During the chronic stage of Schistosoma infection, the female lays fertile eggs, triggering a strong anti‐parasitic type 2 helper T‐cell (Th2) immune response. It is unclear how this Th2 response ...gradually declines even though the worms live for years and continue to produce eggs. Here, we show that Schistosoma mansoni downregulates Th2 differentiation in an antigen‐presenting cell‐independent manner, by modulating the Th2‐specific transcriptional program. Adult schistosomes secrete miRNA‐harboring extracellular vesicles that are internalized by Th cells in vitro. Schistosomal miRNAs are found also in T helper cells isolated from Peyer's patches and mesenteric lymph nodes of infected mice. In T helper cells, the schistosomal miR‐10 targets MAP3K7 and consequently downmodulates NF‐κB activity, a critical transcription factor for Th2 differentiation and function. Our results explain, at least partially, how schistosomes tune down the Th2 response, and provide further insight into the reciprocal geographic distribution between high prevalence of parasitic infections and immune disorders such as allergy. Furthermore, this worm‐host crosstalk mechanism can be harnessed to develop diagnostic and therapeutic approaches for human schistosomiasis and Th2‐associated diseases.
Synopsis
During chronic Schistosoma infection a strong anti‐parasitic Th2‐type immune response is triggered. The parasite counteracts this by releasing extracellular vesicles that contain miRNAs that modulate Th2 differentiation.
Schistosomes preferentially interfere with Th2‐specific differentiation pathways.
Schistosomal miRNAs secreted via extracellular vesicles taken up by T helper cells block Th2 differentiation.
The schistosomal miR‐10 targets MAP3K7 and inhibits NF‐κB activity, essential for Th2 differentiation.
Parasite‐driven silencing of the Th2 pathway may explain the chronicity of schistosomal infection.
During chronic Schistosoma infection a strong anti‐parasitic Th2‐type immune response is triggered. The parasite counteracts this by releasing extracellular vesicles that contain miRNAs that modulate Th2 differentiation.
Malaria is the most serious mosquito‐borne parasitic disease, caused mainly by the intracellular parasite Plasmodium falciparum. The parasite invades human red blood cells and releases extracellular ...vesicles (EVs) to alter its host responses. It becomes clear that EVs are generally composed of sub‐populations. Seeking to identify EV subpopulations, we subject malaria‐derived EVs to size‐separation analysis, using asymmetric flow field‐flow fractionation. Multi‐technique analysis reveals surprising characteristics: we identify two distinct EV subpopulations differing in size and protein content. Small EVs are enriched in complement‐system proteins and large EVs in proteasome subunits. We then measure the membrane fusion abilities of each subpopulation with three types of host cellular membranes: plasma, late and early endosome. Remarkably, small EVs fuse to early endosome liposomes at significantly greater levels than large EVs. Atomic force microscope imaging combined with machine‐learning methods further emphasizes the difference in biophysical properties between the two subpopulations. These results shed light on the sophisticated mechanism by which malaria parasites utilize EV subpopulations as a communication tool to target different cellular destinations or host systems.
Synopsis
Plasmodium falciparum invades human red blood cells and releases two extracellular vesicle subsets secreted by infected cells. These EV subpopulations harbor different protein cargo and have specific mechanical membrane properties, suggesting distinct host cell targets.
Two distinct subsets of malaria‐derived EVs with different sizes are identified using asymmetric flow field‐flow fractionation.
Small EVs are rich in complement system proteins, whereas large EVs contain 20S proteasome subunits.
Small EVs are more efficient in fusing under endosomal conditions as compared to the large subset.
The EV subpopulations possess distinct membrane mechanical properties, suggesting different lipid compositions.
Plasmodium falciparum invades human red blood cells and releases two extracellular vesicle subsets secreted by infected cells. These EV subpopulations harbor different protein cargo and have specific mechanical membrane properties, suggesting distinct host cell targets.
Vesicular transport is a means of communication. While cells can communicate with each other via secretion of extracellular vesicles, less is known regarding organelle‐to organelle communication, ...particularly in the case of mitochondria. Mitochondria are responsible for the production of energy and for essential metabolic pathways in the cell, as well as fundamental processes such as apoptosis and aging. Here, we show that functional mitochondria isolated from Saccharomyces cerevisiae release vesicles, independent of the fission machinery. We isolate these mitochondrial‐derived vesicles (MDVs) and find that they are relatively uniform in size, of about 100 nm, and carry selective protein cargo enriched for ATP synthase subunits. Remarkably, we further find that these MDVs harbor a functional ATP synthase complex. We demonstrate that these vesicles have a membrane potential, produce ATP, and seem to fuse with naive mitochondria. Our findings reveal a possible delivery mechanism of ATP‐producing vesicles, which can potentially regenerate ATP‐deficient mitochondria and may participate in organelle‐to‐organelle communication.
Synopsis
This work isolates mitochondrial‐derived vesicles from yeast and reveals a possible delivery mechanism of ATP‐producing vesicles. MDV‐encapsulated ATP synthase complexes can potentially regenerate ATP‐deficient mitochondria and may participate in organelle‐to‐organelle communication.
Selective mitochondrial proteins are loaded onto mitochondrial‐derived vesicles (MDVs).
MDVs have a membrane potential and carry a functional F1F0 ATP synthase complex with the capability to produce ATP.
MDVs acquire mitochondrial functions and may possibly transfer this ability from one organelle to the other.
MDVs enable the export of proteins from the mitochondria into the cellular environment.
This work isolates mitochondrial‐derived vesicles from yeast and reveals a possible delivery mechanism of ATP producing vesicles. MDV‐encapsulated ATP synthase complexes can potentially regenerate ATP deficient mitochondria and may participate in organelle‐to‐organelle communication.
Pathogens can release extracellular vesicles (EVs) for cell–cell communication and host modulation. EVs from Plasmodium falciparum, the deadliest malaria parasite species, can transfer drug ...resistance genes between parasites. EVs from late‐stage parasite‐infected RBC (iRBC‐EVs) are immunostimulatory and affect endothelial cell permeability, but little is known about EVs from early stage iRBC. We detected the parasite virulence factor PfEMP1, which is responsible for iRBC adherence and a major contributor to disease severity, in EVs, only up to 12‐hr post‐RBC invasion. Furthermore, using PfEMP1 transport knockout parasites, we determined that EVs originated from inside the iRBC rather than the iRBC surface. Proteomic analysis detected 101 parasite and 178 human proteins in iRBC‐EVs. Primary human monocytes stimulated with iRBC‐EVs released low levels of inflammatory cytokines and showed transcriptomic changes. Stimulation with iRBC‐EVs from PfEMP1 knockout parasites induced more gene expression changes and affected pathways involved in defence response, stress response, and response to cytokines, suggesting a novel function of PfEMP1 when present in EVs. We show for the first time the presence of PfEMP1 in early stage P. falciparum iRBC‐EVs and the effects of these EVs on primary human monocytes, uncovering a new mechanism of potential parasite pathogenesis and host interaction.
Extracellular vesicles from Plasmodium falciparum‐infected red blood cells were found to contain PfEMP1, a key parasite virulence factor.
PfEMP1 was only present in vesicles during the first 12 hours of infection.
Vesicles induced transcriptional changes in human monocytes, which were partially dependent on PfEMP1, suggesting a novel mechanism of host modulation by the parasite.
Genetic plasticity of prokaryotic microbial communities is largely dependent on the ongoing exchange of genetic determinants by Horizontal Gene Transfer (HGT). HGT events allow beneficial genetic ...transitions to occur throughout microbial life, thus promoting adaptation to changing environmental conditions. Here, the significance of secreted vesicles in mediating HGT between microorganisms is discussed, while focusing on the benefits gained by vesicle‐mediated gene delivery and its occurrence under different environmental cues. The potential use of secreted DNA‐harboring vesicles as a mechanism of currently unresolved HGT events in eukaryotic microbes is further discussed.
Tuberculosis remains one of the deadliest infectious diseases worldwide. Mycobacterium tuberculosis (M.tb) has developed various mechanisms to manipulate the human host, in particular by disrupting ...the host phagosome and the immune response. It is becoming evident that secreted extracellular vesicles (EVs) are involved in the dynamic crosstalk between M.tb and the host cells. These vesicles shuttle different cargo components, such as RNA, lipids, and proteins, between cells. In this issue of EMBO Reports, Cheng and Schorey describe a previously unknown EV‐mediated process, regulating M.tb RNA loading into EVs and their internalization by naïve macrophages. They identify the mycobacterial Sec2 secretion system as involved in RNA loading into EVs and show that secreted vesicles contain bacterial RNA that not only promotes IFN‐β production upon entry into target cells, but also leads to M.tb clearance via the activation of the host's RIG‐I/MAVS signaling pathway. Importantly, combined treatment with secreted EVs and antibiotics decreases bacterial load in a mouse model, improving lung pathology compared to treatment with antibiotics alone.
The mycobacterial Sec2 secretion system regulates the loading of bacterial RNA into macrophage‐derived extracellular vesicles. Secreted vesicles with mycobacterial RNA promote IFN‐β production upon entry into target cells, and induce bacterial clearance.
Extracellular vesicles (EVs) are emerging as important mediators of cell–cell communication as well as potential disease biomarkers and drug delivery vehicles. However, the mechanical properties of ...these vesicles are largely unknown, and processes leading to microvesicle‐shedding from the plasma membrane are not well understood. Here an in depth atomic force microscopy force spectroscopy study of the mechanical properties of natural EVs is presented. It is found that several natural vesicles of different origin have a different composition of lipids and proteins, but similar mechanical properties. However, vesicles generated by red blood cells (RBC) at different temperatures/incubation times are different mechanically. Quantifying the lipid content of EVs reveals that their stiffness decreases with the increase in their protein/lipid ratio. Further, by maintaining RBC at “extreme” nonphysiological conditions, the cells are pushed to utilize different vesicle generation pathways. It is found that RBCs can generate protein‐rich soft vesicles, possibly driven by protein aggregation, and low membrane–protein content stiff vesicles, likely driven by cytoskeleton‐induced buckling. Since similar cortical cytoskeleton to that of the RBC exists on the membranes of most mammalian cells, our findings help advancing the understanding of the fundamental process of vesicle generation.
A detailed atomic force microscopy force‐spectroscopy study of natural vesicles reveals that the bending modulus of membranes of extracellular vesicles decreases as their total protein content increases. Generation of protein‐rich soft vesicles by red blood cells is possibly driven by protein aggregation, while the generation of low membrane–protein content stiff vesicles is likely driven by cytoskeleton‐induced buckling.
Mature red blood cells (RBCs) lack internal organelles and canonical defense mechanisms, making them both a fascinating host cell, in general, and an intriguing choice for the deadly malaria parasite ...Plasmodium falciparum (Pf), in particular. Pf, while growing inside its natural host, the human RBC, secretes multipurpose extracellular vesicles (EVs), yet their influence on this essential host cell remains unknown. Here we demonstrate that Pf parasites, cultured in fresh human donor blood, secrete within such EVs assembled and functional 20S proteasome complexes (EV-20S). The EV-20S proteasomes modulate the mechanical properties of naïve human RBCs by remodeling their cytoskeletal network. Furthermore, we identify four degradation targets of the secreted 20S proteasome, the phosphorylated cytoskeletal proteins β-adducin, ankyrin-1, dematin and Epb4.1. Overall, our findings reveal a previously unknown 20S proteasome secretion mechanism employed by the human malaria parasite, which primes RBCs for parasite invasion by altering membrane stiffness, to facilitate malaria parasite growth.
Protozoan pathogens secrete nanosized particles called extracellular vesicles (EVs) to facilitate their survival and chronic infection. Here, we show the inhibition by Plasmodium berghei NK65 blood ...stage‐derived EVs of the proliferative response of CD4+ T cells in response to antigen presentation. Importantly, these results were confirmed in vivo by the capacity of EVs to diminish the ovalbumin‐specific delayed type hypersensitivity response. We identified two proteins associated with EVs, the histamine releasing factor (HRF) and the elongation factor 1α (EF‐1α) that were found to have immunosuppressive activities. Interestingly, in contrast to WT parasites, EVs from genetically HRF‐ and EF‐1α‐deficient parasites failed to inhibit T cell responses in vitro and in vivo. At the level of T cells, we demonstrated that EVs from WT parasites dephosphorylate key molecules (PLCγ1, Akt, and ERK) of the T cell receptor signalling cascade. Remarkably, immunisation with EF‐1α alone or in combination with HRF conferred a long‐lasting antiparasite protection and immune memory. In conclusion, we identified a new mechanism by which P. berghei‐derived EVs exert their immunosuppressive functions by altering T cell responses. The identification of two highly conserved immune suppressive factors offers new conceptual strategies to overcome EV‐mediated immune suppression in malaria‐infected individuals.
Necroptosis is a regulated and inflammatory form of cell death. We, and others, have previously reported that necroptotic cells release extracellular vesicles (EVs). We have found that necroptotic ...EVs are loaded with proteins, including the phosphorylated form of the key necroptosis-executing factor, mixed lineage kinase domain-like kinase (MLKL). However, neither the exact protein composition, nor the impact, of necroptotic EVs have been delineated. To characterize their content, EVs from necroptotic and untreated U937 cells were isolated and analyzed by mass spectrometry-based proteomics. A total of 3337 proteins were identified, sharing a high degree of similarity with exosome proteome databases, and clearly distinguishing necroptotic and control EVs. A total of 352 proteins were significantly upregulated in the necroptotic EVs. Among these were MLKL and caspase-8, as validated by immunoblot. Components of the ESCRTIII machinery and inflammatory signaling were also upregulated in the necroptotic EVs, as well as currently unreported components of vesicle formation and transport, and necroptotic signaling pathways. Moreover, we found that necroptotic EVs can be phagocytosed by macrophages to modulate cytokine and chemokine secretion. Finally, we uncovered that necroptotic EVs contain tumor neoantigens, and are enriched with components of antigen processing and presentation. In summary, our study reveals a new layer of regulation during the early stage of necroptosis, mediated by the secretion of specific EVs that influences the microenvironment and may instigate innate and adaptive immune responses. This study sheds light on new potential players in necroptotic signaling and its related EVs, and uncovers the functional tasks accomplished by the cargo of these necroptotic EVs.
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