Friday, April 17, 2015

Inflammasomes in host response to protozoan parasites

2015 May;265(1):156-171. doi: 10.1111/imr.12291.
 
 
Inflammasomes are multimeric complexes of proteins that are assembled in the host cell cytoplasm in response to specific stress signals or contamination of the cytoplasm by microbial molecules. The canonical inflammasomes are composed of at least three main components: an inflammatory caspase (caspase-1, caspase-11), an adapter molecule (such as ASC), and a sensor protein (such as NLRP1, NLRP3, NLRP12, NAIP1, NAIP2, NAIP5, or AIM2). The sensor molecule determines the inflammasome specificity by detecting specific microbial products or cell stress signals. Upon activation, these molecular platforms facilitate restriction of microbial replication and trigger an inflammatory form of cell death called pyroptosis, thus accounting for the genesis of inflammatory processes. Inflammasome activation has been widely reported in response to pathogenic bacteria. However, recent reports have highlighted the important role of the inflammasomes in the host response to the pathogenesis of infections caused by intracellular protozoan parasites. Herein, we review the activation and specific roles of inflammasomes in recognition and host responses to intracellular protozoan parasites such as Trypanosoma cruzi, Toxoplasma gondii, Plasmodium spp., and Leishmania spp.
© 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

KEYWORDS:

Leishmania; Plasmodium; Toxoplasma; Trypanosoma; caspase-1; inflammasomes
PMID:
25879291
[PubMed - as supplied by publisher]

The NLRP1 inflammasomes

2015 May;265(1):22-34. doi: 10.1111/imr.12283.
 
 
Inflammasomes are cytosolic protein complexes that serve as platforms for the recruitment and activation of the pro-inflammatory CASPASE-1 protease. CASPASE-1 activation leads to processing and maturation of the cytokines interleukin-1β and interleukin-18 and a lytic form of cell death termed pyroptosis. Inflammasome assembly is initiated by cytosolic sensors in response to microbial infections. Many of these sensors, including NLRP1 (NLR family, pyrin domain containing 1), are described to form an inflammasome, but until recently, the mechanism of inflammasome activation and its physiological functions in host defense have remained unclear. In the last few years, important advances in our understanding of NLRP1 biology have been achieved. In this review, we discuss the activation of NLRP1 by various stimuli, including Bacillus anthracis lethal toxin, Toxoplasma gondii, muramyl dipeptide, and host intracellular ATP depletion. The role NLRP1 plays in pathogen recognition and resistance during infection is also discussed, as is the regulation of NLRP1 by host and viral proteins. We conclude by discussing the unexpected differences in the mechanism of NLRP1 inflammasome activation, as compared to the activation of other inflammasomes, such as the NAIP (NLR family, apoptosis inhibitory protein)/NLRC4 inflammasomes.
© 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

KEYWORDS:

NLRP1; caspase-1; inflammasome; innate immunity; lethal toxin
PMID:
25879281
[PubMed - as supplied by publisher]

Beyond the association. Toxoplasma gondii in schizophrenia, bipolar disorder, and addiction: systematic review and meta-analysis

2015 Apr 15. doi: 10.1111/acps.12423. [Epub ahead of print]
 

OBJECTIVE:

To perform a meta-analysis on studies reporting prevalence of Toxoplasma gondii (T. gondii) infection in any psychiatric disorder compared with healthy controls. Our secondary objective was to analyze factors possibly moderating heterogeneity.

METHOD:

A systematic search was performed to identify studies into T. gondii infection for all major psychiatric disorders versus healthy controls. Methodological quality, publication bias, and possible moderators were assessed.

RESULTS:

A total of 2866 citations were retrieved and 50 studies finally included. Significant odds ratios (ORs) with IgG antibodies were found in schizophrenia (OR 1.81, P < 0.00001), bipolar disorder (OR 1.52, P = 0.02), obsessive-compulsive disorder (OR 3.4, P < 0.001), and addiction (OR 1.91, P < 0.00001), but not for major depression (OR 1.21, P = 0.28). Exploration of the association between T. gondii and schizophrenia yielded a significant effect of seropositivity before onset and serointensity, but not IgM antibodies or gender. The amplitude of the OR was influenced by region and general seroprevalence. Moderators together accounted for 56% of the observed variance in study effects. After controlling for publication bias, the adjusted OR (1.43) in schizophrenia remained significant.

CONCLUSION:

These findings suggest that T. gondii infection is associated with several psychiatric disorders and that in schizophrenia reactivation of latent T. gondii infection may occur.
© 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

KEYWORDS:

Toxoplasma gondii ; bipolar disorder; meta-analysis; schizophrenia; substance abuse disorder
PMID:
25877655
[PubMed - as supplied by publisher]

Thursday, April 16, 2015

Comprehensive Evaluation of Toxoplasma gondii VEG and Neospora caninum LIV Genomes with Tachyzoite Stage Transcriptome and Proteome Defines Novel Transcript Features

2015 Apr 13;10(4):e0124473. doi: 10.1371/journal.pone.0124473.
 
 
Toxoplasma gondii is an important protozoan parasite that infects all warm-blooded animals and causes opportunistic infections in immuno-compromised humans. Its closest relative, Neospora caninum, is an important veterinary pathogen that causes spontaneous abortion in livestock. Comparative genomics of these two closely related coccidians has been of particular interest to identify genes that contribute to varied host cell specificity and disease. Here, we describe a manual evaluation of these genomes based on strand-specific RNA sequencing and shotgun proteomics from the invasive tachyzoite stages of these two parasites. We have corrected predicted structures of over one third of the previously annotated gene models and have annotated untranslated regions (UTRs) in over half of the predicted protein-coding genes. We observe distinctly long UTRs in both the organisms, almost four times longer than other model eukaryotes. We have also identified a putative set of cis-natural antisense transcripts (cis-NATs) and long intergenic non-coding RNAs (lincRNAs). We have significantly improved the annotation quality in these genomes that would serve as a manually curated dataset for Toxoplasma and Neospora research communities.
PMID:
25875305
[PubMed - as supplied by publisher]

Genotype-associated Arginase 1 Expression in Rat Peritoneal Macrophages induced by Toxoplasma gondii

2015 Apr 14. [Epub ahead of print]
 
 
Toxoplasma gondii induces polarization of mouse macrophages, including both classically activated macrophage (M1) and alternatively activated macrophage (M2) in a genotype-related manner. Here we presented a novel result that the Wh6 strain with type Chinese 1, which is predominantly prevalent in China, induced Arg1 expression in a STAT6 dependent manner in primary rat peritoneal macrophages, compared to the PRU stain with typeⅡ that elicited a high expression of Arg1 in a C/EBPβ dependent manner. Additional experiment revealed that dexamethasone inhibited Arg1 expression in rat macrophage in both contexts. Our data suggested that Arg1 expression, which is abundant in polarized M2 cells, is associated with strain/genotype difference in varying pathways.
PMID:
25872571
[PubMed - as supplied by publisher]

A serine-arginine-rich (SR) splicing factor modulates alternative splicing of over a thousand genes in Toxoplasma gondii

2015 Apr 13. pii: gkv311. [Epub ahead of print]
 
 
Single genes are often subject to alternative splicing, which generates alternative mature mRNAs. This phenomenon is widespread in animals, and observed in over 90% of human genes. Recent data suggest it may also be common in Apicomplexa. These parasites have small genomes, and economy of DNA is evolutionarily favoured in this phylum. We investigated the mechanism of alternative splicing in Toxoplasma gondii, and have identified and localized TgSR3, a homologue of ASF/SF2 (alternative-splicing factor/splicing factor 2, a serine-arginine-rich, or SR protein) to a subnuclear compartment. In addition, we conditionally overexpressed this protein, which was deleterious to growth. qRT-PCR was used to confirm perturbation of splicing in a known alternatively-spliced gene. We performed high-throughput RNA-seq to determine the extent of splicing modulated by this protein. Current RNA-seq algorithms are poorly suited to compact parasite genomes, and hence we complemented existing tools by writing a new program, GeneGuillotine, that addresses this deficiency by segregating overlapping reads into distinct genes. In order to identify the extent of alternative splicing, we released another program, JunctionJuror, that detects changes in intron junctions. Using this program, we identified about 2000 genes that were constitutively alternatively spliced in T. gondii. Overexpressing the splice regulator TgSR3 perturbed alternative splicing in over 1000 genes.
© The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.
PMID:
25870410
[PubMed - as supplied by publisher]

Postdoctoral positions available

We are currently looking for four postdoctoral scholars to work on three different NIH-funded projects investigating host-parasite interactions. We are studying the innate and adaptive immune response to Toxoplasma and, in turn, how Toxoplasma modulates the host cell. We are particularly interested in individual differences in host susceptibility to Toxoplasma and Toxoplasma strain differences in interacting with the host. The Vet School at UC Davis is ranked as the best in the country and our location in the Vet School will allow us to investigate Toxoplasma proteins that play a role in its sexual cycle in cats and to perform genetic crosses to identify the molecular basis for phenotypic strain differences. More information can be found on our website:  www.saeijlab.com

We are looking for team players that are excited about studying host-parasite interactions and have excellent communication skills. Previous experience with molecular biology, biochemistry, immunology, genetics or computational skills is a plus.

UC Davis is located in the town of Davis, which is ranked as one of the best towns to live in in the nation. Davis is close to San Francisco, Sacramento, the Napa/Sonoma wine country and the Lake Tahoe ski area. UC Davis offers excellent benefits and multiple opportunities for professional development for postdocs (see https://gradstudies.ucdavis.edu/postdoctoral).

Interested candidates can submit a current C.V. and names of three references to: Dr. Jeroen Saeij (jsaeij@mit.edu), Associate Professor of Biology.

Tuesday, April 14, 2015

A large scale proteogenomics study of apicomplexan pathogens - Toxoplasma gondii and Neospora caninum

 2015 Apr 13. doi: 10.1002/pmic.201400553. [Epub ahead of print]

Abstract

Proteomics data can supplement genome annotation efforts, for example being used to confirm gene models or correct gene annotation errors. Here we present a large scale proteogenomics study of two important apicomplexan pathogens: Toxoplasma gondii and Neospora caninum. We queried proteomics data against a panel of official and alternate gene models generated directly from RNASeq data, using several newly generated and some previously published MS data sets for this meta-analysis. We identified a total of 201996 and 39953 peptide-spectrum matches (PSMs) for T. gondii and N. caninum respectively at a 1% peptide false discovery rate threshold. This equated to the identification of 30494 distinct peptide sequences and 2921 proteins (matches to official gene models) for T. gondii, and 8911 peptides / 1273 proteins for N. caninum following stringent protein-level thresholding. We have also identified 289 and 140 loci for T. gondii and N. caninum respectively which mapped to RNA-Seq derived gene models used in our analysis and apparently absent from the official annotation (release 10 from EuPathDB) of these species. We present several examples in our study where the RNA-Seq evidence can help in correction of the current gene model and can help in discovery of potential new genes. The findings of this study have been integrated into the EuPathDB. The data have been deposited to the ProteomeXchange with identifiers PXD000297and PXD000298. This article is protected by copyright. All rights reserved.
This article is protected by copyright. All rights reserved.

KEYWORDS: 

Gene annotation; MS/MS; N. caninum; OMSSA; Open-source; Proteogenomics; T. gondii; X!Tandem
PMID:
 
25867681
 
[PubMed - as supplied by publisher]

Forward Genetics Screens Using Macrophages to Identify Toxoplasma gondii Genes Important for Resistance to IFN-γ-Dependent Cell Autonomous Immunity

 2015 Mar 12;(97). doi: 10.3791/52556.

Abstract

Toxoplasma gondii, the causative agent of toxoplasmosis, is an obligate intracellular protozoan pathogen. The parasite invades and replicates within virtually any warm blooded vertebrate cell type. During parasite invasion of a host cell, the parasite creates a parasitophorous vacuole (PV) that originates from the host cell membrane independent of phagocytosis within which the parasite replicates. While IFN-dependent-innate and cell mediated immunity is important for eventual control of infection, innate immune cells, including neutrophils, monocytes and dendritic cells, can also serve as vehicles for systemic dissemination of the parasite early in infection. An approach is described that utilizes the host innate immune response, in this case macrophages, in a forward genetic screen to identify parasite mutants with a fitness defect in infected macrophages following activation but normal invasion and replication in naïve macrophages. Thus, the screen isolates parasite mutants that have a specific defect in their ability to resist the effects of macrophage activation. The paper describes two broad phenotypes of mutant parasites following activation of infected macrophages: parasite stasis versus parasite degradation, often in amorphous vacuoles. The parasite mutants are then analyzed to identify the responsible parasite genes specifically important for resistance to induced mediators of cell autonomous immunity. The paper presents a general approach for the forward genetics screen that, in theory, can be modified to target parasite genes important for resistance to specific antimicrobial mediators. It also describes an approach to evaluate the specific macrophage antimicrobial mediators to which the parasite mutant is susceptible. Activation of infected macrophages can also promote parasite differentiation from the tachyzoite to bradyzoite stage that maintains chronic infection. Therefore, methodology is presented to evaluate the importance of the identified parasite gene to establishment of chronic infection. 
PMID:
 
25867017
 
[PubMed - in process]

Saturday, April 11, 2015

Anti-Toxoplasma Activity of Natural Products: A Review

 2015 Apr 10. [Epub ahead of print]

Abstract

Toxoplasma gondii is an intracellular parasite widely distributed in nature. Infection is asymptomatic in immunocompetent individuals; however, various clinical manifestations may occur in immunocompromised individuals. Although there are medications for the treatment of toxoplasmosis, such as pyrimethamine and sulfonamide, they cannot always be used due to adverse reactions or to therapeutic failures related to intolerance or malabsorption of drugs and to parasite drug resistance. In recent years, the search for new antimicrobial agents derived from plants has intensified because a quarter of synthetic drugs that are currently prescribed have been isolated from a plant source, demonstrating that natural products are important in the development of new drugs. A systematic literature search was conducted to evaluate the use of natural products as an alternative for the treatment of T. gondii infection. The search was conducted for the 2000-2014 period in Science Direct, Scopus, PubMed, EMBASE, and SciELO databases, using the following MeSH terms: anti-Toxoplasma activity, toxoplasma AND natural products, toxoplasma AND plant extracts. Ethnobotanical and experimental evidence (in vitro/in vivo) was found supporting the use of natural products as a source for the discovery of new therapies against T. gondii.
PMID:
 
25858302
 
[PubMed - as supplied by publisher]

Chronic Toxoplasma gondii in Nurr1-Null Heterozygous Mice Exacerbates Elevated Open Field Activity

 2015 Apr 9;10(4):e0119280. doi: 10.1371/journal.pone.0119280. eCollection 2015.

Abstract

Latent infection with Toxoplasma gondii is common in humans (approximately 30% of the global population) and is a significant risk factor for schizophrenia. Since prevalence of T. gondii infection is far greater than prevalence of schizophrenia (0.5-1%), genetic risk factors are likely also necessary to contribute to schizophrenia. To test this concept in an animal model, Nurr1-null heterozygous (+/-) mice and wild-type (+/+) mice were evaluate using an emergence test, activity in an open field and with a novel object, response to bobcat urine and prepulse inhibition of the acoustic startle response (PPI) prior to and 6 weeks after infection with T. gondii. In the emergence test, T. gondii infection significantly decreased the amount of time spent in the cylinder. Toxoplasma gondii infection significantly elevated open field activity in both +/+ and +/- mice but this increase was significantly exacerbated in +/- mice. T. gondii infection reduced PPI in male +/- mice but this was not statistically significant. Aversion to bobcat urine was abolished by T. gondii infection in +/+ mice. In female +/- mice, aversion to bobcat urine remained after T. gondii infection while the male +/- mice showed no aversion to bobcat urine. Antibody titers of infected mice were a critical variable associated with changes in open field activity, such that an inverted U shaped relationship existed between antibody titers and the percent change in open field activity with a significant increase in activity at low and medium antibody titers but no effect at high antibody titers. These data demonstrate that the Nurr1 +/- genotype predisposes mice to T. gondii-induced alterations in behaviors that involve dopamine neurotransmission and are associated with symptoms of schizophrenia. We propose that these alterations in murine behavior were due to further exacerbation of the altered dopamine neurotransmission in Nurr1 +/- mice. 
PMID:
 
25855987
 
[PubMed - in process] 

Thursday, April 09, 2015

α2u-globulins mediate manipulation of host attractiveness in Toxoplasma gondii-Rattus novergicus association

2015 Mar 13. doi: 10.1038/ismej.2015.33. [Epub ahead of print]
 
 
Uninfected female rats (Rattus novergicus) exhibit greater attraction to the males infected with protozoan parasite Toxoplasma gondii. This phenomenon is contrary to the aversion towards infected males observed in multitude of other host-parasite associations. In this report, we describe a proximate mechanism for this anomaly. We demonstrate that T. gondii infection enhances hepatic production and urinary excretion of α2u-globulins in rats. We further demonstrate that α2u-globulins are sufficient to recapitulate male sexual attractiveness akin to effects of the infection. This manipulation possibly results in greater horizontal transmission of this parasite between the infected male and the uninfected female. It supports the notion that in some evolutionary niches parasites can alter host sexual signaling, likely leading to an increased rate of sexual transmission.
PMID:
25853804
[PubMed - as supplied by publisher]

Wednesday, April 08, 2015

AMA1-deficient Toxoplasma parasites transiently colonize mice and trigger an innate immune response that proceeds to long lasting protective immunity

2015 Apr 6. pii: IAI.02606-14. [Epub ahead of print]
 
 
The Apical Membrane Antigen 1 (AMA1) protein was believed to be essential for the perpetuation of two Apicomplexa parasite genera Plasmodium and Toxoplasma, until we genetically engineered viable parasites lacking AMA1. The reduction in invasiveness of the T. gondii RH-AMA1KO tachyzoite population, in vitro, raised key questions about the outcome of these tachyzoites once inoculated in the peritoneal cavity of mice. In this study, we used the AMNIS technology to simultaneously quantify and image the parasitic process driven by AMA1KO tachyzoites. We report their ability to colonize and multiply in mesothelial cells and in both resident and recruited leucocytes. While the RH-AMA1KO population amplification is rapidly lethal in immunocompromised mice, it is controlled in immunocompetent hosts where a combination of immune cells sense parasites and secrete pro-inflammatory cytokines. This innate response further leads to a long-lasting immunoprotective status against a secondary challenge by high inocula of the homologous type I or a distinct type II T. gondii genotypes. While AMA1 is definitively not an essential protein for tachyzoite entry and multiplication in host cells, it clearly assists the expansion of parasite population in vivo.
Copyright © 2015, American Society for Microbiology. All Rights Reserved.
PMID:
25847964
[PubMed - as supplied by publisher]

Distinct contribution of Toxoplasma gondii rhomboid proteases 4 and 5 to micronemal protein protease 1 activity during invasion

2015 Apr 2. doi: 10.1111/mmi.13021. [Epub ahead of print]
 
 
Host cell entry by the Apicomplexa is associated with the sequential secretion of invasion factors from specialized apical organelles. Secretion of micronemal proteins (MICs) complexes by Toxoplasma gondii facilitates parasite gliding motility, host cell attachment and entry, as well as egress from infected cells. The shedding of MICs during these steps is mediated by micronemal protein proteases MPP1, MPP2, and MPP3. The constitutive activity of MPP1 leads to the cleavage of transmembrane MICs and is linked to the surface rhomboid protease 4 (ROM4) and possibly to rhomboid protease 5 (ROM5). To determine their importance and respective contribution to MPP1 activity, in this study ROM4 and ROM5 genes were abrogated using Cre-recombinase and CRISPR-Cas9 nuclease, respectively, and shown to be dispensable for parasite survival. Parasites lacking ROM4 predominantly engage in twirling motility and exhibit enhanced attachment and impaired invasion, whereas intracellular growth and egress are not affected. The substrates MIC2 and MIC6 are not cleaved and accumulate on the rom4-ko parasite surface. In contrast, intramembrane cleavage of AMA1 is reduced but not completely abolished. Shedding of MICs and invasion are not altered in the absence of ROM5 however this protease responsible for the residual cleavage of AMA1, is able to cleave other AMA family members and exhibits a detectable contribution to invasion in the absence of ROM4.
This article is protected by copyright. All rights reserved.

KEYWORDS:

Toxoplasma gondii; invasion; microneme secretion; motility; proteolytic cleavage; rhomboid protease; shedding
PMID:
25846828
[PubMed - as supplied by publisher]

Thursday, April 02, 2015

Infection by Toxoplasma gondii, a severe parasite in neonates and AIDS patients, causes impaired anion secretion in airway epithelia

2015 Mar 23. pii: 201503474. [Epub ahead of print]
 
 
The airway epithelia initiate and modulate the inflammatory responses to various pathogens. The cystic fibrosis transmembrane conductance regulator-mediated Cl- secretion system plays a key role in mucociliary clearance of inhaled pathogens. We have explored the effects of Toxoplasma gondii, an opportunistic intracellular protozoan parasite, on Cl- secretion of the mouse tracheal epithelia. In this study, ATP-induced Cl- secretion indicated the presence of a biphasic short-circuit current (Isc) response, which was mediated by a Ca2+-activated Cl- channel (CaCC) and the cystic fibrosis transmembrane conductance regulator. However, the ATP-evoked Cl- secretion in T. gondii-infected mouse tracheal epithelia and the elevation of [Ca2+]i in T. gondii-infected human airway epithelial cells were suppressed. Quantitative reverse transcription-PCR revealed that the mRNA expression level of the P2Y2 receptor (P2Y2-R) increased significantly in T. gondii-infected mouse tracheal cells. This revealed the influence that pathological changes in P2Y2-R had on the downstream signal, suggesting that P2Y2-R was involved in the mechanism underlying T. gondii infection in airways. These results link T. gondii infection as well as other pathogen infections to Cl- secretion, via P2Y2-R, which may provide new insights for the treatment of pneumonia caused by pathogens including T. gondii.

KEYWORDS:

ATP; Cl− secretion; P2Y2 receptor; cystic fibrosis; pneumonia
PMID:
25831498
[PubMed - as supplied by publisher]