A conserved noncanonical motif in the pseudoactive site of the ROP5 pseudokinase domain mediates its effect on Toxoplasma virulence

J Biol Chem. 2011 Jun 27. [Epub ahead of print]

A conserved noncanonical motif in the pseudoactive site of the ROP5 pseudokinase domain mediates its effect on Toxoplasma virulence.

Reese ML, Boothroyd JC.

SourceStanford University, United States.

Abstract
The ROP5 family is a closely related set of polymorphic pseudokinases that are critical to Toxoplasma's ability to cause disease. ROP5's polymorphisms also make it a major determinant of strain-specific differences in virulence. ROP5 possesses all of the major kinase motifs required for catalysis except for a substitution at the catalytic Asp. We show that this substitution in ROP5's catalytic loop is part of a motif conserved in other pseudokinases of both Toxoplasma and human origin, and that this motif is required for ROP5's full activity in vivo. This suggests evolutionary selection at this site for a biochemical function, rather than simple drift away from catalysis. We present the crystal structures of a virulent isoform of ROP5 both in its ATP-bound and -unbound states and have demonstrated that, in spite of maintaining the canonical ATP-binding motifs, ROP5 binds ATP in a distorted conformation mediated by unusual Mg coordination sites that would not be predicted from the primary sequence. In addition, we have mapped the polymorphisms spread throughout ROP5's primary sequence to two major surfaces, including ROP5's activation segment. This suggests that the pseudoactive site of this class of pseudokinases may have evolved to use the canonical ATP-binding motifs for non-catalytic signaling through allostery.

PMID:21708941[PubMed - as supplied by publisher]

Toxoplasma gondii peroxiredoxin promotes altered macrophage function, caspase-1-dependent IL-1beta secretion enhances parasite replication

Vet Res. 2011 Jun 27;42(1):80. [Epub ahead of print]

Toxoplasma gondii peroxiredoxin promotes altered macrophage function, caspase-1-dependent IL-1beta secretion enhances parasite replication.

Marshall ES, Elsheikha HM, Hakimi MA, Flynn RJ.

Abstract
ABSTRACT: Alternatively activated macrophages (AAM) are a key feature Th2 immunity and have been associated with a variety of roles during helminth infection. The role this cell subset plays in protzoan infection remain relatively unexplored, herein we describe the effects of a redox enzyme (rTgPrx) derived from Toxoplasma gondii on murine macrophage phenotype in vitro. RTgPrx has been previously associated with the maintainence of parasite oxidative balance. Here our experiments show that rTgPrx promotes AAM as indicated by high arginase-1 (arg-1), YM1 and FIZZ expression via both signal transducer and activator of transcription (STAT)6-dependent and -independent mechanisms. Additionally rTgPrx treatment reduced caspase-1 activity and IL-1beta secretion, while simultaneously increasing IL-10 release. Furthermore the in vitro replication of T. gondii (RH strain) was enhanced when macrophages were treated with rTgPrx. This is in contrast with the previously described effects of a Plasmodium berghei ANKA 2-cys-peroxiredoxin that promotes pro-inflammatory cytokine production. These results highlight the role of T. gondii derived redox enzymes as important immune modulators and potentially indicate a role for AAM in modulating immunopathology and promoting parasite replication during T. gondii infection.

PMID:21707997[PubMed - as supplied by publisher]

Identification of plant-like galactolipids in Chromera velia, a photosynthetic relative of malaria parasites

J Biol Chem. 2011 Jun 28. [Epub ahead of print]

Identification of plant-like galactolipids in Chromera velia, a photosynthetic relative of malaria parasites.

Botte CY, Yamaryo-Botte Y, Janouskovec J, Rupasinghe T, Keeling PJ, Crellin P, Coppel R, Marechal E, McConville MJ, McFadden GI.

SourceSchool of Botany, University of Melbourne, France;

Abstract
Apicomplexa are protist parasites that include Plasmodium spp., the causative agents of malaria, and Toxoplasma gondii, responsible for toxoplasmosis. Most Apicomplexa possess a relict plastid, the apicoplast, which was acquired by secondary endosymbiosis of a red alga. Despite being non-photosynthetic, the apicoplast is otherwise metabolically similar to algal and plant plastids and is essential for parasite survival. Previous studies of Toxoplasma gondii identified membrane lipids with some structural features of plastid galactolipids, the major plastid lipid class. However, direct evidence for the plant-like enzymes responsible for galactolipid synthesis in apicomplexan parasites has yet been obtained. Chromera velia (C. velia) is an apicomplexan relative recently discovered in Australian corals. C. velia retains a photosynthetic plastid, providing a unique model to study the evolution of the apicoplast. Here we report the unambiguous presence of plant-like monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol (DGDG) in C. velia, and localize DGDG to the plastid. We also provide evidence for a plant-like biosynthesis pathway and identify candidate galactosyltranferases responsible for galactolipid synthesis. Our study provides new insights in the evolution of these important enzymes in plastid-containing eukaryotes and will help reconstruct the evolution of glycerolipid metabolism in important parasites such as Plasmodium and Toxoplasma.

PMID:21712377

The IFN-γ-Inducible GTPase, Irga6, Protects Mice against Toxoplasma gondii but Not against Plasmodium berghei

PLoS One. 2011;6(6):e20568. Epub 2011 Jun 17

The IFN-γ-Inducible GTPase, Irga6, Protects Mice against Toxoplasma gondii but Not against Plasmodium berghei and Some Other Intracellular Pathogens

Liesenfeld O, Parvanova I, Zerrahn J, Han SJ, Heinrich F, Muñoz M, Kaiser F, Aebischer T, Buch T, Waisman A, Reichmann G, Utermöhlen O, von Stebut E, von Loewenich FD, Bogdan C, Specht S, Saeftel M, Hoerauf A, Mota MM, Könen-Waisman S, Kaufmann SH, Howard JC

SourceInstitute of Microbiology and Hygiene, Charité Universitätsmedizin Berlin, Berlin, Germany.

Abstract
Clearance of infection with intracellular pathogens in mice involves interferon-regulated GTPases of the IRG protein family. Experiments with mice genetically deficient in members of this family such as Irgm1(LRG-47), Irgm3(IGTP), and Irgd(IRG-47) has revealed a critical role in microbial clearance, especially for Toxoplasma gondii. The in vivo role of another member of this family, Irga6 (IIGP, IIGP1) has been studied in less detail. We investigated the susceptibility of two independently generated mouse strains deficient in Irga6 to in vivo infection with T. gondii, Mycobacterium tuberculosis, Leishmania mexicana, L. major, Listeria monocytogenes, Anaplasma phagocytophilum and Plasmodium berghei. Compared with wild-type mice, mice deficient in Irga6 showed increased susceptibility to oral and intraperitoneal infection with T. gondii but not to infection with the other organisms. Surprisingly, infection of Irga6-deficient mice with the related apicomplexan parasite, P. berghei, did not result in increased replication in the liver stage and no Irga6 (or any other IRG protein) was detected at the parasitophorous vacuole membrane in IFN-γ-induced wild-type cells infected with P. berghei in vitro. Susceptibility to infection with T. gondii was associated with increased mortality and reduced time to death, increased numbers of inflammatory foci in the brains and elevated parasite loads in brains of infected Irga6-deficient mice. In vitro, Irga6-deficient macrophages and fibroblasts stimulated with IFN-γ were defective in controlling parasite replication. Taken together, our results implicate Irga6 in the control of infection with T. gondii and further highlight the importance of the IRG system for resistance to this pathogen.

PMID:21698150[PubMed - in process]

Apicoplast isoprenoid precursor synthesis and the molecular basis of fosmidomycin resistance in Toxoplasma gondii

J Exp Med. 2011 Jun 20. [Epub ahead of print]

Apicoplast isoprenoid precursor synthesis and the molecular basis of fosmidomycin resistance in Toxoplasma gondii

Nair SC, Brooks CF, Goodman CD, Strurm A, McFadden GI, Sundriyal S, Anglin JL, Song Y, Moreno SN, Striepen B

SourceDepartment of Cellular Biology and 2 Center for Tropical and Emerging Global Diseases, University of Georgia, Athens, GA, 30602.

Abstract
Apicomplexa are important pathogens that include the causative agents of malaria, toxoplasmosis, and cryptosporidiosis. Apicomplexan parasites contain a relict chloroplast, the apicoplast. The apicoplast is indispensable and an attractive drug target. The apicoplast is home to a 1-deoxy-d-xylulose-5-phosphate (DOXP) pathway for the synthesis of isoprenoid precursors. This pathway is believed to be the most conserved function of the apicoplast, and fosmidomycin, a specific inhibitor of the pathway, is an effective antimalarial. Surprisingly, fosmidomycin has no effect on most other apicomplexans. Using Toxoplasma gondii, we establish that the pathway is essential in parasites that are highly fosmidomycin resistant. We define the molecular basis of resistance and susceptibility, experimentally testing various host and parasite contributions in T. gondii and Plasmodium. We demonstrate that in T. gondii the parasite plasma membrane is a critical barrier to drug uptake. In strong support of this hypothesis, we engineer de novo drug-sensitive T. gondii parasites by heterologous expression of a bacterial transporter protein. Mice infected with these transgenic parasites can now be cured from a lethal challenge with fosmidomycin. We propose that the varied extent of metabolite exchange between host and parasite is a crucial determinator of drug susceptibility and a predictor of future resistance.

PMID:21690250[PubMed - as supplied by publisher]

A Purification Method for Enrichment of the Toxoplasma gondii Cyst Wall

J Neuroparasitology. 2010 Dec;1. pii: N101001

A Purification Method for Enrichment of the Toxoplasma gondii Cyst Wall

Zhang YW, Halonen SK, Ma YF, Tanowtiz HB, Weiss LM.
SourceDepartment of Pathology, Division of Parasitology, Albert Einstein College of Medicine, Bronx, NY 10461, USA.

Abstract
The tissue cyst wall of Toxoplasma gondii is a stage-specific structure that is produced by modification of the bradyzoite-containing parasitophorous vacuole. It is a limiting membrane structure and is critically important for cyst survival and transmission of infection. Studies on the structure and function of the cyst wall should provide new therapeutic strategies for the elimination or prevention of latency during T. gondii infection. The membrane proteins of the T. gondii cyst are an important target for studies of the biochemical and immunological function(s) of the cyst. However, the components of the cyst membrane have been poorly characterized due to the difficulty of purification of these membrane proteins. We developed a lectin DBA (Dolichos biflorus) coated magnetic bead isolation method to isolate T. gondii cyst wall proteins. Our data suggests that this method can isolate cyst wall proteins from both in vitro cell culture or in vivo mouse brain derived tissue cysts. Antibodies to these isolated protein preparations were shown to localize to the cyst wall.

PMID:21687827[PubMed]

CD8 T Cells and Toxoplasma gondii: A New Paradigm

J Parasitol Res. 2011;2011:243796. Epub 2011 May 18

CD8 T Cells and Toxoplasma gondii: A New Paradigm

Gigley JP, Bhadra R, Khan IA.
SourceDepartment of Microbiology, Immunology and Tropical Medicine, George Washington University, Washington, DC 20037, USA.

Abstract
CD8 T cells are essential for control of Toxoplasma gondii infection. Once activated they undergo differentiation into short-lived effector and memory precursor effector cells. As effector cells, CD8 T cells exert immune pressure on the parasite via production of inflammatory cytokines and through their cytolytic activity. Once immune control has been established, the parasite encysts and develops into chronic infection regulated by the memory CD8 T-cell population. Several signals are needed for this process to be initiated and for development of fully differentiated memory CD8 T cells. With newly developed tools including CD8 T-cell tetramers and TCR transgenic mice, dissecting the biology behind T. gondii-specific CD8 T-cell responses can now be more effectively addressed. In this paper, we discuss what is known about the signals required for effective T. gondii-specific CD8 T-cell development, their differentiation, and effector function.

PMID:21687650[PubMed - in process]

1NM-PP1 Treatment of Mice Infected with Toxoplasma gondii

J Vet Med Sci. 2011 Jun 16. [Epub ahead of print]

1NM-PP1 Treatment of Mice Infected with Toxoplasma gondii

Sugi T, Kato K, Kobayashi K, Kurokawa H, Takemae H, Haiyan G, Recuenco FC, Iwanaga T, Horimoto T, Akashi H.
SourceDepartment of Veterinary Microbiology, Graduate School of Agricultural and Life Sciences, The University of Tokyo.

Abstract
Bumped kinase inhibitors (BKIs) target analog-sensitive kinases, which the genomes of mammals rarely encode. Previously, we demonstrated that a BKI effectively suppressed the in vitro replication of Toxoplasma gondii, the causative pathogen of toxoplasmosis, by targeting T. gondii calcium-dependent protein kinase 1 (TgCDPK1) (Eukaryotic Cell, 9:667-670). Here, we examined whether the BKI 1NM-PP1 reduced parasite replication in vivo. A high dose of 1NM-PP1, by intraperitoneal injection, just before the parasite inoculation effectively reduced the parasite load in the brains, livers, and lungs of T. gondii-infected mice, however, a low dose of 1NM-PP1 with oral administration didn't change the survival rates of infected mice.

PMID:21685719[PubMed - as supplied by publisher]

EVIDENCE FOR HOST AS THE MAJOR CONTRIBUTOR TO LIPID IN THE INTRAVACUOLAR NETWORK OF TOXOPLASMA-INFECTED CELLS

Eukaryot Cell. 2011 Jun 17. [Epub ahead of print]

EVIDENCE FOR HOST AS THE MAJOR CONTRIBUTOR TO LIPID IN THE INTRAVACUOLAR NETWORK OF TOXOPLASMA-INFECTED CELLS

Caffaro CE, Boothroyd JC.
SourceDepartment of Microbiology & Immunology, Stanford University School of Medicine, Stanford, CA 94305-5124, USA.

Abstract
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 uncover evidence that strongly supports the host cell as the primary, if not exclusive, source of lipid for the parasite IVN remodeling. Fluorescence Recovery After Photobleaching (FRAP) microscopy experiments reveal 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.

PMID:21685319[PubMed - as supplied by publisher]

Protein Arginine Methylation in Parasitic Protozoa

Eukaryot Cell. 2011 Jun 17. [Epub ahead of print]

Protein Arginine Methylation in Parasitic Protozoa

Fisk JC, Read LK.
SourceDepartment of Microbiology and Immunology, School of Medicine and Biomedical Sciences, University at Buffalo, Buffalo, NY 14214.

Abstract
Protozoa constitute the earliest branch of the eukaryotic lineage, and several groups of protozoans are serious parasites of humans and other animals. Better understanding of biochemical pathways that are either in common with or divergent from with higher eukaryotes is integral in the defense against these parasites. In yeast and humans, the posttranslational methylation of arginine residues in proteins affects myriad cellular processes including transcription, RNA processing, DNA replication and repair, and signal transduction. The protein arginine methyltransferases that catalyze these reactions, which are unique to the eukaryotic kingdom of organisms, first become evident in protozoa. In this review, we focus on the current understanding of arginine methylation in multiple species of parasitic protozoa, including Trichomonas, Entamoeba, Toxoplasma, Plasmodium, and Trypanosoma, and discuss how arginine methylation may play important and unique roles in each type of parasite. We mine available genomic and transcriptomic data to inventory the families of PRMTs in different parasites and the changes in their abundance during the life cycle. We further review the limited functional studies on the roles of arginine methylation in parasites, including epigenetic regulation in Apicomplexa and RNA processing in trypanosomes. Interestingly, each of the parasites considered herein has significantly differing sets of PRMTs, and we speculate on importance of this diversity in aspects of parasite biology such as differentiation and antigenic variation.

PMID:21685318[PubMed - as supplied by publisher]

Proteases as regulators of pathogenesis: Examples from the apicomplexa

Biochim Biophys Acta. 2011 Jun 14. [Epub ahead of print]

Proteases as regulators of pathogenesis: Examples from the apicomplexa.

Li H, Child MA, Bogyo M.

Abstract
The diverse functional roles that proteases play in basic biological processes make them essential for virtually all organisms. Not surprisingly, proteolysis is also a critical process required for many aspects of pathogenesis. In particular, obligate intracellular parasites must precisely coordinate proteolytic events during their highly regulated life cycle inside multiple host cell environments. Advances in chemical, proteomic and genetic tools that can be applied to parasite biology have led to an increased understanding of the complex events centrally regulated by proteases. In this review, we outline recent advances in our knowledge of specific proteolytic enzymes in two medically relevant apicomplexan parasites: Plasmodium falciparum and Toxoplasma gondii. Efforts over the last decade have begun to provide a map of key proteotolyic events that are essential for both parasite survival and propagation inside host cells. These advances in our molecular understanding of proteolytic events involved in parasite pathogenesis provide a foundation for the validation of new networks and enzyme targets that could be exploited for therapeutic purposes. This article is part of a Special Issue entitled: Proteolysis 50years after the discovery of lysosome.

Copyright © 2011. Published by Elsevier B.V.

PMID:21683169[PubMed - as supplied by publisher]

Toxoplasma gondii infection and abdominal hernia: evidence of a new association

Parasit Vectors. 2011 Jun 17;4(1):112. [Epub ahead of print]

Toxoplasma gondii infection and abdominal hernia: evidence of a new association

Alvarado-Esquivel C, Estrada-Martinez S.
Abstract
ABSTRACT:

BACKGROUND: We performed a retrospective, observational study in 1156 adult subjects from the general population of Durango City, Mexico, Fifty five subjects with a history of abdominal hernia repair and 1101 subjects without hernia were examined with enzyme-linked immunoassays for the presence of anti-Toxoplasma IgG and IgM antibodies.

RESULTS: The seroprevalence of anti-Toxoplasma IgG antibodies and IgG titers was significantly higher in subjects with abdominal hernia repair than those without hernia. There was a tendency for subjects with hernia repair to have a higher seroprevalence of anti-Toxoplasma IgM antibodies than subjects without hernia. The seroprevalence of anti-Toxoplasma IgG antibodies in subjects with hernia repair was significantly higher in subjects [greater than or equal to]50 years old than those <50 years old. Further analysis in subjects aged [greater than or equal to]50 years showed that the seroprevalence of anti-Toxoplasma IgG antibodies was also significantly higher in individuals with hernia repair than those without hernia (OR = 2.72; 95% CI: 1.10-6.57). Matching by age and sex further showed that the seroprevalence of Toxoplasma infection was significantly higher in patients with hernia repair than those without hernia (OR: 4.50; 95% CI: 1.22-17.33).

CONCLUSIONS: Results indicate that infection with Toxoplasma is associated with abdominal hernia. The contributing role of infection with Toxoplasma in abdominal hernia was observed mainly in subjects aged [greater than or equal to]50 years old. Our results might have clinical, prevention and treatment implications and warrant for further investigation.

PMID:21682896[PubMed - as supplied by publisher]

Chemical genetic screen identifies Toxoplasma DJ-1 as a regulator of parasite secretion, attachment, and invasion

Proc Natl Acad Sci U S A. 2011 Jun 13. [Epub ahead of print]

Chemical genetic screen identifies Toxoplasma DJ-1 as a regulator of parasite secretion, attachment, and invasion

Hall CI, Reese ML, Weerapana E, Child MA, Bowyer PW, Albrow VE, Haraldsen JD, Phillips MR, Sandoval ED, Ward GE, Cravatt BF, Boothroyd JC, Bogyo M

SourceDepartments of Microbiology and Immunology and Pathology, Stanford University School of Medicine, Stanford, CA 94305.

Abstract
Toxoplasma gondii is a member of the phylum Apicomplexa that includes several important human pathogens, such as Cryptosporidium and Plasmodium falciparum, the causative agent of human malaria. It is an obligate intracellular parasite that can cause severe disease in congenitally infected neonates and immunocompromised individuals. Despite the importance of attachment and invasion to the success of the parasite, little is known about the underlying mechanisms that drive these processes. Here we describe a screen to identify small molecules that block the process of host cell invasion by the T. gondii parasite. We identified a small molecule that specifically and irreversibly blocks parasite attachment and subsequent invasion of host cells. Using tandem orthogonal proteolysis-activity-based protein profiling, we determined that this compound covalently modifies a single cysteine residue in a poorly characterized protein homologous to the human protein DJ-1. Mutation of this key cysteine residue in the native gene sequence resulted in parasites that were resistant to inhibition of host cell attachment and invasion by the compound. Further analysis of the invasion phenotype confirmed that modification of Cys127 on TgDJ-1 resulted in a block of microneme secretion and motility, even in the presence of direct stimulators of calcium release. Together, our results suggest that TgDJ-1 plays an important role that is likely downstream of the calcium flux required for microneme secretion, parasite motility, and subsequent invasion of host cells.

PMID:21670272[PubMed - as supplied by publisher]

ATF6{beta} is a host cellular target of the Toxoplasma gondii virulence factor ROP18

J Exp Med. 2011 Jun 13. [Epub ahead of print]

ATF6{beta} is a host cellular target of the Toxoplasma gondii virulence factor ROP18

Yamamoto M, Ma JS, Mueller C, Kamiyama N, Saiga H, Kubo E, Kimura T, Okamoto T, Okuyama M, Kayama H, Nagamune K, Takashima S, Matsuura Y, Soldati-Favre D, Takeda K

SourceDepartment of Microbiology and Immunology and 2 Department of Cardiovascular Medicine, Graduate School of Medicine, 3 Laboratory of Mucosal Immunology, World Premier International Research Center Immunology Frontier Research Center; and 4 Department of Molecular Virology, Research Institute for Microbial Diseases; Osaka University, Suita, Osaka 565-0871, Japan.

Abstract
The ROP18 kinase has been identified as a key virulence determinant conferring a high mortality phenotype characteristic of type I Toxoplasma gondii strains. This major effector molecule is secreted by the rhoptries into the host cells during invasion; however, the molecular mechanisms by which this kinase exerts its pathogenic action remain poorly understood. In this study, we show that ROP18 targets the host endoplasmic reticulum-bound transcription factor ATF6β. Disruption of the ROP18 gene severely impairs acute toxoplasmosis by the type I RH strain. Because another virulence factor ROP16 kinase modulates immune responses through its N-terminal portion, we focus on the role of the N terminus of ROP18 in the subversion of host cellular functions. The N-terminal extension of ROP18 contributes to ATF6β-dependent pathogenicity by interacting with ATF6β and destabilizing it. The kinase activity of ROP18 is essential for proteasome-dependent degradation of ATF6β and for parasite virulence. Consistent with a key role for ATF6β in resistance against this intracellular pathogen, ATF6β-deficient mice exhibit a high susceptibility to infection by ROP18-deficient parasites. The results reveal that interference with ATF6β-dependent immune responses is a novel pathogenic mechanism induced by ROP18.

PMID:21670204[PubMed - as supplied by publisher]

Toxoplasma polymorphic effectors determine macrophage polarization and intestinal inflammation

Cell Host Microbe. 2011 Jun 16;9(6):472-83

Toxoplasma polymorphic effectors determine macrophage polarization and intestinal inflammation

Jensen KD, Wang Y, Wojno ED, Shastri AJ, Hu K, Cornel L, Boedec E, Ong YC, Chien YH, Hunter CA, Boothroyd JC, Saeij JP

SourceDepartment of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Department of Microbiology and Immunology, Stanford University, Stanford, CA 94305, USA.

Abstract
European and North American strains of the parasite Toxoplasma gondii belong to three distinct clonal lineages, type I, type II, and type III, which differ in virulence. Understanding the basis of Toxoplasma strain differences and how secreted effectors work to achieve chronic infection is a major goal of current research. Here we show that type I and III infected macrophages, a cell type required for host immunity to Toxoplasma, are alternatively activated, while type II infected macrophages are classically activated. The Toxoplasma rhoptry kinase ROP16, which activates STAT6, is responsible for alternative activation. The Toxoplasma dense granule protein GRA15, which activates NF-κB, promotes classical activation by type II parasites. These effectors antagonistically regulate many of the same genes, and mice infected with type II parasites expressing type I ROP16 are protected against Toxoplasma-induced ileitis. Thus, polymorphisms in determinants that modulate macrophage activation influence the ability of Toxoplasma to establish a chronic infection.

Copyright © 2011 Elsevier Inc. All rights reserved.

PMID:21669396[PubMed - in process]

Mol Biochem Parasitol. 2011 Jun 2. [Epub ahead of print]

A transmembrane domain containing pellicle protein of Toxoplasma gondii enhances virulence and invasion after extracellular stress

Payne TM, Lund PJ, Knoll LJ

SourceDepartment of Medical Microbiology and Immunology, University of Wisconsin School of Medicine and Public Health, Microbial Sciences Building Room 3345, 1550 Linden Drive, Madison, WI 53706, United States.

Abstract
To identify Toxoplasma gondii genes important in the establishment of a persistent infection, we previously used signature-tagged mutagenesis to identify mutants with reduced cyst numbers in the brains of mice. One of the mutants, 95C5, has an insertion within a predicted six transmembrane domain protein, which localizes to the parasite pellicle, thus we named it transmembrane pellicle protein 1 (TgTPP1). Although the 95C5 mutant was found be reduced in its ability to form brain cysts, it is defective during acute infection. Addition of TgTPP1 expressed from its endogenous promoter restored the acute lethality of the 95C5 mutant to parental levels. The 95C5 mutant does not have a growth defect in standard tissue culture conditions; however, we found a significant defect in host cell penetration after extracellular stress. Overall, TgTPP1 may function during acute infection by enhancing the parasites ability to invade after extracellular stress.

Copyright © 2011 Elsevier B.V. All rights reserved.

PMID:21669237[PubMed - as supplied by publisher]

The CD8 T-cell road to immunotherapy of toxoplasmosis

Immunotherapy. 2011 Jun;3(6):789-801

The CD8 T-cell road to immunotherapy of toxoplasmosis

Bhadra R, Gigley JP, Khan IA.
SourceDepartment of Microbiology, Immunology & Tropical Medicine, George Washington University, Washington, DC 20037, USA.

Abstract
Toxoplasma gondii infection induces a robust CD8 T-cell immunity that is critical for keeping chronic infection under control. In studies using animal models, it has been demonstrated that the absence of this response can compromise the host ability to keep chronic infection under check. Therapeutic agents that facilitate the induction and maintenance of CD8 T-cell response against the pathogen need to be developed. In the last decade, major strides in understanding the development of effector and memory response, particularly in viral and tumor models, have been made. However, factors involved in the generation of effector or memory response against T. gondii infection have not been extensively investigated. This information will be invaluable in designing immunotherapeutic regimens needed for combating this intracellular pathogen that poses a severe risk for pregnant women and immunocompromised individuals.

PMID:21668315[PubMed - in process]

Cathepsin Proteases in Toxoplasma gondii

Adv Exp Med Biol. 2011;712:49-61.

Cathepsin Proteases in Toxoplasma gondii

Dou Z, Carruthers VB

SourceDepartment of Microbiology and Immunology, University of Michigan School of Medicine, Ann Arbor, Michigan, USA.

Abstract
Cysteine proteases are important for the growth and survival of apicomplexan parasites that infect humans. The apicomplexan Toxoplasma gondii expresses five members of the C1 family of cysteine proteases, including one cathepsin L-like (TgCPL), one cathepsin B-like (TgCPB) and three cathepsin C-like (TgCPC1, 2 and 3) proteases. Recent genetic, biochemical and structural studies reveal that cathepsins function in microneme and rhoptry protein maturation, host cell invasion, replication and nutrient acquisition. here, we review the key features and roles of T. gondii cathepsins and discuss the therapeutic potential for specific inhibitor development.

PMID:21660658[PubMed - in process]

Focus on the ringleader: the role of AMA1 in apicomplexan invasion and replication

Trends Parasitol. 2011 Jun 7. [Epub ahead of print]

Focus on the ringleader: the role of AMA1 in apicomplexan invasion and replication

Tyler JS, Treeck M, Boothroyd JC.

SourceDepartment of Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA 94305, USA.

Abstract
Apicomplexan parasites exhibit an unusual mechanism of host cell penetration. A central player in this process is the protein apical membrane antigen 1 (AMA1). Although essential for invasion, the precise functional roles AMA1 plays have been unclear. Several recent studies have provided important functional insight into its role within the multiprotein complex that comprises the moving junction (MJ). Initially formed at the apical tip of the invading parasite, the MJ represents a ring-like region of contact between the surfaces of the invading parasite and the host cell as the invaginated host plasma membrane is forced inward by the penetrating parasite. This review discusses these and other recent insights into AMA1 with particular emphasis on studies conducted in Plasmodium and Toxoplasma.

Copyright © 2011 Elsevier Ltd. All rights reserved.

PMID:21659001[PubMed - as supplied by publisher]

Involvement of a Toxoplasma gondii Chromatin Remodeling Complex Ortholog in Developmental Regulation

PLoS One. 2011;6(5):e19570. Epub 2011 May 31

Involvement of a Toxoplasma gondii Chromatin Remodeling Complex Ortholog in Developmental Regulation

Rooney PJ, Neal LM, Knoll LJ

SourceDepartment of Medical Microbiology and Immunology, University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin, United States of America.

Abstract
The asexual cycle of the parasite Toxoplasma gondii has two developmental stages: a rapidly replicating form called a tachyzoite and a slow growing cyst form called a bradyzoite. While the importance of ATP-independent histone modifications for gene regulation in T. gondii have been demonstrated, ATP-dependent chromatin remodeling pathways have not been examined. In this study we characterized C9, an insertional mutant showing reduced expression of bradyzoite differentiation marker BAG1, in cultured human fibroblasts. This mutant contains an insertion in the gene encoding TgRSC8, which is homologous to the Saccharomyces cerevisiae proteins Rsc8p (remodel the structure of chromatin complex subunit 8) and Swi3p (switch/sucrose non-fermentable [SWI/SNF]) of ATP-dependent chromatin-remodeling complexes. In the C9 mutant, TgRSC8 is the downstream open reading frame on a dicistronic transcript. Though protein was expressed from the downstream gene of the dicistron, TgRSC8 levels were decreased in C9 from those of wild-type parasites, as determined by western immunoblot and flow cytometry. As TgRSC8 localized to the parasite nucleus, we postulated a role in gene regulation. Transcript levels of several markers were assessed by quantitative PCR to test this hypothesis. The C9 mutant displayed reduced steady state transcript levels of bradyzoite-induced genes BAG1, LDH2, SUSA1, and ENO1, all of which were significantly increased with addition of TgRSC8 to the mutant. Transcript levels of some bradyzoite markers were unaltered in C9, or unable to be increased by complementation with TgRSC8, indicating multiple pathways control bradyzoite-upregulated genes. Together, these data suggest a role for TgRSC8 in control of bradyzoite-upregulated gene expression. Thus chromatin remodeling, by both ATP-independent and dependent mechanisms, is an important mode of gene regulation during stage differentiation in parasites.

PMID:21655329[PubMed - in process]

Parasite mitogen-activated protein kinases as drug discovery targets to treat human protozoan pathogens

J Signal Transduct. 2011;2011:971968. Epub 2011 Feb 27

Parasite mitogen-activated protein kinases as drug discovery targets to treat human protozoan pathogens

Brumlik MJ, Pandeswara S, Ludwig SM, Murthy K, Curiel TJ

SourceDepartment of Medicine, School of Medicine, and Program in Immunology and Microbiology, Graduate School of Biomedical Sciences, University of Texas Health Science Center, San Antonio, TX 78229, USA.

Abstract
Protozoan pathogens are a highly diverse group of unicellular organisms, several of which are significant human pathogens. One group of protozoan pathogens includes obligate intracellular parasites such as agents of malaria, leishmaniasis, babesiosis, and toxoplasmosis. The other group includes extracellular pathogens such as agents of giardiasis and amebiasis. An unfortunate unifying theme for most human protozoan pathogens is that highly effective treatments for them are generally lacking. We will review targeting protozoan mitogen-activated protein kinases (MAPKs) as a novel drug discovery approach towards developing better therapies, focusing on Plasmodia, Leishmania, and Toxoplasma, about which the most is known.

PMID:21637385[PubMed - in process]

Polyparasitism Is Associated with Increased Disease Severity in Toxoplasma gondii-Infected Marine Sentinel Species

PLoS Negl Trop Dis. 2011 May;5(5):e1142. Epub 2011 May 24.

Polyparasitism Is Associated with Increased Disease Severity in Toxoplasma gondii-Infected Marine Sentinel Species

Gibson AK, Raverty S, Lambourn DM, Huggins J, Magargal SL, Grigg ME.
SourceLaboratory of Parasitic Diseases, National Institutes of Health, National Institute of Allergy and Infectious Diseases (NIAID), Bethesda, Maryland, United States of America.

Abstract
In 1995, one of the largest outbreaks of human toxoplasmosis occurred in the Pacific Northwest region of North America. Genetic typing identified a novel Toxoplasma gondii strain linked to the outbreak, in which a wide spectrum of human disease was observed. For this globally-distributed, water-borne zoonosis, strain type is one variable influencing disease, but the inability of strain type to consistently explain variations in disease severity suggests that parasite genotype alone does not determine the outcome of infection. We investigated polyparasitism (infection with multiple parasite species) as a modulator of disease severity by examining the association of concomitant infection of T. gondii and the related parasite Sarcocystis neurona with protozoal disease in wild marine mammals from the Pacific Northwest. These hosts ostensibly serve as sentinels for the detection of terrestrial parasites implicated in water-borne epidemics of humans and wildlife in this endemic region. Marine mammals (151 stranded and 10 healthy individuals) sampled over 6 years were assessed for protozoal infection using multi-locus PCR-DNA sequencing directly from host tissues. Genetic analyses uncovered a high prevalence and diversity of protozoa, with 147/161 (91%) of our sampled population infected. From 2004 to 2009, the relative frequency of S. neurona infections increased dramatically, surpassing that of T. gondii. The majority of T. gondii infections were by genotypes bearing Type I lineage alleles, though strain genotype was not associated with disease severity. Significantly, polyparasitism with S. neurona and T. gondii was common (42%) and was associated with higher mortality and more severe protozoal encephalitis. Our finding of widespread polyparasitism among marine mammals indicates pervasive contamination of waterways by zoonotic agents. Furthermore, the significant association of concomitant infection with mortality and protozoal encephalitis identifies polyparasitism as an important factor contributing to disease severity in marine mammals.

PMID:21629726[PubMed - in process]

Migratory activation of primary cortical microglia upon infection with Toxoplasma gondii

Infect Immun. 2011 May 31. [Epub ahead of print]

Migratory activation of primary cortical microglia upon infection with Toxoplasma gondii

Dellacasa-Lindberg I, Fuks JM, Arrighi RB, Lambert H, Wallin RP, Chambers BJ, Barragan A.
SourceCenter for Infectious Medicine, Dept. of Medicine, Karolinska Institutet, 171 77 Stockholm, Sweden.

Abstract
Disseminated Toxoplasmosis in the central nervous system (CNS) is often accompanied by lethal outcome. Studies in murine models of infection have focused on the role of systemic immunity for control of toxoplasmic encephalitis while knowledge remains limited on the contribution of resident cells with immune functions in the CNS. Here, the role of glia cells was addressed in the setting of recrudescent Toxoplasma infection in mice. Activated astrocytes and microglia were observed in close vicinity of foci with replicating parasites in situ in the brain parenchyma. Toxoplasma gondii tachyzoites were allowed to infect primary microglia and astrocytes in vitro. Microglia were permissive to parasite replication and infected microglia readily transmigrated across transwell membranes and cell monolayers. Thus, infected microglia, but not astrocytes, exhibited a hypermotility phenotype reminiscent of that recently described for infected dendritic cells. In contrast to interferon γ-activated microglia, Toxoplasma-infected microglia did not upregulate MHC class II and the co-stimulatory molecule CD86. Yet, Toxoplasma-infected microglia and astrocytes exhibited increased sensitivity to T cell-mediated killing, leading to rapid parasite transfer to effector T cells in vitro. We hypothesize that glia and T cells, besides their role in triggering anti-parasite immunity, may also act as 'Trojan horses' paradoxically facilitating dissemination of Toxoplasma within the CNS. To our knowledge, this constitutes the first report of migratory activation of a resident CNS cell by an intracellular parasite.

PMID:21628522[PubMed - as supplied by publisher]