Lipopolysaccharide structure impacts the entry kinetics of bacterial outer membrane vesicles into host cells

O’Donoghue, Eloise J.; Sirisaengtaksin, Natalie; Browning, Douglas F.; Bielska, Ewa; Hadis, Mohammed; Fernández-Trillo, Francisco; Alderwick, Luke J.; Jabbari, Sara; Krachler, Anne Marie

doi:10.1371/journal.ppat.1006760

Cited by 62 publications

(92 citation statements)

References 48 publications

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“…2A), in line with previous observations (10,13). Fusion of bacterial MVs to host membranes is involved in transmission of virulence factors in non-phagocytic cells (15) as well as in reprogramming of phagosomes in phagocytic macrophage (32,33). The homogenous binding of DiO-labelled bacterial MVs to the reconstituted lipid membranes mimicking the outer-leaflet of the host cell membrane ( Fig.…”

Section: Discussionsupporting

confidence: 90%

“…2A). The kinetics of binding of bacterial MVs to the host membrane comprising outer leaflet lipids appeared to be fast as reported earlier in different cell lines (15). We next sought to investigate whether the interaction of the bacterial MVs with the host membrane results in changes in the fluidity and dipole potential (Ψd) of the host membrane.…”

Section: Bacterial Membrane Vesicle (Mv) Fusion Increases the Fluiditmentioning

confidence: 62%

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Lipid Specificity of the Fusion of Bacterial Extracellular Vesicles with the Host Membrane

Prince

Tiwari

Mandal

et al. 2019

Preprint

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Bacterial membrane vesicles (MVs) facilitate long-distance delivery of virulence factors crucial for pathogenicity. The entry and trafficking mechanisms of virulence factors inside host cells is recently emerging, however, if bacterial MVs modulate the physicochemical properties of the host lipid membrane remains unknown. Here we quantitatively show that bacterial MV interaction increases the fluidity, dipole potential and elasticity of a biologically relevant multi-component host model membrane. The presence of lipids containing head-groups such as phosphatidylcholine, phosphatidylglycerol and phosphatidylinositol and a moderate acyl chain length of C16 helps the MV interaction. While significant binding of bacterial MVs to the raft-like lipid membranes with phase separated regions of the membrane was observed, however, the elevated levels of cholesterol tend to hinder the interaction of bacterial MVs. We further quantify the change in excess Gibbs free energy of mixing of bacterial MVs with host lipid membranes driving the modulation of host membrane parameters.

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Section: Discussionsupporting

confidence: 90%

Section: Bacterial Membrane Vesicle (Mv) Fusion Increases the Fluiditmentioning

confidence: 62%

Lipid Specificity of the Fusion of Bacterial Extracellular Vesicles with the Host Membrane

Prince

Tiwari

Mandal

et al. 2019

Preprint

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“…To determine if loss of a functional type III secretion system would affect the kinetics of OMV entry into host cells, we used a CCF2-AM based reporter assay as previously described (30). This assay uses a vesicle-targeted beta-lactamase reporter (ClyA-Bla) to measure the fast entry kinetics of OMVs into the host cell cytoplasm in real time.…”

Section: Resultsmentioning

confidence: 99%

“…This assay uses a vesicle-targeted beta-lactamase reporter (ClyA-Bla) to measure the fast entry kinetics of OMVs into the host cell cytoplasm in real time. Epithelial host cells were pre-loaded with the fluorescent beta-lactamase substrate CCF2-AM, which upon enzymatic cleavage shifts from green to blue fluorescence emission (30, 35). The ClyA-Bla reporter was expressed in either EHEC NCTC 12900 wild type cells or the isogenic Δ escN mutant.…”

Section: Resultsmentioning

confidence: 99%

“…EHEC-derived OMVs play important roles during infection: they constitute a decoy protecting bacteria against antimicrobial peptides (24) and have been shown to enter host cells (29). EHEC OMVs traffic a wide range of cargoes into host cells, including lipopolysaccharide (30, 31), Shiga-like toxins, CdtV and Hly (29, 32), which have been shown to reach the target cells in a biologically active form and contribute to pathogenesis (29, 31). However, localization and trafficking of T3SS components by OMVs has not been studied.…”

Section: Introductionmentioning

confidence: 99%

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Bacterial outer membrane vesicles provide an alternative pathway for trafficking of type III secreted effectors into epithelial cells

Sirisaengtaksin

O’Donoghue

Jabbari

et al. 2018

Preprint

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21Outer membrane vesicles (OMVs) are proteo-liposomes universally shed by Gram-negative bacteria. 22 Their secretion is significantly enhanced by the transition into the intra-host milieu and OMVs have been 23 shown to play critical roles during pathogenesis. Enterohemorrhagic Escherichia coli O157 (EHEC), 24 causes diarrheal disease in humans, and soluble toxins including Shiga-like toxins that contribute to 25 disease severity and clinical complications including hemolytic uremic syndrome, have been shown to be 26 OMV associated. In addition to Shiga-like toxins, EHEC produces a type III secretion system (T3SS), and 27 T3SS effectors are associated with colonization and disease severity in vivo. Here, we show that type III 28 secreted substrates including translocators and effectors are incorporated into OMVs independent of type 29 III secretion activity. EHEC strains with non-functional type III secretion systems shed more OMVs and 30 vesicles enter host cells with accelerated kinetics compared to vesicles shed from wild type EHEC. The 31 T3SS effector translocated intimin receptor (Tir) is trafficked from OMVs into host cells and localizes to 32 the membrane. However, its clustering on the host membrane and co-localization with bacterial pedestals 33 is intimin-dependent. We further show that OMV-delivered Tir can cross-complement an effector-34 deficient EHEC strain, demonstrating that OMV-associated effectors reach the host cell in a biologically 35 intact form. Finally, we observe that the non-LEE encoded E3 ubiquitin ligase effector NleL is also 36 trafficked to host cells via OMVs, where it ubiquitinylates its target kinase JNK. Together, these data 37 demonstrate that trafficking of OMV-associated effectors is a novel and T3SS-independent pathway for 38 the delivery of active effectors to host cells. 41Enterohemorrhagic Escherichia coli (EHEC) are a leading cause of food-borne diarrheal disease world-42 wide. In some cases, gastrointestinal symptoms can be complicated by the development of hemolytic 43 uremic syndrome (HUS), which is linked with increased morbidity and mortality (1). Secreted toxins, 44 chiefly Shiga-like toxins, lead to the development of HUS, and the accessory toxins cytolethal distending 45 toxin V (CdtV) and hemolysin (Hly) are thought to contribute to HUS pathology (2, 3). However, many 46 more virulence factors are associated with primary colonization of the gastrointestinal tract, in particular 47 the locus of enterocyte effacement (LEE), which encodes for a type III secretion system (T3SS) and 48 associated effectors (4, 5). The T3SS is a needle-like conduit that translocates effector proteins into the 49 host cell where they manipulate host cellular signaling machinery, induce cytoskeletal rearrangements and 50 modulate immunity to facilitate infection. The T3SS needle is formed by the structural protein EspA (6), 51 and secretion activity is driven by the ATPase EscN (7). EscN directly interacts with T3SS chaperones as 52 well as secreted effectors (8). The T3SS in...

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Bacterial Extracellular Vesicles and the Gut‐Microbiota Brain Axis: Emerging Roles in Communication and Potential as Therapeutics

2021

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Bacterial extracellular vesicles (BEVs) have emerged as candidate signaling vectors for long‐distance interkingdom communication within the gut‐microbiota brain axis. Most bacteria release these nanosized vesicles, capable of signaling to the brain via their abundant protein and small RNA cargo, possibly directly via crossing the blood‐brain barrier. BEVs have been shown to regulate brain gene expression and induce pathology at most stages of neuroinflammation and neurodegeneration, and thus they may play a causal role in diseases such as Alzheimer's, Parkinson's, and depression/anxiety. On the other hand, BEVs have intrinsic therapeutic properties that may be relevant to probiotic therapy and can also be engineered to function as drug delivery vehicles and vaccines. Thus, BEVs may be both a cause of and solution to neuropathological conditions. In this review, current knowledge of the physiological roles of BEVs as well as state of the art pertaining to the development of therapeutic BEVs in the context of the microbiome‐gut‐brain axis are summarized.

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Lipopolysaccharide structure impacts the entry kinetics of bacterial outer membrane vesicles into host cells

Cited by 62 publications

References 48 publications

Lipid Specificity of the Fusion of Bacterial Extracellular Vesicles with the Host Membrane

Lipid Specificity of the Fusion of Bacterial Extracellular Vesicles with the Host Membrane

Bacterial outer membrane vesicles provide an alternative pathway for trafficking of type III secreted effectors into epithelial cells

Bacterial Extracellular Vesicles and the Gut‐Microbiota Brain Axis: Emerging Roles in Communication and Potential as Therapeutics

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