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PloS One 2021The foodborne pathogen Listeria monocytogenes is able to survive across a wide range of intra- and extra-host environments by appropriately modulating gene expression...
The foodborne pathogen Listeria monocytogenes is able to survive across a wide range of intra- and extra-host environments by appropriately modulating gene expression patterns in response to different stimuli. Positive Regulatory Factor A (PrfA) is the major transcriptional regulator of virulence gene expression in L. monocytogenes. It has long been known that activated charcoal is required to induce the expression of PrfA-regulated genes in complex media, such as Brain Heart Infusion (BHI), but not in chemically defined media. In this study, we show that the expression of the PrfA-regulated hly, which encodes listeriolysin O, is induced 5- and 8-fold in L. monocytogenes cells grown in Chelex-treated BHI (Ch-BHI) and in the presence of activated charcoal (AC-BHI), respectively, relative to cells grown in BHI medium. Specifically, we show that metal ions present in BHI broth plays a role in the reduced expression of the PrfA regulon. In addition, we show that expression of hly is induced when the levels of bioavailable extra- or intercellular iron are reduced. L. monocytogenes cells grown Ch-BHI and AC-BHI media showed similar levels of resistance to the iron-activated antibiotic, streptonigrin, indicating that activated charcoal reduces the intracellular labile iron pool. Metal depletion and exogenously added glutathione contributed synergistically to PrfA-regulated gene expression since glutathione further increased hly expression in metal-depleted BHI but not in BHI medium. Analyses of transcriptional reporter fusion expression patterns revealed that genes in the PrfA regulon are differentially expressed in response to metal depletion, metal excess and exogenous glutathione. Our results suggest that metal ion abundance plays a role in modulating expression of PrfA-regulated virulence genes in L. monocytogenes.
Topics: Bacterial Proteins; Bacterial Toxins; Charcoal; Culture Media; Gene Expression Regulation, Bacterial; Glutathione; Heat-Shock Proteins; Hemolysin Proteins; Iron; Listeria monocytogenes; Peptide Termination Factors; Polystyrenes; Polyvinyls; Streptonigrin; Virulence; Zinc
PubMed: 33914817
DOI: 10.1371/journal.pone.0250989 -
Veterinary Research Mar 2021Iron is essential for most bacteria to survive, but excessive iron leads to damage by the Fenton reaction. Therefore, the concentration of intracellular free iron must...
Iron is essential for most bacteria to survive, but excessive iron leads to damage by the Fenton reaction. Therefore, the concentration of intracellular free iron must be strictly controlled in bacteria. Riemerella anatipestifer (R. anatipestifer), a Gram-negative bacterium, encodes the iron uptake system. However, the iron homeostasis mechanism remains largely unknown. In this study, it was shown that compared with the wild type R. anatipestifer CH-1, R. anatipestifer CH-1Δfur was more sensitive to streptonigrin, and this effect was alleviated when the bacteria were cultured in iron-depleted medium, suggesting that the fur mutant led to excess iron accumulation inside cells. Similarly, compared with R. anatipestifer CH-1∆recA, R. anatipestifer CH-1∆recAΔfur was more sensitive to HO-induced oxidative stress when the bacteria were grown in iron-rich medium rather than iron-depleted medium. Accordingly, it was shown that R. anatipestifer CH-1∆recAΔfur produced more intracellular ROS than R. anatipestifer CH-1∆recA in iron-rich medium. Electrophoretic mobility shift assays showed that R. anatipestifer CH-1 Fur suppressed the transcription of putative iron uptake genes through binding to their promoter regions. Finally, it was shown that compared with the wild type, R. anatipestifer CH-1Δfur was significantly attenuated in ducklings and that the colonization ability of R. anatipestifer CH-1Δfur in various tissues or organs was decreased. All these results suggested that Fur is important for iron homeostasis in R. anatipestifer and its pathogenic mechanism.
Topics: Bacterial Proteins; Iron; Oxidative Stress; Riemerella; Virulence
PubMed: 33741064
DOI: 10.1186/s13567-021-00919-9 -
Cells Jan 2021Circulating extracellular DNA (ecDNA) is known to worsen the outcome of many diseases. ecDNA released from neutrophils during infection or inflammation is present in the...
Circulating extracellular DNA (ecDNA) is known to worsen the outcome of many diseases. ecDNA released from neutrophils during infection or inflammation is present in the form of neutrophil extracellular traps (NETs). It has been shown that higher ecDNA concentration occurs in a number of inflammatory diseases including inflammatory bowel disease (IBD). Enzymes such as peptidyl arginine deiminases (PADs) are crucial for NET formation. We sought to describe the dynamics of ecDNA concentrations and fragmentation, along with NETosis during a mouse model of chemically induced colitis. Plasma ecDNA concentration was highest on day seven of dextran sulfate sodium (DSS) intake and the increase was time-dependent. This increase correlated with the percentage of cells undergoing NETosis and other markers of disease activity. Relative proportion of nuclear ecDNA increased towards more severe colitis; however, absolute amount decreased. In colon explant medium, the highest concentration of ecDNA was on day three of DSS consumption. Early administration of PAD4 inhibitors did not alleviate disease activity, but lowered the ecDNA concentration. These results uncover the biological characteristics of ecDNA in IBD and support the role of ecDNA in intestinal inflammation. The therapeutic intervention aimed at NETs and/or nuclear ecDNA has yet to be fully investigated.
Topics: Animals; Biomarkers; Colitis; DNA; DNA, Mitochondrial; Deoxyribonucleases; Dextran Sulfate; Endoscopy; Extracellular Space; Extracellular Traps; Inflammation; Intestinal Mucosa; Intestines; Mice, Inbred C57BL; Ornithine; Protein-Arginine Deiminase Type 4; Severity of Illness Index; Streptonigrin; Mice
PubMed: 33418977
DOI: 10.3390/cells10010081 -
Scientific Reports Jun 2020Elevated levels of plasma alpha1-antitrypsin (AAT) correlate with a poor prognosis of various cancers. Herein, we investigated effects of exogenous AAT on non-small lung...
Elevated levels of plasma alpha1-antitrypsin (AAT) correlate with a poor prognosis of various cancers. Herein, we investigated effects of exogenous AAT on non-small lung cancer cell lines with high (H1975) and very low (H661) baseline expression of SERPINA1 gene encoding AAT protein. Comparison of cells grown for 3 weeks in a regular medium versus medium supplemented with 2 mg/ml of AAT revealed that in the presence of AAT cells acquire better proliferative properties, resistance to staurosporine (STS)-induced apoptosis, and show higher expression of CLU, a pro-tumorigenic gene coding clusterin protein. Similarly, the co-administration of STS with AAT or addition of AAT to the cells pre-treated with STS abrogated effects of STS in both cell lines. Following experiments with H1975 cells have shown that AAT blocks critical steps in STS-induced cell death: inhibition of AKT/MAPK pathways, and activation of caspase 3 and autophagy. AAT does not inhibit apoptosis-triggered by chloroquine (inhibitor of autophagy) or streptonigrin (inducer of p53 pathway). The anti-apoptotic effects of AAT were unaffected by lipopolysaccharide (LPS). However, AAT induced TLR4 levels and enhanced LPS effects on the production of IL-6, a tumor-promoting cytokine. Our data provide further evidence that AAT plays a significant role in the tumorigenesis.
Topics: Apoptosis; Autophagy; Cell Movement; Cell Proliferation; Cytokines; Drug Resistance, Neoplasm; Enzyme Inhibitors; Humans; Lipopolysaccharides; Lung Neoplasms; Protective Agents; Serine Proteinase Inhibitors; Staurosporine; Tumor Cells, Cultured; alpha 1-Antitrypsin
PubMed: 32533048
DOI: 10.1038/s41598-020-66825-w -
Journal of Bacteriology Jun 2020Group A streptococcus (GAS) produces millions of infections worldwide, including mild mucosal infections, postinfection sequelae, and life-threatening invasive diseases....
Group A streptococcus (GAS) produces millions of infections worldwide, including mild mucosal infections, postinfection sequelae, and life-threatening invasive diseases. During infection, GAS readily acquires nutritional iron from host heme and hemoproteins. Here, we identified a new heme importer, named SiaFGH, and investigated its role in GAS pathophysiology. The SiaFGH proteins belong to a group of transporters with an unknown ligand from the recently described family of energy coupling factors (ECFs). A deletion mutant exhibited high streptonigrin resistance compared to the parental strain, suggesting that iron ions or an iron complex is the likely ligand. Iron uptake and inductively coupled plasma mass spectrometry (ICP-MS) studies showed that the loss of did not impact GAS import of ferric or ferrous iron, but the mutant was impaired in using hemoglobin iron for growth. Analysis of cells growing on hemoglobin iron revealed a substantial decrease in the cellular heme content in the mutant compared to the complemented strain. The induction of the genes in resulted in the induction of heme uptake. The mutant exhibited a significant impairment in murine models of mucosal colonization and systemic infection. Together, the data show that SiaFGH is a new type of heme importer that is key for GAS use of host hemoproteins and that this system is imperative for bacterial colonization and invasive infection. ECF systems are new transporters that take up various vitamins, cobalt, or nickel with a high affinity. Here, we establish the GAS SiaFGH proteins as a new ECF module that imports heme and demonstrate its importance in virulence. SiaFGH is the first heme ECF system described in bacteria. We identified homologous systems in the genomes of related pathogens from the phylum. Notably, GAS and other pathogens that use a SiaFGH-type importer rely on host hemoproteins for a source of iron during infection. Hence, recognizing the function of this noncanonical ABC transporter in heme acquisition and the critical role that it plays in disease has broad implications.
Topics: Animals; Bacterial Proteins; Biological Transport; Female; Gene Expression Regulation, Bacterial; Heme; Humans; Iron; Membrane Transport Proteins; Mice; Streptococcal Infections; Streptococcus pyogenes; Virulence
PubMed: 32393520
DOI: 10.1128/JB.00205-20 -
Scientific Reports Feb 2020Heterochromatin is essential for regulating global gene transcription and protecting genome stability, and may play a role in tumor suppression. Drugs promoting...
Heterochromatin is essential for regulating global gene transcription and protecting genome stability, and may play a role in tumor suppression. Drugs promoting heterochromatin are potential cancer therapeutics but very few are known. In order to identify drugs that can promote heterochromatin, we used a cell-based method and screened NCI drug libraries consisting of oncology drugs and natural compounds. Since heterochromatin is originally defined as intensely stained chromatin in the nucleus, we estimated heterochromatin contents of cells treated with different drugs by quantifying the fluorescence intensity of nuclei stained with Hoechst DNA dye. We used HeLa cells and screened 231 FDA-approved oncology and natural substance drugs included in two NCI drug libraries representing a variety of chemical structures. Among these drugs, streptonigrin most prominently caused an increase in Hoechst-stained nuclear fluorescence intensity. We further show that streptonigrin treated cells exhibit compacted DNA foci in the nucleus that co-localize with Heterochromatin Protein 1 alpha (HP1α), and exhibit an increase in total levels of the heterochromatin mark, H3K9me3. Interestingly, we found that streptonigrin promotes heterochromatin at a concentration as low as one nanomolar, and at this concentration there were no detectable effects on cell proliferation or viability. Finally, in line with a previous report, we found that streptonigrin inhibits STAT3 phosphorylation, raising the possibility that non-canonical STAT function may contribute to the effects of streptonigrin on heterochromatin. These results suggest that, at low concentrations, streptonigrin may primarily enhance heterochromatin formation with little toxic effects on cells, and therefore might be a good candidate for epigenetic cancer therapy.
Topics: Antibiotics, Antineoplastic; Cell Nucleus; Cell Proliferation; Chromatin Assembly and Disassembly; Chromobox Protein Homolog 5; Chromosomal Proteins, Non-Histone; HeLa Cells; Heterochromatin; Histones; Humans; Phosphorylation; STAT3 Transcription Factor; Streptonigrin
PubMed: 32103104
DOI: 10.1038/s41598-020-60469-6 -
Molecules (Basel, Switzerland) Nov 2019Natural 5,8-quinolinedione antibiotics exhibit a broad spectrum of activities including anticancer, antibacterial, antifungal, and antimalarial activities. The... (Review)
Review
Natural 5,8-quinolinedione antibiotics exhibit a broad spectrum of activities including anticancer, antibacterial, antifungal, and antimalarial activities. The structure-activity research showed that the 5,8-quinolinedione scaffold is responsible for its biological effect. The subject of this review report is a presentation of the pharmacological activity of synthetic 5,8-quinolinedione compounds containing different groups at C-6 and/or C-7 positions. The relationship between the activity and the mechanism of action is included if these data have been included in the original literature. The review mostly covers the period between 2000 and 2019. Previously published literature data were used to present historical points.
Topics: Molecular Structure; Quinolines; Streptonigrin; Structure-Activity Relationship
PubMed: 31739496
DOI: 10.3390/molecules24224115