Apicomplexa in Mammalian Cells: Trafficking to the Parasitophorous Vacuole

Cesbron-Delauw, Marie-France; Gendrin, Claire; Travier, Laetitia; Ruffiot, Pauline; Mercier, Corinne

doi:10.1111/j.1600-0854.2008.00728.x

Cited by 66 publications

(53 citation statements)

References 74 publications

(106 reference statements)

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“…No identifiable ortholog can be discerned even in closely related Conoidasida such as Cryptosporidium or Eimeria or Aconoidasida such as Plasmodium, which do not form tissue cysts (10). Our results suggest that in addition to GRA4 and GRA6 other members of the prominent GRA protein family may also play critical roles in biology underlying successful cyst development and latent infection.…”

Section: Vol 10 2011 Analysis Of Cyst Development and Latent Infectmentioning

confidence: 81%

“…GRA15 plays a significant role in host cell NF-B nuclear translocation and NF-B-mediated transcription and regulates the induction of IL-12 secretion in infected mouse macrophages (54). Various potential biological roles for GRA proteins have been previously proposed and significant studies have been performed on GRA protein secretion, traffic, and interaction biology (10,49). At the tachyzoite stage, most of the GRA proteins associate with the PV membranes, meaning either the PVM and its digitations or the intravacuolar network of membranous tubules (49).…”

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confidence: 99%

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Type II Toxoplasma gondiiKU80Knockout Strains Enable Functional Analysis of Genes Required for Cyst Development and Latent Infection

Fox¹,

Falla²,

Rommereim³

et al. 2011

Eukaryot Cell

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Type II Toxoplasma gondii KU80 knockouts (⌬ku80) deficient in nonhomologous end joining were developed to delete the dominant pathway mediating random integration of targeting episomes. Gene targeting frequency in the type II ⌬ku80 ⌬hxgprt strain measured at the orotate (OPRT) and the uracil (UPRT) phosphoribosyltransferase loci was highly efficient. To assess the potential of the type II ⌬ku80 ⌬hxgprt strain to examine gene function affecting cyst biology and latent stages of infection, we targeted the deletion of four parasite antigen genes (GRA4, GRA6, ROP7, and tgd057) that encode characterized CD8 ؉ T cell epitopes that elicit corresponding antigen-specific CD8 ؉ T cell populations associated with control of infection. Cyst development in these type II mutant strains was not found to be strictly dependent on antigen-specific CD8 ؉ T cell host responses. In contrast, a significant biological role was revealed for the dense granule proteins GRA4 and GRA6 in cyst development since brain tissue cyst burdens were drastically reduced specifically in mutant strains with GRA4 and/or GRA6 deleted. Complementation of the ⌬gra4 and ⌬gra6 mutant strains using a functional allele of the deleted GRA coding region placed under the control of the endogenous UPRT locus was found to significantly restore brain cyst burdens. These results reveal that GRA proteins play a functional role in establishing cyst burdens and latent infection. Collectively, our results suggest that a type II ⌬ku80 ⌬hxgprt genetic background enables a higher-throughput functional analysis of the parasite genome to reveal fundamental aspects of parasite biology controlling virulence, pathogenesis, and transmission.

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Section: Vol 10 2011 Analysis Of Cyst Development and Latent Infectmentioning

confidence: 81%

mentioning

confidence: 99%

Type II Toxoplasma gondiiKU80Knockout Strains Enable Functional Analysis of Genes Required for Cyst Development and Latent Infection

Fox¹,

Falla²,

Rommereim³

et al. 2011

Eukaryot Cell

Self Cite

199

253

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“…Rhoptry proteins have also been shown to associate with the PVM, where ROP2's N terminus and ROP5's C terminus are exposed to the host cell cytosol (2,15). While ROP2 family proteins (including ROP5) were initially thought to be transmembrane proteins, it now appears that they may instead be soluble proteins that are secreted into the host cell and subsequently attach to the cytoplasmic face of the PVM by an unknown mechanism (9,15). GRA and ROP proteins that localize to the PVM have been directly implicated in interacting with their host cells.…”

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confidence: 99%

Intervacuolar Transport and Unique Topology of GRA14, a Novel Dense Granule Protein inToxoplasma gondii

et al. 2008

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Toxoplasma gondii is an obligate intracellular parasite that resides in the cytoplasm of its host in a unique membrane-bound vacuole known as the parasitophorous vacuole (PV). The membrane surrounding the parasite is remodeled by the dense granules, secretory organelles that release an array of proteins into the vacuole and to the PV membrane (PVM). Only a small portion of the protein constituents of the dense granules have been identified, and little is known regarding their roles in infection or how they are trafficked within the infected host cell. In this report, we identify a novel secreted dense granule protein, GRA14, and show that it is targeted to membranous structures within the vacuole known as the intravacuolar network and to the vacuolar membrane surrounding the parasite. We disrupted GRA14 and exploited the knockout strain to show that GRA14 can be transferred between vacuoles in a coinfection experiment with wild-type parasites. We also show that GRA14 has an unexpected topology in the PVM with its C terminus facing the host cytoplasm and its N terminus facing the vacuolar lumen. These findings have important implications both for the trafficking of GRA proteins to their ultimate destinations and for expectations of functional domains of GRA proteins at the host-parasite interface.Capable of infecting essentially any warm-blooded vertebrate, Toxoplasma gondii is one of the most successful pathogens on the planet (20, 39). Toxoplasma infects nearly onethird of the human population and causes potentially fatal disease in immunocompromised individuals and congenitally infected neonates (20). This protozoan parasite also causes ocular disease in immunocompetent individuals who are either congenitally or postnatally infected (46). As an obligate intracellular parasite, Toxoplasma enters the host cell into a nonfusogenic vacuole (the parasitophorous vacuole [PV]), in which the parasite replicates in the cytoplasm of its host. The PV membrane (PVM) is porous to small molecules (less than 1,300 Da) but otherwise serves as a boundary between the host and parasite during its intracellular survival (36).Toxoplasma invasion is mediated by a trio of specialized secretory organelles, named the micronemes, rhoptries, and dense granules, which contribute to the parasite's ability to initiate and sustain infection within its host. The first proteins secreted are from the micronemes, which release molecular adhesins that interact with the parasite's actin-myosin motor to provide the driving force for invasion (24). The rhoptries are then released and help to establish the nascent PV and modulate host cell processes (4). Lastly, proteins from the dense granules that are implicated in the remodeling and maintenance of the PV for intracellular survival are secreted (29).The precise role of dense granule proteins (GRAs) in the T. gondii life cycle is still largely unknown. To date, two groups of GRA proteins have been identified. The first group contains proteins that lack homology to organisms other than closely relat...

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“…However, the contents of the parasite's apical organelles are secreted during invasion, and it is possible that some of this material is also incorporated into the nascent PVM (8). Since the RBC lacks the machinery to synthesize new lipids or proteins as the parasite grows over the next 48 h from 1 to 8 m, the PVM is expanded and modified by the parasite, providing a customized environment for development (5). The PVM is closely associated with the parasite plasma membrane, so it is difficult to isolate the two membranes for direct proteomic and functional analysis, but a number of PVM-associated proteins have been identified by immunofluorescence assays (IFAs) and immunoelectron microscopy (IEM) (30,32,34).…”

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confidence: 99%

Protein Targeting to the Parasitophorous Vacuole Membrane of Plasmodium falciparum

Ekşi

2011

Eukaryot Cell

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Red blood cell (RBC) invasion and parasitophorous vacuole (PV) formation by Plasmodium falciparum are critical for the development and pathogenesis of malaria, a continuing global health problem. Expansion of the PV membrane (PVM) during growth is orchestrated by the parasite. This is particularly important in mature RBCs, which lack internal organelles and no longer actively synthesize membranes. Pfs16, a 16-kDa integral PVM protein expressed by gametocytes, was chosen as a model for studying the trafficking of material from the parasite across the PV space to the PVM. The locations of Pfs16-green fluorescent protein (GFP) reporter proteins containing distinct regions of Pfs16 were tracked from RBC invasion to emergence. Inclusion of the 53 C-terminal amino acids (aa) of Pfs16 to a GFP reporter construct already containing the N-terminal secretory signal sequence was sufficient for targeting to and retention on the PVM. An amino acid motif identified in this region was also found in seven other known PVM proteins. Removal of the 11 C-terminal aa did not affect PVM targeting, but membrane retention was decreased. Additionally, during emergence from the PVM and RBC, native Pfs16 and the full-length Pfs16-GFP reporter protein were found to concentrate on the ends of the gametocyte. Capping was not observed in constructs lacking the amino acids between the N-terminal secretory signal sequence and the transmembrane domain, suggesting that this region, which is not required for PVM targeting, is involved in capping. This is the first report to define the amino acid domains required for targeting to the P. falciparum PVM.

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Apicomplexa in Mammalian Cells: Trafficking to the Parasitophorous Vacuole

Cited by 66 publications

References 74 publications

Type II Toxoplasma gondiiKU80Knockout Strains Enable Functional Analysis of Genes Required for Cyst Development and Latent Infection

Type II Toxoplasma gondiiKU80Knockout Strains Enable Functional Analysis of Genes Required for Cyst Development and Latent Infection

Intervacuolar Transport and Unique Topology of GRA14, a Novel Dense Granule Protein inToxoplasma gondii

Protein Targeting to the Parasitophorous Vacuole Membrane of Plasmodium falciparum

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