Secreted Effectors Modulating Immune Responses to Toxoplasma gondii

Toxoplasma gondii is an obligate intracellular parasite that chronically infects a third of humans. It can cause life-threatening encephalitis in immune-compromised individuals. Congenital infection also results in blindness and intellectual disabilities. In the intracellular milieu, parasites encounter various immunological effectors that have been shaped to limit parasite infection. Parasites not only have to suppress these anti-parasitic inflammatory responses but also ensure the host organism’s survival until their subsequent transmission. Recent advancements in T. gondii research have revealed a plethora of parasite-secreted proteins that suppress as well as activate immune responses. This mini-review will comprehensively examine each secreted immunomodulatory effector based on the location of their actions. The first section is focused on secreted effectors that localize to the parasitophorous vacuole membrane, the interface between the parasites and the host cytoplasm. Murine hosts are equipped with potent IFNγ-induced immune-related GTPases, and various parasite effectors subvert these to prevent parasite elimination. The second section examines several cytoplasmic and ER effectors, including a recently described function for matrix antigen 1 (MAG1) as a secreted effector. The third section covers the repertoire of nuclear effectors that hijack transcription factors and epigenetic repressors that alter gene expression. The last section focuses on the translocation of dense-granule effectors and effectors in the setting of T. gondii tissue cysts (the bradyzoite parasitophorous vacuole).

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

Infection and Immunity ◽

10.1128/iai.00782-08 ◽

2008 ◽

Vol 76 (11) ◽

pp. 4865-4875 ◽

Cited By ~ 60

Author(s):

Michael E. Rome ◽

Josh R. Beck ◽

Jay M. Turetzky ◽

Paul Webster ◽

Peter J. Bradley

Keyword(s):

Toxoplasma Gondii ◽

Parasitophorous Vacuole ◽

Dense Granule ◽

Obligate Intracellular ◽

Dense Granules ◽

Granule Protein ◽

Host Cytoplasm ◽

C Terminus ◽

Membrane Bound ◽

Host Parasite

ABSTRACT 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.

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Regulated secretion of multi-lamellar vesicles leads to formation of a tubulo-vesicular network in host-cell vacuoles occupied by Toxoplasma gondii

Journal of Cell Science ◽

10.1242/jcs.108.4.1669 ◽

1995 ◽

Vol 108 (4) ◽

pp. 1669-1677 ◽

Cited By ~ 1

Author(s):

L.D. Sibley ◽

I.R. Niesman ◽

S.F. Parmley ◽

M.F. Cesbron-Delauw

Keyword(s):

Toxoplasma Gondii ◽

Host Cell ◽

Soluble Protein ◽

Parasitophorous Vacuole ◽

Dense Granule ◽

Host Cell Invasion ◽

Cell Fractionation ◽

Obligate Intracellular ◽

Dense Granules ◽

Storage Granules

Toxoplasma gondii is an obligate intracellular parasite that actively invades virtually all types of nucleated cells, surviving within a specialized vacuole called the parasitophorous vacuole. Shortly after invasion, the parasite modifies this vacuole by secreting a variety of proteins from electron-dense storage granules. Additionally, the parasite forms a network of membranous tubules within the lumen of the vacuole and connecting with the vacuolar membrane. We have used immunolabeling and cell fractionation to examine the secretion of two dense granule proteins, GRA1 and GRA2, which are involved in formation of the intravacuolar network. Following host-cell invasion, GRA1 was secreted into the lumen of the vacuole as a soluble protein that subsequently became peripherally associated with the network. In addition to being secreted as a soluble protein from dense granules, GRA2 was secreted within multi-lamellar vesicles released from a specialized posterior invagination of the parasite. The multi-lamellar vesicles assemble to form the intravacuolar network, which contains an integral membrane form of GRA2. These findings indicate that Toxoplasma has a highly developed regulated exocytosis pathway that modifies the parasitophorous vacuole by secretion of soluble proteins and by a novel process of membrane secretion.

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The Toxoplasma gondii Dense Granule Protein GRA7 Is Phosphorylated upon Invasion and Forms an Unexpected Association with the Rhoptry Proteins ROP2 and ROP4

Infection and Immunity ◽

10.1128/iai.01667-07 ◽

2008 ◽

Vol 76 (12) ◽

pp. 5853-5861 ◽

Cited By ~ 59

Author(s):

Joe Dan Dunn ◽

Sandeep Ravindran ◽

Seon-Kyeong Kim ◽

John C. Boothroyd

Keyword(s):

Toxoplasma Gondii ◽

Host Cell ◽

Parasitophorous Vacuole ◽

Opportunistic Pathogen ◽

Dense Granule ◽

Host Cells ◽

Obligate Intracellular ◽

Dense Granules ◽

Obligate Intracellular Parasite ◽

Granule Protein

ABSTRACT The obligate intracellular parasite Toxoplasma gondii infects warm-blooded animals throughout the world and is an opportunistic pathogen of humans. As it invades a host cell, Toxoplasma forms a novel organelle, the parasitophorous vacuole, in which it resides during its intracellular development. The parasite modifies the parasitophorous vacuole and its host cell with numerous proteins delivered from rhoptries and dense granules, which are secretory organelles unique to the phylum Apicomplexa. For the majority of these proteins, little is known other than their localization. Here we show that the dense granule protein GRA7 is phosphorylated but only in the presence of host cells. Within 10 min of invasion, GRA7 is present in strand-like structures in the host cytosol that contain rhoptry proteins. GRA7 strands also contain GRA1 and GRA3. Independently of its phosphorylation state, GRA7 associates with the rhoptry proteins ROP2 and ROP4 in infected host cells. This is the first report of interactions between proteins secreted from rhoptries and dense granules.

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A Patatin-Like Protein Protects Toxoplasma gondii from Degradation in a Nitric Oxide-Dependent Manner

Infection and Immunity ◽

10.1128/iai.05543-11 ◽

2011 ◽

Vol 80 (1) ◽

pp. 55-61 ◽

Cited By ~ 37

Author(s):

Crystal M. Tobin ◽

Laura J. Knoll

Keyword(s):

Nitric Oxide ◽

Toxoplasma Gondii ◽

Parasitophorous Vacuole ◽

Dense Granule ◽

Dependent Manner ◽

Activated Macrophages ◽

Content Type ◽

Obligate Intracellular ◽

Exogenous Addition ◽

Inos Inhibitor

ABSTRACTToxoplasma gondiiis an obligate intracellular parasite that uses immune cells to disseminate throughout its host.T. gondiican persist and even slowly replicate in activated host macrophages by reducing the antimicrobial effects of molecules such as nitric oxide (NO). AT. gondiipatatin-like protein called TgPL1 was previously shown to be important for survival in activated macrophages. Here we show that aT. gondiimutant with a deletion of theTgPL1gene (ΔTgPL1) is degraded in activated macrophages. This degradation phenotype is abolished by the removal of NO by the use of an inducible NO synthase (iNOS) inhibitor or iNOS-deficient macrophages. The exogenous addition of NO to macrophages results in reduced parasite growth but not the degradation of ΔTgPL1 parasites. These results suggest that NO is necessary but not sufficient for the degradation of ΔTgPL1 parasites in activated macrophages. While some patatin-like proteins have phospholipase A2(PLA2) activity, recombinant TgPL1 purified fromEscherichia colidoes not have phospholipase activity. This result was not surprising, as TgPL1 contains a G-to-S change at the predicted catalytic serine residue. An epitope-tagged version of TgPL1 partially colocalized with a dense granule protein in the parasitophorous vacuole space. These results may suggest that TgPL1 moves to the parasitophorous vacuole to protect parasites from nitric oxide by an undetermined mechanism.

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Toxoplasma gondii Development of Its Replicative Niche: in Its Host Cell and Beyond

Eukaryotic Cell ◽

10.1128/ec.00081-14 ◽

2014 ◽

Vol 13 (8) ◽

pp. 965-976 ◽

Cited By ~ 32

Author(s):

Ira J. Blader ◽

Anita A. Koshy

Keyword(s):

Toxoplasma Gondii ◽

Host Cell ◽

Host Cells ◽

Content Type ◽

Obligate Intracellular ◽

Cellular Processes ◽

High Prevalence ◽

Life Threatening ◽

Cell Processes ◽

Cellular Pathways

ABSTRACTIntracellular pathogens can replicate efficiently only after they manipulate and modify their host cells to create an environment conducive to replication. While diverse cellular pathways are targeted by different pathogens, metabolism, membrane and cytoskeletal architecture formation, and cell death are the three primary cellular processes that are modified by infections.Toxoplasma gondiiis an obligate intracellular protozoan that infects ∼30% of the world's population and causes severe and life-threatening disease in developing fetuses, in immune-comprised patients, and in certain otherwise healthy individuals who are primarily found in South America. The high prevalence ofToxoplasmain humans is in large part a result of its ability to modulate these three host cell processes. Here, we highlight recent work defining the mechanisms by whichToxoplasmainteracts with these processes. In addition, we hypothesize why some processes are modified not only in the infected host cell but also in neighboring uninfected cells.

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Innate, adaptive, and cell-autonomous immunity against Toxoplasma gondii infection

Experimental & Molecular Medicine ◽

10.1038/s12276-019-0353-9 ◽

2019 ◽

Vol 51 (12) ◽

pp. 1-10 ◽

Cited By ~ 7

Author(s):

Miwa Sasai ◽

Masahiro Yamamoto

Keyword(s):

Immune Response ◽

Toxoplasma Gondii ◽

Immune Responses ◽

Parasitophorous Vacuole ◽

Interferon Γ ◽

Protective Role ◽

Ifn Γ ◽

Acquired Immune Responses ◽

Toxoplasma Gondii Infection ◽

Antimicrobial Immunity

AbstractHosts have been fighting pathogens throughout the evolution of all infectious diseases. Toxoplasma gondii is one of the most common infectious agents in humans but causes only opportunistic infection in healthy individuals. Similar to antimicrobial immunity against other organisms, the immune response against T. gondii activates innate immunity and in turn induces acquired immune responses. After activation of acquired immunity, host immune cells robustly produce the proinflammatory cytokine interferon-γ (IFN-γ), which activates a set of IFN-γ-inducible proteins, including GTPases. IFN-inducible GTPases are essential for cell-autonomous immunity and are specialized for effective clearance and growth inhibition of T. gondii by accumulating in parasitophorous vacuole membranes. Recent studies suggest that the cell-autonomous immune response plays a protective role in host defense against not only T. gondii but also various intracellular bacteria. Moreover, the negative regulatory mechanisms of such strong immune responses are also important for host survival after infection. In this review, we will discuss in detail recent advances in the understanding of host defenses against T. gondii and the roles played by cell-autonomous immune responses.

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RHΔgra17Δnpt1 Strain of Toxoplasma gondii Elicits Protective Immunity Against Acute, Chronic and Congenital Toxoplasmosis in Mice

Microorganisms ◽

10.3390/microorganisms8030352 ◽

2020 ◽

Vol 8 (3) ◽

pp. 352 ◽

Cited By ~ 3

Author(s):

Qin-Li Liang ◽

Li-Xiu Sun ◽

Hany M. Elsheikha ◽

Xue-Zhen Cao ◽

Lan-Bi Nie ◽

...

Keyword(s):

Toxoplasma Gondii ◽

Cellular Response ◽

Protective Efficacy ◽

Interleukin 2 ◽

Congenital Toxoplasmosis ◽

Congenital Infection ◽

Dense Granule ◽

Lethal Challenge ◽

Double Deletion ◽

Brain Cysts

In the present study, a dense granule protein 17 (gra17) and novel putative transporter (npt1) double deletion mutant of Toxoplasma gondii RH strain was engineered. The protective efficacy of vaccination using RHΔgra17Δnpt1 tachyzoites against acute, chronic, and congenital toxoplasmosis was studied in a mouse model. Immunization using RHΔgra17Δnpt1 induced a strong humoral and cellular response, as indicated by the increased levels of anti-T. gondii specific IgG, interleukin 2 (IL-2), IL-10, IL-12, and interferon-gamma (IFN-γ). Vaccinated mice were protected against a lethal challenge dose (103 tachyzoites) of wild-type homologous (RH) strain and heterologous (PYS and TgC7) strains, as well as against 100 tissue cysts or oocysts of Pru strain. Vaccination also conferred protection against chronic infection with 10 tissue cysts or oocysts of Pru strain, where the numbers of brain cysts in the vaccinated mice were significantly reduced compared to those detected in the control (unvaccinated + infected) mice. In addition, vaccination protected against congenital infection with 10 T. gondii Pru oocysts (administered orally on day 5 of gestation) as shown by the increased litter size, survival rate and the bodyweight of pups born to vaccinated dams compared to those born to unvaccinated + infected dams. The brain cyst burden of vaccinated dams was significantly lower than that of unvaccinated dams infected with oocysts. Our data show that T. gondii RHΔgra17Δnpt1 mutant strain can protect mice against acute, chronic, and congenital toxoplasmosis by balancing inflammatory response with immunogenicity.

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Coimmunoprecipitation with MYR1 Identifies Three Additional Proteins within the Toxoplasma gondii Parasitophorous Vacuole Required for Translocation of Dense Granule Effectors into Host Cells

mSphere ◽

10.1128/msphere.00858-19 ◽

2020 ◽

Vol 5 (1) ◽

Cited By ~ 9

Author(s):

Alicja M. Cygan ◽

Terence C. Theisen ◽

Alma G. Mendoza ◽

Nicole D. Marino ◽

Michael W. Panas ◽

...

Keyword(s):

Toxoplasma Gondii ◽

Protein Translocation ◽

Affinity Purification ◽

Parasitophorous Vacuole ◽

Dense Granule ◽

Effector Proteins ◽

Host Cells ◽

Content Type ◽

Cyclin E1 ◽

Infected Cells

ABSTRACT Toxoplasma gondii is a ubiquitous, intracellular protozoan that extensively modifies infected host cells through secreted effector proteins. Many such effectors must be translocated across the parasitophorous vacuole (PV), in which the parasites replicate, ultimately ending up in the host cytosol or nucleus. This translocation has previously been shown to be dependent on five parasite proteins: MYR1, MYR2, MYR3, ROP17, and ASP5. We report here the identification of several MYR1-interacting and novel PV-localized proteins via affinity purification of MYR1, including TGGT1_211460 (dubbed MYR4), TGGT1_204340 (dubbed GRA54), and TGGT1_270320 (PPM3C). Further, we show that three of the MYR1-interacting proteins, GRA44, GRA45, and MYR4, are essential for the translocation of the Toxoplasma effector protein GRA16 and for the upregulation of human c-Myc and cyclin E1 in infected cells. GRA44 and GRA45 contain ASP5 processing motifs, but like MYR1, processing at these sites appears to be nonessential for their role in protein translocation. These results expand our understanding of the mechanism of effector translocation in Toxoplasma and indicate that the process is highly complex and dependent on at least eight discrete proteins. IMPORTANCE Toxoplasma is an extremely successful intracellular parasite and important human pathogen. Upon infection of a new cell, Toxoplasma establishes a replicative vacuole and translocates parasite effectors across this vacuole to function from the host cytosol and nucleus. These effectors play a key role in parasite virulence. The work reported here newly identifies three parasite proteins that are necessary for protein translocation into the host cell. These results significantly increase our knowledge of the molecular players involved in protein translocation in Toxoplasma-infected cells and provide additional potential drug targets.

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In vivo expression and distribution of dense granule protein 7 (GRA7) in the exoenteric (tachyzoite, bradyzoite) and enteric (coccidian) forms of Toxoplasma gondii

Parasitology ◽

10.1017/s0031182099004692 ◽

1999 ◽

Vol 119 (3) ◽

pp. 259-265 ◽

Cited By ~ 26

Author(s):

D. J. P. FERGUSON ◽

D. JACOBS ◽

E. SAMAN ◽

J-F. DUBREMETZ ◽

S. E. WRIGHT

Keyword(s):

Toxoplasma Gondii ◽

Staining Pattern ◽

Parasitophorous Vacuole ◽

Dense Granule ◽

Parasite Development ◽

Double Labelling ◽

Granule Protein ◽

Specific Variation ◽

Strong Staining

The in vivo expression and distribution of the dense granule protein GRA7 was examined in both the exoenteric (tachyzoite and bradyzoite) and enteric (coccidian) forms of Toxoplasma gondii by immunocytochemistry. There was strong staining of GRA7 in granules within all the infectious stages (tachyzoite, bradyzoite, merozoite and sporozoite). During tachyzoite development, GRA7 was secreted and was associated with the parasitophorous vacuole. In contrast, although there was staining of granules within the bradyzoites of more mature cysts, there appeared to be little staining of the tissue cyst wall or host cell. The apparent stage-specific variation in secretion of GRA7 between tachyzoites and bradyzoites was confirmed by double labelling using stage-specific markers (SAG1 and BAG1). In the enteric forms in the cat gut there was strong labelling of the PV containing early asexual and sexual stages and staining of a few granules in the apical cytoplasm of the merozoite. The positive enteric staining pattern differentiates GRA7 from the other GRA proteins (GRA1–6) which were absent in the merozoites and enteric stages. The staining pattern of GRA7 with strong staining during tachyzoite and enteric development and reduced staining in the tissue cysts is similar to that seen for NTPases. The function of GRA7 is unknown but it is unique among the dense granule proteins in being expressed in all the infectious forms of T. gondii which would point to a basic role in the vacuolar adaptations required for active parasite development.

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