Lon Protease Activity Causes Down-Regulation of Salmonella Pathogenicity Island 1 Invasion Gene Expression after Infection of Epithelial Cells

Abstract

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.

Signal Pathway in Salt-Activated Expression of theSalmonellaPathogenicity Island 1 Type III Secretion System inSalmonella entericaSerovar Typhimurium

“…Similar results were observed when 100 mM NaCl was replaced with 200 mM sucrose (data not shown). These results are consistent with previous reports indicating that HilA, HilC, and HilD regulate SPI1 gene transcription and that HilC and HilD activate hilA transcription equally (3,7,8,9,10,11,19,33,34,37,40,41,47,49). To determine if the salt response required HilC and HilD equally, we performed an epistasis analysis.…”

“…In contrast, the network responsible for controlling expression, assembly, and function of the SPI1 T3SS is still being actively investigated. It is quite clear that HilA is the proximal activator of SPI1 genes (4,7,8,26,33,34,43,49) and that hilA transcription is controlled by two other regulators, HilD and HilC (3,9,10,11,19,37,39,41,42,47,52). These regulators, in turn, have been reported to be controlled by four two-component signal transduction pathways.…”

Regulation ofSalmonella entericaPathogenicity Island 1 by DNA Adenine Methylation

“…The ferric uptake regulatory protein, Fur, and the BarA/SirA two-component system are SPI-1 activators (Fortune et al 2006;Ellermeier and Slauch 2008). In turn, HilE (Fahlen et al 2000) and Lon (Takaya et al 2003;Boddicker and Jones 2004) are negative regulators of SPI-1.…”

Targeted Protein Degradation by Salmonella under Phagosome-mimicking Culture Conditions Investigated Using Comparative Peptidomics

“…Bacterial histone-like proteins have similar DNA-binding extensions (54,55), and three (hupA (56), stpA (57), and ymoA (58)) are known Lon substrates. Paradoxically, although Lon down-regulates the expression of invasion genes and the efficiency of host-cell invasion (59,60), the study of Lon-null mutants has demonstrated that Lon is essential for systemic infection (61). Because the availability of nutrients within the phagosome is limited, ribosomal protein degradation may foster pathogenesis by generating much needed amino acids, thereby increasing the survivability of phagocytosed Salmonella.…”