In vitro and in vivo effects of the phytohormone inhibitor fluridone against Neospora caninum infection

Parasitol Int. 2016 Mar 25. pii: S1383-5769(16)30034-4. doi: 10.1016/j.parint.2016.03.009. [Epub ahead of print]

Ybañez RH¹, Leesombun A², Nishimura M², Matsubara R³, Kojima M⁴, Sakakibara H⁴, Nagamune K⁵, Nishikawa Y⁶.

Neospora caninum causes abortion and stillbirth in cattle. Identification of effective drugs against this parasite remains a challenge. Previous studies have suggested that disruption of abscisic acid (ABA)-mediated signaling in apicomplexan parasites such as Toxoplasma gondii offers a new drug target. In this study, the ABA inhibitor, fluridone (FLU), was evaluated for its action against N. caninum. Production of endogenous ABA within N. caninum was confirmed by ultra-performance liquid chromatography-tandem quadruple mass spectrometry. Subsequently, FLU treatment efficacy was assessed using in vitro. Results revealed that FLU inhibited the growth of N. caninum and T. gondii in vitro (IC₅₀ 143.1±43.96μM and 330.6±52.38μM, respectively). However, FLU did not affect parasite replication at 24h post-infection, but inhibited egress of N. caninum thereafter. To evaluate the effect of FLU in vivo, N. caninum-infected mice were treated with FLU for 15days. FLU treatment appeared to ameliorate acute neosporosis induced by lethal parasite challenge. Together, our data shows that ABA might control egress in N. caninum. Therefore, FLU has potential as a candidate drug for the treatment of acute neosporosis.
Copyright © 2015. Published by Elsevier Ireland Ltd.

KEYWORDS:

Abscisic acid; Fluridone; Neospora caninum

PMID:

27021922

[PubMed - as supplied by publisher]

Toxoplasma gondii ROP18: potential to manipulate host cell mitochondrial apoptosis

Parasitol Res. 2016 Mar 28. [Epub ahead of print]

Wu L^1,2, Wang X², Li Y², Liu Y², Su D², Fu T², Guo F², Gu L², Jiang X², Chen S³, Cao J⁴.

Toxoplasma gondii is an obligate intracellular parasite that may manipulate host cell mitochondrial apoptosis pathways. In our experiment, 293T cells were transfected with the p3×FLAG-CMV-Myc-ROP18 vector and expressed the ROP18-Myc fusion protein. Cell apoptosis was induced by 0.5 μg/mL actinomycin D (ActD) and was detected by Annexin V-FITC/PI assay. The cell mitochondrial membrane potential was determined by JC-1. Cytochrome c (Cyto-c) from mitochondria and the cytoplasm was measured by Western blot. The Bcl-2 and Bax coding gene expression levels were detected by real-time PCR. We found, in vitro, that T. gondii ROP18 significantly suppressed 293T cell apoptosis induced by ActD and maintained mitochondrial membrane potential and integrity, thereby preventing the release of Cyto-c from mitochondria into the cytoplasm. The ratio of Bcl-2/Bax in ROP18-overexpressing cells was significantly higher than that of the negative control. Therefore, we speculate that ROP18 could suppress host cell apoptosis via the mitochondrial apoptosis pathway in vitro.

KEYWORDS:

Apoptosis; Mitochondrial apoptosis pathway; ROP18; Toxoplasma gondii

PMID:

27021182

[PubMed - as supplied by publisher]

Role of the ER and Golgi in protein export by Apicomplexa

Curr Opin Cell Biol. 2016 Mar 25;41:18-24. doi: 10.1016/j.ceb.2016.03.007. [Epub ahead of print]

Coffey MJ1, Jennison C1, Tonkin CJ2, Boddey JA3.

Author information

Abstract

Apicomplexan parasites cause diseases of medical and agricultural importance linked to dramatic changes they impart upon infected host cells. Following invasion, the malaria parasite Plasmodium falciparum renovates the host erythrocyte using mechanisms previously believed to be malaria-specific. This involves proteolytic cleavage of effectors in the endoplasmic reticulum that licences proteins for translocation into the host cell. Recently, it was demonstrated that the related parasite Toxoplasma gondii, responsible for disease in immunocompromised individuals and congenital birth defects, has an analogous pathway with some differences, including proteolytic processing in the Golgi. Here we review the similarities and distinctions in export mechanisms between these and other Apicomplexan parasites to reconcile how this group of pathogens modify their host cells to survive and proliferate.

Copyright © 2016 Elsevier Ltd. All rights reserved.

PMID:

 

27019341

 

[PubMed - as supplied by publisher]

Migratory activation of parasitized dendritic cells by the protozoan Toxoplasma gondii 14-3-3 protein

Cell Microbiol. 2016 Mar 28. doi: 10.1111/cmi.12595. [Epub ahead of print]

Weidner JM1,2, Kanatani S1,2, Uchtenhagen H3, Varas-Godoy M4,5, Schulte T3, Engelberg K6, Gubbels MJ6, Sun HS7, Harrison RE7, Achour A3, Barragan A1,2.

Author information

Abstract

The obligate intracellular parasite Toxoplasma gondii exploits cells of the immune system to disseminate. Upon infection, parasitized dendritic cells (DCs) and microglia exhibit a hypermigratory phenotype in vitro that has been associated with enhancing parasite dissemination in vivo in mice. One unresolved question is how parasites commandeer parasitized cells to achieve systemic dissemination by a ´Trojan horse´ mechanism. By chromatography and mass spectrometry analyses, we identified an orthologue of the 14-3-3 protein family, T. gondii 14-3-3 (Tg14-3-3), as mediator of DC hypermotility. We demonstrate that parasite-derived polypeptide fractions enriched for Tg14-3-3 or recombinant Tg14-3-3 are sufficient to induce the hypermotile phenotype when introduced by protein transfection into murine DCs, human DCs or microglia. Further, gene transfer of Tg14-3-3 by lentiviral transduction induced hypermotility in primary human DCs. In parasites expressing Tg14-3-3 in a ligand regulatable fashion, over-expression of Tg14-3-3 was correlated with induction of hypermotility in parasitized DCs. Localization studies in infected DCs identified Tg14-3-3 within the parasitophorous vacuolar space and a rapid recruitment of host cell 14-3-3 to the parasitophorous vacuole membrane. The present work identifies a determinant role for Tg14-3-3 in the induction of the migratory activation of immune cells by T. gondii. Collectively, the findings reveal Tg14-3-3 as a novel target for an intracellular pathogen that acts by hijacking the host cell's migratory properties to disseminate.

This article is protected by copyright. All rights reserved.

PMID:

 

27018989

 

[PubMed - as supplied by publisher]

Chronic Toxoplasma gondii infection enhances β-amyloid phagocytosis and clearance by recruited monocytes

Acta Neuropathol Commun. 2016 Mar 16;4(1):25. doi: 10.1186/s40478-016-0293-8.

Möhle L1, Israel N2, Paarmann K3,4,5, Krohn M3, Pietkiewicz S6, Müller A2,7, Lavrik IN6, Buguliskis JS8, Schott BH9,10,11, Schlüter D1,9, Gundelfinger ED9,12,13,4, Montag D5, Seifert U2, Pahnke J3,14,15, Dunay IR16,17.

Author information

Abstract

INTRODUCTION: 

Alzheimer's disease (AD) is associated with the accumulation of β-amyloid (Aβ) as senile plaques in the brain, thus leading to neurodegeneration and cognitive impairment. Plaque formation depends not merely on the amount of generated Aβ peptides, but more importantly on their effective removal. Chronic infections with neurotropic pathogens, most prominently the parasite Toxoplasma (T.) gondii, are frequent in the elderly, and it has been suggested that the resulting neuroinflammation may influence the course of AD. In the present study, we investigated how chronic T. gondii infection and resulting neuroinflammation affect plaque deposition and removal in a mouse model of AD.

RESULTS: 

Chronic infection with T. gondii was associated with reduced Aβ and plaque load in 5xFAD mice. Upon infection, myeloid-derived CCR2(hi) Ly6C(hi) monocytes, CCR2(+) Ly6C(int), and CCR2(+) Ly6C(low) mononuclear cells were recruited to the brain of mice. Compared to microglia, these recruited mononuclear cells showed highly increased phagocytic capacity of Aβ ex vivo. The F4/80(+) Ly6C(low) macrophages expressed high levels of Triggering Receptor Expressed on Myeloid cells 2 (TREM2), CD36, and Scavenger Receptor A1 (SCARA1), indicating phagocytic activity. Importantly, selective ablation of CCR2(+) Ly6C(hi) monocytes resulted in an increased amount of Aβ in infected mice. Elevated insulin-degrading enzyme (IDE), matrix metalloproteinase 9 (MMP9), as well as immunoproteasome subunits β1i/LMP2, β2i/MECL-1, and β5i/LMP7 mRNA levels in the infected brains indicated increased proteolytic Aβ degradation. Particularly, LMP7 was highly expressed by the recruited mononuclear cells in the brain, suggesting a novel mechanism of Aβ clearance.

CONCLUSIONS: 

Our results indicate that chronic Toxoplasma infection ameliorates β-amyloidosis in a murine model of AD by activation of the immune system, specifically by recruitment of Ly6C(hi) monocytes and by enhancement of phagocytosis and degradation of soluble Aβ. Our findings provide evidence for a modulatory role of inflammation-induced Aβ phagocytosis and degradation by newly recruited peripheral immune cells in the pathophysiology of AD.

KEYWORDS: 

Alzheimer’s disease; Aβ clearance; Chronic infection; Ly6Chi monocytes; Toxoplasma gondii

PMID:

 

26984535

 

[PubMed - in process] 

A three-dimensional culture system recapitulates placental syncytiotrophoblast development and microbial resistance

Sci Adv. 2016 Mar 4;2(3):e1501462. doi: 10.1126/sciadv.1501462. eCollection 2016.

McConkey CA¹, Delorme-Axford E¹, Nickerson CA², Kim KS³, Sadovsky Y⁴, Boyle JP⁵, Coyne CB⁴.

In eutherians, the placenta acts as a barrier and conduit at the maternal-fetal interface. Syncytiotrophoblasts, the multinucleated cells that cover the placental villous tree surfaces of the human placenta, are directly bathed in maternal blood and are formed by the fusion of progenitor cytotrophoblasts that underlie them. Despite their crucial role in fetal protection, many of the events that govern trophoblast fusion and protection from microbial infection are unknown. We describe a three-dimensional (3D)-based culture model using human JEG-3 trophoblast cells that develop syncytiotrophoblast phenotypes when cocultured with human microvascular endothelial cells. JEG-3 cells cultured in this system exhibit enhanced fusogenic activity and morphological and secretory activities strikingly similar to those of primary human syncytiotrophoblasts. RNASeq analyses extend the observed functional similarities to the transcriptome, where we observed significant overlap between syncytiotrophoblast-specific genes and 3D JEG-3 cultures. Furthermore, JEG-3 cells cultured in 3D are resistant to infection by viruses and Toxoplasma gondii, which mimics the high resistance of syncytiotrophoblasts to microbial infections in vivo. Given that this system is genetically manipulatable, it provides a new platform to dissect the mechanisms involved in syncytiotrophoblast development and microbial resistance.

KEYWORDS:

3-D cell culture; Placenta; RWV bioreactor; Toxoplasma gondii; trophoblast

PMID:

26973875

[PubMed]

Oxidative stress generated during monensin treatment contributes to altered Toxoplasma gondii mitochondrial function

Sci Rep. 2016 Mar 15;6:22997. doi: 10.1038/srep22997.

Charvat RA¹, Arrizabalaga G^1,2.

The ionophore monensin displays potent activities against several coccidian parasites of veterinary and medical importance including the opportunistic pathogen of humans, Toxoplasma gondii. While monensin is used widely in animals, toxicity impedes its use in humans. Nonetheless, given its potency, understanding its mode of action would reveal vulnerable aspects of the parasite that can be exploited for drug development. We previously established that monensin induces Toxoplasma to undergo cell cycle arrest and an autophagy-like cell death. Interestingly, these effects are dependent on the mitochondrion-localized TgMSH-1 protein, suggesting that monensin disrupts mitochondrial function. We demonstrate that monensin treatment results in decreased mitochondrial membrane potential and altered morphology. These effects are mitigated by the antioxidant compound N-acetyl-cysteine suggesting that monensin causes an oxidative stress, which was indeed the case based on direct detection of reactive oxygen species. Moreover, over-expression of the antioxidant proteins glutaredoxin and peroxiredoxin 2 protect Toxoplasma from the deleterious effects of monensin. Thus, our studies show that the effects of monensin on Toxoplasma are due to a disruption of mitochondrial function caused by the induction of an oxidative stress and implicate parasite redox biology as a viable target for the development of drugs against Toxoplasma and related pathogenic parasites.

PMID:

26976749

[PubMed - in process]

Impact of environmental factors on the emergence, transmission and distribution of Toxoplasma gondii

Parasit Vectors. 2016 Mar 10;9(1):137. doi: 10.1186/s13071-016-1432-6.

Yan C^1,2, Liang LJ^2,3, Zheng KY⁴, Zhu XQ^5,6,7.

Toxoplasma gondii is an obligate intracellular protozoan that poses a great threat to human health and economic well-being worldwide. The effects of environmental factors such as changing climate and human activities on the ecology of this protozoan are being discovered. Accumulated evidence shows that changes of these environmental factors can exert influence on the occurrence, transmission and distribution of T. gondii. This article reviews studies from different geographical regions with varying climates, social cultures and animal welfare standards. It aims to illustrate how these environmental factors work, highlighting their importance in influencing the ecology of T. gondii, as well as providing clues which may contribute to preventing transmission of this important zoonotic pathogen.

KEYWORDS:

Changing climate; Ecology; Human activities; Oocysts; Toxoplasma gondii

PMID:

26965989

[PubMed - in process]

PMCID:

PMC4785633

Anti-NMDA receptor autoantibodies and associated neurobehavioral pathology in mice are dependent on age of first exposure to Toxoplasma gondii

Neurobiol Dis. 2016 Mar 8. pii: S0969-9961(16)30049-3. doi: 10.1016/j.nbd.2016.03.005. [Epub ahead of print]

Kannan G¹, Crawford JA², Yang C², Gressitt KL³, Ihenatu C³, Krasnova IN⁴, Cadet JL⁴, Yolken RH³, Severance EG³, Pletnikov MV⁵.

BACKGROUND:

Toxoplasma gondii is a pathogen implicated in psychiatric disorders. As elevated antibodies to T. gondii are also present in non-symptomatic individuals, we hypothesized that the age during first exposure to the pathogen may affect symptom manifestation. We tested this hypothesis by evaluating neurobehavioral abnormalities and the immune response in mice following adolescent or adult T. gondii infection.

METHODS:

Mice were infected with T. gondii at postnatal day 33 (adolescent/juvenile) or 61 (adult). At 8weeks post-infection (wpi), pre-pulse inhibition of the acoustic startle (PPI) in mice administered MK-801 (0.1 and 0.3mg/kg) and amphetamine (5 and 10mg/kg) was assessed. Peripheral (anti-T. gondii, C1q-associated IgG and anti-GLUN2 antibodies) and central (C1q and Iba1) markers of the immune response were also evaluated. In addition, regional brain expression of N-methyl-d-aspartate receptor (NMDAR) subunits (GLUN1 and GLUN2A), glutamatergic (vGLUT1, PSD95) and GABAergic (GAD67) markers, and monoamines (DA, NE, 5-HT) and their metabolites were measured.

RESULTS:

Juvenile and adult infected mice exhibited opposite effects of MK-801 on PPI, with decreased PPI in juveniles and increased PPI in adults. There was a significantly greater elevation of GLUN2 autoantibodies in juvenile-compared to adult-infected mice. In addition, age-dependent differences were found in regional expression of NMDAR subunits and markers of glutamatergic, GABAergic, and monoaminergic systems. Activated microglia and C1q elevations were found in both juvenile- and adult-T. gondii infected mice.

CONCLUSIONS:

Our study demonstrates that the age at first exposure to T. gondii is an important factor in shaping distinct behavioral and neurobiological abnormalities. Elevation in GLUN2 autoantibodies or complement protein C1q may be a potential underlying mechanism. A better understanding of these age-related differences may lead to more efficient treatments of behavioral disorders associated with T. gondii infection.
Copyright © 2015. Published by Elsevier Inc.

KEYWORDS:

Autoantibody; C1q; NMDAR; Neurodevelopment; Schizophrenia; Toxoplasma

PMID:

26969530

[PubMed - as supplied by publisher]

Phosphatidic Acid-Mediated Signaling Regulates Microneme Secretion in Toxoplasma

Cell Host Microbe. 2016 Mar 9;19(3):349-360. doi: 10.1016/j.chom.2016.02.006.

Bullen HE1, Jia Y1, Yamaryo-Botté Y2, Bisio H1, Zhang O3, Jemelin NK1, Marq JB1, Carruthers V3, Botté CY2, Soldati-Favre D4.

Author information

Abstract

The obligate intracellular lifestyle of apicomplexan parasites necessitates an invasive phase underpinned by timely and spatially controlled secretion of apical organelles termed micronemes. In Toxoplasma gondii, extracellular potassium levels and other stimuli trigger a signaling cascade culminating in phosphoinositide-phospholipase C (PLC) activation, which generates the second messengers diacylglycerol (DAG) and IP3 and ultimately results in microneme secretion. Here we show that a delicate balance between DAG and its downstream product, phosphatidic acid (PA), is essential for controlling microneme release. Governing this balance is the apicomplexan-specific DAG-kinase-1, which interconverts PA and DAG, and whose depletion impairs egress and causes parasite death. Additionally, we identify an acylated pleckstrin-homology (PH) domain-containing protein (APH) on the microneme surface that senses PA during microneme secretion and is necessary for microneme exocytosis. As APH is conserved in Apicomplexa, these findings highlight a potentially widely used mechanism in which key lipid mediators regulate microneme exocytosis.

Copyright © 2016 Elsevier Inc. All rights reserved.

KEYWORDS: 

Apicomplexa; Toxoplasma gondii; diacylglycerol kinase; invasion; microneme; phosphatidic acid; phosphatidic acid phosphatase; phosphoinositide-phospholipase C; pleckstrin homology domain; propranolol; regulated exocytosis

PMID:

 

26962945

 

[PubMed - as supplied by publisher]

Identification of Novel O-Linked Glycosylated Toxoplasma Proteins by Vicia villosa Lectin Chromatography

PLoS One. 2016 Mar 7;11(3):e0150561. doi: 10.1371/journal.pone.0150561. eCollection 2016.

Wang K¹, Peng ED¹, Huang AS¹, Xia D², Vermont SJ², Lentini G³, Lebrun M³, Wastling JM⁴, Bradley PJ¹.

Toxoplasma gondii maintains its intracellular life cycle using an extraordinary arsenal of parasite-specific organelles including the inner membrane complex (IMC), rhoptries, micronemes, and dense granules. While these unique compartments play critical roles in pathogenesis, many of their protein constituents have yet to be identified. We exploited the Vicia villosa lectin (VVL) to identify new glycosylated proteins that are present in these organelles. Purification of VVL-binding proteins by lectin affinity chromatography yielded a number of novel proteins that were subjected to further study, resulting in the identification of proteins from the dense granules, micronemes, rhoptries and IMC. We then chose to focus on three proteins identified by this approach, the SAG1 repeat containing protein SRS44, the rhoptry neck protein RON11 as well as a novel IMC protein we named IMC25. To assess function, we disrupted their genes by homologous recombination or CRISPR/Cas9. The knockouts were all successful, demonstrating that these proteins are not essential for invasion or intracellular survival. We also show that IMC25 undergoes substantial proteolytic processing that separates the C-terminal domain from the predicted glycosylation site. Together, we have demonstrated that lectin affinity chromatography is an efficient method of identifying new glycosylated parasite-specific proteins.

PMID:

26950937

[PubMed - in process]

Serum Albumin Stimulates Protein Kinase G-dependent Microneme Secretion in Toxoplasma gondii

J Biol Chem. 2016 Mar 1. pii: jbc.M115.700518. [Epub ahead of print]

Brown KM¹, Lourido S², Sibley LD³.

Microneme secretion is essential for motility, invasion, and egress in apicomplexan parasites. Although previous studies indicate that Ca2+ and cGMP control microneme secretion, little is known about how these pathways are naturally activated. Here we have developed genetically-encoded indicators for Ca2+ and microneme secretion to better define the signaling pathways that regulate these processes in Toxoplasma gondii. We found that microneme secretion was triggered in vitro by exposure to a single host protein, serum albumin. The natural agonist serum albumin induced microneme secretion in a protein kinase G-dependent manner that correlated with increased cGMP levels. Surprisingly, serum albumin acted independently of elevated Ca2+ and yet it was augmented by artificial agonists that raise Ca2+, such as ethanol. Furthermore, although ethanol elevated intracellular Ca2+, it alone was unable to trigger secretion without the presence of serum or serum albumin. This dichotomy was recapitulated by zaprinast, a phosphodiesterase inhibitor that elevated cGMP and separately increased Ca2+ in a protein kinase G-independent manner leading to microneme secretion. Taken together, these findings reveal that microneme secretion is centrally controlled by protein kinase G and that this pathway is further augmented by elevation of intracellular Ca2+.
Copyright © 2016, The American Society for Biochemistry and Molecular Biology.

KEYWORDS:

calcium; calcium imaging; calcium intracellular release; cyclic GMP (cGMP); protein kinase G (PKG); protein secretion; second messenger

PMID:

26933037

[PubMed - as supplied by publisher]

Using a Genetically Encoded Sensor to Identify Inhibitors of Toxoplasma gondii Ca2+ Signalling

J Biol Chem. 2016 Mar 1. pii: jbc.M115.703546. [Epub ahead of print]

Sidik SM¹, Hortua Triana MA², Paul AS³, El Bakkouri M⁴, Hackett CG¹, Tran F⁵, Westwood NJ⁵, Hui R⁴, Zuercher WJ⁶, Duraisingh MT³, Moreno SN², Lourido S⁷.

The lifecycles of apicomplexan parasites progress in accordance with fluxes in cytosolic Ca²⁺. Such fluxes are necessary for events like motility and egress from host cells. We used genetically-encoded Ca²⁺ indicators (GCaMPs) to develop a cell-based phenotypic screen for compounds that modulate Ca²⁺ signalling in the model apicomplexan ^{Toxoplasma gondii}. In doing so, we took advantage of the phosphodiesterase inhibitor zaprinast, which we show acts in part through cGMP-dependent protein kinase (PKG) to raise levels of cytosolic Ca²⁺. We define the pool of Ca²⁺ regulated by PKG to be a neutral store distinct from the endoplasmic reticulum. Screening a library of 823 ATP mimetics, we identify both inhibitors and enhancers of Ca²⁺ signalling. Two such compounds constitute novel PKG inhibitors, and prevent zaprinast from increasing cytosolic Ca²⁺. The enhancers identified are capable of releasing intracellular Ca²⁺ stores independently of zaprinast or PKG. One of these enhancers blocks parasite egress and invasion, and shows strong antiparasitic activity against T. gondii. The same compound inhibits invasion of the most lethal malaria parasite, Plasmodium falciparum. Inhibition of Ca²⁺-related phenotypes in these two apicomplexan parasites suggests that depletion of intracellular Ca²⁺ stores by the enhancer may be an effective antiparasitic strategy. These results establish a powerful new strategy for identifying compounds that modulate the essential parasite signalling pathways regulated by Ca²⁺, underscoring the importance of these pathways and the therapeutic potential of their inhibition.
Copyright © 2016, The American Society for Biochemistry and Molecular Biology.

KEYWORDS:

calcium; calcium intracellular release; drug screening; parasitology; protein kinase G (PKG); signal transduction

PMID:

26933036

[PubMed - as supplied by publisher]

The antigen-specific response to Toxoplasma gondii profilin, a TLR11/12 ligand, depends on its intrinsic adjuvant properties

Med Microbiol Immunol. 2016 Mar 2. [Epub ahead of print]

Hedhli D¹, Moiré N¹, Akbar H¹, Laurent F², Héraut B¹, Dimier-Poisson I¹, Mévélec MN³.

Agonists that activate Toll-like receptors (TLR) are potential vaccine adjuvants. In particular, Toxoplasma gondii profilin (TgPRF) is recognized by TLR11/12 to generate an inflammatory response. Unlike most TLR ligands, TgPRF is also a protein and can therefore simultaneously induce innate and adaptive immune responses. We found that variations in the conformation of TgPRF can affect its ability to induce a TLR11/12-dependent inflammatory response. The secreted recombinant T. gondii (S2-profilin), produced by Schneider 2 cells, has lost its ability to generate an IL-12 response. Reduction of the intramolecular disulfide bonds in S2-profilin rescued the TLR11/12-dependent IL-12 response. Immunization of mice with reduced S2-profilin induced strong cellular and humoral responses compared to mice immunized with unreduced S2-profilin. A mixed Th1/Th2 response was induced with both S2-profilins. However, a more polarized Th1-type response, which was consistent with the IgG2a-polarized humoral response, was observed with reduced S2-profilin. In conclusion, the intrinsic adjuvant properties of TgPRF had significant consequences on the immune response against TgPRF.

KEYWORDS:

Adjuvant properties; Immunization; Profilin; T. gondii; TLR

PMID:

26935827

[PubMed - as supplied by publisher]

The immunity-related GTPase Irga6 dimerizes in a parallel head-to-head fashion

BMC Biol. 2016 Mar 2;14(1):14. doi: 10.1186/s12915-016-0236-7.

Schulte K¹, Pawlowski N^2,3, Faelber K¹, Fröhlich C¹, Howard J^4,5, Daumke O⁶.

BACKGROUND:

The immunity-related GTPases (IRGs) constitute a powerful cell-autonomous resistance system against several intracellular pathogens. Irga6 is a dynamin-like protein that oligomerizes at the parasitophorous vacuolar membrane (PVM) of Toxoplasma gondii leading to its vesiculation. Based on a previous biochemical analysis, it has been proposed that the GTPase domains of Irga6 dimerize in an antiparallel fashion during oligomerization.

RESULTS:

We determined the crystal structure of an oligomerization-impaired Irga6 mutant bound to a non-hydrolyzable GTP analog. Contrary to the previous model, the structure shows that the GTPase domains dimerize in a parallel fashion. The nucleotides in the center of the interface participate in dimerization by forming symmetric contacts with each other and with the switch I region of the opposing Irga6 molecule. The latter contact appears to activate GTP hydrolysis by stabilizing the position of the catalytic glutamate 106 in switch I close to the active site. Further dimerization contacts involve switch II, the G4 helix and the trans stabilizing loop.

CONCLUSIONS:

The Irga6 structure features a parallel GTPase domain dimer, which appears to be a unifying feature of all dynamin and septin superfamily members. This study contributes important insights into the assembly and catalytic mechanisms of IRG proteins as prerequisite to understand their anti-microbial action.

PMID:

26934976

[PubMed - in process

Profiles of low complexity regions in Apicomplexa

BMC Evol Biol. 2016 Feb 29;16(1):47. doi: 10.1186/s12862-016-0625-0.

Battistuzzi FU¹, Schneider KA², Spencer MK³, Fisher D⁴, Chaudhry S^5,6, Escalante AA⁷.

BACKGROUND:

Low complexity regions (LCRs) are a ubiquitous feature in genomes and yet their evolutionary history and functional roles are unclear. Previous studies have shown contrasting evidence in favor of both neutral and selective mechanisms of evolution for different sets of LCRs suggesting that modes of identification of these regions may play a role in our ability to discern their evolutionary history. To further investigate this issue, we used a multiple threshold approach to identify species-specific profiles of proteome complexity and, by comparing properties of these sets, determine the influence that starting parameters have on evolutionary inferences.

RESULTS:

We find that, although qualitatively similar, quantitatively each species has a unique LCR profile which represents the frequency of these regions within each genome. Inferences based on these profiles are more accurate in comparative analyses of genome complexity as they allow to determine the relative complexity of multiple genomes as well as the type of repetitiveness that is most common in each. Based on the multiple threshold LCR sets obtained, we identified predominant evolutionary mechanisms at different complexity levels, which show neutral mechanisms acting on highly repetitive LCRs (e.g., homopolymers) and selective forces becoming more important as heterogeneity of the LCRs increases.

CONCLUSIONS:

Our results show how inferences based on LCRs are influenced by the parameters used to identify these regions. Sets of LCRs are heterogeneous aggregates of regions that include homo- and heteropolymers and, as such, evolve according to different mechanisms. LCR profiles provide a new way to investigate genome complexity across species and to determine the driving mechanism of their evolution.

PMID:

26923229

[PubMed - in process]

Induction of Autophagy interferes the tachyzoite to bradyzoite transformation of Toxoplasma gondii

Parasitology. 2016 Mar 1:1-7. [Epub ahead of print]

Li X¹, Chen DI², Hua Q³, Wan Y¹, Zheng L¹, Liu Y⁴, Lin J⁴, Pan C¹, Hu X⁴, Tan F¹.

Autophagy process in Toxoplasma gondii plays a vital role in regulating parasite survival or death. Thus, once having an understanding of certain effects of autophagy on the transformation of tachyzoite to bradyzoite this will allow us to elucidate the function of autophagy during parasite development. Herein, we used three TgAtg proteins involved in Atg8 conjugation system, TgAtg3, TgAtg7 and TgAtg8 to evaluate the autophagy level in tachyzoite and bradyzoite of Toxoplasma in vitro based on Pru TgAtg7-HA transgenic strains. We showed that both TgAtg3 and TgAtg8 were expressed at a significantly lower level in bradyzoites than in tachyzoites. Importantly, the number of parasites containing fluorescence-labelled TgAtg8 puncta was significantly reduced in bradyzoites than in tachyzoites, suggesting that autophagy is downregulated in Toxoplasma bradyzoite in vitro. Moreover, after treatment with drugs, bradyzoite-specific gene BAG1 levels decreased significantly in rapamycin-treated bradyzoites and increased significantly in 3-MA-treated bradyzoites in comparison with control bradyzoites, indicating that Toxoplasma autophagy is involved in the transformation of tachyzoite to bradyzoite in vitro. Together, it is suggested that autophagy may serve as a potential strategy to regulate the transformation.

KEYWORDS:

Toxoplasma gondii ; autophagy; bradyzoite; tachyzoite; transformation

PMID:

26928609

[PubMed - as supplied by publisher]

Effect of Toxoplasma gondii infection on the junctional complex of retinal pigment epithelial cells

Parasitology. 2016 Mar 1:1-8. [Epub ahead of print]

Nogueira AR¹, Leve F², Morgado-Diaz J², Tedesco RC³, Pereira MC¹.

Ocular toxoplasmosis is the most frequent cause of uveitis, leading to partial or total loss of vision, with the retina the main affected structure. The cells of the retinal pigment epithelium (RPE) play an important role in the physiology of the retina and formation of the blood-retinal barrier. Several pathogens induce barrier dysfunction by altering tight junction (TJ) integrity. Here, we analysed the effect of infection by Toxoplasma gondii on TJ integrity in ARPE-19 cells. Loss of TJ integrity was demonstrated in T. gondii-infected ARPE-19 cells, causing increase in paracellular permeability and disturbance of the barrier function of the RPE. Confocal microscopy also revealed alteration in the TJ protein occludin induced by T. gondii infection. Disruption of junctional complex was also evidenced by scanning and transmission electron microscopy. Cell-cell contact loss was noticed in the early stages of infection by T. gondii with the visualization of small to moderate intercellular spaces. Large gaps were mostly observed with the progression of the infection. Thus, our data suggest that the alterations induced by T. gondii in the structural organization of the RPE may contribute to retinal injury evidenced by ocular toxoplasmosis.

KEYWORDS:

Toxoplasma gondii ; junctional complex; retinal pigment epithelial cells

PMID:

26928468

[PubMed - as supplied by publisher]

The kynurenine pathway and parasitic infections that affect CNS function

Neuropharmacology. 2016 Feb 25. pii: S0028-3908(16)30064-8. doi: 10.1016/j.neuropharm.2016.02.029. [Epub ahead of print]

Hunt NH¹, Too LK², Khaw LT², Guo J², Hee L², Mitchell AJ², Grau GE², Ball HJ².

The kynurenine pathway of tryptophan metabolism has been implicated in brain function, immunoregulation, anti-microbial mechanisms and pregnancy. Some of these actions are due to depletion of tryptophan and others to the formation of biologically active metabolites. This review focuses on the roles of the kynurenine pathway in host responses during two parasitic diseases of major health and economic importance, malaria and toxoplasmosis, with an emphasis on their impacts on CNS function.
Copyright © 2016. Published by Elsevier Ltd.

KEYWORDS:

Malaria; Toxoplasma; cerebral malaria; indoleamine 2,3-dioxygenase; interferon-γ; kynurenine pathway; tryptophan

PMID:

26924710

[PubMed - as supplied by publisher]

The possible association between Toxoplasma gondii infection and risk of anxiety and cognitive disorders in BALB/c mice

Pathog Glob Health. 2015 Dec;109(8):369-76. doi: 10.1080/20477724.2015.1117742.

Mahmoudvand H¹, Ziaali N, Aghaei I, Sheibani V, Shojaee S, Keshavarz H, Shabani M.

There are conflicting reports concerning the association of Toxoplasma gondii infection with increased risk of mental disorders. This investigation will provide a good understanding about defining the possible association between T. gondii exposure and risk of anxiety and cognitive alterations. Besides, a secondary objective of this study was to determine the effect of pioglitazone administration on the possible alterations induced by T. gondii exposure. Male BALB/c mice were used for this study. The animal model of Toxoplasma infection was established by the intraperitoneal inoculation of 20-25 tissue cysts from Tehran strain of T. gondii. Pioglitazone (20 mg/kg, i.p.1/day) was administered to the animals for 2 weeks before behavioural tests. Behavioural tests including open-field, elevated plus-maze and passive avoidance learning were evaluated in the groups. Since cytokines were implicated as a contributing factor for mood disorders, the mRNA levels of TNF-α, IL-1β, IL-6 as well as inducible nitric oxide synthase (iNOs) were examined by real-time PCR. Findings demonstrated that T. gondii caused anxiety-like symptoms and impaired cognitive functions of the infected BALB/c mice, whereas pioglitazone, a peroxisome proliferator-activated receptor agonist, showed a promising effect against the cognitive impairments induced by Toxoplasma infection. The results also revealed that the mRNA levels of the aforementioned cytokines were significantly (p < 0.05) increased in the infected mice compared to the uninfected BALB/c ones. Pioglitazone can be offered as a potential neuroprotective agent in the treatment of patients with T. gondii infection that manifests anxiety and cognitive impairments; however, further studies are needed to clarify the exact mechanisms.

KEYWORDS:

Anxiety; Cognitive function; Cytokine; Learning; Pioglitazone; Toxoplasma gondii

PMID:

26924347

[PubMed - in process]

Host Mitochondrial Association Evolved in the Human Parasite Toxoplasma gondii via Neofunctionalization of a Gene Duplicate

Genetics. 2016 Feb 26. pii: genetics.115.186270. [Epub ahead of print]

Adomako-Ankomah Y1, English ED1, Danielson JJ1, Pernas LF2, Parker ML3, Boulanger MJ3, Dubey JP4, Boyle JP5.

Author information

Abstract

In Toxoplasma gondii, an intracellular parasite of humans and other animals, host mitochondrial association (HMA) is driven by a gene family that encodes multiple mitochondrial association factor 1 (MAF1) proteins. However, the importance of MAF1 gene duplication in the evolution of HMA is not understood, nor is the impact of HMA on parasite biology. Here we used within- and between-species comparative analysis to determine that the MAF1 locus is duplicated in T. gondii and its nearest extant relative Hammondia hammondi, but not another close relative, Neospora caninum. Using cross-species complementation, we determined that the MAF1 locus harbors multiple distinct paralogs that differ in their ability to mediate HMA, and that only T. gondii and H. hammondi harbor HMA+ paralogs. Additionally, we found that exogenous expression of an HMA+ paralog in T. gondii strains that do not normally exhibit HMA provides a competitive advantage over their wildtype counterparts during a mouse infection. These data indicate that HMA likely evolved by neofunctionalization of a duplicate MAF1 copy in the common ancestor of T. gondii and H. hammondi, and that the neofunctionalized gene duplicate is selectively advantageous.

Copyright © 2016, The Genetics Society of America.

KEYWORDS: 

Hammondia hammondi; Neospora caninum; Toxoplasma gondii; gene duplication; neofunctionalization

PMID:

 

26920761

 

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