Salmonella enterica strains survive and propagate in macrophages by both circumventing and resisting the antibacterial effectors normally delivered to the phagosome. An important aspect of Salmonella resistance is the production of periplasmic superoxide dismutase to combat phagocytic superoxide. S. enterica serovar Typhimurium strain 14028 produces two periplasmic superoxide dismutases: SodCI and SodCII. Both enzymes are produced during infection, but only SodCI contributes to virulence in the animal. Although 60% identical to SodCII at the amino acid level with very similar enzymatic properties, SodCI is dimeric, protease resistant, and tethered within the periplasm via a noncovalent interaction. In contrast, SodCII is monomeric and protease sensitive and is released from the periplasm normally by osmotic shock. We have constructed an enzymatically active monomeric SodCI enzyme by site-directed mutagenesis. The resulting protein was released by osmotic shock and sensitive to protease and could not complement the loss of wild-type dimeric SodCI during infection. To distinguish which property is most critical during infection, we cloned and characterized related SodC proteins from a variety of bacteria. Brucella abortus SodC was monomeric and released by osmotic shock but was protease resistant and could complement SodCI in the animal. These data suggest that protease resistance is a critical property that allows SodCI to function in the harsh environment of the phagosome to combat phagocytic superoxide. We propose a model to account for the various properties of SodCI and how they contribute to bacterial survival in the phagosome.Cu/Zn superoxide dismutases (SODs) are metalloproteins that dismute toxic superoxide radicals to H 2 O 2 and O 2 through the alternate oxidation and reduction of the copper(II) ion in the active site. Among bacteria, Cu/Zn SODs (referred to as SodCs) are located in the periplasms of certain gram-negative bacteria (2, 25) and anchored to the surfaces of some grampositive bacteria (13). Salmonella enterica serovar Typhimurium strain 14028 produces two Cu/Zn SODs-SodCI and SodCII. SodCII is the ortholog of Escherichia coli SodC, while SodCI is encoded by the Gifsy-2 prophage. The ability of these two enzymes to contribute to virulence has been studied extensively (12,14,15,24,34,36). Uzzau et al. (36) showed that only SodCI is required for the full virulence of serovar Typhimurium and that SodCII does not contribute to virulence even in the absence of SodCI. Our previous work has confirmed this result, and we have shown that this disparity in virulence phenotypes is due mainly to some differences at the protein level rather than in the regulation of the genes (24). Indeed, SodCI under the control of the more weakly in vivo-induced sodCII promoter (20) was fully capable of complementing wild-type SodCI in an animal infection. These and other data suggested that any differences in enzymatic activity or stability of the active site were insufficient to explain the differential roles of SodC...
Intrauterine infection is a major detriment for maternal-child health and occurs despite local mechanisms that protect the maternal-fetal interface from a wide variety of pathogens. The bacterial pathogen Listeria monocytogenes causes spontaneous abortion, stillbirth, and preterm labor in humans and serves as a model for placental pathogenesis. Given the unique immunological environment of the maternal-fetal interface, we hypothesized that virulence determinants with placental tropism are required for infection of this tissue. We performed a genomic screen in pregnant guinea pigs that led to the identification of 201 listerial genes important for infection of the placenta but not maternal liver. Among these genes was lmrg1778 (lmo2470), here named inlP, predicted to encode a secreted protein that belongs to the internalin family. InlP is conserved in virulent L. monocytogenes strains but absent in Listeria species that are nonpathogenic for humans. The intracellular life cycle of L. monocytogenes deficient in inlP (⌬inlP) was not impaired. In guinea pigs and mice, InlP increased the placental bacterial burden by a factor of 3 log 10 while having only a minor role in other maternal organs. Furthermore, the ⌬inlP strain was attenuated in intracellular growth in primary human placental organ cultures and trophoblasts. InlP is a novel virulence factor for listeriosis with a strong tropism for the placenta. This virulence factor represents a tool for the development of new modalities to prevent and treat infection-related pregnancy complications.T he immunological environment of the maternal-fetal interface is unique because protection of the fetus from pathogens has to be balanced with tolerance of the fetus by the maternal immune system (1, 2). How this is accomplished is one of the major enigmas of mammalian reproduction. Contrary to the long-standing hypothesis that the pregnant mother is immunocompromised (3), recent evidence suggests that the maternal immune system is intricately regulated during pregnancy, and the placenta is well guarded against infection (4-6). A few predominantly intracellular microbes are able to infect the placenta and cause pregnancy complications such as preterm labor, fetal damage, and death (5, 7). Given the unique immunological environment of the maternal-fetal interface and the inability of many pathogens to colonize the placenta, we hypothesized that specific virulence determinants are required for microbes to survive and replicate in this tissue.Listeria monocytogenes is a facultative intracellular bacterial pathogen that causes spontaneous abortion, preterm labor, and stillbirth in humans and other mammals (8, 9). There are ϳ1,600 human cases in the United States per year, and about one-third of these cases are pregnancy associated (10). L. monocytogenes is also extremely amenable to experimental analysis and therefore has been exploited over the past 5 decades to understand host-pathogen interactions of intracellular microbes (11, 12). L. monocytogenes can infect a wide variety ...
Salmonella enterica serovar Typhimurium replicates in macrophages, where it is subjected to antimicrobial substances, including superoxide, antimicrobial peptides, and proteases. The bacterium produces two periplasmic superoxide dismutases, SodCI and SodCII. Although both are expressed during infection, only SodCI contributes to virulence in the mouse by combating phagocytic superoxide. The differential contribution to virulence is at least partially due to inherent differences in the SodCI and SodCII proteins that are independent of enzymatic activity. SodCII is protease sensitive, and like other periplasmic proteins, it is released by osmotic shock. In contrast, SodCI is protease resistant and is retained within the periplasm after osmotic shock, a phenomenon that we term "tethering." We hypothesize that in the macrophage, antimicrobial peptides transiently disrupt the outer membrane. SodCII is released and/or phagocytic proteases gain access to the periplasm, and SodCII is degraded. SodCI is tethered within the periplasm and is protease resistant, thereby remaining to combat superoxide. Here we test aspects of this model. SodCII was released by the antimicrobial peptide polymyxin B or a mouse macrophage antimicrobial peptide (CRAMP), while SodCI remained tethered within the periplasm. A Salmonella pmrA constitutive mutant no longer released SodCII in vitro. Moreover, in the constitutive pmrA background, SodCII could contribute to survival of Salmonella during infection. SodCII also provided a virulence benefit in mice genetically defective in production of CRAMP. Thus, consistent with our model, protecting the outer membrane against antimicrobial peptides allows SodCII to contribute to virulence in vivo. These data also suggest direct in vivo cooperative interactions between macrophage antimicrobial effectors.
Background: During intracellular replication, Legionella pneumophila produces effector proteins that exploit host cell processes. Results:The effector GobX is an E3 ubiquitin ligase that targets to the Golgi where it is post-translationally modified with a palmitoyl moiety. Conclusion: Bacteria can hijack host-mediated S-palmitoylation for subcellular localization of their effectors. Significance: Pharmacological inhibition of S-palmitoylation enzymes may interfere with effector function and thus microbial virulence.
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