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Zhang Y, Zhang J, Chen J, Qi X, Zhang Z, Chen H, Wu B, Zhan L, Chen J. Sporadic Listeriosis Cases, Genotypic Characteristics, and Antibiotic Resistance of Associated Listeria monocytogenes Isolates in Zhejiang Province, China (2016 - 2022). Foodborne Pathog Dis 2025. [PMID: 39992218 DOI: 10.1089/fpd.2024.0083] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/25/2025] Open
Abstract
Listeria monocytogenes (L. monocytogenes) is an important foodborne pathogen. In this study, 41 sporadic listeriosis cases were collected during 2016-2022, including 92.7% of invasive cases and 56.1% of pregnancy-associated cases. The age of cases ranged from 0 days to 88 years, with the majority occurring in individuals aged 20 to <30 years. Serotype 1/2 b was most prevalent among 43 L. monocytogenes isolates, followed by 1/2 a, 4 b, and 3a. Sixteen clonal complexes (CCs) were determined. CC87 occupied the top slot. Genome sequencing-based phylogeny results indicated that Chinese CC1, CC8, and CC87 isolates mostly clustered in clades separating from isolates from other countries. Meanwhile, a few Chinese isolates participated in cocirculating CC1, CC8, and CC87 in Asia, Europe, Africa, South America, North America, and Oceania. All isolates harbored LIPI-1 while LIPI-2 was absent. LIPI-3 and LIPI-4 exhibited an apparent relationship with lineage and CCs. It was notably that CC4, CC224, and CC619 carried both LIPI-3 and LIPI-4. inlB, inlC, inlH, inlK, ipeA, srtA, dltA, lap, ami, fbpA, stp, oatA, intA, prsA2, lgt, hpt, iplA1, bsh, mdrT, mdrM, and brtA existed in all isolates. The percentages of inlA, inlF, inlJ, aut, vip were 97.7%, 97.7%, 97.7%, 83.7%, and 83.7%. A premature stop codon mutation of position 1474(C→T) was detected, resulting in a truncated InlA with 491 aa. High susceptibility to penicillin (100%), ampicillin (100%), gentamicin (100%), erythromycin (100%), daptomycin (100%), meropenem (100%), trimethoprim-sulfamethoxazole (100%), vancomycin (97.7%), tetacycline (97.7%), chloramphenicol (97.7%), and ciprofloxacin (90.7%) was demonstrated. lin and fosX were present in 93.0% of the isolates, respectively. tetM and ermB were also detected. This comprehensive study enriched the understanding of listeriosis cases and diversity of clinical isolates, meanwhile, indicated the spread characteristics of CC1, CC8, and CC87 in China based on phylogeny analysis, providing fundamental data for developing targeting food safety interventions to prevent listeriosis.
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Affiliation(s)
- Yunyi Zhang
- Department of Microbiology, Zhejiang Provincial Center for Disease Control and Prevention, Hangzhou, China
| | - Junyan Zhang
- Department of Microbiology, Zhejiang Provincial Center for Disease Control and Prevention, Hangzhou, China
| | - Jiancai Chen
- Department of Microbiology, Zhejiang Provincial Center for Disease Control and Prevention, Hangzhou, China
| | - Xiaojuan Qi
- Department of Microbiology, Zhejiang Provincial Center for Disease Control and Prevention, Hangzhou, China
| | - Zhen Zhang
- Department of Microbiology, Zhejiang Provincial Center for Disease Control and Prevention, Hangzhou, China
| | - Honghu Chen
- Department of Microbiology, Zhejiang Provincial Center for Disease Control and Prevention, Hangzhou, China
| | - Beibei Wu
- Department of Microbiology, Zhejiang Provincial Center for Disease Control and Prevention, Hangzhou, China
| | - Li Zhan
- Department of Microbiology, Zhejiang Provincial Center for Disease Control and Prevention, Hangzhou, China
| | - Jie Chen
- Zhejiang Provincial Center for Animal Disease Prevention and Control, Hangzhou, China
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Radhakrishnan P, Theriot JA. Listeria monocytogenes cell-to-cell spread bypasses nutrient limitation for replicating intracellular bacteria. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2025:2025.01.31.635960. [PMID: 39975404 PMCID: PMC11838505 DOI: 10.1101/2025.01.31.635960] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 02/21/2025]
Abstract
Listeria monocytogenes is an intracellular bacterial pathogen that obtains nutrients from the mammalian host cell to fuel its replication in cytosol. Sparse infection of epithelial monolayers by L. monocytogenes results in the formation of distinct infectious foci, where each focus originates from the initial infection of a single host cell followed by multiple rounds of active bacterial cell-to-cell spread into neighboring host cells in the monolayer. We used time-lapse microscopy to measure changes in bacterial growth rate in individual foci over time and found that intracellular bacteria initially replicate exponentially, but then bacterial growth rate slows later in infection, particularly in the center of the infectious focus. We found that the intracellular replication rate of L. monocytogenes is measurably decreased by limiting host cell glucose availability, by decreasing the rate of intracellular bacterial oligopeptide import, and, most interestingly, by alterations in host cell junctional proteins that limit bacterial spread into neighboring cells without directly affecting bacterial growth or metabolism. By measuring the carrying capacity of individual host cells, we found that the nutritional density of cytoplasm is comparable to rich medium. Taken together, our results indicate that the rate of intracellular L. monocytogenes replication is governed by a balance of the rate of nutrient depletion by the bacteria, the rate of nutrient replenishment by the metabolically active host cells, and the rate of bacterial cell-to-cell spread which enables the bacteria to seek out "greener pastures" before nutrient availability in a single host cell becomes limiting.
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Affiliation(s)
- Prathima Radhakrishnan
- Department of Biology and Howard Hughes Medical Institute, University of Washington, Seattle, WA 98195-1800
| | - Julie A Theriot
- Department of Biology and Howard Hughes Medical Institute, University of Washington, Seattle, WA 98195-1800
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3
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Heissler SM, Chinthalapudi K. Structural and functional mechanisms of actin isoforms. FEBS J 2025; 292:468-482. [PMID: 38779987 PMCID: PMC11796330 DOI: 10.1111/febs.17153] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/06/2024] [Revised: 04/01/2024] [Accepted: 04/23/2024] [Indexed: 05/25/2024]
Abstract
Actin is a highly conserved and fundamental protein in eukaryotes and participates in a broad spectrum of cellular functions. Cells maintain a conserved ratio of actin isoforms, with muscle and non-muscle actins representing the main actin isoforms in muscle and non-muscle cells, respectively. Actin isoforms have specific and redundant functional roles and display different biochemistries, cellular localization, and interactions with myosins and actin-binding proteins. Understanding the specific roles of actin isoforms from the structural and functional perspective is crucial for elucidating the intricacies of cytoskeletal dynamics and regulation and their implications in health and disease. Here, we review how the structure contributes to the functional mechanisms of actin isoforms with a special emphasis on the questions of how post-translational modifications and disease-linked mutations affect actin isoforms biochemistry, function, and interaction with actin-binding proteins and myosin motors.
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Affiliation(s)
- Sarah M. Heissler
- Department of Physiology and Cell Biology, Dorothy M. Davis Heart & Lung Research InstituteThe Ohio State UniversityColumbusOHUSA
| | - Krishna Chinthalapudi
- Department of Physiology and Cell Biology, Dorothy M. Davis Heart & Lung Research InstituteThe Ohio State UniversityColumbusOHUSA
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4
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Fayoud H, Belousov MV, Antonets KS, Nizhnikov AA. Pathogenesis-Associated Bacterial Amyloids: The Network of Interactions. BIOCHEMISTRY. BIOKHIMIIA 2024; 89:2107-2132. [PMID: 39865026 DOI: 10.1134/s0006297924120022] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 09/13/2024] [Revised: 10/20/2024] [Accepted: 10/21/2024] [Indexed: 01/28/2025]
Abstract
Amyloids are protein fibrils with a characteristic cross-β structure that is responsible for the unusual resistance of amyloids to various physical and chemical factors, as well as numerous pathogenic and functional consequences of amyloidogenesis. The greatest diversity of functional amyloids was identified in bacteria. The majority of bacterial amyloids are involved in virulence and pathogenesis either via facilitating formation of biofilms and adaptation of bacteria to colonization of a host organism or through direct regulation of toxicity. Recent studies have shown that, beside their commonly known activity, amyloids may be involved in the spatial regulation of proteome by modulating aggregation of other amyloidogenic proteins with multiple functional or pathological effects. Although the studies on the role of microbiome-produced amyloids in the development of amyloidoses in humans and animals have only been started, it is clear that humans as holobionts contain amyloids encoded not only by the host genome, but also by microorganisms that constitute the microbiome. Amyloids acquired from external sources (e.g., food) can interact with holobiont amyloids and modulate the effects of bacterial and host amyloids, thus adding another level of complexity to the holobiont-associated amyloid network. In this review, we described bacterial amyloids directly or indirectly involved in disease pathogenesis in humans and discussed the significance of bacterial amyloids in the three-component network of holobiont-associated amyloids.
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Affiliation(s)
- Haidar Fayoud
- Faculty of Biology, St. Petersburg State University, St. Petersburg, 199034, Russia
- All-Russia Research Institute for Agricultural Microbiology, St. Petersburg, 196608, Russia
| | - Mikhail V Belousov
- Faculty of Biology, St. Petersburg State University, St. Petersburg, 199034, Russia
- All-Russia Research Institute for Agricultural Microbiology, St. Petersburg, 196608, Russia
| | - Kirill S Antonets
- Faculty of Biology, St. Petersburg State University, St. Petersburg, 199034, Russia
- All-Russia Research Institute for Agricultural Microbiology, St. Petersburg, 196608, Russia
| | - Anton A Nizhnikov
- Faculty of Biology, St. Petersburg State University, St. Petersburg, 199034, Russia. ARRAY(0x5ae2b7af6df8)
- All-Russia Research Institute for Agricultural Microbiology, St. Petersburg, 196608, Russia
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Brown SRB, Sun L, Gensler CA, D'Amico DJ. The Impact of Subinhibitory Concentrations of Ɛ-polylysine, Hydrogen Peroxide, and Lauric Arginate on Listeria monocytogenes Virulence. J Food Prot 2024; 87:100385. [PMID: 39427815 DOI: 10.1016/j.jfp.2024.100385] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/12/2024] [Revised: 10/10/2024] [Accepted: 10/15/2024] [Indexed: 10/22/2024]
Abstract
Recent studies on the use of plant-derived and other bioactive compounds and antimicrobials in food have challenged the idea that exposure to antimicrobials at sublethal or subinhibitory concentrations (SICs) increases the virulence potential of bacterial pathogens including Listeria monocytogenes. The objective of this study was to determine the effect of exposure to SICs of Ɛ-polylysine (EPL), hydrogen peroxide (HP), and lauric arginate (LAE) on L. monocytogenes virulence. For all assays, L. monocytogenes strains Scott A and 2014L-6025 were grown to mid-log phase in the presence of SICs of EPL, HP, or LAE. Motility was determined by spot inoculating cultures on soft brain heart infusion agar (0.3% agar). Cultures grown in SICs of antimicrobials were also inoculated onto Caco-2 cells (10:1 MOI) to determine the effects on subsequent adhesion and invasion. Last, the relative expression of key virulence genes (prfA, plcB, hlyA, actA, inlA, inlB, sigB, and virR) following growth in SICs was determined by RT-qPCR. Results indicate that L. monocytogenes growth in the presence of SICs of EPL, HP, or LAE did not affect the motility, adhesion, or invasion capacity of either strain. Changes in gene expression were observed for both L. monocytogenes strains. More specifically, SICs of EPL and LAE reduced hlyA expression in Scott A, whereas SICs of EPL and HP increased the expression of virR. The upregulation of sigB and actA in the presence of EPL and LAE, respectively, was observed in strain 2014L-6025. These findings indicate that exposure to SICs of these antimicrobials has varying effects on L. monocytogenes that differ by strain. Although no phenotypic effects were observed in terms of motility, adhesion, and invasion, the observed changes in virulence gene expression warrant further investigation.
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Affiliation(s)
- Stephanie R B Brown
- Dept. of Animal Science, University of Connecticut, Ag. Biotechnology Laboratory, 1390 Storrs Road, U-4163, Storrs CT, 06269-4163, USA
| | - Lang Sun
- Dept. of Animal Science, University of Connecticut, Ag. Biotechnology Laboratory, 1390 Storrs Road, U-4163, Storrs CT, 06269-4163, USA
| | - Catherine A Gensler
- Dept. of Animal Science, University of Connecticut, Ag. Biotechnology Laboratory, 1390 Storrs Road, U-4163, Storrs CT, 06269-4163, USA
| | - Dennis J D'Amico
- Dept. of Animal Science, University of Connecticut, Ag. Biotechnology Laboratory, 1390 Storrs Road, U-4163, Storrs CT, 06269-4163, USA.
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Sousa M, Magalhães R, Ferreira V, Teixeira P. Current methodologies available to evaluate the virulence potential among Listeria monocytogenes clonal complexes. Front Microbiol 2024; 15:1425437. [PMID: 39493856 PMCID: PMC11528214 DOI: 10.3389/fmicb.2024.1425437] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/29/2024] [Accepted: 09/26/2024] [Indexed: 11/05/2024] Open
Abstract
Listeria monocytogenes is a foodborne pathogen that causes listeriosis in humans, the severity of which depends on multiple factors, including intrinsic characteristics of the affected individuals and the pathogen itself. Additionally, emerging evidence suggests that epigenetic modifications may also modulate host susceptibility to infection. Therefore, different clinical outcomes can be expected, ranging from self-limiting gastroenteritis to severe central nervous system and maternal-neonatal infections, and bacteremia. Furthermore, L. monocytogenes is a genetically and phenotypically diverse species, resulting in a large variation in virulence potential between strains. Multilocus sequence typing (MLST) has been widely used to categorize the clonal structure of bacterial species and to define clonal complexes (CCs) of genetically related isolates. The combination of MLST and epidemiological data allows to distinguish hypervirulent CCs, which are notably more prevalent in clinical cases and typically associated with severe forms of the disease. Conversely, other CCs, termed hypovirulent, are predominantly isolated from food and food processing environments and are associated with the occurrence of listeriosis in immunosuppressed individuals. Reports of genetic traits associated with this diversity have been described. The Food and Agriculture Organization (FAO) is encouraging the search for virulence biomarkers to rapidly identify the main strains of concern to reduce food waste and economical losses. The aim of this review is to comprehensively collect, describe and discuss the methodologies used to discriminate the virulence potential of L. monocytogenes CCs. From the exploration of in vitro and in vivo models to the study of expression of virulence genes, each approach is critically explored to better understand its applicability and efficiency in distinguishing the virulence potential of the pathogen.
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Affiliation(s)
| | | | | | - Paula Teixeira
- Universidade Católica Portuguesa, CBQF - Centro de Biotecnologia e Química Fina – Laboratório Associado, Escola Superior de Biotecnologia, Rua Diogo Botelho 1327, Porto, Portugal
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7
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Velle KB. mSphere of Influence: The power of observational research. mSphere 2024; 9:e0017624. [PMID: 38953618 PMCID: PMC11288056 DOI: 10.1128/msphere.00176-24] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 07/04/2024] Open
Abstract
Katrina Velle is a cell biologist who uses microscopy to study amoebae. In this mSphere of Influence article, she reflects on how a classic paper on Listeria by Tilney and Portnoy made an impact on her by highlighting how much we can learn from simply looking at cells.
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Affiliation(s)
- Katrina B. Velle
- Department of Biology, University of Massachusetts Dartmouth, North Dartmouth, Massachusetts, USA
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8
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Coscia SM, Moore AS, Thompson CP, Tirrito CF, Ostap EM, Holzbaur ELF. An interphase actin wave promotes mitochondrial content mixing and organelle homeostasis. Nat Commun 2024; 15:3793. [PMID: 38714822 PMCID: PMC11076292 DOI: 10.1038/s41467-024-48189-1] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/05/2023] [Accepted: 04/22/2024] [Indexed: 05/10/2024] Open
Abstract
Across the cell cycle, mitochondrial dynamics are regulated by a cycling wave of actin polymerization/depolymerization. In metaphase, this wave induces actin comet tails on mitochondria that propel these organelles to drive spatial mixing, resulting in their equitable inheritance by daughter cells. In contrast, during interphase the cycling actin wave promotes localized mitochondrial fission. Here, we identify the F-actin nucleator/elongator FMNL1 as a positive regulator of the wave. FMNL1-depleted cells exhibit decreased mitochondrial polarization, decreased mitochondrial oxygen consumption, and increased production of reactive oxygen species. Accompanying these changes is a loss of hetero-fusion of wave-fragmented mitochondria. Thus, we propose that the interphase actin wave maintains mitochondrial homeostasis by promoting mitochondrial content mixing. Finally, we investigate the mechanistic basis for the observation that the wave drives mitochondrial motility in metaphase but mitochondrial fission in interphase. Our data indicate that when the force of actin polymerization is resisted by mitochondrial tethering to microtubules, as in interphase, fission results.
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Affiliation(s)
- Stephen M Coscia
- Department of Physiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA, USA
- Pennsylvania Muscle Institute, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA, USA
- Cell and Molecular Biology Graduate Group, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA, USA
| | - Andrew S Moore
- Howard Hughes Medical Institute, Janelia Research Campus, Ashburn, VA, USA
| | - Cameron P Thompson
- Department of Physiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA, USA
- Pennsylvania Muscle Institute, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA, USA
- Biochemistry and Molecular Biophysics Graduate Group, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA, USA
| | - Christian F Tirrito
- Raymond G. Perelman Center for Cellular and Molecular Therapeutics, The Children's Hospital of Philadelphia, Philadelphia, PA, USA
- Department of Pathology and Laboratory Medicine, University of Pennsylvania, Philadelphia, PA, USA
| | - E Michael Ostap
- Department of Physiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA, USA
- Pennsylvania Muscle Institute, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA, USA
| | - Erika L F Holzbaur
- Department of Physiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA, USA.
- Pennsylvania Muscle Institute, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA, USA.
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9
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Chevée V, Hullahalli K, Dailey KG, Güereca L, Zhang C, Waldor MK, Portnoy DA. Temporal and spatial dynamics of Listeria monocytogenes central nervous system infection in mice. Proc Natl Acad Sci U S A 2024; 121:e2320311121. [PMID: 38635627 PMCID: PMC11046682 DOI: 10.1073/pnas.2320311121] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/18/2023] [Accepted: 02/22/2024] [Indexed: 04/20/2024] Open
Abstract
Listeria monocytogenes is a bacterial pathogen that can cause life-threatening central nervous system (CNS) infections. While mechanisms by which L. monocytogenes and other pathogens traffic to the brain have been studied, a quantitative understanding of the underlying dynamics of colonization and replication within the brain is still lacking. In this study, we used barcoded L. monocytogenes to quantify the bottlenecks and dissemination patterns that lead to cerebral infection. Following intravenous (IV) inoculation, multiple independent invasion events seeded all parts of the CNS from the blood, however, only one clone usually became dominant in the brain. Sequential IV inoculations and intracranial inoculations suggested that clones that had a temporal advantage (i.e., seeded the CNS first), rather than a spatial advantage (i.e., invaded a particular brain region), were the main drivers of clonal dominance. In a foodborne model of cerebral infection with immunocompromised mice, rare invasion events instead led to a highly infected yet monoclonal CNS. This restrictive bottleneck likely arose from pathogen transit into the blood, rather than directly from the blood to the brain. Collectively, our findings provide a detailed quantitative understanding of the L. monocytogenes population dynamics that lead to CNS infection and a framework for studying the dynamics of other cerebral infections.
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Affiliation(s)
- Victoria Chevée
- Department of Molecular and Cell Biology, University of California, Berkeley, CA94720
| | - Karthik Hullahalli
- Division of Infectious Diseases, Brigham and Women’s Hospital, Boston, MA02115
- Department of Microbiology, Harvard Medical School, Boston, MA02115
- HHMI, Bethesda, MD20815
| | - Katherine G. Dailey
- Division of Infectious Diseases, Brigham and Women’s Hospital, Boston, MA02115
- Department of Microbiology, Harvard Medical School, Boston, MA02115
- HHMI, Bethesda, MD20815
| | - Leslie Güereca
- Department of Molecular and Cell Biology, University of California, Berkeley, CA94720
| | - Chenyu Zhang
- Department of Molecular and Cell Biology, University of California, Berkeley, CA94720
| | - Matthew K. Waldor
- Division of Infectious Diseases, Brigham and Women’s Hospital, Boston, MA02115
- Department of Microbiology, Harvard Medical School, Boston, MA02115
- HHMI, Bethesda, MD20815
| | - Daniel A. Portnoy
- Department of Molecular and Cell Biology, University of California, Berkeley, CA94720
- Department of Plant and Microbial Biology, University of California, Berkeley, CA94720
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10
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Smith HB, Lee K, Freeman MJ, Stevenson DM, Amador-Noguez D, Sauer JD. Listeria monocytogenes requires DHNA-dependent intracellular redox homeostasis facilitated by Ndh2 for survival and virulence. Infect Immun 2023; 91:e0002223. [PMID: 37754681 PMCID: PMC10580952 DOI: 10.1128/iai.00022-23] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/12/2023] [Accepted: 08/09/2023] [Indexed: 09/28/2023] Open
Abstract
Listeria monocytogenes is a remarkably well-adapted facultative intracellular pathogen that can thrive in a wide range of ecological niches. L. monocytogenes maximizes its ability to generate energy from diverse carbon sources using a respiro-fermentative metabolism that can function under both aerobic and anaerobic conditions. Cellular respiration maintains redox homeostasis by regenerating NAD+ while also generating a proton motive force. The end products of the menaquinone (MK) biosynthesis pathway are essential to drive both aerobic and anaerobic cellular respirations. We previously demonstrated that intermediates in the MK biosynthesis pathway, notably 1,4-dihydroxy-2-naphthoate (DHNA), are required for the survival and virulence of L. monocytogenes independent of their role in respiration. Furthermore, we found that restoration of NAD+/NADH ratio through expression of water-forming NADH oxidase could rescue phenotypes associated with DHNA deficiency. Here, we extend these findings to demonstrate that endogenous production or direct supplementation of DHNA restored both the cellular redox homeostasis and metabolic output of fermentation in L. monocytogenes. Furthermore, exogenous supplementation of DHNA rescues the in vitro growth and ex vivo virulence of L. monocytogenes DHNA-deficient mutants. Finally, we demonstrate that exogenous DHNA restores redox balance in L. monocytogenes specifically through the recently annotated NADH dehydrogenase Ndh2, independent of its role in the extracellular electron transport pathway. These data suggest that the production of DHNA may represent an additional layer of metabolic adaptability by L. monocytogenes to drive energy metabolism in the absence of respiration-favorable conditions.
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Affiliation(s)
- Hans B. Smith
- Department of Medical Microbiology and Immunology, University of Wisconsin-Madison, Madison, Wisconsin, USA
| | - Kijeong Lee
- Department of Medical Microbiology and Immunology, University of Wisconsin-Madison, Madison, Wisconsin, USA
| | - Matthew J. Freeman
- Department of Medical Microbiology and Immunology, University of Wisconsin-Madison, Madison, Wisconsin, USA
| | - David M. Stevenson
- Department of Bacteriology, University of Wisconsin-Madison, Madison, Wisconsin, USA
| | - Daniel Amador-Noguez
- Department of Bacteriology, University of Wisconsin-Madison, Madison, Wisconsin, USA
| | - John-Demian Sauer
- Department of Medical Microbiology and Immunology, University of Wisconsin-Madison, Madison, Wisconsin, USA
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11
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Ding YD, Shu LZ, He RS, Chen KY, Deng YJ, Zhou ZB, Xiong Y, Deng H. Listeria monocytogenes: a promising vector for tumor immunotherapy. Front Immunol 2023; 14:1278011. [PMID: 37868979 PMCID: PMC10587691 DOI: 10.3389/fimmu.2023.1278011] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/15/2023] [Accepted: 09/25/2023] [Indexed: 10/24/2023] Open
Abstract
Cancer receives enduring international attention due to its extremely high morbidity and mortality. Immunotherapy, which is generally expected to overcome the limits of traditional treatments, serves as a promising direction for patients with recurrent or metastatic malignancies. Bacteria-based vectors such as Listeria monocytogenes take advantage of their unique characteristics, including preferential infection of host antigen presenting cells, intracellular growth within immune cells, and intercellular dissemination, to further improve the efficacy and minimize off-target effects of tailed immune treatments. Listeria monocytogenes can reshape the tumor microenvironment to bolster the anti-tumor effects both through the enhancement of T cells activity and a decrease in the frequency and population of immunosuppressive cells. Modified Listeria monocytogenes has been employed as a tool to elicit immune responses against different tumor cells. Currently, Listeria monocytogenes vaccine alone is insufficient to treat all patients effectively, which can be addressed if combined with other treatments, such as immune checkpoint inhibitors, reactivated adoptive cell therapy, and radiotherapy. This review summarizes the recent advances in the molecular mechanisms underlying the involvement of Listeria monocytogenes vaccine in anti-tumor immunity, and discusses the most concerned issues for future research.
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Affiliation(s)
- Yi-Dan Ding
- Medical College, Nanchang University, Nanchang, China
| | - Lin-Zhen Shu
- Medical College, Nanchang University, Nanchang, China
| | - Rui-Shan He
- Medical College, Nanchang University, Nanchang, China
| | - Kai-Yun Chen
- Office of Clinical Trials Administration, The Fourth Affiliated Hospital of Nanchang University, Nanchang, China
| | - Yan-Juan Deng
- Department of Pathology, The Fourth Affiliated Hospital of Nanchang University, Nanchang, China
- Tumor Immunology Institute, Nanchang University, Nanchang, China
| | - Zhi-Bin Zhou
- Department of Pathology, The Fourth Affiliated Hospital of Nanchang University, Nanchang, China
- Tumor Immunology Institute, Nanchang University, Nanchang, China
| | - Ying Xiong
- Department of General Medicine, The Second Affiliated Hospital of Nanchang University, Nanchang, China
| | - Huan Deng
- Department of Pathology, The Fourth Affiliated Hospital of Nanchang University, Nanchang, China
- Tumor Immunology Institute, Nanchang University, Nanchang, China
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12
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Matsuda Y, Yamauchi H, Hara H. Activation of inflammasomes and mechanisms for intracellular recognition of Listeria monocytogenes. Microbiol Immunol 2023; 67:429-437. [PMID: 37461376 DOI: 10.1111/1348-0421.13091] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/28/2023] [Accepted: 07/03/2023] [Indexed: 10/06/2023]
Abstract
The high mortality rate associated with Listeria monocytogenes can be attributed to its ability to invade the body systemically and to activate inflammasomes. Both of these processes are facilitated by expressing a major virulence factor known as listeriolysin O, a 56 kDa pore-forming protein encoded by the hly gene. Listeriolysin O plays a crucial role in the pathogenesis of the bacterium by facilitating the escape of the pathogen from the phagosome into the cytosol. This process is essential for the successful establishment of infection. In addition, listeriolysin O is known as an immunomodulator that activates host signal transduction. In addition to listeriolysin O, Listeria expresses a variety of bacterial ligands, such as lipoteichoic acid, nucleotide, and flagellin, that are recognized by host intracellular pattern-recognition receptors including Nod-like receptors, AIM2-like receptors, and RIG-I-like receptors. This review introduces intracellular recognition of Listeria monocytogenes since recent studies have revealed that the activation of inflammasome exacerbates Gram-positive bacteria infection.
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Affiliation(s)
- Yasuyuki Matsuda
- Department of Infectious Diseases, Division of Microbiology and Immunochemistry, Asahikawa Medical University, Asahikawa, Japan
| | - Hajime Yamauchi
- Department of Infectious Diseases, Division of Microbiology and Immunochemistry, Asahikawa Medical University, Asahikawa, Japan
| | - Hideki Hara
- Department of Infectious Diseases, Division of Microbiology and Immunochemistry, Asahikawa Medical University, Asahikawa, Japan
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13
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Cossart P. Raising a Bacterium to the Rank of a Model System: The Listeria Paradigm. Annu Rev Microbiol 2023; 77:1-22. [PMID: 37713460 DOI: 10.1146/annurev-micro-110422-112841] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 09/17/2023]
Abstract
My scientific career has resulted from key decisions and reorientations, sometimes taken rapidly but not always, guided by discussions or collaborations with amazing individuals from whom I learnt a lot scientifically and humanly. I had never anticipated that I would accomplish so much in what appeared as terra incognita when I started to interrogate the mechanisms underlying the virulence of the bacterium Listeria monocytogenes. All this has been possible thanks to a number of talented team members who ultimately became friends.
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Affiliation(s)
- Pascale Cossart
- Department of Cell Biology and Infection, Institut Pasteur, Paris, France;
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14
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Stögerer T, Silva-Barrios S, Carmona-Pérez L, Swaminathan S, Mai LT, Leroux LP, Jaramillo M, Descoteaux A, Stäger S. Leishmania donovani Exploits Tunneling Nanotubes for Dissemination and Propagation of B Cell Activation. Microbiol Spectr 2023; 11:e0509622. [PMID: 37404188 PMCID: PMC10434010 DOI: 10.1128/spectrum.05096-22] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/12/2022] [Accepted: 06/08/2023] [Indexed: 07/06/2023] Open
Abstract
Polyclonal B cell activation and the resulting hypergammaglobulinemia are a detrimental consequence of visceral leishmaniasis (VL); however, the mechanisms underlying this excessive production of nonprotective antibodies are still poorly understood. Here, we show that a causative agent of VL, Leishmania donovani, induces CD21-dependent formation of tunneling nanotubule (TNT)-like protrusions in B cells. These intercellular connections are used by the parasite to disseminate among cells and propagate B cell activation, and close contact both among the cells and between B cells and parasites is required to achieve this activation. Direct contact between cells and parasites is also observed in vivo, as L. donovani can be detected in the splenic B cell area as early as 14 days postinfection. Interestingly, Leishmania parasites can also glide from macrophages to B cells via TNT-like protrusions. Taken together, our results suggest that, during in vivo infection, B cells may acquire L. donovani from macrophages via TNT-like protrusions, and these connections are subsequently exploited by the parasite to disseminate among B cells, thus propagating B cell activation and ultimately leading to polyclonal B cell activation. IMPORTANCE Leishmania donovani is a causative agent of visceral leishmaniasis, a potentially lethal disease characterized by strong B cell activation and the subsequent excessive production of nonprotective antibodies, which are known to worsen the disease. How Leishmania activates B cells is still unknown, particularly because this parasite mostly resides inside macrophages and would not have access to B cells during infection. In this study, we describe for the first time how the protozoan parasite Leishmania donovani induces and exploits the formation of protrusions that connect B lymphocytes with each other or with macrophages and glides on these structures from one cell to another. In this way, B cells can acquire Leishmania from macrophages and become activated upon contact with the parasites. This activation will then lead to antibody production. These findings provide an explanation for how the parasite may propagate B cell activation during infection.
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Affiliation(s)
- Tanja Stögerer
- Institut National de la Recherche Scientifique (INRS) – Centre Armand-Frappier Santé Biotechnologie and Infectiopôle INRS, Laval, Quebec, Canada
| | - Sasha Silva-Barrios
- Institut National de la Recherche Scientifique (INRS) – Centre Armand-Frappier Santé Biotechnologie and Infectiopôle INRS, Laval, Quebec, Canada
| | - Liseth Carmona-Pérez
- Institut National de la Recherche Scientifique (INRS) – Centre Armand-Frappier Santé Biotechnologie and Infectiopôle INRS, Laval, Quebec, Canada
| | - Sharada Swaminathan
- Institut National de la Recherche Scientifique (INRS) – Centre Armand-Frappier Santé Biotechnologie and Infectiopôle INRS, Laval, Quebec, Canada
| | - Linh Thuy Mai
- Institut National de la Recherche Scientifique (INRS) – Centre Armand-Frappier Santé Biotechnologie and Infectiopôle INRS, Laval, Quebec, Canada
| | - Louis-Philippe Leroux
- Institut National de la Recherche Scientifique (INRS) – Centre Armand-Frappier Santé Biotechnologie and Infectiopôle INRS, Laval, Quebec, Canada
| | - Maritza Jaramillo
- Institut National de la Recherche Scientifique (INRS) – Centre Armand-Frappier Santé Biotechnologie and Infectiopôle INRS, Laval, Quebec, Canada
| | - Albert Descoteaux
- Institut National de la Recherche Scientifique (INRS) – Centre Armand-Frappier Santé Biotechnologie and Infectiopôle INRS, Laval, Quebec, Canada
| | - Simona Stäger
- Institut National de la Recherche Scientifique (INRS) – Centre Armand-Frappier Santé Biotechnologie and Infectiopôle INRS, Laval, Quebec, Canada
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15
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Oliveira AH, Tiensuu T, Guerreiro D, Tükenmez H, Dessaux C, García-del Portillo F, O’Byrne C, Johansson J. The Virulence and Infectivity of Listeria monocytogenes Are Not Substantially Altered by Elevated SigB Activity. Infect Immun 2023; 91:e0057122. [PMID: 37125941 PMCID: PMC10269059 DOI: 10.1128/iai.00571-22] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/19/2022] [Accepted: 03/26/2023] [Indexed: 05/02/2023] Open
Abstract
Listeria monocytogenes is a bacterial pathogen capable of causing severe infections but also thriving outside the host. To respond to different stress conditions, L. monocytogenes mainly utilizes the general stress response regulon, which largely is controlled by the alternative sigma factor Sigma B (SigB). In addition, SigB is important for virulence gene expression and infectivity. Upon encountering stress, a large multicomponent protein complex known as the stressosome becomes activated, ultimately leading to SigB activation. RsbX is a protein needed to reset a "stressed" stressosome and prevent unnecessary SigB activation in nonstressed conditions. Consequently, absence of RsbX leads to constitutive activation of SigB even without prevailing stress stimulus. To further examine the involvement of SigB in the virulence of this pathogen, we investigated whether a strain with constitutively active SigB would be affected in virulence factor expression and/or infectivity in cultured cells and in a chicken embryo infection model. Our results suggest that increased SigB activity does not substantially alter virulence gene expression compared with the wild-type (WT) strain at transcript and protein levels. Bacteria lacking RsbX were taken up by phagocytic and nonphagocytic cells at a similar frequency to WT bacteria, both in stressed and nonstressed conditions. Finally, the absence of RsbX only marginally affected the ability of bacteria to infect chicken embryos. Our results suggest only a minor role of RsbX in controlling virulence factor expression and infectivity under these conditions.
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Affiliation(s)
- Ana H. Oliveira
- Laboratory for Molecular Infection Medicine Sweden, Umeå University, Umeå, Sweden
- Department of Molecular Biology, Umeå University, Umeå, Sweden
- Umeå Centre of Microbial Research, Umeå University, Umeå, Sweden
| | - Teresa Tiensuu
- Laboratory for Molecular Infection Medicine Sweden, Umeå University, Umeå, Sweden
- Department of Molecular Biology, Umeå University, Umeå, Sweden
- Umeå Centre of Microbial Research, Umeå University, Umeå, Sweden
| | - Duarte Guerreiro
- Laboratory for Molecular Infection Medicine Sweden, Umeå University, Umeå, Sweden
- Department of Molecular Biology, Umeå University, Umeå, Sweden
- Umeå Centre of Microbial Research, Umeå University, Umeå, Sweden
- Bacterial Stress Response Group, Microbiology, School of Biological and Chemical Sciences, University of Galway, Galway, Ireland
| | - Hasan Tükenmez
- Laboratory for Molecular Infection Medicine Sweden, Umeå University, Umeå, Sweden
- Department of Molecular Biology, Umeå University, Umeå, Sweden
- Umeå Centre of Microbial Research, Umeå University, Umeå, Sweden
- Department of Chemistry, Umeå University, Umeå, Sweden
| | - Charlotte Dessaux
- Laboratory of Intracellular Bacterial Pathogens, National Center of Biotechnology, (CNB)-CSIC, Madrid, Spain
| | | | - Conor O’Byrne
- Bacterial Stress Response Group, Microbiology, School of Biological and Chemical Sciences, University of Galway, Galway, Ireland
| | - Jörgen Johansson
- Laboratory for Molecular Infection Medicine Sweden, Umeå University, Umeå, Sweden
- Department of Molecular Biology, Umeå University, Umeå, Sweden
- Umeå Centre of Microbial Research, Umeå University, Umeå, Sweden
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16
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Wiktorczyk-Kapischke N, Skowron K, Wałecka-Zacharska E. Genomic and pathogenicity islands of Listeria monocytogenes-overview of selected aspects. Front Mol Biosci 2023; 10:1161486. [PMID: 37388250 PMCID: PMC10300472 DOI: 10.3389/fmolb.2023.1161486] [Citation(s) in RCA: 15] [Impact Index Per Article: 7.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/08/2023] [Accepted: 06/01/2023] [Indexed: 07/01/2023] Open
Abstract
Listeria monocytogenes causes listeriosis, a disease characterized by a high mortality rate (up to 30%). Since the pathogen is highly tolerant to changing conditions (high and low temperature, wide pH range, low availability of nutrients), it is widespread in the environment, e.g., water, soil, or food. L. monocytogenes possess a number of genes that determine its high virulence potential, i.e., genes involved in the intracellular cycle (e.g., prfA, hly, plcA, plcB, inlA, inlB), response to stress conditions (e.g., sigB, gadA, caspD, clpB, lmo1138), biofilm formation (e.g., agr, luxS), or resistance to disinfectants (e.g., emrELm, bcrABC, mdrL). Some genes are organized into genomic and pathogenicity islands. The islands LIPI-1 and LIPI-3 contain genes related to the infectious life cycle and survival in the food processing environment, while LGI-1 and LGI-2 potentially ensure survival and durability in the production environment. Researchers constantly have been searching for new genes determining the virulence of L. monocytogenes. Understanding the virulence potential of L. monocytogenes is an important element of public health protection, as highly pathogenic strains may be associated with outbreaks and the severity of listeriosis. This review summarizes the selected aspects of L. monocytogenes genomic and pathogenicity islands, and the importance of whole genome sequencing for epidemiological purposes.
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Affiliation(s)
- Natalia Wiktorczyk-Kapischke
- Department of Microbiology, Ludwik Rydygier Collegium Medicum in Bydgoszcz, Nicolaus Copernicus University, Toruń, Poland
| | - Krzysztof Skowron
- Department of Microbiology, Ludwik Rydygier Collegium Medicum in Bydgoszcz, Nicolaus Copernicus University, Toruń, Poland
| | - Ewa Wałecka-Zacharska
- Department of Food Hygiene and Consumer Health, Wrocław University of Environmental and Life Sciences, Wrocław, Poland
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17
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Tucker JS, Cho J, Albrecht TM, Ferrell JL, D’Orazio SEF. Egress of Listeria monocytogenes from Mesenteric Lymph Nodes Depends on Intracellular Replication and Cell-to-Cell Spread. Infect Immun 2023; 91:e0006423. [PMID: 36916918 PMCID: PMC10112146 DOI: 10.1128/iai.00064-23] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/15/2023] [Accepted: 02/20/2023] [Indexed: 03/15/2023] Open
Abstract
The mesenteric lymph nodes (MLN) function as a barrier to systemic spread for both commensal and pathogenic bacteria in the gut. Listeria monocytogenes, a facultative intracellular foodborne pathogen, readily overcomes this barrier and spreads into the bloodstream, causing life-threatening systemic infections. We show here that intracellular replication protected L. monocytogenes from clearance by monocytes and neutrophils and promoted colonization of the small intestine-draining MLN (sMLN) but was not required for dissemination to the colon-draining MLN (cMLN). Intestinal tissue had enough free lipoate to support LplA2-dependent extracellular growth of L. monocytogenes, but exogenous lipoate in the MLN was severely limited, and so the bacteria could replicate only inside cells, where they used LplA1 to scavenge lipoate from host peptides. When foodborne infection was manipulated to allow ΔlplA1 L. monocytogenes to colonize the MLN to the same extent as wild-type bacteria, the mutant was still never recovered in the spleen or liver of any animal. We found that intracellular replication in the MLN promoted actin-based motility and cell-to-cell spread of L. monocytogenes and that rapid efficient exit from the MLN was actA dependent. We conclude that intracellular replication of L. monocytogenes in intestinal tissues is not essential and serves primarily to amplify bacterial burdens above a critical threshold needed to efficiently colonize the cMLN. In contrast, intracellular replication in the MLN is absolutely required for further systemic spread and serves primarily to promote ActA-mediated cell-to-cell spread.
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Affiliation(s)
- Jamila S. Tucker
- Department of Microbiology, Immunology, and Molecular Genetics, University of Kentucky, Lexington, Kentucky, USA
| | - Jooyoung Cho
- Department of Microbiology, Immunology, and Molecular Genetics, University of Kentucky, Lexington, Kentucky, USA
| | - Taylor M. Albrecht
- Department of Microbiology, Immunology, and Molecular Genetics, University of Kentucky, Lexington, Kentucky, USA
| | - Jessica L. Ferrell
- Department of Microbiology, Immunology, and Molecular Genetics, University of Kentucky, Lexington, Kentucky, USA
| | - Sarah E. F. D’Orazio
- Department of Microbiology, Immunology, and Molecular Genetics, University of Kentucky, Lexington, Kentucky, USA
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18
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Mason ER, Soni DM, Chu S. Microglial Phagocytosis/Cell Health High-Content Assay. Curr Protoc 2023; 3:e724. [PMID: 36971657 PMCID: PMC10433541 DOI: 10.1002/cpz1.724] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 03/29/2023]
Abstract
We report a microglial phagocytosis/cell health high-content assay that has been used to test small molecule chemical probes and support our drug discovery projects targeting microglia for Alzheimer's disease therapy. The assay measures phagocytosis and cell health (cell count and nuclear intensity) simultaneously in 384-well plates processed with an automatic liquid handler. The mix-and-read live cell imaging assay is highly reproducible with capacity to meet drug discovery research needs. Assay procedures take 4 days including plating cells, treating cells, adding pHrodo-myelin/membrane debris to cells for phagocytosis, staining cell nuclei before performing high-content imaging, and analysis. Three selected parameters are measured from cells: 1) mean total fluorescence intensity per cell of pHrodo-myelin/membrane debris in phagocytosis vesicles to quantify phagocytosis; 2) cell counts per well (measuring compound effects on proliferation and cell death); and 3) average nuclear intensity (measuring compound induced apoptosis). The assay has been used on HMC3 cells (an immortalized human microglial cell line), BV2 cells (an immortalized mouse microglial cell line), and primary microglia isolated from mouse brains. Simultaneous measurements of phagocytosis and cell health allow for the distinction of compound effects on regulation of phagocytosis from cellular stress/toxicity related changes, a distinguishing feature of the assay. The combination of cell counts and nuclear intensity as indicators of cell health is also an effective way to measure cell stress and compound cytotoxicity, which may have broad applications as simultaneous profiling measurements for other phenotypic assays. © 2023 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol: Microglial phagocytosis/cell health high-content assay protocol Support Protocol: Procedures to isolate myelin/membrane debris from mouse brain and label with pHrodo.
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Affiliation(s)
- Emily R Mason
- Division of Clinical Pharmacology, Department of Medicine, IUSM-Purdue TREAT-AD Center, Indiana University School of Medicine, Indianapolis, Indiana
| | - Disha M Soni
- Department of Radiology & Imaging Sciences, Stark Neurosciences Research Institute, Indiana University School of Medicine, Indianapolis, Indiana
| | - Shaoyou Chu
- Division of Clinical Pharmacology, Department of Medicine, IUSM-Purdue TREAT-AD Center, Indiana University School of Medicine, Indianapolis, Indiana
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19
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Mills MK, McCabe LG, Rodrigue EM, Lechtreck KF, Starai VJ. Wbm0076, a candidate effector protein of the Wolbachia endosymbiont of Brugia malayi, disrupts eukaryotic actin dynamics. PLoS Pathog 2023; 19:e1010777. [PMID: 36800397 PMCID: PMC9980815 DOI: 10.1371/journal.ppat.1010777] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/01/2022] [Revised: 03/02/2023] [Accepted: 01/31/2023] [Indexed: 02/18/2023] Open
Abstract
Brugia malayi, a parasitic roundworm of humans, is colonized by the obligate intracellular bacterium, Wolbachia pipientis. The symbiosis between this nematode and bacterium is essential for nematode reproduction and long-term survival in a human host. Therefore, identifying molecular mechanisms required by Wolbachia to persist in and colonize B. malayi tissues will provide new essential information regarding the basic biology of this endosymbiosis. Wolbachia utilize a Type IV secretion system to translocate so-called "effector" proteins into the cytosol of B. malayi cells to promote colonization of the eukaryotic host. However, the characterization of these Wolbachia secreted proteins has remained elusive due to the genetic intractability of both organisms. Strikingly, expression of the candidate Wolbachia Type IV-secreted effector protein, Wbm0076, in the surrogate eukaryotic cell model, Saccharomyces cerevisiae, resulted in the disruption of the yeast actin cytoskeleton and inhibition of endocytosis. Genetic analyses show that Wbm0076 is a member of the family of Wiskott-Aldrich syndrome proteins (WAS [p]), a well-conserved eukaryotic protein family required for the organization of actin skeletal structures. Thus, Wbm0076 likely plays a central role in the active cell-to-cell movement of Wolbachia throughout B. malayi tissues during nematode development. As most Wolbachia isolates sequenced to date encode at least partial orthologs of wBm0076, we find it likely that the ability of Wolbachia to directly manipulate host actin dynamics is an essential requirement of all Wolbachia endosymbioses, independent of host cell species.
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Affiliation(s)
- Michael K. Mills
- Department of Microbiology, University of Georgia, Athens, Georgia, United States of America
| | - Lindsey G. McCabe
- Department of Microbiology, University of Georgia, Athens, Georgia, United States of America
| | - Eugenie M. Rodrigue
- Department of Microbiology, University of Georgia, Athens, Georgia, United States of America
| | - Karl F. Lechtreck
- Department of Cellular Biology, University of Georgia, Athens, Georgia, United States of America
| | - Vincent J. Starai
- Department of Microbiology, University of Georgia, Athens, Georgia, United States of America
- Department of Infectious Diseases, University of Georgia, Athens, Georgia, United States of America
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20
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Smith HB, Lee K, Stevenson DM, Amador-Noguez D, Sauer JD. Listeria monocytogenes requires DHNA-dependent intracellular redox homeostasis facilitated by Ndh2 for survival and virulence. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2023:2023.01.13.524026. [PMID: 36711537 PMCID: PMC9882099 DOI: 10.1101/2023.01.13.524026] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Indexed: 01/16/2023]
Abstract
Listeria monocytogenes is a remarkably well-adapted facultative intracellular pathogen that can thrive in a wide range of ecological niches. L. monocytogenes maximizes its ability to generate energy from diverse carbon sources using a respiro-fermentative metabolism that can function under both aerobic and anaerobic conditions. Cellular respiration maintains redox homeostasis by regenerating NAD + while also generating a proton motive force (PMF). The end products of the menaquinone (MK) biosynthesis pathway are essential to drive both aerobic and anaerobic cellular respiration. We previously demonstrated that intermediates in the MK biosynthesis pathway, notably 1,4-dihydroxy-2-naphthoate (DHNA), are required for the survival and virulence of L. monocytogenes independent of their role in respiration. Furthermore, we found that restoration of NAD + /NADH ratio through expression of water-forming NADH oxidase (NOX) could rescue phenotypes associated with DHNA deficiency. Here we extend these findings to demonstrate that endogenous production or direct supplementation of DHNA restored both the cellular redox homeostasis and metabolic output of fermentation in L. monocytogenes . Further, exogenous supplementation of DHNA rescues the in vitro growth and ex vivo virulence of L. monocytogenes DHNA-deficient mutants. Finally, we demonstrate that exogenous DHNA restores redox balance in L. monocytogenes specifically through the recently annotated NADH dehydrogenase Ndh2, independent of the extracellular electron transport (EET) pathway. These data suggest that the production of DHNA may represent an additional layer of metabolic adaptability by L. monocytogenes to drive energy metabolism in the absence of respiration-favorable conditions.
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Affiliation(s)
- Hans B. Smith
- Department of Medical Microbiology and Immunology, University of Wisconsin-Madison, Madison, WI, 53706, United States of America
| | - Kijeong Lee
- Department of Medical Microbiology and Immunology, University of Wisconsin-Madison, Madison, WI, 53706, United States of America
| | - David M. Stevenson
- Department of Bacteriology, University of Wisconsin-Madison, Madison, WI, 53706, United States of America
| | - Daniel Amador-Noguez
- Department of Bacteriology, University of Wisconsin-Madison, Madison, WI, 53706, United States of America
| | - John-Demian Sauer
- Department of Medical Microbiology and Immunology, University of Wisconsin-Madison, Madison, WI, 53706, United States of America
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21
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Singh V, Rai R, Mathew BJ, Chourasia R, Singh AK, Kumar A, Chaurasiya SK. Phospholipase C: underrated players in microbial infections. Front Cell Infect Microbiol 2023; 13:1089374. [PMID: 37139494 PMCID: PMC10149971 DOI: 10.3389/fcimb.2023.1089374] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/04/2022] [Accepted: 03/21/2023] [Indexed: 05/05/2023] Open
Abstract
During bacterial infections, one or more virulence factors are required to support the survival, growth, and colonization of the pathogen within the host, leading to the symptomatic characteristic of the disease. The outcome of bacterial infections is determined by several factors from both host as well as pathogen origin. Proteins and enzymes involved in cellular signaling are important players in determining the outcome of host-pathogen interactions. phospholipase C (PLCs) participate in cellular signaling and regulation by virtue of their ability to hydrolyze membrane phospholipids into di-acyl-glycerol (DAG) and inositol triphosphate (IP3), which further causes the activation of other signaling pathways involved in various processes, including immune response. A total of 13 PLC isoforms are known so far, differing in their structure, regulation, and tissue-specific distribution. Different PLC isoforms have been implicated in various diseases, including cancer and infectious diseases; however, their roles in infectious diseases are not clearly understood. Many studies have suggested the prominent roles of both host and pathogen-derived PLCs during infections. PLCs have also been shown to contribute towards disease pathogenesis and the onset of disease symptoms. In this review, we have discussed the contribution of PLCs as a determinant of the outcome of host-pathogen interaction and pathogenesis during bacterial infections of human importance.
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Affiliation(s)
- Vinayak Singh
- Molecular Signalling Lab, Department of Biological Science and Engineering, Maulana Azad National Institute of Technology, Bhopal, Madhya Pradesh, India
| | - Rupal Rai
- Molecular Signalling Lab, Department of Biological Science and Engineering, Maulana Azad National Institute of Technology, Bhopal, Madhya Pradesh, India
| | - Bijina J. Mathew
- Molecular Signalling Lab, Department of Biological Science and Engineering, Maulana Azad National Institute of Technology, Bhopal, Madhya Pradesh, India
| | - Rashmi Chourasia
- Department of Chemistry, IES University, Bhopal, Madhya Pradesh, India
| | - Anirudh K. Singh
- School of Sciences, SAM Global University, Raisen, Madhya Pradesh, India
| | - Awanish Kumar
- Department of Biotechnology, National Institute of Technology, Raipur, Chhattisgarh, India
| | - Shivendra K. Chaurasiya
- Molecular Signalling Lab, Department of Biological Science and Engineering, Maulana Azad National Institute of Technology, Bhopal, Madhya Pradesh, India
- *Correspondence: Shivendra K. Chaurasiya,
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22
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In through the Out Exit: the Role of the Exocyst in Listeria monocytogenes Cell Entry. Infect Immun 2022; 90:e0048422. [PMID: 36394320 PMCID: PMC9753639 DOI: 10.1128/iai.00484-22] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/19/2022] Open
Abstract
The intracellular pathogen Listeria monocytogenes is one of the leading causes of death from foodborne illness in the United States. Internalin A is the key surface protein that drives Listeria uptake by epithelial cells expressing E-cadherin. G. C. Gyanwali, T. U. B. Herath, A. Gianfelice, and K. Ireton (Infect Immun 90:e00326-22, 2022, https://doi.org/10.1128/iai.00326-22) unravel the close relationship between internalin A and the exocyst, adding another layer of complexity to the bacterial internalization process.
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23
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Listeria monocytogenes-How This Pathogen Uses Its Virulence Mechanisms to Infect the Hosts. Pathogens 2022; 11:pathogens11121491. [PMID: 36558825 PMCID: PMC9783847 DOI: 10.3390/pathogens11121491] [Citation(s) in RCA: 29] [Impact Index Per Article: 9.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/26/2022] [Revised: 11/23/2022] [Accepted: 12/05/2022] [Indexed: 12/13/2022] Open
Abstract
Listeriosis is a serious food-borne illness, especially in susceptible populations, including children, pregnant women, and elderlies. The disease can occur in two forms: non-invasive febrile gastroenteritis and severe invasive listeriosis with septicemia, meningoencephalitis, perinatal infections, and abortion. Expression of each symptom depends on various bacterial virulence factors, immunological status of the infected person, and the number of ingested bacteria. Internalins, mainly InlA and InlB, invasins (invasin A, LAP), and other surface adhesion proteins (InlP1, InlP4) are responsible for epithelial cell binding, whereas internalin C (InlC) and actin assembly-inducing protein (ActA) are involved in cell-to-cell bacterial spread. L. monocytogenes is able to disseminate through the blood and invade diverse host organs. In persons with impaired immunity, the elderly, and pregnant women, the pathogen can also cross the blood-brain and placental barriers, which results in the invasion of the central nervous system and fetus infection, respectively. The aim of this comprehensive review is to summarize the current knowledge on the epidemiology of listeriosis and L. monocytogenes virulence mechanisms that are involved in host infection, with a special focus on their molecular and cellular aspects. We believe that all this information is crucial for a better understanding of the pathogenesis of L. monocytogenes infection.
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24
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Ayariga JA, Ibrahim I, Gildea L, Abugri J, Villafane R. Microbiota in a long survival discourse with the human host. Arch Microbiol 2022; 205:5. [PMID: 36441284 DOI: 10.1007/s00203-022-03342-6] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/21/2022] [Revised: 11/14/2022] [Accepted: 11/16/2022] [Indexed: 11/29/2022]
Abstract
The relationship between human health and gut microbiota is becoming more apparent. It is now widely believed that healthy gut flora plays a vital role in the overall well-being of the individual. There are spatial and temporal variations in the distribution of microbes from the esophagus to the rectum throughout an individual's lifetime. Through the development of genome sequencing technologies, scientists have been able to study the interactions between different microorganisms and their hosts to improve the health and disease of individuals. The normal gut microbiota provides various functions to the host, whereas the host, in turn, provides nutrients and promotes the development of healthy and resilient microbiota communities. Thus, the microbiota provides and maintains the gut's structural integrity and protects the gut against pathogens. The development of the normal gut microbiota is influenced by various factors. Some of these include the mode of delivery, diet, and antibiotics. In addition, the environment can also affect the development of the gut microbiota. For example, one of the main concerns of antibiotic use is the alteration of the gut microbiota, which could lead to the development of multidrug-resistant organisms. When microbes are disturbed, it can potentially lead to various diseases. Depending on the species' ability to adapt to the human body's environment, the fate of the microbes in the host and their relationship with the human body are decided. This review aims to provide a comprehensive analysis of microbe, microbes-host immune interactions, and factors that can disturb their interactions.
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Affiliation(s)
- Joseph A Ayariga
- Department of Biological Sciences, Microbiology PhD. Program, College of Science, Technology, Engineering and Mathematics (C-STEM), Alabama State University, 1627 Hall Street Montgomery, Montgomery, AL, 36104, USA.
| | - Iddrisu Ibrahim
- Department of Biological Sciences, Microbiology PhD. Program, College of Science, Technology, Engineering and Mathematics (C-STEM), Alabama State University, 1627 Hall Street Montgomery, Montgomery, AL, 36104, USA
| | - Logan Gildea
- Department of Biological Sciences, Microbiology PhD. Program, College of Science, Technology, Engineering and Mathematics (C-STEM), Alabama State University, 1627 Hall Street Montgomery, Montgomery, AL, 36104, USA
| | - James Abugri
- Department of Biochemistry and Forensic Sciences, School of Chemical and Biochemical Sciences, C. K. Tedam University of Technology and Applied Sciences, Navrongo, Ghana.
| | - Robert Villafane
- Department of Biological Sciences, Microbiology PhD. Program, College of Science, Technology, Engineering and Mathematics (C-STEM), Alabama State University, 1627 Hall Street Montgomery, Montgomery, AL, 36104, USA
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Selection of Listeria monocytogenes InlA-Binding Peptides Using Phage Display—Novel Compounds for Diagnostic Applications? Appl Microbiol 2022. [DOI: 10.3390/applmicrobiol2040070] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/12/2022]
Abstract
Listeria monocytogenes is a pathogenic, gram-positive bacterium causing foodborne infections and listeriosis, an infection responsible for serious medical conditions, especially for pregnant women, newborns, or people with a weak immune system. Even after antibiotic treatment, 30% of clinical infections result in death. L. monocytogenes is able to enter and multiply in mammalian cells. Invasion into epithelial cells in the human intestine is mediated by the interaction of the bacterial surface protein internalin A (InlA) with the host cell receptor E-cadherin (E-cad). We have used phage display to select InlA-specific peptides consisting of 12 amino acids using a randomized, recombinant peptide library. We could demonstrate that the selected peptides bound to recombinant InlA protein as well as to L. monocytogenes cells. In vitro, some of the peptides inhibited the interaction between recombinant InlA and human E-cad. As far as we know, this is the first publication on the development of InlA-specific peptide ligands. In the future, our peptides might be used for the development of innovative diagnostic tools or even therapeutic approaches.
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Cho SY, Na HW, Oh H, Kwak YM, Song WS, Park S, Jeon WJ, Cho H, Oh BC, Park J, Kang S, Lee GS, Yoon SI. Structural basis of flagellar motility regulation by the MogR repressor and the GmaR antirepressor in Listeria monocytogenes. Nucleic Acids Res 2022; 50:11315-11330. [PMID: 36283692 PMCID: PMC9638930 DOI: 10.1093/nar/gkac815] [Citation(s) in RCA: 11] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/25/2021] [Revised: 08/01/2022] [Accepted: 09/11/2022] [Indexed: 08/27/2023] Open
Abstract
The pathogenic Listeria monocytogenes bacterium produces the flagellum as a locomotive organelle at or below 30°C outside the host, but it halts flagellar expression at 37°C inside the human host to evade the flagellum-induced immune response. Listeria monocytogenes GmaR is a thermosensor protein that coordinates flagellar expression by binding the master transcriptional repressor of flagellar genes (MogR) in a temperature-responsive manner. To understand the regulatory mechanism whereby GmaR exerts the antirepression activity on flagellar expression, we performed structural and mutational analyses of the GmaR-MogR system. At or below 30°C, GmaR exists as a functional monomer and forms a circularly enclosed multidomain structure via an interdomain interaction. GmaR in this conformation recognizes MogR using the C-terminal antirepressor domain in a unique dual binding mode and mediates the antirepressor function through direct competition and spatial restraint mechanisms. Surprisingly, at 37°C, GmaR rapidly forms autologous aggregates that are deficient in MogR neutralization capabilities.
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Affiliation(s)
- So Yeon Cho
- Division of Biomedical Convergence, College of Biomedical Science, Kangwon National University, Chuncheon 24341, Republic of Korea
| | - Hye-won Na
- Division of Biomedical Convergence, College of Biomedical Science, Kangwon National University, Chuncheon 24341, Republic of Korea
| | - Han Byeol Oh
- Division of Biomedical Convergence, College of Biomedical Science, Kangwon National University, Chuncheon 24341, Republic of Korea
| | - Yun Mi Kwak
- Division of Biomedical Convergence, College of Biomedical Science, Kangwon National University, Chuncheon 24341, Republic of Korea
| | - Wan Seok Song
- Institute of Bioscience and Biotechnology, Kangwon National University, Chuncheon 24341, Republic of Korea
| | - Sun Cheol Park
- Division of Biomedical Convergence, College of Biomedical Science, Kangwon National University, Chuncheon 24341, Republic of Korea
| | - Wook-Jong Jeon
- Department of Biological Sciences, College of Natural Sciences, Sungkyunkwan University, Suwon 16419, Republic of Korea
| | - Hongbaek Cho
- Department of Biological Sciences, College of Natural Sciences, Sungkyunkwan University, Suwon 16419, Republic of Korea
| | - Byung-Chul Oh
- Lee Gil Ya Cancer and Diabetes Institute, College of Medicine, Gachon University, Incheon 406-840, Republic of Korea
| | - Jeongho Park
- College of Veterinary Medicine, Kangwon National University, Chuncheon 24341, Republic of Korea
| | - Seung Goo Kang
- Division of Biomedical Convergence, College of Biomedical Science, Kangwon National University, Chuncheon 24341, Republic of Korea
| | - Geun-Shik Lee
- College of Veterinary Medicine, Kangwon National University, Chuncheon 24341, Republic of Korea
| | - Sung-il Yoon
- Division of Biomedical Convergence, College of Biomedical Science, Kangwon National University, Chuncheon 24341, Republic of Korea
- Institute of Bioscience and Biotechnology, Kangwon National University, Chuncheon 24341, Republic of Korea
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The Transcriptional Regulator SpxA1 Influences the Morphology and Virulence of Listeria monocytogenes. Infect Immun 2022; 90:e0021122. [PMID: 36102657 PMCID: PMC9584327 DOI: 10.1128/iai.00211-22] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022] Open
Abstract
Listeria monocytogenes is a Gram-positive facultative anaerobe and an excellent model pathogen for investigating regulatory changes that occur during infection of a mammalian host. SpxA1 is a widely conserved transcriptional regulator that induces expression of peroxide-detoxifying genes in L. monocytogenes and is thus required for aerobic growth. SpxA1 is also required for L. monocytogenes virulence, although the SpxA1-dependent genes important in this context remain to be identified. Here, we sought to investigate the role of SpxA1 in a tissue culture model of infection and made the surprising discovery that ΔspxA1 cells are dramatically elongated during growth in the host cytosol. Quantitative microscopy revealed that ΔspxA1 cells also form elongated filaments extracellularly during early exponential phase in rich medium. Scanning and transmission electron microscopy analysis found that the likely cause of this morphological phenotype is aberrantly placed division septa localized outside cell midpoints. Quantitative mass spectrometry of whole-cell lysates identified SpxA1-dependent changes in protein abundance, including a significant number of motility and flagellar proteins that were depleted in the ΔspxA1 mutant. Accordingly, we found that both the filamentation and the lack of motility contributed to decreased phagocytosis of ΔspxA1 cells by macrophages. Overall, we identify a novel role for SpxA1 in regulating cell elongation and motility, both of which impact L. monocytogenes virulence.
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Listeria monocytogenes Infection Alters the Content and Function of Extracellular Vesicles Produced by Trophoblast Stem Cells. Infect Immun 2022; 90:e0034722. [PMID: 36154271 DOI: 10.1128/iai.00347-22] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Placental immunity is critical for fetal health during pregnancy, as invading pathogens spread from the parental blood to the fetus through this organ. However, inflammatory responses in the placenta can adversely affect both the fetus and the pregnant person, and the balance between protective placental immune response and detrimental inflammation is poorly understood. Extracellular vesicles (EVs) are membrane-enclosed vesicles that play a critical role in placental immunity. EVs produced by placental trophoblasts mediate immune tolerance to the fetus and to the placenta itself, but these EVs can also activate detrimental inflammatory responses. The regulation of these effects is not well characterized, and the role of trophoblast EVs (tEVs) in the response to infection has yet to be defined. The Gram-positive bacterial pathogen Listeria monocytogenes infects the placenta, serving as a model to study tEV function in this context. We investigated the effect of L. monocytogenes infection on the production and function of tEVs, using a trophoblast stem cell (TSC) model. We found that tEVs from infected TSCs can induce the production of the proinflammatory cytokine tumor necrosis factor alpha (TNF-α) in recipient cells. Surprisingly, this tEV treatment could confer increased susceptibility to subsequent L. monocytogenes infection, which has not been reported previously as an effect of EVs. Proteomic analysis and RNA sequencing revealed that tEVs from infected TSCs had altered cargo compared with those from uninfected TSCs. However, no L. monocytogenes proteins were detected in tEVs from infected TSCs. Together, these results suggest an immunomodulatory role for tEVs during prenatal infection.
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Izquierdo-Serrano R, Fernández-Delgado I, Moreno-Gonzalo O, Martín-Gayo E, Calzada-Fraile D, Ramírez-Huesca M, Jorge I, Camafeita E, Abián J, Vicente-Manzanares M, Veiga E, Vázquez J, Sánchez-Madrid F. Extracellular vesicles from Listeria monocytogenes-infected dendritic cells alert the innate immune response. Front Immunol 2022; 13:946358. [PMID: 36131943 PMCID: PMC9483171 DOI: 10.3389/fimmu.2022.946358] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/17/2022] [Accepted: 08/11/2022] [Indexed: 11/13/2022] Open
Abstract
Communication through cell-cell contacts and extracellular vesicles (EVs) enables immune cells to coordinate their responses against diverse types of pathogens. The function exerted by EVs in this context depends on the proteins and nucleic acids loaded into EVs, which elicit specific responses involved in the resolution of infection. Several mechanisms control protein and nucleic acid loading into EVs; in this regard, acetylation has been described as a mechanism of cellular retention during protein sorting to exosomes. HDAC6 is a deacetylase involved in the control of cytoskeleton trafficking, organelle polarity and cell migration, defense against Listeria monocytogenes (Lm) infection and other immune related functions. Here, we show that the protein content of dendritic cells (DCs) and their secreted EVs (DEVs) vary during Lm infection, is enriched in proteins related to antiviral functions compared to non-infected cells and depends on HDAC6 expression. Analyses of the post-translational modifications revealed an alteration of the acetylation and ubiquitination profiles upon Lm infection both in DC lysates and DEVs. Functionally, EVs derived from infected DCs upregulate anti-pathogenic genes (e.g. inflammatory cytokines) in recipient immature DCs, which translated into protection from subsequent infection with vaccinia virus. Interestingly, absence of Listeriolysin O in Lm prevents DEVs from inducing this anti-viral state. In summary, these data underscore a new mechanism of communication between bacteria-infected DC during infection as they alert neighboring, uninfected DCs to promote antiviral responses.
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Affiliation(s)
- Raúl Izquierdo-Serrano
- Vascular Pathophysiology Area, Centro Nacional Investigaciones Cardiovasculares (CNIC) Carlos III, Madrid, Spain
| | - Irene Fernández-Delgado
- Vascular Pathophysiology Area, Centro Nacional Investigaciones Cardiovasculares (CNIC) Carlos III, Madrid, Spain
- Department of Immunology, Instituto Investigación Sanitaria Hospital Universitario La Princesa (IIS-HUP), Universidad Autónoma de Madrid (UAM), Madrid, Spain
| | - Olga Moreno-Gonzalo
- Vascular Pathophysiology Area, Centro Nacional Investigaciones Cardiovasculares (CNIC) Carlos III, Madrid, Spain
- Department of Immunology, Instituto Investigación Sanitaria Hospital Universitario La Princesa (IIS-HUP), Universidad Autónoma de Madrid (UAM), Madrid, Spain
| | - Enrique Martín-Gayo
- Department of Immunology, Instituto Investigación Sanitaria Hospital Universitario La Princesa (IIS-HUP), Universidad Autónoma de Madrid (UAM), Madrid, Spain
| | - Diego Calzada-Fraile
- Vascular Pathophysiology Area, Centro Nacional Investigaciones Cardiovasculares (CNIC) Carlos III, Madrid, Spain
| | - Marta Ramírez-Huesca
- Vascular Pathophysiology Area, Centro Nacional Investigaciones Cardiovasculares (CNIC) Carlos III, Madrid, Spain
- Department of Immunology, Instituto Investigación Sanitaria Hospital Universitario La Princesa (IIS-HUP), Universidad Autónoma de Madrid (UAM), Madrid, Spain
| | - Inmaculada Jorge
- Vascular Pathophysiology Area, Centro Nacional Investigaciones Cardiovasculares (CNIC) Carlos III, Madrid, Spain
- Centro de Investigación Biomédica en Red de Enfermedades Cardiovasculares (CIBERCV), Madrid, Spain
| | - Emilio Camafeita
- Vascular Pathophysiology Area, Centro Nacional Investigaciones Cardiovasculares (CNIC) Carlos III, Madrid, Spain
- Centro de Investigación Biomédica en Red de Enfermedades Cardiovasculares (CIBERCV), Madrid, Spain
| | - Joaquín Abián
- Biological and Environmental Proteomics, Institut d’Investigacions Biomèdiques de Barcelona, Consejo Superior de Investigaciones Científicas (IIBB-CSIC), Institut d’Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Barcelona, Spain
| | - Miguel Vicente-Manzanares
- Molecular Mechanisms Program, Centro de Investigación del Cáncer and Instituto de Biología Molecular y Celular del Cáncer, Consejo Superior de Investigaciones Científicas (CSIC)-Universidad de Salamanca, Salamanca, Spain
| | - Esteban Veiga
- Department of Molecular & Cellular Biology, Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Científicas (CNB-CSIC), Madrid, Spain
| | - Jesús Vázquez
- Vascular Pathophysiology Area, Centro Nacional Investigaciones Cardiovasculares (CNIC) Carlos III, Madrid, Spain
- Centro de Investigación Biomédica en Red de Enfermedades Cardiovasculares (CIBERCV), Madrid, Spain
| | - Francisco Sánchez-Madrid
- Vascular Pathophysiology Area, Centro Nacional Investigaciones Cardiovasculares (CNIC) Carlos III, Madrid, Spain
- Department of Immunology, Instituto Investigación Sanitaria Hospital Universitario La Princesa (IIS-HUP), Universidad Autónoma de Madrid (UAM), Madrid, Spain
- Centro de Investigación Biomédica en Red de Enfermedades Cardiovasculares (CIBERCV), Madrid, Spain
- *Correspondence: Francisco Sánchez-Madrid,
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Structural and biochemical analyses of the flagellar expression regulator DegU from Listeria monocytogenes. Sci Rep 2022; 12:10856. [PMID: 35798759 PMCID: PMC9263151 DOI: 10.1038/s41598-022-14459-5] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/13/2021] [Accepted: 06/07/2022] [Indexed: 11/24/2022] Open
Abstract
Listeria monocytogenes is a pathogenic bacterium that produces flagella, the locomotory organelles, in a temperature-dependent manner. At 37 °C inside humans, L. monocytogenes employs MogR to repress the expression of flagellar proteins, thereby preventing the production of flagella. However, in the low-temperature environment outside of the host, the antirepressor GmaR inactivates MogR, allowing flagellar formation. Additionally, DegU is necessary for flagellar expression at low temperatures. DegU transcriptionally activates the expression of GmaR and flagellar proteins by binding the operator DNA in the fliN-gmaR promoter as a response regulator of a two-component regulatory system. To determine the DegU-mediated regulation mechanism, we performed structural and biochemical analyses on the recognition of operator DNA by DegU. The DegU-DNA interaction is primarily mediated by a C-terminal DNA-binding domain (DBD) and can be fortified by an N-terminal receiver domain (RD). The DegU DBD adopts a tetrahelical helix-turn-helix structure and assembles into a dimer. The DegU DBD dimer recognizes the operator DNA using a positive patch. Unexpectedly, unlike typical response regulators, DegU interacts with operator DNA in both unphosphorylated and phosphorylated states with similar binding affinities. Therefore, we conclude that DegU is a noncanonical response regulator that is constitutively active irrespective of phosphorylation.
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Dhanda AS, Guttman JA. Localization of host endocytic and actin-associated proteins during Shigella flexneri intracellular motility and intercellular spreading. Anat Rec (Hoboken) 2022; 306:1088-1110. [PMID: 35582740 DOI: 10.1002/ar.24955] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/28/2022] [Revised: 04/24/2022] [Accepted: 04/27/2022] [Indexed: 11/10/2022]
Abstract
Shigella flexneri (S. flexneri), the causative agent of bacillary dysentery, uses an effector-mediated strategy to hijack host cells and cause disease. To propagate and spread within human tissues, S. flexneri bacteria commandeer the host actin cytoskeleton to generate slender actin-rich comet tails to move intracellularly, and later, plasma membrane actin-based protrusions to move directly between adjacent host cells. To facilitate intercellular bacterial spreading, large micron-sized endocytic-like membrane invaginations form at the periphery of neighboring host cells that come into contact with S. flexneri-containing membrane protrusions. While S. flexneri comet tails and membrane protrusions consist primarily of host actin cytoskeletal proteins, S. flexneri membrane invaginations remain poorly understood with only clathrin and the clathrin adapter epsin-1 localized to the structures. Tangentially, we recently reported that Listeria monocytogenes, another actin-hijacking pathogen, exploits an assortment of caveolar and actin-bundling proteins at their micron-sized membrane invaginations formed during their cell-to-cell movement. Thus, to further characterize the S. flexneri disease process, we set out to catalog the distribution of a variety of actin-associated and caveolar proteins during S. flexneri actin-based motility and cell-to-cell spreading. Here we show that actin-associated proteins found at L. monocytogenes comet tails and membrane protrusions mimic those present at S. flexneri comet tails with the exception of α-actinins 1 and 4, which were shed from S. flexneri membrane protrusions. We also demonstrate that all known host endocytic components found at L. monocytogenes membrane invaginations are also present at those formed during S. flexneri infections.
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Affiliation(s)
- Aaron Singh Dhanda
- Department of Biological Sciences, Centre for Cell Biology, Development, and Disease, Simon Fraser University, Burnaby, British Columbia, Canada
| | - Julian Andrew Guttman
- Department of Biological Sciences, Centre for Cell Biology, Development, and Disease, Simon Fraser University, Burnaby, British Columbia, Canada
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Listeria monocytogenes TcyKLMN Cystine/Cysteine Transporter Facilitates Glutathione Synthesis and Virulence Gene Expression. mBio 2022; 13:e0044822. [PMID: 35435705 PMCID: PMC9239247 DOI: 10.1128/mbio.00448-22] [Citation(s) in RCA: 7] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Bacterial pathogens sense the repertoire of metabolites in the mammalian niche and use this information to shift into the pathogenic state to accomplish a successful infection. Glutathione is a virulence-activating signal that is synthesized by
L. monocytogenes
during infection of mammalian cells.
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Yadav V, Gopalakrishnan M. Force-velocity relation and load-sharing in the linear polymerization ratchet revisited: the effects of barrier diffusion. THE EUROPEAN PHYSICAL JOURNAL. E, SOFT MATTER 2022; 45:35. [PMID: 35416551 DOI: 10.1140/epje/s10189-022-00190-6] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 12/20/2021] [Accepted: 03/29/2022] [Indexed: 06/14/2023]
Abstract
We study the velocity-force (V-F) relation for a Brownian ratchet consisting of a linear rigid polymer growing against a diffusing barrier, acted upon by a opposing constant force (F). Using a careful mathematical analysis, we derive the V-F relations in the extreme limits of fast and slow barrier diffusion. In the first case, V depends exponentially on the load F, in agreement with the well-known formula proposed by Peskin, Odell and Oster (1993), while the relationship becomes linear in the second case. For a bundle of two filaments growing against a common barrier, equal sharing of load in the corresponding V-F relation is predicted by a mean-field argument in both limits. However, the scaling behaviour of velocity with the number of filaments is different for the two cases. In the limit of large D, the validity of the mean-field approach is tested, and partially supported by a detailed and rigorous analysis. Our principal predictions are also verified in numerical simulations.
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Affiliation(s)
- Vandana Yadav
- Department of Physics, Indian Institute of Technology Madras, Chennai, 600036, India
| | - Manoj Gopalakrishnan
- Department of Physics, Indian Institute of Technology Madras, Chennai, 600036, India.
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Maudet C, Kheloufi M, Levallois S, Gaillard J, Huang L, Gaultier C, Tsai YH, Disson O, Lecuit M. Bacterial inhibition of Fas-mediated killing promotes neuroinvasion and persistence. Nature 2022; 603:900-906. [PMID: 35296858 DOI: 10.1038/s41586-022-04505-7] [Citation(s) in RCA: 18] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/20/2020] [Accepted: 02/03/2022] [Indexed: 12/19/2022]
Abstract
Infections of the central nervous system are among the most serious infections1,2, but the mechanisms by which pathogens access the brain remain poorly understood. The model microorganism Listeria monocytogenes (Lm) is a major foodborne pathogen that causes neurolisteriosis, one of the deadliest infections of the central nervous system3,4. Although immunosuppression is a well-established host risk factor for neurolisteriosis3,5, little is known about the bacterial factors that underlie the neuroinvasion of Lm. Here we develop a clinically relevant experimental model of neurolisteriosis, using hypervirulent neuroinvasive strains6 inoculated in a humanized mouse model of infection7, and we show that the bacterial surface protein InlB protects infected monocytes from Fas-mediated cell death by CD8+ T cells in a manner that depends on c-Met, PI3 kinase and FLIP. This blockade of specific anti-Lm cellular immune killing lengthens the lifespan of infected monocytes, and thereby favours the transfer of Lm from infected monocytes to the brain. The intracellular niche that is created by InlB-mediated cell-autonomous immune resistance also promotes Lm faecal shedding, which accounts for the selection of InlB as a core virulence gene of Lm. We have uncovered a specific mechanism by which a bacterial pathogen confers an increased lifespan to the cells it infects by rendering them resistant to cell-mediated immunity. This promotes the persistence of Lm within the host, its dissemination to the central nervous system and its transmission.
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Affiliation(s)
- Claire Maudet
- Institut Pasteur, Université de Paris, Inserm U1117, Biology of Infection Unit, Paris, France
| | - Marouane Kheloufi
- Institut Pasteur, Université de Paris, Inserm U1117, Biology of Infection Unit, Paris, France
| | - Sylvain Levallois
- Institut Pasteur, Université de Paris, Inserm U1117, Biology of Infection Unit, Paris, France
| | - Julien Gaillard
- Institut Pasteur, Université de Paris, Inserm U1117, Biology of Infection Unit, Paris, France
| | - Lei Huang
- Institut Pasteur, Université de Paris, Inserm U1117, Biology of Infection Unit, Paris, France
| | - Charlotte Gaultier
- Institut Pasteur, Université de Paris, Inserm U1117, Biology of Infection Unit, Paris, France
| | - Yu-Huan Tsai
- Institut Pasteur, Université de Paris, Inserm U1117, Biology of Infection Unit, Paris, France.,Institute of Microbiology and Immunology, National Yang-Ming University, Taipei, Taiwan
| | - Olivier Disson
- Institut Pasteur, Université de Paris, Inserm U1117, Biology of Infection Unit, Paris, France
| | - Marc Lecuit
- Institut Pasteur, Université de Paris, Inserm U1117, Biology of Infection Unit, Paris, France. .,Institut Pasteur, National Reference Center and WHO Collaborating Center Listeria, Paris, France. .,Necker-Enfants Malades University Hospital, Division of Infectious Diseases and Tropical Medicine, APHP, Institut Imagine, Paris, France.
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Regulator of Actin-Based Motility (RoaM) Downregulates Actin Tail Formation by Rickettsia rickettsii and Is Negatively Selected in Mammalian Cell Culture. mBio 2022; 13:e0035322. [PMID: 35285700 PMCID: PMC9040884 DOI: 10.1128/mbio.00353-22] [Citation(s) in RCA: 15] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/05/2023] Open
Abstract
The etiological agent of Rocky Mountain spotted fever, Rickettsia rickettsii, is an obligately intracellular pathogen that induces the polymerization of actin filaments to propel the bacterium through the cytoplasm and spread to new host cells. Cell-to-cell spread via actin-based motility is considered a key virulence determinant for spotted fever group rickettsiae, as interruption of sca2, the gene directly responsible for actin polymerization, has been shown to reduce fever in guinea pigs. However, little is known about how, or if, motility is regulated by the bacterium itself. We isolated a hyperspreading variant of R. rickettsii Sheila Smith that produces actin tails at an increased rate. A1G_06520 (roaM [regulator of actin-based motility]) was identified as a negative regulator of actin tail formation. Disruption of RoaM significantly increased the number of actin tails compared to the wild-type strain but did not increase virulence in guinea pigs; however, overexpression of RoaM dramatically decreased the presence of actin tails and moderated fever response. Localization experiments suggest that RoaM is not secreted, while reverse transcription-quantitative PCR (RT-qPCR) data show that various levels of RoaM do not significantly affect the expression of the known rickettsial actin-regulating proteins sca2, sca4, and rickA. Taken together, the data suggest a previously unrecognized level of regulation of actin-based motility in spotted fever group rickettsiae. Although this gene is intact in many isolates of spotted fever, transitional, and ancestral group Rickettsia spp., it is often ablated in highly passaged laboratory strains. Serial passage experiments revealed strong negative selection of roaM in Vero 76 cells.
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Kim J, Park J, Choi Z, Hong M. Structure-based molecular characterization of the LltR transcription factor from Listeria monocytogenes. Biochem Biophys Res Commun 2022; 600:142-149. [PMID: 35219103 DOI: 10.1016/j.bbrc.2022.02.067] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/16/2022] [Accepted: 02/18/2022] [Indexed: 12/18/2022]
Abstract
Listeria monocytogenes is a psychrotrophic food-borne pathogenic bacterium that causes listeriosis. Due to its unusual adaptation, an ability to grow at extended temperatures ranging from 4 to 45 °C, L. monocytogenes is notoriously hard to control in food-manufacturing processes. In addition, the growing number of antibiotic-resistant L. monocytogenes strains have made listeriosis steadily refractory to clinical treatments and can lead to serious life-threatening diseases, such as sepsis and meningitis, in immunocompromised persons and neonates. Transcription factors that belong to the PadR family play a key role in bacterial survival against unfavorable environmental stresses. The LltR protein from L. monocytogenes was identified as a PadR-type transcription factor and was shown to be required for bacterial growth adaptation at low temperatures. Despite the functional significance of LltR in listeria survival and pathogenesis, our molecular understanding of the LltR-mediated transcriptional regulation is highly limited. Here, we report the crystal structure of LltR and reveal the operator DNA recognition mechanism used by LltR. LltR dimerizes into an isosceles triangle-like shape and requires a winged helix-turn-helix motif for dsDNA recognition. Indeed, LltR and putative operator dsDNA binding was observed and suggests a transcriptional repression of the llfR-lmo0600-lmo0601 operon by direct interaction between the LltR transcription factor and its promoter region. Structure-based comparative and mutational analyses showed that LltR interacts with dsDNA via a unique strategy that combines both LltR-specific and PadR family-common mechanisms.
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Affiliation(s)
- Junghun Kim
- Division of Biological Science and Technology, Yonsei University, Wonju, 26493, Republic of Korea
| | - Jaewan Park
- Division of Biological Science and Technology, Yonsei University, Wonju, 26493, Republic of Korea
| | - Zion Choi
- Division of Biological Science and Technology, Yonsei University, Wonju, 26493, Republic of Korea
| | - Minsun Hong
- Division of Biological Science and Technology, Yonsei University, Wonju, 26493, Republic of Korea.
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Chalenko Y, Kolbasova O, Pivova E, Abdulkadieva M, Povolyaeva O, Kalinin E, Kolbasov D, Ermolaeva S. Listeria monocytogenes Invasion Into Sheep Kidney Epithelial Cells Depends on InlB, and Invasion Efficiency Is Modulated by Phylogenetically Defined InlB Isoforms. Front Microbiol 2022; 13:825076. [PMID: 35197955 PMCID: PMC8859113 DOI: 10.3389/fmicb.2022.825076] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/29/2021] [Accepted: 01/05/2022] [Indexed: 11/13/2022] Open
Abstract
The facultative intracellular pathogen Listeria monocytogenes is of major veterinary importance in small ruminants. Nevertheless, details of L. monocytogenes interactions with cells of small ruminants are not fully established. To study the potential of L. monocytogenes to infect sheep cells, we used the finite sheep kidney cell line (shKEC), which was infected with the wild-type L. monocytogenes strain EGDe. The invasion efficiency was 0.015 ± 0.004%. The invasion factor InlB was critically important for invasion, and inlB gene deletion almost prevented L. monocytogenes invasion into shKEC cells. Comparison of the potential of phylogenetically defined InlB isoforms to restore the invasive phenotype of the EGDeΔinlB strain demonstrated that although all InlB isoforms restored invasion of the EGDeΔinlB strain into shKEC cells, the InlB isoforms typical of highly virulent ruminant strains of the clonal complexes CC1 and CC7 were more efficient than isoforms typical of CC2 and CC9 strains (which are less virulent toward ruminants) in supporting invasion. Listeria monocytogenes effectively multiplied with a doubling of time in about 90 min after they entered the sheep cells. Intracellular bacteria moved using the well-known actin polymerization mechanism. Cell-to-cell spreading was restricted to the infection of a few tens of neighboring cells for 7 days. Overall, the obtained results demonstrated that (i) InlB is required for invasion into sheep cells, (ii) InlB isoforms might be important for hypervirulence of certain clonal groups toward ruminants, and (iii) L. monocytogenes effectively multiplies in ovine cells once entered.
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Affiliation(s)
- Yaroslava Chalenko
- Laboratory of Ecology of Pathogenic Bacteria, Gamaleya Research Center of Epidemiology and Microbiology, Moscow, Russia
- Yaroslava Chalenko,
| | - Olga Kolbasova
- Federal Research Center for Virology and Microbiology (FRCVM), Volginsky, Russia
| | - Elena Pivova
- Federal Research Center for Virology and Microbiology (FRCVM), Volginsky, Russia
| | - Mariam Abdulkadieva
- Department of Dusty Plasma, Joint Institute for High Temperatures, Russian Academy of Sciences, Moscow, Russia
| | - Olga Povolyaeva
- Federal Research Center for Virology and Microbiology (FRCVM), Volginsky, Russia
| | - Egor Kalinin
- Laboratory of Ecology of Pathogenic Bacteria, Gamaleya Research Center of Epidemiology and Microbiology, Moscow, Russia
| | - Denis Kolbasov
- Federal Research Center for Virology and Microbiology (FRCVM), Volginsky, Russia
| | - Svetlana Ermolaeva
- Laboratory of Ecology of Pathogenic Bacteria, Gamaleya Research Center of Epidemiology and Microbiology, Moscow, Russia
- Federal Research Center for Virology and Microbiology (FRCVM), Nizhny Novgorod Research Veterinary Institute Branch, Nizhny Novgorod, Russia
- *Correspondence: Svetlana Ermolaeva,
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Cheng C, Han X, Xu J, Sun J, Li K, Han Y, Chen M, Song H. YjbH mediates the oxidative stress response and infection by regulating SpxA1 and the phosphoenolpyruvate-carbohydrate phosphotransferase system (PTS) in Listeria monocytogenes. Gut Microbes 2022; 13:1-19. [PMID: 33573432 PMCID: PMC7889195 DOI: 10.1080/19490976.2021.1884517] [Citation(s) in RCA: 9] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/04/2023] Open
Abstract
The foodborne pathogen Listeria monocytogenes relies on its ability to fine-tune the expression of virulence factors and stress regulators in response to rapidly changing environments. Here, we reveal that YjbH, a putative thioredoxin family oxidoreductase, plays a pivotal role in bacterial adaption to oxidative stress and host infection. YjbH directly interacts with SpxA1, an ArsC family oxidative stress response regulator, and the deletion of YjbH compromised the oxidative stress tolerance of L. monocytogenes. Also, YjbH is required for the bacterial spread in host cells and proliferation in mouse organs, thereby contributing to virulence. Transcriptomic analysis of strains treated with Cd2+ revealed that most virulence genes and phosphoenolpyruvate-carbohydrate phosphotransferase system (PTS) genes were significantly downregulated in the absence of YjbH. However, YjbH inhibits PrfA expression when bacteria were grown in the media, suggesting that YjbH participates in regulating the virulence genes via a complicated regulatory network involving PrfA and PTS. Collectively, these findings provide a valuable model for clarifying the roles of thioredoxins from foodborne pathogens regarding improving survival in the external environment and, more importantly, successfully establishing infection within the host.
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Affiliation(s)
- Changyong Cheng
- College of Animal Science and Technology & College of Veterinary Medicine of Zhejiang A&F University, Key Laboratory of Applied Technology on Green-Eco-Healthy Animal Husbandry of Zhejiang Province, China-Australia Joint Laboratory for Animal Health Big Data Analytics, Zhejiang Provincial Engineering Laboratory for Animal Health Inspection & Internet Technology, Hangzhou, Zhejiang, P. R. China
| | - Xiao Han
- College of Animal Science and Technology & College of Veterinary Medicine of Zhejiang A&F University, Key Laboratory of Applied Technology on Green-Eco-Healthy Animal Husbandry of Zhejiang Province, China-Australia Joint Laboratory for Animal Health Big Data Analytics, Zhejiang Provincial Engineering Laboratory for Animal Health Inspection & Internet Technology, Hangzhou, Zhejiang, P. R. China
| | - Jiali Xu
- College of Animal Science and Technology & College of Veterinary Medicine of Zhejiang A&F University, Key Laboratory of Applied Technology on Green-Eco-Healthy Animal Husbandry of Zhejiang Province, China-Australia Joint Laboratory for Animal Health Big Data Analytics, Zhejiang Provincial Engineering Laboratory for Animal Health Inspection & Internet Technology, Hangzhou, Zhejiang, P. R. China
| | - Jing Sun
- College of Animal Science and Technology & College of Veterinary Medicine of Zhejiang A&F University, Key Laboratory of Applied Technology on Green-Eco-Healthy Animal Husbandry of Zhejiang Province, China-Australia Joint Laboratory for Animal Health Big Data Analytics, Zhejiang Provincial Engineering Laboratory for Animal Health Inspection & Internet Technology, Hangzhou, Zhejiang, P. R. China
| | - Kang Li
- College of Animal Science and Technology & College of Veterinary Medicine of Zhejiang A&F University, Key Laboratory of Applied Technology on Green-Eco-Healthy Animal Husbandry of Zhejiang Province, China-Australia Joint Laboratory for Animal Health Big Data Analytics, Zhejiang Provincial Engineering Laboratory for Animal Health Inspection & Internet Technology, Hangzhou, Zhejiang, P. R. China
| | - Yue Han
- College of Animal Science and Technology & College of Veterinary Medicine of Zhejiang A&F University, Key Laboratory of Applied Technology on Green-Eco-Healthy Animal Husbandry of Zhejiang Province, China-Australia Joint Laboratory for Animal Health Big Data Analytics, Zhejiang Provincial Engineering Laboratory for Animal Health Inspection & Internet Technology, Hangzhou, Zhejiang, P. R. China
| | - Mianmian Chen
- College of Animal Science and Technology & College of Veterinary Medicine of Zhejiang A&F University, Key Laboratory of Applied Technology on Green-Eco-Healthy Animal Husbandry of Zhejiang Province, China-Australia Joint Laboratory for Animal Health Big Data Analytics, Zhejiang Provincial Engineering Laboratory for Animal Health Inspection & Internet Technology, Hangzhou, Zhejiang, P. R. China
| | - Houhui Song
- College of Animal Science and Technology & College of Veterinary Medicine of Zhejiang A&F University, Key Laboratory of Applied Technology on Green-Eco-Healthy Animal Husbandry of Zhejiang Province, China-Australia Joint Laboratory for Animal Health Big Data Analytics, Zhejiang Provincial Engineering Laboratory for Animal Health Inspection & Internet Technology, Hangzhou, Zhejiang, P. R. China,CONTACT Houhui Song College of Animal Science and Technology & College of Veterinary Medicine of Zhejiang A&F University, Key Laboratory of Applied Technology on Green-Eco-Healthy Animal Husbandry of Zhejiang Province, China-Australia Joint Laboratory for Animal Health Big Data Analytics, Zhejiang Provincial Engineering Laboratory for Animal Health Inspection & Internet Technology, Hangzhou, Zhejiang311300, P. R. China
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Abstract
The defining characteristic of eukaryotic cells is the segregation of critical cellular functions within various membrane bound cellular organelles, including the nucleus, endoplasmic reticulum, Golgi apparatus, lysosomes, and mitochondria. Cell biologists therefore have extensively utilized organelle specific counterstains to help identify the localization of specific proteins or other targets of interest in order to garner an understanding of either their potential functions or their effects on the cell. There currently is a wide array of fluorescent dyes and reagents that can be utilized in live and fixed cells to identify organelles, thereby creating challenges in both choosing between the plethora of options and optimizing their use. Here we present a discussion of commonly utilized commercially available organelle dyes and summarize the factors that influence selection of the various dyes for: a given organelle; live versus fixed cellular conditions; adaptation to a specific protocol; spectral multiplexing; or matching excitation/emission spectra to available imaging equipment. Also presented are recommended protocols for a typical example reagent that can be reliably utilized to visualize its target cellular organelle.
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Affiliation(s)
- Timothy Paul Foster
- Department of Microbiology, Immunology & Parasitology, Louisiana State University Health Sciences Center, New Orleans, LA, USA.
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Mancinelli AM, Vichich JM, Zinnen AD, Hugon AM, Bondarenko V, Metzger JM, Simmons HA, Golos TG, Emborg ME. Acute Exposure to the Food-Borne Pathogen Listeria monocytogenes Does Not Induce α-Synuclein Pathology in the Colonic ENS of Nonhuman Primates. J Inflamm Res 2021; 14:7265-7279. [PMID: 34992416 PMCID: PMC8710837 DOI: 10.2147/jir.s337549] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/03/2021] [Accepted: 12/01/2021] [Indexed: 11/23/2022] Open
Abstract
INTRODUCTION Gastrointestinal (GI) inflammation elicited by environmental factors is proposed to trigger Parkinson's disease (PD) by stimulating accumulation of pathological α-synuclein (α-syn) in the enteric nervous system (ENS), which then propagates to the central nervous system via the vagus nerve. The goal of this study was to model, in nonhuman primates, an acute exposure to a common food-borne pathogen in order to assess whether the related acute GI inflammation could initiate persistent α-syn pathology in the ENS, ultimately leading to PD. METHODS Adult female cynomolgus macaques were inoculated by oral gavage with 1×108 colony-forming units (CFUs) Listeria monocytogenes (LM, n=10) or vehicle (mock, n=3) and euthanized 2 weeks later. Evaluations included clinical monitoring, blood and fecal shedding of LM, and postmortem pathological analysis of colonic and cecal tissues. RESULTS LM inoculation of healthy adult cynomolgus macaques induced minimal to mild clinical signs of infection; LM shedding in feces was not seen in any of the animals nor was bacteremia detected. Colitis varied from none to moderate in LM-treated subjects and none to minimal in mock-treated subjects. Expression of inflammatory markers (HLA-DR, CD3, CD20), oxidative stress (8-OHDG), α-syn, and phosphorylated-α-syn in the enteric ganglia was not significantly different between treatment groups. DISCUSSION Our results demonstrate that cynomolgus macaques orally inoculated with LM present with a clinical response that resembles human LM exposure. They also suggest that acute exposure to food-borne pathogens is not sufficient to induce significant and persistent α-syn changes in healthy adult female subjects. Based on the results of this limited experimental setting, we propose that, if LM has a role in PD pathology, other underlying factors or conditions, such as male sex, inflammatory bowel disease, exposure to toxins, dysbiosis, and/or aging, are needed to be present.
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Affiliation(s)
- Anthony M Mancinelli
- Wisconsin National Primate Research Center, University of Wisconsin-Madison, Madison, WI, USA
| | - Jonathan M Vichich
- Wisconsin National Primate Research Center, University of Wisconsin-Madison, Madison, WI, USA
| | - Alexandra D Zinnen
- Wisconsin National Primate Research Center, University of Wisconsin-Madison, Madison, WI, USA
| | - Anna Marie Hugon
- Department of Pathology and Laboratory Medicine, University of Wisconsin-Madison, Madison, WI, USA
| | - Viktoriya Bondarenko
- Wisconsin National Primate Research Center, University of Wisconsin-Madison, Madison, WI, USA
| | - Jeanette M Metzger
- Wisconsin National Primate Research Center, University of Wisconsin-Madison, Madison, WI, USA
| | - Heather A Simmons
- Wisconsin National Primate Research Center, University of Wisconsin-Madison, Madison, WI, USA
| | - Thaddeus G Golos
- Wisconsin National Primate Research Center, University of Wisconsin-Madison, Madison, WI, USA
- Department of Comparative Biosciences, University of Wisconsin-Madison, Madison, WI, USA
- Department of Obstetrics and Gynecology, University of Wisconsin-Madison, Madison, WI, USA
| | - Marina E Emborg
- Wisconsin National Primate Research Center, University of Wisconsin-Madison, Madison, WI, USA
- Department of Medical Physics, University of Wisconsin-Madison, Madison, WI, USA
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Quereda JJ, Morón-García A, Palacios-Gorba C, Dessaux C, García-del Portillo F, Pucciarelli MG, Ortega AD. Pathogenicity and virulence of Listeria monocytogenes: A trip from environmental to medical microbiology. Virulence 2021; 12:2509-2545. [PMID: 34612177 PMCID: PMC8496543 DOI: 10.1080/21505594.2021.1975526] [Citation(s) in RCA: 90] [Impact Index Per Article: 22.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/04/2020] [Revised: 08/23/2021] [Accepted: 08/25/2021] [Indexed: 01/02/2023] Open
Abstract
Listeria monocytogenes is a saprophytic gram-positive bacterium, and an opportunistic foodborne pathogen that can produce listeriosis in humans and animals. It has evolved an exceptional ability to adapt to stress conditions encountered in different environments, resulting in a ubiquitous distribution. Because some food preservation methods and disinfection protocols in food-processing environments cannot efficiently prevent contaminations, L. monocytogenes constitutes a threat to human health and a challenge to food safety. In the host, Listeria colonizes the gastrointestinal tract, crosses the intestinal barrier, and disseminates through the blood to target organs. In immunocompromised individuals, the elderly, and pregnant women, the pathogen can cross the blood-brain and placental barriers, leading to neurolisteriosis and materno-fetal listeriosis. Molecular and cell biology studies of infection have proven L. monocytogenes to be a versatile pathogen that deploys unique strategies to invade different cell types, survive and move inside the eukaryotic host cell, and spread from cell to cell. Here, we present the multifaceted Listeria life cycle from a comprehensive perspective. We discuss genetic features of pathogenic Listeria species, analyze factors involved in food contamination, and review bacterial strategies to tolerate stresses encountered both during food processing and along the host's gastrointestinal tract. Then we dissect host-pathogen interactions underlying listerial pathogenesis in mammals from a cell biology and systemic point of view. Finally, we summarize the epidemiology, pathophysiology, and clinical features of listeriosis in humans and animals. This work aims to gather information from different fields crucial for a comprehensive understanding of the pathogenesis of L. monocytogenes.
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Affiliation(s)
- Juan J. Quereda
- Departamento de Producción y Sanidad Animal, Salud Pública Veterinaria y Ciencia y Tecnología de los Alimentos, Facultad de Veterinaria, Universidad Cardenal Herrera-CEU, CEU Universities. Valencia, Spain
| | - Alvaro Morón-García
- Departamento de Biología Celular. Facultad de Ciencias Biológicas, Universidad Complutense de Madrid. Madrid, Spain
| | - Carla Palacios-Gorba
- Departamento de Producción y Sanidad Animal, Salud Pública Veterinaria y Ciencia y Tecnología de los Alimentos, Facultad de Veterinaria, Universidad Cardenal Herrera-CEU, CEU Universities. Valencia, Spain
| | - Charlotte Dessaux
- Departamento de Biotecnología Microbiana, Centro Nacional de Biotecnología (CNB)- Consejo Superior De Investigaciones Científicas (CSIC), Madrid, Spain
| | - Francisco García-del Portillo
- Departamento de Biotecnología Microbiana, Centro Nacional de Biotecnología (CNB)- Consejo Superior De Investigaciones Científicas (CSIC), Madrid, Spain
| | - M. Graciela Pucciarelli
- Departamento de Biotecnología Microbiana, Centro Nacional de Biotecnología (CNB)- Consejo Superior De Investigaciones Científicas (CSIC), Madrid, Spain
- Centro de Biología Molecular ‘Severo Ochoa’. Departamento de Biología Molecular, Facultad de Ciencias, Universidad Autónoma de Madrid. Madrid, Spain
| | - Alvaro D. Ortega
- Departamento de Biología Celular. Facultad de Ciencias Biológicas, Universidad Complutense de Madrid. Madrid, Spain
- Departamento de Biotecnología Microbiana, Centro Nacional de Biotecnología (CNB)- Consejo Superior De Investigaciones Científicas (CSIC), Madrid, Spain
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mDia1 Assembles a Linear F-Actin Coat at Membrane Invaginations To Drive Listeria monocytogenes Cell-to-Cell Spreading. mBio 2021; 12:e0293921. [PMID: 34781738 PMCID: PMC8593688 DOI: 10.1128/mbio.02939-21] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/24/2022] Open
Abstract
Direct cell-to-cell spreading of Listeria monocytogenes requires the bacteria to induce actin-based finger-like membrane protrusions in donor host cells that are endocytosed through caveolin-rich membrane invaginations by adjacent receiving cells. An actin shell surrounds these endocytic sites; however, its structure, composition, and functional significance remain elusive. Here, we show that the formin mDia1, but surprisingly not the Arp2/3 complex, is enriched at the membrane invaginations generated by L. monocytogenes during HeLa and Jeg-3 cell infections. Electron microscopy reveals a band of linear actin filaments that run along the longitudinal axis of the invagination membrane. Mechanistically, mDia1 expression is vital for the assembly of this F-actin shell. mDia1 is also required for the recruitment of Filamin A, a caveola-associated F-actin cross-linking protein, and caveolin-1 to the invaginations. Importantly, mixed-cell infection assays show that optimal caveolin-based L. monocytogenes cell-to-cell spreading correlates with the formation of the linear actin filament-containing shell by mDia1. IMPORTANCE Listeria monocytogenes spreads from one cell to another to colonize tissues. This cell-to-cell movement requires the propulsive force of an actin-rich comet tail behind the advancing bacterium, which ultimately distends the host plasma membrane into a slender bacterium-containing membrane protrusion. These membrane protrusions induce a corresponding invagination in the membrane of the adjacent host cell. The host cell that receives the protrusion utilizes caveolin-based endocytosis to internalize the structures, and filamentous actin lines these membrane invaginations. Here, we set out to determine the structure and function of this filamentous actin "shell." We demonstrate that the formin mDia1, but not the Arp2/3 complex, localizes to the invaginations. Morphologically, we show that this actin is organized into linear arrays and not branched dendritic networks. Mechanistically, we show that the actin shell is assembled by mDia1 and that mDia1 is required for efficient cell-to-cell transfer of L. monocytogenes.
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Ireton K, Mortuza R, Gyanwali GC, Gianfelice A, Hussain M. Role of internalin proteins in the pathogenesis of Listeria monocytogenes. Mol Microbiol 2021; 116:1407-1419. [PMID: 34704304 DOI: 10.1111/mmi.14836] [Citation(s) in RCA: 30] [Impact Index Per Article: 7.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/19/2021] [Accepted: 10/22/2021] [Indexed: 12/15/2022]
Abstract
Listeria monocytogenes is a food-borne bacterium that causes gastroenteritis, meningitis, or abortion. L. monocytogenes induces its internalization (entry) into human cells and either spreads laterally in tissues or transcytoses to traverse anatomical barriers. In this review, we discuss mechanisms by which five structurally related proteins of the "internalin" family of L. monocytogenes (InlA, InlB, InlC, InlF, and InlP) interact with distinct host receptors to promote infection of human cells and/or crossing of the intestinal, blood-brain, or placental barriers. We focus on recent results demonstrating that the internalin proteins InlA, InlB, and InlC exploit exocytic pathways to stimulate transcytosis, entry, or cell-to-cell spread, respectively. We also discuss evidence that InlA-mediated transcytosis contributes to traversal of the intestinal barrier, whereas InlF promotes entry into endothelial cells to breach the blood-brain barrier. InlB also facilitates the crossing of the blood-brain barrier, but does so by extending the longevity of infected monocytes that may subsequently act as a "Trojan horse" to transfer bacteria to the brain. InlA, InlB, and InlP each contribute to fetoplacental infection by targeting syncytiotrophoblast or cytotrophoblast layers of the placenta. This work highlights the diverse functions of internalins and the complex mechanisms by which these structurally related proteins contribute to disease.
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Affiliation(s)
- Keith Ireton
- Department of Microbiology and Immunology, University of Otago, Dunedin, New Zealand
| | - Roman Mortuza
- Department of Microbiology and Immunology, University of Otago, Dunedin, New Zealand
| | | | - Antonella Gianfelice
- Department of Microbiology and Immunology, University of Otago, Dunedin, New Zealand
| | - Mazhar Hussain
- Department of Biochemistry, University of Otago, Dunedin, New Zealand
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Kelliher JL, Grunenwald CM, Abrahams RR, Daanen ME, Lew CI, Rose WE, Sauer JD. PASTA kinase-dependent control of peptidoglycan synthesis via ReoM is required for cell wall stress responses, cytosolic survival, and virulence in Listeria monocytogenes. PLoS Pathog 2021; 17:e1009881. [PMID: 34624065 PMCID: PMC8528326 DOI: 10.1371/journal.ppat.1009881] [Citation(s) in RCA: 15] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/03/2021] [Revised: 10/20/2021] [Accepted: 09/27/2021] [Indexed: 02/01/2023] Open
Abstract
Pathogenic bacteria rely on protein phosphorylation to adapt quickly to stress, including that imposed by the host during infection. Penicillin-binding protein and serine/threonine-associated (PASTA) kinases are signal transduction systems that sense cell wall integrity and modulate multiple facets of bacterial physiology in response to cell envelope stress. The PASTA kinase in the cytosolic pathogen Listeria monocytogenes, PrkA, is required for cell wall stress responses, cytosolic survival, and virulence, yet its substrates and downstream signaling pathways remain incompletely defined. We combined orthogonal phosphoproteomic and genetic analyses in the presence of a β-lactam antibiotic to define PrkA phosphotargets and pathways modulated by PrkA. These analyses synergistically highlighted ReoM, which was recently identified as a PrkA target that influences peptidoglycan (PG) synthesis, as an important phosphosubstrate during cell wall stress. We find that deletion of reoM restores cell wall stress sensitivities and cytosolic survival defects of a ΔprkA mutant to nearly wild-type levels. While a ΔprkA mutant is defective for PG synthesis during cell wall stress, a double ΔreoM ΔprkA mutant synthesizes PG at rates similar to wild type. In a mouse model of systemic listeriosis, deletion of reoM in a ΔprkA background almost fully restored virulence to wild-type levels. However, loss of reoM alone also resulted in attenuated virulence, suggesting ReoM is critical at some points during pathogenesis. Finally, we demonstrate that the PASTA kinase/ReoM cell wall stress response pathway is conserved in a related pathogen, methicillin-resistant Staphylococcus aureus. Taken together, our phosphoproteomic analysis provides a comprehensive overview of the PASTA kinase targets of an important model pathogen and suggests that a critical role of PrkA in vivo is modulating PG synthesis through regulation of ReoM to facilitate cytosolic survival and virulence. Many antibiotics target bacterial cell wall biosynthesis, justifying continued study of this process and the ways bacteria respond to cell wall insults during infection. Penicillin-binding protein and serine/threonine-associated (PASTA) kinases are master regulators of cell wall stress responses in bacteria and are conserved in several major pathogens, including Listeria monocytogenes, Staphylococcus aureus, and Mycobacterium tuberculosis. We previously showed that the PASTA kinase in L. monocytogenes, PrkA, is essential for the response to cell wall stress and for virulence. In this work, we combined proteomic and genetic approaches to identify PrkA substrates in L. monocytogenes. We show that regulation of one candidate from both screens, ReoM, increases synthesis of the cell wall component peptidoglycan and that this regulation is required for pathogenesis. We also demonstrate that the PASTA kinase-ReoM pathway regulates cell wall stress responses in another significant pathogen, methicillin-resistant S. aureus. Additionally, we uncover a PrkA-independent role for ReoM in vivo in L. monocytogenes, suggesting a need for nuanced modulation of peptidoglycan synthesis during infection. Cumulatively, this study provides new insight into how bacterial pathogens control cell wall synthesis during infection.
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Affiliation(s)
- Jessica L. Kelliher
- Department of Medical Microbiology and Immunology, University of Wisconsin-Madison, Madison, Wisconsin, United States of America
| | - Caroline M. Grunenwald
- Department of Medical Microbiology and Immunology, University of Wisconsin-Madison, Madison, Wisconsin, United States of America
| | - Rhiannon R. Abrahams
- Department of Medical Microbiology and Immunology, University of Wisconsin-Madison, Madison, Wisconsin, United States of America
| | - McKenzie E. Daanen
- Department of Medical Microbiology and Immunology, University of Wisconsin-Madison, Madison, Wisconsin, United States of America
| | - Cassandra I. Lew
- School of Pharmacy, University of Wisconsin-Madison, Madison, Wisconsin, United States of America
| | - Warren E. Rose
- School of Pharmacy, University of Wisconsin-Madison, Madison, Wisconsin, United States of America
| | - John-Demian Sauer
- Department of Medical Microbiology and Immunology, University of Wisconsin-Madison, Madison, Wisconsin, United States of America
- * E-mail:
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Abstract
Bacteria have developed a large array of motility mechanisms to exploit available resources and environments. These mechanisms can be broadly classified into swimming in aqueous media and movement over solid surfaces. Swimming motility involves either the rotation of rigid helical filaments through the external medium or gyration of the cell body in response to the rotation of internal filaments. On surfaces, bacteria swarm collectively in a thin layer of fluid powered by the rotation of rigid helical filaments, they twitch by assembling and disassembling type IV pili, they glide by driving adhesins along tracks fixed to the cell surface and, finally, non-motile cells slide over surfaces in response to outward forces due to colony growth. Recent technological advances, especially in cryo-electron microscopy, have greatly improved our knowledge of the molecular machinery that powers the various forms of bacterial motility. In this Review, we describe the current understanding of the physical and molecular mechanisms that allow bacteria to move around.
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Howell LM, Forbes NS. Bacteria-based immune therapies for cancer treatment. Semin Cancer Biol 2021; 86:1163-1178. [PMID: 34547442 DOI: 10.1016/j.semcancer.2021.09.006] [Citation(s) in RCA: 17] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/14/2021] [Revised: 09/03/2021] [Accepted: 09/12/2021] [Indexed: 12/23/2022]
Abstract
Engineered bacterial therapies that target the tumor immune landscape offer a new class of cancer immunotherapy. Salmonella enterica and Listeria monocytogenes are two species of bacteria that have been engineered to specifically target tumors and serve as delivery vessels for immunotherapies. Therapeutic bacteria have been engineered to deliver cytokines, gene silencing shRNA, and tumor associated antigens that increase immune activation. Bacterial therapies stimulate both the innate and adaptive immune system, change the immune dynamics of the tumor microenvironment, and offer unique strategies for targeting tumors. Bacteria have innate adjuvant properties, which enable both the delivered molecules and the bacteria themselves to stimulate immune responses. Bacterial immunotherapies that deliver cytokines and tumor-associated antigens have demonstrated clinical efficacy. Harnessing the diverse set of mechanisms that Salmonella and Listeria use to alter the tumor-immune landscape has the potential to generate many new and effective immunotherapies.
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Affiliation(s)
- Lars M Howell
- Department of Chemical Engineering, University of Massachusetts, Amherst, United States
| | - Neil S Forbes
- Department of Chemical Engineering, University of Massachusetts, Amherst, United States.
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Shahid AD, Lu Y, Iqbal MA, Lin L, Huang S, Jiang X, Chen S. Listeria monocytogenes crosses blood brain barrier through Rho GTPases induced migration of macrophages and inflammatory interleukin expression. Microb Pathog 2021; 159:105143. [PMID: 34400281 DOI: 10.1016/j.micpath.2021.105143] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/17/2021] [Revised: 08/09/2021] [Accepted: 08/12/2021] [Indexed: 10/20/2022]
Abstract
Listeria monocytogenes crossing the blood-brain barrier in the form of "Trojan Horse" is of great significance for the establishment of bacterial encephalitis and meningitis. Induction of cell migration and crossing the blood-brain barrier is very important to understand the Listeria pathogenesis. The Rho GTPases family is considered a key factor in regulating cell migration. This study was designed to investigate the expression of Rho GTPases and their effect on the behavior of cell migration and the stimulation of immune factors. Selective Rho GTPases were investigated by real-time PCR and Western blot. Among these, the expression of RhoA was significantly increased following the infection of Listeria monocytogenes in macrophages. Further, we found that RhoA improves the migration of macrophages and expression of IL-1β, IL-6, and TNF-α. The expression of IL-1β, IL-6 and TNF-α possibly facilitates the migration and adhesion of macrophages to cross the blood-brain barrier. This study provides preliminary ground to investigate the detailed mechanism of Listeria monocytogenes crossing the blood-brain barrier.
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Affiliation(s)
| | - Ye Lu
- School of Medicine, Jiangsu University, Zhenjiang, 212013, China; Department of Clinical Laboratory, Yixing People's Hospital, Affiliated Jiangsu University, Wuxi, 214200, China
| | | | - Lin Lin
- School of Medicine, Jiangsu University, Zhenjiang, 212013, China
| | - Shuang Huang
- School of Medicine, Jiangsu University, Zhenjiang, 212013, China
| | - Xugan Jiang
- School of Medicine, Jiangsu University, Zhenjiang, 212013, China
| | - Shengxia Chen
- School of Medicine, Jiangsu University, Zhenjiang, 212013, China.
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Hofbauer Cells Spread Listeria monocytogenes among Placental Cells and Undergo Pro-Inflammatory Reprogramming while Retaining Production of Tolerogenic Factors. mBio 2021; 12:e0184921. [PMID: 34399615 PMCID: PMC8406333 DOI: 10.1128/mbio.01849-21] [Citation(s) in RCA: 14] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022] Open
Abstract
Pregnant women are highly susceptible to infection by the bacterial pathogen Listeria monocytogenes, leading to miscarriage, premature birth, and neonatal infection. L. monocytogenes is thought to breach the placental barrier by infecting trophoblasts at the maternal/fetal interface. However, the fate of L. monocytogenes within chorionic villi and how infection reaches the fetus are unsettled. Hofbauer cells (HBCs) are fetal placental macrophages and the only leukocytes residing in healthy chorionic villi, forming a last immune barrier protecting fetal blood from infection. Little is known about the HBCs’ antimicrobial responses to pathogens. Here, we studied L. monocytogenes interaction with human primary HBCs. Remarkably, despite their M2 anti-inflammatory phenotype at basal state, HBCs phagocytose and kill non-pathogenic bacteria like Listeria innocua and display low susceptibility to infection by L. monocytogenes. However, L. monocytogenes can exploit HBCs to spread to surrounding placental cells. Transcriptomic analyses by RNA sequencing revealed that HBCs undergo pro-inflammatory reprogramming upon L. monocytogenes infection, similarly to macrophages stimulated by the potent M1-polarizing agents lipopolysaccharide (LPS)/interferon gamma (IFN-γ). Infected HBCs also express pro-inflammatory chemokines known to promote placental infiltration by maternal leukocytes. However, HBCs maintain the expression of a collection of tolerogenic genes and secretion of tolerogenic cytokines, consistent with their tissue homeostatic role in prevention of fetal rejection. In conclusion, we propose a previously unrecognized model in which HBCs promote the spreading of L. monocytogenes among placental cells and transition to a pro-inflammatory state likely to favor innate immune responses, while maintaining the expression of tolerogenic factors known to prevent maternal anti-fetal adaptive immunity.
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Halsey CR, Glover RC, Thomason MK, Reniere ML. The redox-responsive transcriptional regulator Rex represses fermentative metabolism and is required for Listeria monocytogenes pathogenesis. PLoS Pathog 2021; 17:e1009379. [PMID: 34398937 PMCID: PMC8389512 DOI: 10.1371/journal.ppat.1009379] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/08/2021] [Revised: 08/26/2021] [Accepted: 07/27/2021] [Indexed: 11/30/2022] Open
Abstract
The Gram-positive bacterium Listeria monocytogenes is the causative agent of the foodborne disease listeriosis, one of the deadliest bacterial infections known. In order to cause disease, L. monocytogenes must properly coordinate its metabolic and virulence programs in response to rapidly changing environments within the host. However, the mechanisms by which L. monocytogenes senses and adapts to the many stressors encountered as it transits through the gastrointestinal (GI) tract and disseminates to peripheral organs are not well understood. In this study, we investigated the role of the redox-responsive transcriptional regulator Rex in L. monocytogenes growth and pathogenesis. Rex is a conserved canonical transcriptional repressor that monitors the intracellular redox state of the cell by sensing the ratio of reduced and oxidized nicotinamide adenine dinucleotides (NADH and NAD+, respectively). Here, we demonstrated that L. monocytogenes Rex represses fermentative metabolism and is therefore required for optimal growth in the presence of oxygen. We also show that in vitro, Rex represses the production of virulence factors required for survival and invasion of the GI tract, as a strain lacking rex was more resistant to acidified bile and invaded host cells better than wild type. Consistent with these results, Rex was dispensable for colonizing the GI tract and disseminating to peripheral organs in an oral listeriosis model of infection. However, Rex-dependent regulation was required for colonizing the spleen and liver, and L. monocytogenes lacking the Rex repressor were nearly sterilized from the gallbladder. Taken together, these results demonstrated that Rex functions as a repressor of fermentative metabolism and suggests a role for Rex-dependent regulation in L. monocytogenes pathogenesis. Importantly, the gallbladder is the bacterial reservoir during listeriosis, and our data suggest redox sensing and Rex-dependent regulation are necessary for bacterial survival and replication in this organ. Listeriosis is a foodborne illness caused by Listeria monocytogenes and is one of the deadliest bacterial infections known, with a mortality rate of up to 30%. Following ingestion of contaminated food, L. monocytogenes disseminates from the gastrointestinal (GI) tract to peripheral organs, including the spleen, liver, and gallbladder. In this work, we investigated the role of the redox-responsive regulator Rex in L. monocytogenes growth and pathogenesis. We demonstrated that alleviation of Rex repression coordinates expression of genes necessary in the GI tract during infection, including fermentative metabolism, bile resistance, and invasion of host cells. Accordingly, Rex was dispensable for colonizing the GI tract of mice during an oral listeriosis infection. Interestingly, Rex-dependent regulation was required for bacterial replication in the spleen, liver, and gallbladder. Taken together, our results demonstrate that Rex-mediated redox sensing and transcriptional regulation are important for L. monocytogenes metabolic adaptation and virulence.
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Affiliation(s)
- Cortney R. Halsey
- Department of Microbiology, University of Washington School of Medicine, Seattle, Washington, United States of America
| | - Rochelle C. Glover
- Department of Microbiology, University of Washington School of Medicine, Seattle, Washington, United States of America
| | - Maureen K. Thomason
- Department of Microbiology, University of Washington School of Medicine, Seattle, Washington, United States of America
| | - Michelle L. Reniere
- Department of Microbiology, University of Washington School of Medicine, Seattle, Washington, United States of America
- * E-mail:
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50
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Halladin DK, Ortega FE, Ng KM, Footer MJ, Mitić NS, Malkov SN, Gopinathan A, Huang KC, Theriot JA. Entropy-driven translocation of disordered proteins through the Gram-positive bacterial cell wall. Nat Microbiol 2021; 6:1055-1065. [PMID: 34326523 DOI: 10.1038/s41564-021-00942-8] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/17/2020] [Accepted: 06/28/2021] [Indexed: 02/07/2023]
Abstract
In Gram-positive bacteria, a thick cross-linked cell wall separates the membrane from the extracellular space. Some surface-exposed proteins, such as the Listeria monocytogenes actin nucleation-promoting factor ActA, remain associated with the bacterial membrane but somehow thread through tens of nanometres of cell wall to expose their amino terminus to the exterior. Here, we report that entropy enables the translocation of disordered transmembrane proteins through the Gram-positive cell wall. We build a physical model, which predicts that the entropic constraint imposed by a thin periplasm is sufficient to drive the translocation of an intrinsically disordered protein such as ActA across a porous barrier similar to a peptidoglycan cell wall. We experimentally validate our model and show that ActA translocation depends on the cell-envelope dimensions and disordered-protein length, and that translocation is reversible. We also show that disordered regions of eukaryotic proteins can translocate Gram-positive cell walls via entropy. We propose that entropic forces are sufficient to drive the translocation of specific proteins to the outer surface.
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Affiliation(s)
- David K Halladin
- Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA, USA.,Department of Biochemistry and Howard Hughes Medical Institute, Stanford University School of Medicine, Stanford, CA, USA
| | - Fabian E Ortega
- Department of Biochemistry and Howard Hughes Medical Institute, Stanford University School of Medicine, Stanford, CA, USA
| | - Katharine M Ng
- Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA, USA.,Department of Bioengineering, Stanford University, Stanford, CA, USA
| | - Matthew J Footer
- Department of Biochemistry and Howard Hughes Medical Institute, Stanford University School of Medicine, Stanford, CA, USA.,Department of Biology and Howard Hughes Medical Institute, University of Washington, Seattle, WA, USA
| | - Nenad S Mitić
- Faculty of Mathematics, University of Belgrade, Belgrade, Serbia
| | - Saša N Malkov
- Faculty of Mathematics, University of Belgrade, Belgrade, Serbia
| | - Ajay Gopinathan
- Department of Physics, University of California, Merced, CA, USA
| | - Kerwyn Casey Huang
- Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA, USA. .,Department of Bioengineering, Stanford University, Stanford, CA, USA. .,Chan Zuckerberg Biohub, San Francisco, CA, USA.
| | - Julie A Theriot
- Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA, USA. .,Department of Biochemistry and Howard Hughes Medical Institute, Stanford University School of Medicine, Stanford, CA, USA. .,Department of Biology and Howard Hughes Medical Institute, University of Washington, Seattle, WA, USA.
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