Identification ofSalmonellaSPI-2 secretion system components required for SpvB-mediated cytotoxicity in macrophages and virulence in mice

Abstract

The Salmonella SpvB protein possesses ADP-ribosyl transferase activity. SpvB, acting as an intracellular toxin, covalently modifies monomeric actin, leading to loss of F-actin filaments in Salmonella-infected human macrophages. Using defined Salmonella mutants, different functional components of the SPI-2 type three secretion system (TTSS), ssaV, spiC, sseB, sseC, and sseD, were found to be required for SpvB-mediated actin depolymerization in human macrophages. Expression of SpvB protein in Salmonella was not affected by any of the SPI-2 mutants and the effects of these loci were not due to reduced numbers of intracellular bacteria. Interestingly, the major SPI-2 virulence effector, SifA, is not required for SpvB action. Further, caspase-3 activation is an additional marker of cytotoxicity in Salmonella-infected human macrophages. Caspase-3 activity depended on SpvB and SPI-2 TTSS function, but not on SifA. These human macrophage cell culture results were corroborated by virulence studies in mice. Using competitive infection of mice with mixed inocula of single and double mutants, spvBmut1 mutation did not have an effect independent of ssaJ mutation, essential for SPI-2 TTSS function. In contrast, competitive infection studies in mice confirmed that SpvB and SifA have independent virulence effects, as predicted by the macrophage studies.

Impact ofSalmonella entericaType III Secretion System Effectors on the Eukaryotic Host Cell

“…One report showed that SpvB could be secreted into a culture medium mimicking the intracellular iron concentrations of eukaryotic cells in a T3SS-independent manner [316]. However, several components of the T3SS2, have been shown to be essential for SpvB-induced actin depolymerization and delayed cytotoxicity, strongly supporting the idea that SpvB is translocated into the host cell through T3SS2 [314,317].…”

Diarrhea and Colitis in Mice Require the Salmonella Pathogenicity Island 2-Encoded Secretion Function but Not SifA or Spv Effectors

“…SpvB is an ADP-ribosylating toxin expressed within the SCV and translocated by SPI-2 into the cytoplasm, where it depolymerizes F actin and causes macrophage apoptosis (9,33). SpvB is encoded by a gene that is part of the spv operon that we inactivated directly by a deletion and indirectly by making a mutation in spvR, which abrogates expression of all genes in the operon (19).…”

“…All mutations were transduced into a single 14028s lineage by use of P22 phage. Salmonella SPI-1, SPI-2, and spv mutants have been described previously (9).…”

A Salmonella enterica serovar Typhi plasmid induces rapid and massive apoptosis in infected macrophages

“…Intracellular expression of the SpvB protein induces not only disruption of actin filaments, but also apoptotic cell death in eukaryotic cells. 3,24,25 We found a gene sequence that is homologous to spvB on the pR ST98 plasmid. However, pR ST98 is a large chimeric plasmid containing complex sequences of unknown functions.…”