scholarly journals Listeria monocytogenescell-to-cell spread in epithelia is heterogeneous and dominated by rare pioneer bacteria

2018 ◽  
Author(s):  
Fabian E. Ortega ◽  
Elena F. Koslover ◽  
Julie A. Theriot
Keyword(s):  
Bacterial Infection ◽  
Statistical Modeling ◽  
Population Level ◽  
Quantitative Model ◽  
Epithelial Monolayer ◽  
Spreading Behavior ◽  
Intercellular Spread ◽  
Cell Spread ◽  
Intracellular Motility

ABSTRACTL. monocytogeneshijacks host actin to promote its intracellular motility and intercellular spread. WhileL. monocytogenesvirulence hinges on cell-to-cell spread, little is known about the dynamics of bacterial spread in epithelia at a population level. Here, we use live microscopy and statistical modeling to demonstrate thatL. monocytogenescell-to-cell spread proceeds anisotropically in an epithelial monolayer in culture. We show that boundaries of infection foci are irregular and dominated by rare pioneer bacteria that spread farther than the rest. We extend our quantitative model for bacterial spread to show that heterogeneous spreading behavior can improve the chances of creating a persistentL. monocytogenesinfection in an actively extruding epithelium. Thus, our results indicate thatL. monocytogenescell-to-cell spread is heterogeneous, and that rare pioneer bacteria determine the frontier of infection foci and may promote bacterial infection persistence in dynamic epithelia.

scholarly journals Listeria monocytogenes cell-to-cell spread in epithelia is heterogeneous and dominated by rare pioneer bacteria

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Fabian E Ortega ◽  
Elena F Koslover ◽  
Julie A Theriot
Keyword(s):  
Listeria Monocytogenes ◽  
Bacterial Infection ◽  
Population Level ◽  
Quantitative Model ◽  
Editorial Note ◽  
Editorial Process ◽  
Spreading Behavior ◽  
Intercellular Spread ◽  
Cell Spread ◽  
Intracellular Motility

Listeria monocytogenes hijacks host actin to promote its intracellular motility and intercellular spread. While L. monocytogenes virulence hinges on cell-to-cell spread, little is known about the dynamics of bacterial spread in epithelia at a population level. Here, we use live microscopy and statistical modeling to demonstrate that L. monocytogenes cell-to-cell spread proceeds anisotropically in an epithelial monolayer in culture. We show that boundaries of infection foci are irregular and dominated by rare pioneer bacteria that spread farther than the rest. We extend our quantitative model for bacterial spread to show that heterogeneous spreading behavior can improve the chances of creating a persistent L. monocytogenes infection in an actively extruding epithelium. Thus, our results indicate that L. monocytogenes cell-to-cell spread is heterogeneous, and that rare pioneer bacteria determine the frontier of infection foci and may promote bacterial infection persistence in dynamic epithelia.Editorial note: This article has been through an editorial process in which the authors decide how to respond to the issues raised during peer review. The Reviewing Editor's assessment is that all the issues have been addressed (<xref ref-type="decision-letter" rid="SA1">see decision letter</xref>).

scholarly journals Key Role for DsbA in Cell-to-Cell Spread ofShigella flexneri, Permitting Secretion of Ipa Proteins into Interepithelial Protrusions

2000 ◽  
Vol 68 (11) ◽  
pp. 6449-6456 ◽  
Author(s):  
Jun Yu ◽  
Bryn Edwards-Jones ◽  
Olivier Neyrolles ◽  
J. Simon Kroll
Keyword(s):  
Confocal Microscopy ◽  
Disulfide Bond ◽  
Active Site ◽  
Shigella Flexneri ◽  
Wild Type ◽  
Pathogenic Potential ◽  
Previous Demonstration ◽  
Intercellular Spread ◽  
Cell Spread

ABSTRACT DsbA, a disulfide bond catalyst, is necessary for realization of the pathogenic potential of Shigella flexneri. Sh42, a mutant strain differing from wild-type M90TS solely because it expresses nonfunctional DsbA33G (substitution for 33C at the active site), secreted less IpaB and IpaC than M90TS in response to various stimuli in vitro. A kinetic study demonstrated that Sh42 responded more slowly to Congo red than M90TS. By modulating relative concentrations of functional and nonfunctional DsbA within bacteria, functional enzyme has been shown to be necessary for intercellular spread. By confocal microscopy, M90TS dividing in protrusions was shown to secrete Ipa proteins from the septation furrow, anticipating lysis of protrusions, while Sh42 showed minimal Ipa secretion in this location. In the light of a previous demonstration that DsbA is not necessary for entry of epithelial cells, we conclude that a role in virulence of this disulfide bond catalyst lies in facilitating secretion of Ipa proteins specifically within epithelial protrusions, in turn allowing cell-to-cell spread of S. flexneri.

scholarly journals A plant receptor-like kinase promotes cell-to-cell spread of RNAi and is targeted by a virus

2017 ◽  
Author(s):  
Tabata Rosas-Diaz ◽  
Dan Zhang ◽  
Pengfei Fan ◽  
Liping Wang ◽  
Xue Ding ◽  
...  
Keyword(s):  
Cell Movement ◽  
Plant Cells ◽  
Tomato Yellow Leaf ◽  
Yellow Leaf ◽  
Systemic Spread ◽  
Receptor Like Kinase ◽  
Intercellular Spread ◽  
Leaf Curl Virus ◽  
Redundant Role ◽  
Cell Spread

ABSTRACTRNA interference (RNAi) in plants can move from cell to cell, allowing for systemic spread of an anti-viral immune response. How this cell-to-cell spread of silencing is regulated is currently unknown. Here, we describe that the C4 protein from Tomato yellow leaf curl virus has the ability to inhibit the intercellular spread of RNAi. Using this viral protein as a probe, we have identified the receptor-like kinase (RLK) BARELY ANY MERISTEM 1 (BAM1) as a positive regulator of the cell-to-cell movement of RNAi, and determined that BAM1 and its closest homologue, BAM2, play a redundant role in this process. C4 interacts with the intracellular domain of BAM1 and BAM2 at the plasma membrane and plasmodesmata, the cytoplasmic connections between plant cells, interfering with the function of these RLKs in the cell-to-cell spread of RNAi. Our results identify BAM1 as an element required for the cell-to-cell spread of RNAi and highlight that signalling components have been co-opted to play multiple functions in plants.

scholarly journals An in situ high-throughput screen identifies inhibitors of intracellularBurkholderia pseudomalleiwith therapeutic efficacy

2019 ◽  
Vol 116 (37) ◽  
pp. 18597-18606 ◽  
Author(s):  
Philip L. Bulterys ◽  
Isabelle J. Toesca ◽  
Michael H. Norris ◽  
Jeffrey P. Maloy ◽  
Sorel T. Fitz-Gibbon ◽  
...  
Keyword(s):  
Small Molecules ◽  
High Throughput ◽  
Type Vi Secretion System ◽  
Type Vi Secretion ◽  
Tier 1 ◽  
Attractive Therapeutic Target ◽  
Human Infections ◽  
Intercellular Spread ◽  
Cell Spread

Burkholderia pseudomallei(Bp) andBurkholderia mallei(Bm) are Tier-1 Select Agents that cause melioidosis and glanders, respectively. These are highly lethal human infections with limited therapeutic options. Intercellular spread is a hallmark ofBurkholderiapathogenesis, and its prominent ties to virulence make it an attractive therapeutic target. We developed a high-throughput cell-based phenotypic assay and screened ∼220,000 small molecules for their ability to disrupt intercellular spread byBurkholderia thailandensis, a closely related BSL-2 surrogate. We identified 268 hits, and cross-species validation found 32 hits that also disrupt intercellular spread byBpand/orBm. Among these were a fluoroquinolone analog, which we named burkfloxacin (BFX), which potently inhibits growth of intracellularBurkholderia, and flucytosine (5-FC), an FDA-approved antifungal drug. We found that 5-FC blocks the intracellular life cycle at the point of type VI secretion system 5 (T6SS-5)-mediated cell–cell spread. Bacterial conversion of 5-FC to 5-fluorouracil and subsequently to fluorouridine monophosphate is required for potent and selective activity against intracellularBurkholderia. In a murine model of fulminant respiratory melioidosis, treatment with BFX or 5-FC was significantly more effective than ceftazidime, the current antibiotic of choice, for improving survival and decreasing bacterial counts in major organs. Our results demonstrate the utility of cell-based phenotypic screening for Select Agent drug discovery and warrant the advancement of BFX and 5-FC as candidate therapeutics for melioidosis in humans.

Incorporating Population-Level Variability in Orthopedic Biomechanical Analysis: A Review

2014 ◽  
Vol 136 (2) ◽  
Author(s):  
Jeffrey E. Bischoff ◽  
Yifei Dai ◽  
Casey Goodlett ◽  
Brad Davis ◽  
Marc Bandi
Keyword(s):  
Statistical Modeling ◽  
Principal Component ◽  
Population Level ◽  
Biomechanical Analysis ◽  
Patient Specific ◽  
Data Sets ◽  
Joint Function ◽  
Tissue Properties ◽  
Joint Kinetics ◽  
Biomechanical Models

Effectively addressing population-level variability within orthopedic analyses requires robust data sets that span the target population and can be greatly facilitated by statistical methods for incorporating such data into functional biomechanical models. Data sets continue to be disseminated that include not just anatomical information but also key mechanical data including tissue or joint stiffness, gait patterns, and other inputs relevant to analysis of joint function across a range of anatomies and physiologies. Statistical modeling can be used to establish correlations between a variety of structural and functional biometrics rooted in these data and to quantify how these correlations change from health to disease and, finally, to joint reconstruction or other clinical intervention. Principal component analysis provides a basis for effectively and efficiently integrating variability in anatomy, tissue properties, joint kinetics, and kinematics into mechanistic models of joint function. With such models, bioengineers are able to study the effects of variability on biomechanical performance, not just on a patient-specific basis but in a way that may be predictive of a larger patient population. The goal of this paper is to demonstrate the broad use of statistical modeling within orthopedics and to discuss ways to continue to leverage these techniques to improve biomechanical understanding of orthopedic systems across populations.

scholarly journals Listeria monocytogenes exploits host exocytosis to promote cell-to-cell spread

2020 ◽  
Vol 117 (7) ◽  
pp. 3789-3796 ◽  
Author(s):  
Georgina C. Dowd ◽  
Roman Mortuza ◽  
Manmeet Bhalla ◽  
Hoan Van Ngo ◽  
Yang Li ◽  
...  
Keyword(s):  
Listeria Monocytogenes ◽  
Actin Polymerization ◽  
Intracellular Pathogen ◽  
Filamentous Actin ◽  
Microscopy Imaging ◽  
Genetic Studies ◽  
Exocyst Complex ◽  
Host Plasma Membrane ◽  
Intercellular Spread ◽  
Cell Spread

The facultative intracellular pathogen Listeria monocytogenes uses an actin-based motility process to spread within human tissues. Filamentous actin from the human cell forms a tail behind bacteria, propelling microbes through the cytoplasm. Motile bacteria remodel the host plasma membrane into protrusions that are internalized by neighboring cells. A critical unresolved question is whether generation of protrusions by Listeria involves stimulation of host processes apart from actin polymerization. Here we demonstrate that efficient protrusion formation in polarized epithelial cells involves bacterial subversion of host exocytosis. Confocal microscopy imaging indicated that exocytosis is up-regulated in protrusions of Listeria in a manner that depends on the host exocyst complex. Depletion of components of the exocyst complex by RNA interference inhibited the formation of Listeria protrusions and subsequent cell-to-cell spread of bacteria. Additional genetic studies indicated important roles for the exocyst regulators Rab8 and Rab11 in bacterial protrusion formation and spread. The secreted Listeria virulence factor InlC associated with the exocyst component Exo70 and mediated the recruitment of Exo70 to bacterial protrusions. Depletion of exocyst proteins reduced the length of Listeria protrusions, suggesting that the exocyst complex promotes protrusion elongation. Collectively, these results demonstrate that Listeria exploits host exocytosis to stimulate intercellular spread of bacteria.

Myosin light chain kinase plays an essential role in S. flexneri dissemination

2000 ◽  
Vol 113 (19) ◽  
pp. 3375-3386
Author(s):  
M. Rathman ◽  
P. de Lanerolle ◽  
H. Ohayon ◽  
P. Gounon ◽  
P. Sansonetti
Keyword(s):  
Host Cell ◽  
Light Chain ◽  
Myosin Light Chain Kinase ◽  
Myosin Light Chain ◽  
Shigella Flexneri ◽  
Bacterial Pathogen ◽  
Inhibitory Effect ◽  
Junctional Complex ◽  
Intercellular Spread ◽  
Cell Spread

Shigella flexneri, the causitive agent of bacillary dysentery, has been shown to disseminate in colonic epithelial cells via protrusions that extend from infected cells and are endocytosed by adjacent cells. This phenomenon occurs in the region of the eukaryotic cell's adherens junctions and is inhibited by pharmacological reagents or host cell mutations that completely disrupt the junctional complex. In this study, inhibitors of the myosin light chain kinase (MLCK) were shown to dramatically decrease intercellular spread of S. flexneri but to have no inhibitory effect on bacterial entry, multiplication or actin-based motility within the host cell. Furthermore, cell-to-cell spread of Listeria monocytogenes, another bacterial pathogen that uses an actin-based mechanism to move within the eukaryotic cytoplasm and to spread from cell to cell, was not affected by the MLCK inhibitors, indicating that (1) the inhibition of S. flexneri cell-to-cell spread in treated cells is not due to a complete break down of cell-cell contacts, which was subsequently confirmed by confocal microscopy, and (2) MLCK plays a role in a S. flexneri-specific mechanism of dissemination. Myosin has been shown to play a role in a variety of membrane-based phenomena. The work presented here suggests that activation of this molecule via phosphorylation by MLCK, at the very least participates in the formation of the bacteria-containing protrusion, and could also contribute to the endocytosis of this structure by neighboring cells.

scholarly journals A virus-targeted plant receptor-like kinase promotes cell-to-cell spread of RNAi

2018 ◽  
Vol 115 (6) ◽  
pp. 1388-1393 ◽  
Author(s):  
Tabata Rosas-Diaz ◽  
Dan Zhang ◽  
Pengfei Fan ◽  
Liping Wang ◽  
Xue Ding ◽  
...  
Keyword(s):  
Cell Movement ◽  
Tomato Yellow Leaf ◽  
Antiviral Immune Response ◽  
Systemic Spread ◽  
Receptor Like Kinase ◽  
Intercellular Spread ◽  
Leaf Curl Virus ◽  
Signaling Components ◽  
Redundant Role ◽  
Cell Spread

RNA interference (RNAi) in plants can move from cell to cell, allowing for systemic spread of an antiviral immune response. How this cell-to-cell spread of silencing is regulated is currently unknown. Here, we describe that the C4 protein from Tomato yellow leaf curl virus can inhibit the intercellular spread of RNAi. Using this viral protein as a probe, we have identified the receptor-like kinase (RLK) BARELY ANY MERISTEM 1 (BAM1) as a positive regulator of the cell-to-cell movement of RNAi, and determined that BAM1 and its closest homolog, BAM2, play a redundant role in this process. C4 interacts with the intracellular domain of BAM1 and BAM2 at the plasma membrane and plasmodesmata, the cytoplasmic connections between plant cells, interfering with the function of these RLKs in the cell-to-cell spread of RNAi. Our results identify BAM1 as an element required for the cell-to-cell spread of RNAi and highlight that signaling components have been coopted to play multiple functions in plants.

scholarly journals RECON-Dependent Inflammation in Hepatocytes EnhancesListeria monocytogenesCell-to-Cell Spread

mBio ◽  
2018 ◽  
Vol 9 (3) ◽  
Author(s):  
Adelle P. McFarland ◽  
Thomas P. Burke ◽  
Alexie A. Carletti ◽  
Rochelle C. Glover ◽  
Hannah Tabakh ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Enzymatic Activity ◽  
Host Cells ◽  
Nitric Oxide Production ◽  
Content Type ◽  
Oxide Production ◽  
Cyclic Dinucleotides ◽  
Intercellular Spread ◽  
Oxide Synthase ◽  
Cell Spread

ABSTRACTThe oxidoreductase RECON is a high-affinity cytosolic sensor of bacterium-derived cyclic dinucleotides (CDNs). CDN binding inhibits RECON’s enzymatic activity and subsequently promotes inflammation. In this study, we sought to characterize the effects of RECON on the infection cycle of the intracellular bacteriumListeria monocytogenes, which secretes cyclic di-AMP (c-di-AMP) into the cytosol of infected host cells. Here, we report that during infection of RECON-deficient hepatocytes, which exhibit hyperinflammatory responses,L. monocytogenesexhibits significantly enhanced cell-to-cell spread. Enhanced bacterial spread could not be attributed to alterations in PrfA or ActA, two virulence factors critical for intracellular motility and intercellular spread. Detailed microscopic analyses revealed that in the absence of RECON,L. monocytogenesactin tail lengths were significantly longer and there was a larger number of faster-moving bacteria. Complementation experiments demonstrated that the effects of RECON onL. monocytogenesspread and actin tail lengths were linked to its enzymatic activity. RECON enzyme activity suppresses NF-κB activation and is inhibited by c-di-AMP. Consistent with these previous findings, we found that augmented NF-κB activation in the absence of RECON caused enhancedL. monocytogenescell-to-cell spread and thatL. monocytogenesspread correlated with c-di-AMP secretion. Finally, we discovered that, remarkably, increased NF-κB-dependent inducible nitric oxide synthase expression and nitric oxide production were responsible for promotingL. monocytogenescell-to-cell spread. The work presented here supports a model wherebyL. monocytogenessecretion of c-di-AMP inhibits RECON’s enzymatic activity, drives augmented NF-κB activation and nitric oxide production, and ultimately enhances intercellular spread.IMPORTANCETo date, bacterial CDNs in eukaryotes are solely appreciated for their capacity to activate cytosolic sensing pathways in innate immunity. However, it remains unclear whether pathogens that actively secrete CDNs benefit from this process. Here, we provide evidence that secretion of CDNs leads to enhancement ofL. monocytogenescell-to-cell spread. This is a heretofore-unknown role of these molecules and suggestsL. monocytogenesmay benefit from their secretion in certain contexts. Molecular characterization revealed that, surprisingly, nitric oxide was responsible for the enhanced spread. Pathogens act to prevent nitric oxide production or, likeL. monocytogenes, they have evolved to resist its direct antimicrobial effects. This study provides evidence that intracellular bacterial pathogens not only tolerate nitric oxide, which is inevitably encountered during infection, but can also capitalize on the changes this pleiotropic molecule enacts on the host cell.

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