Exocytosis ofToxoplasma gondii dense granules into the parasitophorous vacuole after host cell invasion

Leriche, M. A.; Dubremetz, Jean François

doi:10.1007/bf00932560

Cited by 82 publications

(25 citation statements)

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“…ImmunoEM studies indicate that dense granule proteins are likely synthesized on membranebound ribosomes, imported into the ER, and translocated to the Golgi for packaging (Charif et al, 1990;Sibley et al, 1995) (Coppens et al, and CesbronDelauw, unpublished results). Dense granule secretion in Toxoplasma occurs by a process that resembles classical exocytosis with the release of amorphous material into the PV (Leriche and Dubremetz, 1990). Ac- (A) GRA5-HA9 (anti-HA11, 18-nm gold particles) was found concentrated within the same population of dense granules as GRA1 (TG17-43, 12-nm gold particles).…”

Section: Discussionmentioning

confidence: 99%

Transmembrane Insertion of theToxoplasma gondiiGRA5 Protein Occurs after Soluble Secretion into the Host Cell

Lecordier

Mercier

Sibley

et al. 1999

MBoC

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The intracellular parasite Toxoplasma gondii resides within a specialized compartment, the parasitophorous vacuole (PV), that resists fusion with host cell endocytic and lysosomal compartments. The PV is extensively modified by secretion of parasite proteins, including the dense granule protein GRA5 that is specifically targeted to the delimiting membrane of the PV (PVM). We show here that GRA5 is present both in a soluble form and in hydrophobic aggregates. GRA5 is secreted as a soluble form into the PV after which it becomes stably associated with the PVM. Topological studies demonstrated that GRA5 was inserted into the PVM as a transmembrane protein with its N-terminal domain extending into the cytoplasm and its C terminus in the vacuole lumen. Deletion of 8 of the 18 hydrophobic amino acids of the single predicted transmembrane domain resulted in the failure of GRA5 to associate with the PVM; yet it remained correctly packaged in the dense granules and was secreted as a soluble protein into the PV.Collectively, these studies demonstrate that the secretory pathway in Toxoplasma is unusual in two regards; it allows soluble export of proteins containing typical transmembrane domains and provides a mechanism for their insertion into a host cell membrane after secretion from the parasite. INTRODUCTIONAmong the varied lifestyles adopted by intracellular parasites is the residence in host cell vacuoles that do not fuse with lysosomes, a strategy exhibited by the protozoan parasite Toxoplasma and the bacterial pathogens Legionella, Chlamydia, and Mycobacteria (Garcia delPortillo and Finlay, 1995). The Toxoplasma-containing vacuole, called the parasitophorous vacuole (PV) 1 , forms by active penetration of the parasite causing invagination of the plasma membrane bilayer (Suss-Toby et al., 1996). This compartment subsequently resists fusion with endosomes and lysosomes by a mechanism that is poorly understood; however, it is clear that the fate of the vacuole is determined at the time of formation (Sibley et al., 1985;Joiner, 1991;Mordue and Sibley, 1997). As the parasite grows and replicates within the cell, it forms a network of tubular membranes that are continuous with the delimiting membrane of the vacuole (Nichols et al., 1983;Sibley et al., 1995).Secretion of proteins stored in apical organelles (micronemes, rhoptries, and dense granules) is a prominent feature of Toxoplasma invasion and presumably contributes both to the nonfusigenic status of the PV and subsequent nutrient uptake by the parasite (Carruthers and Sibley, 1997). Segregated from the endocytic system and cut off from the extracellular fluid, § Corresponding author. E-mail address: mcesbron@infobiogen. fr. † Present address: Laboratory of Molecular Parasitology, Université Libre de Bruxelles, Rhode Saint Genese, Belgium. 1 Abbreviations used: EM, electron microscopy; FBS, fetal bovine serum; F/T, freeze/thaw; HFF, human foreskin fibroblasts; HSP, high-speed pellet; HSS, high-speed supernatant; LSP, lowspeed pellet; LSS, low-speed supernatant; NP-40...

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Section: Discussionmentioning

confidence: 99%

Transmembrane Insertion of theToxoplasma gondiiGRA5 Protein Occurs after Soluble Secretion into the Host Cell

Lecordier

Mercier

Sibley

et al. 1999

MBoC

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“…Western blots were probed with MAbs using either a 1:50 dilution of tissue culture supernatant or a 1:1,000 to 1:2,000 dilution of ascites fluid in PBS. The T. gondii-specific MAbs used included MAbs against AMA1 (CL22), MIC2 (6D10) (44), ROP2, ROP3, and ROP4 (T34A7) (22), and GRA3 (2H11) (23).…”

Section: Peptide Synthesis Two Peptides Corresponding To Ser-21 Thromentioning

confidence: 99%

Toxoplasma gondii Homologue of Plasmodium Apical Membrane Antigen 1 Is Involved in Invasion of Host Cells

et al. 2000

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Proteins with constitutive or transient localization on the surface of Apicomplexa parasites are of particular interest for their potential role in the invasion of host cells. We describe the identification and characterization of TgAMA1, the Toxoplasma gondii homolog of the Plasmodium apical membrane antigen 1 (AMA1), which has been shown to elicit a protective immune response against merozoites dependent on the correct pairing of its numerous disulfide bonds. TgAMA1 shows between 19% (Plasmodium berghei) and 26% (Plasmodium yoelii) overall identity to the different Plasmodium AMA1 homologs and has a conserved arrangement of 16 cysteine residues and a putative transmembrane domain, indicating a similar architecture. The single-copy TgAMA1 gene is interrupted by seven introns and is transcribed into an mRNA of ϳ3.3 kb. The TgAMA1 protein is produced during intracellular tachyzoite replication and initially localizes to the micronemes, as determined by immunofluorescence assay and immunoelectron microscopy. Upon release of mature tachyzoites, TgAMA1 is found distributed predominantly on the apical end of the parasite surface. A ϳ54-kDa cleavage product of the large ectodomain is continuously released into the medium by extracellular parasites. Mouse antiserum against recombinant TgAMA1 blocked invasion of new host cells by approximately 40%. This and our inability to produce a viable TgAMA1 knock-out mutant indicate that this phylogenetically conserved protein fulfills a key function in the invasion of host cells by extracellular T. gondii tachyzoites.

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“…A sequential secretion of components from secretory organelles begins at the time of tachyzoites adhesion to the host cell membrane and through the release of MIC proteins from micronemes , followed by the penetration of the host cell membrane through secretion of ROP proteins from rhoptries, and with the consecutive internalization within a parasitophorous vacuole (PV). Once inside the PV, GRA proteins from dense granules are released to modify the PV membrane molecular topography and to conform an intravacuolar vesicle-tubular network (Werk, 1985;Sibley et al, 1986;Leriche and Dubremetz, 1990;Mercier et al, 2005). Once tachyzoites have proliferated inside the PV, they leave the host cell following similar dynamic and secretory events as those achieved during invasion, such as motility activation, conoid extrusion and rhoptries secretion (Endo et al, 1982;Nichols et al, 1983;Moudy et al, 2001).…”

Section: Introductionmentioning

confidence: 99%

Induction and regulation of conoid extrusion inToxoplasma gondii

Carmen

Mondragón

González

et al. 2009

Cellular Microbiology

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SummaryCell invasion by the intracellular parasite Toxoplasma gondii occurs through an active process that involves dynamic events, such as gliding motility and conoid extrusion, followed by a sequential secretion from specialized secretory organelles. Increase of intracellular Ca2+ by ionophores induces conoid extrusion, although in an irreversible way, thus limiting the characterization of the regulatory pathways. In this report we studied the effect of different activating conoid conditions to characterize the regulatory mechanisms involved. Exposure of tachyzoites to ethanol, a well-known activator of microneme secretion through the increase of intracellular Ca 2+, induced conoid extrusion without affecting parasite viability nor its in vitro invasive capability, in a process that could be completely reverted and repeatedly reactivated. A temporal relationship between conoid extrusion and microneme secretion was here studied. Under this condition, signal transduction pathways and the precise role of the parasite cytoskeleton were characterized. Our results indicate that phospholipase C, Ca 2+ released through channels sensitive to inositol-3-phosphate and ryanodine, as well as myosin together with actin filaments, but not microtubules, all participate in conoid extrusion. Specific inhibitors for serine-threonine kinases blocked conoid extrusion; in contrast, calmodulin inhibitors did not affect the induction. A regulatory model for conoid activation is here proposed.

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Exocytosis ofToxoplasma gondii dense granules into the parasitophorous vacuole after host cell invasion

Cited by 82 publications

References 9 publications

Transmembrane Insertion of theToxoplasma gondiiGRA5 Protein Occurs after Soluble Secretion into the Host Cell

Transmembrane Insertion of theToxoplasma gondiiGRA5 Protein Occurs after Soluble Secretion into the Host Cell

Toxoplasma gondii Homologue of Plasmodium Apical Membrane Antigen 1 Is Involved in Invasion of Host Cells

Induction and regulation of conoid extrusion inToxoplasma gondii

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