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1.
We present a short insight into the problem of parasitophorous vacuole (PV) formation as a most peculiar kind of cell vacuolization occurring in the course of intracellular development of coccidian pathogens of the genera Eimeria, Isospora, Toxoplasma, Sarcocystis, Cryptosporidium, Epieimeria, and Karyolysus. The review focuses on the morpho-functional diversity of PVs in these parasites. By the present time, the PVs containing different parasite genera and species have been examined to different extent. The membrane of the PV (PVM) obviously derives from the host cell plasmalemma. But soon after parasite penetration, the morphofunctional organization and biochemical composition of the PVM drastically changes: its proteins are selectively excluded and those of the parasite are incorporated. As the result, the PV becomes not fusigenic for lysosomes or any other vacuoles or vesicles, because host cell surface markers necessary for membrane fusion are eliminated from the PVM during parasite invasion.The pattern of the PVs is parasite specific and demonstrates a broad diversity within the same genera and species and even at different stages of the endogenous development. The PV is far from being an indifferent membrane vesicle containing the parasite. Instead, it represents a dynamic system that reflects the innermost events of host-parasite relationships, thus promoting the accomplishing of the parasite life cycle, which, in its turn, is a necessary prerequisite of the parasite eventual survival as a species.  相似文献   

2.
The human malaria parasite Plasmodium falciparum develops in a parasitophorous vacuolar membrane (PVM) within the mature red cell and extensively modifies structural and antigenic properties of this host cell. Recent studies shed significant new, mechanistic perspective on the underlying processes. There is finally, definitive evidence that despite the absence of endocytosis, transmembrane proteins in the host red cell membrane are imported in to the PVM. These are not major erythrocyte proteins but components that reside in detergent resistant membrane (DRM) rafts in red cell membrane and are detected in rafts in the PVM. Disruption of either erythrocyte or vacuolar rafts is detrimental to infection suggesting that raft proteins and lipids are essential for the parasitization of the red cell. On secretory export of parasite proteins: an ER secretory signal (SS) sequence is required for protein secretion to the PV. Proteins carrying an additional plastid targeting sequence (PTS) are also detected in the PV but subsequently delivered to the plastid organelle within the parasite, suggesting that the PTS may have a second function as an endocytic sorting signal. A distinct but yet undefined peptidic motif underlies protein transport across the PVM to the red cell (although all of the published data does not yet fit this model). Further multiple exported proteins transit through secretory 'cleft' structures, suggesting that clefts may be sorting compartments assembled by the parasite in the red cell.  相似文献   

3.
Trypanosoma cruzi, an obligate intracellular protozoan parasite, exhibits developmental regulation of virulence. Although both noninfective epimastigote and infective trypomastigote stages of T. cruzi enter phagocytic cells via the formation of a parasitophorous vacuole (PV), only the latter developmental stages survive ingestion and perpetuate the infection. To determine whether the membrane composition of PV surrounding these different stages might contribute to differences in the outcome of infection, we identified selected membrane constituents by immunofluorescence and intracellular radioiodination, and studied their incorporation into PV. Complement receptors (CR3) are incorporated preferentially into the PV membrane surrounding serum-opsonized epimastigotes but not culture-derived metacyclic trypomastigotes. FcR are not preferentially incorporated into PV membranes unless epimastigotes or culture-derived metacyclic trypomastigotes are opsonized with anti-T. cruzi antibody. PV surrounding either parasite stage contain beta 1 integrins and lysosomal membrane glycoproteins (lgp). These results indicate that the plasma membrane glycoproteins incorporated into the surrounding PV membrane differ depending upon the stage of parasite being internalized, and that these differences reflect, at least in part, selective ligation of cell surface receptors mediating uptake. Furthermore, they imply that although virulent trypomastigote stages may avoid host cell uptake by conventional phagocytic receptors, i.e., CR3 or FcR, they do not escape fusion with an lgp-containing vacuole where they could still be exposed to lysosomal antimicrobial mechanisms.  相似文献   

4.
The obligate intracellular parasite Toxoplasma gondii secretes a vast variety of effector molecules from organelles known as rhoptries (ROPs) and dense granules (GRAs). ROP proteins are released into the cytosol of the host cell where they are directed to the cell nucleus or to the parasitophorous vacuole (PV) membrane. ROPs secrete proteins that enable host cell penetration and vacuole formation by the parasites, as well as hijacking host-immune responses. After invading host cells, T. gondii multiplies within a PV that is maintained by the parasite proteins secreted from GRAs. Most GRA proteins remain within the PV, but some are known to access the host cytosol across the PV membrane, and a few are able to traffic into the host-cell nucleus. These effectors bind to host cell proteins and affect host cell signaling pathways to favor the parasite. Studies on host–pathogen interactions have identified many infection-altered host signal transductions. Notably, the relationship between individual parasite effector molecules and the specific targeting of host-signaling pathways is being elucidated through the advent of forward and reverse genetic strategies. Understanding the complex nature of the host–pathogen interactions underlying how the host-signaling pathway is manipulated by parasite effectors may lead to new molecular biological knowledge and novel therapeutic methods for toxoplasmosis. In this review, we discuss how T. gondii modulates cell signaling pathways in the host to favor its survival.  相似文献   

5.
Ultrastructural observations on the invasion and early development of merozoites (bradyzoites) of Sarcocystis muris in Madin-Darby canine kidney (MDCK) cells are presented. Invading merozoites cause the host cell plasmalemma to invaginate; they form a membrane junction (moving junction) and move into the host cell where they are enclosed in a primary parasitophorous vacuole (PV). Within 30–45 min after becoming intracellular, merozoites begin to vacate the newly established primary PV and move, forming a new membrane junction, into a secondary PV. Simultaneously with the movement of the parasite, the contents of dense granules in the apical part of the merozoites are shed by exocytosis into the lumen of the developing secondary PV. A lamella of the endoplasmic reticulum of the host cell becomes attached to the PV membrane, forming a PV limited by three host cell membranes.  相似文献   

6.
Most Apicomplexa are obligatory intracellular parasites that multiply inside a so-called parasitophorous vacuole (PV) formed upon parasite entry into the host cell. Plasmodium , the agent of malaria and the Apicomplexa most deadly to humans, multiplies in both hepatocytes and erythrocytes in the mammalian host. Although much has been learned on how Apicomplexa parasites invade host cells inside a PV, little is known of how they rupture the PV membrane and egress host cells. Here, we characterize a Plasmodium protein, called LISP1 ( li ver- s pecific p rotein 1), which is specifically involved in parasite egress from hepatocytes. LISP1 is expressed late during parasite development inside hepatocytes and locates at the PV membrane. Intracellular parasites deficient in LISP1 develop into hepatic merozoites, which display normal infectivity to erythrocytes. However, LISP1-deficient liver-stage parasites do not rupture the membrane of the PV and remain trapped inside hepatocytes. LISP1 is the first Plasmodium protein shown by gene targeting to be involved in the lysis of the PV membrane.  相似文献   

7.
Ultrastructural observations on the invasion and early development of merozoites (bradyzoites) of Sarcocystis muris in Madin-Darby canine kidney (MDCK) cells are presented. Invading merozoites cause the host cell plasmalemma to invaginate; they form a membrane junction (moving junction) and move into the host cell where they are enclosed in a primary parasitophorous vacuole (PV). Within 30-45 min after becoming intracellular, merozoites begin to vacate the newly established primary PV and move, forming a new membrane junction, into a secondary PV. Simultaneously with the movement of the parasite, the contents of dense granules in the apical part of the merozoites are shed by exocytosis into the lumen of the developing secondary PV. A lamella of the endoplasmic reticulum of the host cell becomes attached to the PV membrane, forming a PV limited by three host cell membranes.  相似文献   

8.
Data on parasitophorous vacuole (PV) formation in host cells (HC) harbouring different intracellular protozoan parasites have been reviewed and critically analysed, with special reference to the main representatives of the Coccidia. The vacuole membrane (PVM) is the interface between host and parasite, playing a role in nutrient acquisition by the parasite from the HC. The PV phenomenon is regarded as a generalized HC response to the introduction of alien bodies (microorganisms), which eventually reflects the evolutionary established host-parasite relationships at cellular, subcellular and molecular levels. Special attention has been paid to the existing morpho-functional diversity of the PVs within the same genera and species of parasites, and even at different stages of the parasite life cycle. The PVM is generally considered to derive from the HC plasmalemma, whose biochemical composition undergoes significant changes as the intravacuolar parasite grows. The original HC proteins are selectively excluded from the PVM, while those of the parasite are incorporated. As the result, the changed PVM becomes not fusigenic for HC lysosomes. For Toxoplasma gondii and other cyst-forming coccidia (Isospora, Sarcocystis), a definite correlation has been noticed between the extent of rhoptry and dense granule secrets released by a zoite during HC internalization, on the one hand, and the pattern of the PV that forms, on the other one. In T. gondii, tachyzoites, known to discharge abundant secrets, commonly force the development of PVs limited with a single unit membrane and equipped with a tubulovesicular network in the lumen. Unlike, bradyzoites known to be deficient in secretory materials trigger the formation of PVs with a three-membrane lining composed of the changed invaginated plasmalemma in addition to two membranes of endoplasmic reticulum. The two different types of PV harbour, respectively, exoenteric and enteric stages of T. gondii, the latter being confined to the cat intestine only. Unlike, all endogenous stages of the classic intestinal coccidia (Eimeria spp.) develop within PVs limited with a single membrane, with some invaginations extending into the PV lumen. Unusual PV patterns are characteristic of the extracytoplasmic eimerian coccidia (Cryptosporidium, Epieimeria) and adeleid haemogreagarines (Karyolysus). In cyst-forming coccidia, the PVM is actively involved in tissue cyst wall formation, thus protecting the encysted parasites from recognition by the host immune system. All this strongly suggests that the PV is far from being an indifferent membraneous vesicle containing a parasite, but represents a metabolically active compartment in infected cells. Since all the coccidia are obligate intracellular parasites, the mode of their intimate interaction with the HC, largely accomplished via the PV and its membrane, is vital for their survival as biological species.  相似文献   

9.
Single and nested polymerase chain reaction (PCR) assays were developed for the detection of the microsporidian parasite Microsporidium seriolae, which is responsible for emaciation and even death in farmed Japanese yellowtail. Extremely high rDNA identities exist between this parasite and other members of the as yet unclassified genus, necessitating the design of generic, rather than species-specific primer sets. The nested PCR was several orders of magnitude more sensitive than the standard single PCRs, with visible target product amplified from as little as 0.01 pg of parasite DNA (equivalent to that extracted from a single spore). The specificity of the assays was tested against a range of potential host fishes and 6 other microsporidians infecting either fish or the musculature of their hosts. Single PCRs were found to be specific to the target genus, but the nested PCR replicated rDNA from several different microsporidian genera, limiting its utility. This study highlights problems associated with the use of the rRNA gene for PCR assays of certain microsporidians, but nevertheless provides a rapid and sensitive means for the detection of pre-spore forms not possible by current staining methods. Consequently, these assays may be employed for further studies on the portals of entry, migration to the musculature and transmission of this economically important pathogen.  相似文献   

10.
In two variants of Loma salmonae that have specificity for rainbow trout Oncorhynchus mykiss (OA variant) and specificity to brook trout Salvelinus fontinalis (SV variant), the parasitophorous vacuole forms at the onset of sporogony. In the OA variant the merogonial stage is bound by a single plasma membrane in direct contact with host cytoplasm. The parasitophorous vacuole formation is initiated by the host cell surrounding the merogonial stages with endoplasmic reticulum (ER) as occurs in autophagy. Of the two host ER membranes surrounding the parasite, one remains in close association with the plasma membrane of the meront, while the other forms the limiting membrane of the vacuole. The sporogonial stage is bounded by two closely apposed membranes, giving the appearance of a thick electron dense plasmalemma. The observations from this study support the novel hypothesis that this microsporidian uses the intracellular process of autophagy to aid formation of a parasitophorous vacuole. The morphology of the SV‐variant is consistent with that of the OA‐variant suggesting that it uses the same mechanism for development.  相似文献   

11.
Plasmodium sporozoites can enter host cells by two distinct pathways, either through disruption of the plasma membrane followed by parasite transmigration through cells, or by formation of a parasitophorous vacuole (PV) where the parasite further differentiates into a replicative exo-erythrocytic form (EEF). We now provide evidence that following invasion without PV formation, transmigrating Plasmodium falciparum and Plasmodium yoelii sporozoites can partially develop into EEFs inside hepatocarcinoma cell nuclei. We also found that rodent P. yoelii sporozoites can infect both mouse and human hepatocytes, while human P. falciparum sporozoites infect human but not mouse hepatocytes. We have previously reported that the host tetraspanin CD81 is required for PV formation by P. falciparum and P. yoelii sporozoites. Here we show that expression of human CD81 in CD81-knockout mouse hepatocytes is sufficient to confer susceptibility to P. yoelii but not P. falciparum sporozoite infection, showing that the narrow P. falciparum host tropism does not rely on CD81 only. Also, expression of CD81 in a human hepatocarcinoma cell line is sufficient to promote the formation of a PV by P. yoelii but not P. falciparum sporozoites. These results highlight critical differences between P. yoelii and P. falciparum sporozoite infection, and suggest that in addition to CD81, other molecules are specifically required for PV formation during infection by the human malaria parasite.  相似文献   

12.
The intracellular parasite Toxoplasma gondii develops inside a parasitophorous vacuole (PV) that derives from the host cell plasma membrane during invasion. Previous electron micrograph images have shown that the membrane of this vacuole undergoes an extraordinary remodeling with an extensive network of thin tubules and vesicles, the intravacuolar network (IVN), which fills the lumen of the PV. While dense granule proteins, secreted during and after invasion, are the main factors for the organization and tubulation of the network, little is known about the source of lipids used for this remodeling. By selectively labeling host cell or parasite membranes, we uncovered evidence that strongly supports the host cell as the primary, if not exclusive, source of lipids for parasite IVN remodeling. Fluorescence recovery after photobleaching (FRAP) microscopy experiments revealed that lipids are surprisingly dynamic within the parasitophorous vacuole and are continuously exchanged or replenished by the host cell. The results presented here suggest a new model for development of the parasitophorous vacuole whereby the host provides a continuous stream of lipids to support the growth and maturation of the PVM and IVN.  相似文献   

13.
Apicomplexa in mammalian cells: trafficking to the parasitophorous vacuole   总被引:2,自引:0,他引:2  
Most Apicomplexa reside and multiply in the cytoplasm of their host cell, within a parasitophorous vacuole (PV) originating from both parasite and host cell components. Trafficking of parasite-encoded proteins destined to membrane compartments beyond the confine of the parasite plasma membrane is a process that offers a rich territory to explore novel mechanisms of protein–membrane interactions. Here, we focus on the PVs formed by the asexual stages of two pathogens of medical importance, Plasmodium and Toxoplasma . We compare the PVs of both parasites, with a particular emphasis on their evolutionary divergent compartmentalization within the host cell. We also discuss the existence of peculiar export mechanisms and/or sorting determinants that are potentially involved in the post-secretory targeting of parasite proteins to the PV subcompartments.  相似文献   

14.
Microsporidia are intracellular parasites, frequently infecting the planktonic crustacean Daphnia. Questioning the ability to detect and identify microsporidia with conventional microscopic techniques, we applied molecular methods in order to investigate the distribution and co-infection patterns of this parasite among 8 communities of the Daphnia longispina hybrid complex. Eight microsporidian taxa were detected, including 3 that previously had not been characterized genetically. Microsporidian communities from nearby lakes were found to be more similar to each other, apparently due to short distance dispersal via secondary hosts. Moreover, we detected seasonal (but not interannual) changes in microsporidian community structure. With some microsporidia being host-specific, these changes might have resulted from seasonal changes in host taxon and clonal composition. The 2 dominant and closely related parasite species were found mainly in single infections, whereas another pair of related microsporidians was found predominantly in co-infections; suggesting species-level differences in the ability to colonize infected hosts. By applying molecular methods, we were not only able to unambiguously identify parasite taxa but also to reveal multiple infections that otherwise would have remained undetected. Given the increased level of accuracy and sensitivity, we highly recommend molecular approaches in future parasite surveys of Daphnia infections.  相似文献   

15.
Microsporidia are obligate intracellular pathogens which enter host cells by the discharge of a hollow tube through which the sporoplasma is extruded into the host cell. Since this invasion mechanism is very different from common entry strategies, the formation of the parasitophorous vacuole (PV) in Encephalitozoon species is likely to be distinct from known principles. We investigated the origin of the nascent Encephalitozoon cuniculi PV membrane with the aid of fluorescent lipid probes. When Bodipy 500/510-C(12)-HPC-labeled spores were used for infection, the emerging PV membrane was unlabeled, suggesting that sporoplasma-derived lipids do not significantly contribute to the formation of the PV membrane. In contrast, when raft and nonraft microdomains of the host cell plasma membrane were selectively labeled with DiIC(16) and Speedy DiO, both tracers were detectable in the nascent PV membrane shortly after infection, indicating that the bulk lipids of the PV membrane are host cell derived. Time-lapse fluorescence microscopy revealed that the formation of the PV membrane is a fast event (<1.3 s), which occurred simultaneously with the extrusion of the sporoplasma. The portion of the discharged tube which is in contact with the host cell was found to be coated with labeled host cell lipids, which might be an indication for a plasma membrane invagination at the contact site. To investigate the presence of pores in the E. cuniculi PV membrane, we microinjected fluorescent dyes of different sizes into infected host cells. A 0.5-kDa dextran as well as 0.8- to 1.1-kDa peptides could rapidly enter the PV, while a 10-kDa dextran was stably excluded from the PV lumen, indicating that the PV membrane possesses pores with an exclusion size of <10 kDa, which should allow metabolite exchange.  相似文献   

16.
Host cell invasion in the Apicomplexa is unique in its dependency on a parasite actin-driven machinery and in the exclusion of most host cell membrane proteins during parasitophorous vacuole (PV) formation. This exclusion occurs at a junction between host cell and parasite plasma membranes that has been called the moving junction, a circumferential zone which forms at the apical tip of the parasite, moves backward and eventually pinches the PV from the host cell membrane. Despite having been described by electron microscopic studies 30 years ago, the molecular nature of this singular structure is still enigmatic. We have obtained a monoclonal antibody that recognizes the moving junction of invading tachyzoites of Toxoplasma gondii, in a pattern clearly distinct from those described so far for microneme and rhoptry proteins. The protein recognized by this antibody has been affinity purified. Mass spectrometry analysis showed that it is a rhoptry neck protein (RON4), a hypothetical protein with homologues restricted to Apicomplexa. Our findings reveals for the first time the participation of rhoptry neck proteins in moving junction formation and strongly suggest the conservation of this structure at the molecular level among Apicomplexa.  相似文献   

17.
Coxiella burnetii is a gram‐negative intracellular bacterium that forms a large, lysosome‐like parasitophorous vacuole (PV) essential for bacterial replication. Host membrane lipids are critical for the formation and maintenance of this intracellular niche, yet the mechanisms by which Coxiella manipulates host cell lipid metabolism, trafficking and signalling are unknown. Oxysterol‐binding protein‐related protein 1 long (ORP1L) is a mammalian lipid‐binding protein that plays a dual role in cholesterol‐dependent endocytic trafficking as well as interactions between endosomes and the endoplasmic reticulum (ER). We found that ORP1L localized to the Coxiella PV within 12 h of infection through a process requiring the Coxiella Dot/Icm Type 4B secretion system, which secretes effector proteins into the host cell cytoplasm where they manipulate trafficking and signalling pathways. The ORP1L N‐terminal ankyrin repeats were necessary and sufficient for PV localization, indicating that ORP1L binds a PV membrane protein. Strikingly, ORP1L simultaneously co‐localized with the PV and ER, and electron microscopy revealed membrane contact sites between the PV and ER membranes. In ORP1L‐depleted cells, PVs were significantly smaller than PVs from control cells. These data suggest that ORP1L is specifically recruited by the bacteria to the Coxiella PV, where it influences PV membrane dynamics and interactions with the ER.  相似文献   

18.
Parasitophorous vacuoles (PV) that harbour Leishmania parasites acquire some characteristics from fusion with host cell vesicles. Recent studies have shown that PVs acquire and display resident endoplasmic reticulum (ER) molecules. We investigated the importance of ER molecules to PV biology by assessing the consequence of blocking the fusion of PVs with vesicles that originate from the early secretory pathway. This was achieved by targeting the N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs) that mediate the fusion of early secretory vesicles. In the presence of dominant negative variants of sec22b or some of its known cognate partners, D12 and syntaxin 18, PVs failed to distend and harboured fewer parasites. These observations were confirmed in studies in which each of the SNAREs listed above including the intermediate compartment ER/Golgi SNARE, syntaxin 5, was knocked down. The knock-down of these SNARES had little or no measurable effect on the morphology of the ER or on activated secretion even though they resulted in a more significant reduction of PV size. Moreover, the knock-down of the ER/Golgi SNAREs resulted in significant reduction in parasite replication. Taken together, these studies provide further evidence that PVs acquire ER components by fusing with vesicles derived from the early secretory pathway; disruption of this interaction results in inhibition of the development of PVs as well as the limitation of parasite replication within infected cells.  相似文献   

19.
A critical step in infection by the apicomplexan parasite Toxoplasma gondii is the formation of a membrane-bound compartment within which the parasite proliferates. This process relies on a set of secretory organelles that discharge their contents into the host cell upon invasion. Among these organelles, the dense granules are specialized in the export of transmembrane (TM) GRA proteins, which are major components of the mature parasitophorous vacuole (PV) membrane. How eukaryotic pathogens export and sort membrane-bound proteins destined for the host cell is still poorly understood at the mechanistic level. In this study, we show that soluble trafficking of the PV-targeted GRA5 TM protein is parasite specific: when expressed in mammalian cells, GRA5 is targeted to the plasma membrane and behaves as an integral membrane protein with a type I toplogy. We also demonstrate the dual role of the GRA5 N-terminal ectodomain, which is sufficient to prevent membrane integration within the parasite and is essential for both sorting and post-secretory membrane insertion into the vacuolar membrane. These results contrast with the general rule that states that information contained within the cytoplasmic tail and/or the TM domain of integral membrane proteins dictates their cellular localization. They also highlight the diversity of sorting mechanisms that leads to the specialization of secretory processes uniquely adapted to intracellular parasitism.  相似文献   

20.
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