Bacterial and host factors governing enterocyte exit of Salmonella

沙门氏菌致病的特点是能够穿透完整的肠道上皮屏障，到达固有层并引起炎症反应。上皮屏障的穿透需要非吞噬细胞的入侵、细胞内的存活、顶端到基底侧的迁移和细胞的退出。尽管在过去的几十年里，细胞侵袭和细胞内存活已经被广泛研究，但沙门氏菌移行和退出上皮细胞的过程并没有得到太多的关注。在前人工作的基础上，本研究拟对肠上皮细胞底部和根尖出口进行功能研究，筛选和分析影响肠上皮细胞出口的细菌因素，并探讨顶端肠细胞出口及其在体内宿主传播中的作用。为了实现这些目标，将使用生命成像和先进的电子显微镜、转座子突变和迭代共培养筛选来确定促进细胞退出的基因、定向细菌突变和遗传命运图谱，并将结合在Transwell插入物上生长的极化肠上皮m-ICcl2细胞、肠上皮干细胞器体和已建立的新生小鼠感染模型的体外共培养模型进行结合。这些结果有望极大地拓宽我们对感染早期复杂宿主-微生物相互作用的理解，并可能为未来的预防或治疗策略提供分子靶点。

Hallmark of the pathogenesis of Salmonella is the ability to penetrate the intact intestinal epithelial barrier, reach the underlying lamina propria and evoke an inflammatory response. Penetration of the epithelial barrier requires invasion of non-phagocytic cells, intracellular survival, apical to basolateral transmigration and cell exit. Whereas cell invasion and intracellular survival have been extensively investigated during the last decades, the process the Salmonella transmigration and exit from epithelial cells has not obtained much attention. Based on our previous work, here we plan to functionally characterize the basolateral and apical enterocyte exit, screen for and analyze bacterial factors influencing enterocyte exit, and investigate apical enterocyte exit and its role in vivo in host transmission. To achieve these goals, life imaging and advanced electron microscopy, transposon mutagenesis and an iterative co-culture screen to identify genes promoting cell exit, targeted bacterial mutagenesis, and genetic fate mapping will be employed in combination with an in vitro co-culture model of polarized intestinal epithelial m-ICcl2 cells grown on transwell inserts, intestinal epithelial stem cell organoids and an established neonatal mouse infection model. The results are expected to significantly broaden our understanding of the intricate host-microbe interaction during the early course of the infection and may provide molecular targets for future preventive or therapeutic strategies.