Boddicker Jennifer D, Jones Bradley D
Department of Microbiology, Roy J. and Lucille A. Carver School of Medicine, University of Iowa, Iowa City, Iowa 52242, USA.
Infect Immun. 2004 Apr;72(4):2002-13. doi: 10.1128/IAI.72.4.2002-2013.2004.
Salmonella enterica serovar Typhimurium causes self-limiting gastroenteritis in humans and a typhoid-like disease in mice that serves as a model for typhoid infections in humans. A critical step in Salmonella pathogenesis is the invasion of enterocytes and M cells of the small intestine via expression of a type III secretion system, encoded on Salmonella pathogenicity island 1 (SPI-1), that secretes effector proteins into host cells, leading to engulfment of the bacteria within large membrane ruffles. The in vitro regulation of invasion genes has been the subject of much scientific investigation. Transcription of the hilA gene, which encodes an OmpR/ToxR-type transcriptional activator of downstream invasion genes, is increased during growth under high-osmolarity and low-oxygen conditions, which presumably mimic the environment found within the small intestine. Several negative regulators of invasion gene expression have been identified, including HilE, Hha, and Lon protease. Mutations within the respective genes increase the expression of hilA when the bacteria are grown under environmental conditions that are not favorable for hilA expression and invasion. In this study, the intracellular expression of invasion genes was examined, after bacterial invasion of HEp-2 epithelial cells, using Salmonella strains containing plasmid-encoded short-half-life green fluorescent protein reporters of hilA, hilD, hilC, or sicA expression. Interestingly, the expression of SPI-1 genes was down-regulated after invasion, and this was important for the intracellular survival of the bacteria. In addition, the effects of mutations in genes encoding negative regulators of invasion on intracellular hilA expression were examined. Our results indicate that Lon protease is important for down-regulation of hilA expression and intracellular survival after the invasion of epithelial cells.
肠炎沙门氏菌鼠伤寒血清型可引起人类自限性肠胃炎以及小鼠的伤寒样疾病，后者可作为人类伤寒感染的模型。沙门氏菌致病的关键步骤是通过表达Ⅲ型分泌系统侵入小肠的肠上皮细胞和M细胞，该系统由沙门氏菌致病岛1（SPI-1）编码，可将效应蛋白分泌到宿主细胞中，导致细菌被大的膜皱褶吞噬。侵袭基因的体外调控一直是众多科学研究的主题。hilA基因编码下游侵袭基因的OmpR/ToxR型转录激活因子，在高渗透压和低氧条件下生长时其转录增加，这可能模拟了小肠内的环境。已鉴定出几种侵袭基因表达的负调控因子，包括HilE、Hha和Lon蛋白酶。当细菌在不利于hilA表达和侵袭的环境条件下生长时，相应基因内的突变会增加hilA的表达。在本研究中，使用含有质粒编码的hilA、hilD、hilC或sicA表达的短半衰期绿色荧光蛋白报告基因的沙门氏菌菌株，在细菌侵袭HEp-2上皮细胞后检测侵袭基因的细胞内表达。有趣的是，侵袭后SPI-1基因的表达下调，这对细菌的细胞内存活很重要。此外，还检测了编码侵袭负调控因子的基因突变对细胞内hilA表达的影响。我们的结果表明，Lon蛋白酶对于上皮细胞侵袭后hilA表达的下调和细胞内存活很重要。
