导致宿主清除入侵肠道细菌的能力下降。因此，肠道上皮细胞脱落是宿主应对外源细菌侵染的有机组成部分，与抗菌肽的激活一起构筑起了肠道上皮防御。本项目进一步通过转录组学技术分析获得了可能参与上皮细胞脱落的基因；使用GFP-Relish敲入品系，我们优化了发现Relish直接结合的下游基因区域以及Relish互作蛋白的方法，为全面揭示NFkB因子Relish发挥作用的蓝图打下了基础。本项目取得的结果为天然免疫研究领域理解免疫通路在调节免疫和其它非免疫过程之间开关的机制提供了基本框架，项目揭示的调控上皮细胞脱落的机制对理解宿主与病原微生物的互作具有重要理论意义。