Infectivity of Trypanosoma cruzi strains is associated with differential expression of surface glycoproteins with differential Ca2+ signalling activity.
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ABSTRACT:
SUBMITTER: Ruiz RC
PROVIDER: S-EPMC1219166 | biostudies-other | 1998 Feb
REPOSITORIES: biostudies-other
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Project description:BackgroundTSSA (Trypomastigote Small Surface Antigen) is an antigenic, adhesion molecule displayed on the surface of Trypanosoma cruzi trypomastigotes. TSSA displays substantial sequence identity to members of the TcMUC gene family, which code for the trypomastigote mucins (tGPI-mucins). In addition, TSSA bears sequence polymorphisms among parasite strains; and two TSSA variants expressed as recombinant molecules (termed TSSA-CL and TSSA-Sy) were shown to exhibit contrasting features in their host cell binding and signaling properties.Methods/principle findingsHere we used a variety of approaches to get insights into TSSA structure/function. We show that at variance with tGPI-mucins, which rely on their extensive O-glycoslylation to achieve their protective function, TSSA seems to be displayed on the trypomastigote coat as a hypo-glycosylated molecule. This has a functional correlate, as further deletion mapping experiments and cell binding assays indicated that exposition of at least two peptidic motifs is critical for the engagement of the 'adhesive' TSSA variant (TSSA-CL) with host cell surface receptor(s) prior to trypomastigote internalization. These motifs are not conserved in the 'non-adhesive' TSSA-Sy variant. We next developed transgenic lines over-expressing either TSSA variant in different parasite backgrounds. In strict accordance to recombinant protein binding data, trypomastigotes over-expressing TSSA-CL displayed improved adhesion and infectivity towards non-macrophagic cell lines as compared to those over-expressing TSSA-Sy or parental lines. These phenotypes could be specifically counteracted by exogenous addition of peptides spanning the TSSA-CL adhesion motifs. In addition, and irrespective of the TSSA variant, over-expression of this molecule leads to an enhanced trypomastigote-to-amastigote conversion, indicating a possible role of TSSA also in parasite differentiation.Conclusion/significanceIn this study we provided novel evidence indicating that TSSA plays an important role not only on the infectivity and differentiation of T. cruzi trypomastigotes but also on the phenotypic variability displayed by parasite strains.
| S-EPMC5568413 | biostudies-literature
Project description:Trypanosoma cruzi is a protozoan parasite and an etiological agent of Chagas disease. There is a wide variability in the clinical outcome of its infection, ranging from asymptomatic individuals to those with chronic fatal mega syndromes. Both parasite and host factors, as well as their interplay, are thought to be involved in the process.To evaluate the resistance to complement-mediated killing in two T. cruzi TcI strains with differential virulence and the subsequent effect on their infectivity in mammalian cells.Tissue-culture derived trypomastigotes of both strains were incubated in guinea pig serum and subjected to flow cytometry in order to determine their viability and complement activations. Trypomastigotes were also incubated on host cells monolayers in the presence of serum, and infectivity was evaluated under different conditions of complement pathway inhibition. Relative expression of the main parasite-specific complement receptors between the two strains was assessed by quantitative real-time polymerase chain reaction.In this work, we showed that two TcI strains, one with lower virulence (Ninoa) compared to the other (Qro), differ in their resistance to the lytic activity of complement system, hence causing a compromised ability of Ninoa strain to invade mammalian cells. These results correlate with the three-fold lower messenger RNA (mRNA) levels of complement regulatory protein (CRP), trypomastigote-decay acceleration factor (T-DAF), and complement C2 receptor inhibitor trispanning (CRIT) in Ninoa compared to those in Qro. On the other hand, calreticulin (CRT) mRNA and surface protein levels were higher in Ninoa strain and promoted its infectivity when the lectin pathway of the complement system was inhibited.This work suggests the complex interplay of CRP, T-DAF, CRIT, and CRT, and the diagnostic value of mRNA levels in the assessment of virulence potential of T. cruzi strains, particularly when dealing with isolates with similar genetic background.
| S-EPMC5851054 | biostudies-literature
Project description:BackgroundDiversity of T. cruzi strains is a central problem in Chagas disease research because of its correlation with the wide range of clinical manifestations and the biogeographical parasite distribution. The role played by parasite microdiversity in Chagas disease epidemiology is still debatable. Also awaits clarification whether such diversity is associated with the outcome of oral T. cruzi infection, responsible for frequent outbreaks of acute Chagas disease.Methods and findingsWe addressed the impact of microdiversity in oral T. cruzi infection, by comparative analysis of two strains, Y30 and Y82, both derived from Y strain, a widely used experimental model. Network genealogies of four nuclear genes (SSU rDNA, actin, DHFR-TS, EF1?) revealed that Y30 is closely related to Discrete Typing Unit TcII while Y82 is more closely related to TcVI, a group containing hybrid strains. Nevertheless, excepting one A-G transition at position 1463, Y30 and Y82 SSU rDNAs were identical. Y82 strain, expressing the surface molecule gp82, infected mice orally more efficiently than Y30, which expresses a related gp30 molecule. Both molecules are involved in lysosome exocytosis-dependent host cell invasion, but exhibit differential gastric mucin-binding capacity, a property critical for parasite migration toward the gastric mucosal epithelium. Upon oral infection of mice, the number of Y30 and Y82 parasites in gastric epithelial cells differed widely.ConclusionsWe conclude that metacyclic forms of gp82-expressing Y82 strain, closely related to TcVI, are better adapted than Y30 strain (TcII) to traverse the stomach mucous layer and establish oral route infection. The efficiency to infect target cell is the same because gp82 and gp30 strains have similar invasion-promoting properties. Unknown is whether differences in Y30 and Y82 are natural parasite adaptations or a product of lab-induced evolution by differential selection along the 60 years elapsed since the Y strain isolation.
| S-EPMC3464286 | biostudies-literature
Project description:Trypomastigote forms of Trypanosoma cruzi, the causative agent of Chagas Disease, shed extracellular vesicles (EVs) enriched with glycoproteins of the gp85/trans-sialidase (TS) superfamily and other ?-galactosyl (?-Gal)-containing glycoconjugates, such as mucins. Here, purified vesicles from T. cruzi strains (Y, Colombiana, CL-14 and YuYu) were quantified according to size, intensity and concentration. Qualitative analysis revealed differences in their protein and ?-galactosyl contents. Later, those polymorphisms were evaluated in the modulation of immune responses (innate and in the chronic phase) in C57BL/6 mice. EVs isolated from YuYu and CL-14 strains induced in macrophages higher levels of proinflammatory cytokines (TNF-? and IL-6) and nitric oxide via TLR2. In general, no differences were observed in MAPKs activation (p38, JNK and ERK 1/2) after EVs stimulation. In splenic cells derived from chronically infected mice, a different modulation pattern was observed, where Colombiana (followed by Y strain) EVs were more proinflammatory. This modulation was independent of the T. cruzi strain used in the mice infection. To test the functional importance of this modulation, the expression of intracellular cytokines after in vitro exposure was evaluated using EVs from YuYu and Colombiana strains. Both EVs induced cytokine production with the appearance of IL-10 in the chronically infected mice. A high frequency of IL-10 in CD4+ and CD8+ T lymphocytes was observed. A mixed profile of cytokine induction was observed in B cells with the production of TNF-? and IL-10. Finally, dendritic cells produced TNF-? after stimulation with EVs. Polymorphisms in the vesicles surface may be determinant in the immunopathologic events not only in the early steps of infection but also in the chronic phase.
| S-EPMC4662668 | biostudies-literature
Project description:BackgroundTrypanosoma cruzi strains are currently classified into six discrete typing units (DTUs) named TcI to VI. It is known that these DTUs have different geographical distribution, as well as biological features. TcI and TcII are major DTUs found in patients from northern and southern Latin America, respectively. Our hypothesis is that upon infection of human peripheral blood cells, Y strain (Tc II) and Col cl1.7 (Tc I), cause distinct immunological changes, which might influence the clinical course of Chagas disease.Methodology/principal findingsWe evaluated the infectivity of CFSE-stained trypomastigotes of Col cl1.7 and Y strain in human monocytes for 15 and 72 hours, and determined the immunological profile of lymphocytes and monocytes exposed to the different isolates using multiparameter flow cytometry. Our results showed a similar percentage and intensity of monocyte infection by Y and Col cl1.7. We also observed an increased expression of CD80 and CD86 by monocytes infected with Col cl1.7, but not Y strain. IL-10 was significantly higher in monocytes infected with Col cl1.7, as compared to Y strain. Moreover, infection with Col cl1.7, but not Y strain, led to an increased expression of IL-17 by CD8+ T cells. On the other hand, we observed a positive correlation between the expression of TNF-alpha and granzyme A only after infection with Y strain.Conclusion/significanceOur study shows that while Col cl1.7 induces higher monocyte activation and, at the same time, production of IL-10, infection with Y strain leads to a lower monocyte activation but higher inflammatory profile. These results show that TcI and TcII have a distinct immunological impact on human cells during early infection, which might influence disease progression.
| S-EPMC4492932 | biostudies-literature
Project description:We describe the characterization, purification, expression, and location of a 52-kDa protein secreted during interaction between the metacyclic form of Trypanosoma cruzi and its target host cell. The protein, which we have named MASP52, belongs to the family of mucin-associated surface proteins (MASPs). The highest levels of expression of both the protein and mRNA occur during the metacyclic and bloodstream trypomastigote stages, the forms that infect the vertebrate host cells. The protein is located in the plasma membrane and in the flagellar pockets of the epimastigote, metacyclic, and trypomastigote forms and is secreted into the medium at the point of contact between the parasite and the cell membrane, as well as into the host-cell cytosol during the amastigote stage. IgG antibodies specific against a synthetic peptide corresponding to the catalytic zone of MASP52 significantly reduce the parasite's capacity to infect the host cells. Furthermore, when the protein is adsorbed onto inert particles of bentonite and incubated with a nonphagocytic cell culture, the particles are able to induce endocytosis in the cells, which seems to demonstrate that MASP52 plays a role in a process whereby the trypomastigote forms of the parasite invade the host cell.
| S-EPMC3187265 | biostudies-literature
Project description:TcSMUG L products were recently identified as novel mucin-type glycoconjugates restricted to the surface of insect-dwelling epimastigote forms of Trypanosoma cruzi, the etiological agent of Chagas disease. The remarkable conservation of their predicted mature N-terminal region, which is exposed to the extracellular milieu, suggests that TcSMUG L products may be involved in structural and/or functional aspects of the interaction with the insect vector.Here, we investigated the putative roles of TcSMUG L mucins in both in vivo development and ex vivo attachment of epimastigotes to the luminal surface of the digestive tract of Rhodnius prolixus. Our results indicate that the exogenous addition of TcSMUG L N-terminal peptide, but not control T. cruzi mucin peptides, to the infected bloodmeal inhibited the development of parasites in R. prolixus in a dose-dependent manner. Pre-incubation of insect midguts with the TcSMUG L peptide impaired the ex vivo attachment of epimastigotes to the luminal surface epithelium, likely by competing out TcSMUG L binding sites on the luminal surface of the posterior midgut, as revealed by fluorescence microscopy.Together, these observations indicate that TcSMUG L mucins are a determinant of both adhesion of T. cruzi epimastigotes to the posterior midgut epithelial cells of the triatomine, and the infection of the insect vector, R. prolixus.
| S-EPMC3828161 | biostudies-other
Project description:Autophagy is a cell process that in normal conditions serves to recycle cytoplasmic components and aged or damaged organelles. The autophagic pathway has been implicated in many physiological and pathological situations, even during the course of infection by intracellular pathogens. Many compounds are currently used to positively or negatively modulate the autophagic response. Recently it was demonstrated that the polyamine spermidine is a physiological inducer of autophagy in eukaryotic cells. We have previously shown that the etiological agent of Chagas disease, the protozoan parasite Trypanosoma cruzi, interacts with autophagic compartments during host cell invasion and that preactivation of autophagy significantly increases host cell colonization by this parasite. In the present report we have analyzed the effect of polyamine depletion on the autophagic response of the host cell and on T. cruzi infectivity. Our data showed that depleting intracellular polyamines by inhibiting the biosynthetic enzyme ornithine decarboxylase with difluoromethylornithine (DFMO) suppressed the induction of autophagy in response to starvation or rapamycin treatment in two cell lines. This effect was associated with a decrease in the levels of LC3 and ATG5, two proteins required for autophagosome formation. As a consequence of inhibiting host cell autophagy, DFMO impaired T. cruzi colonization, indicating that polyamines and autophagy facilitate parasite infection. Thus, our results point to DFMO as a novel autophagy inhibitor. While other autophagy inhibitors such as wortmannin and 3-methyladenine are nonspecific and potentially toxic, DFMO is an FDA-approved drug that may have value in limiting autophagy and the spread of the infection in Chagas disease and possibly other pathological settings.
| S-EPMC3722317 | biostudies-literature
Project description:Trypanosoma cruzi, the etiologic agent of Chagas disease, is currently divided into six discrete typing units (DTUs), named TcI-TcVI. CL Brener, the reference strain of the T. cruzi genome project, is a hybrid with a genome assembled into 41 putative chromosomes. Gene copy number variation (CNV) is well documented as an important mechanism to enhance gene expression and variability in T. cruzi. Chromosomal CNV (CCNV) is another level of gene CNV in which whole blocks of genes are expanded simultaneously. Although the T. cruzi karyotype is not well defined, several studies have demonstrated a significant variation in the size and content of chromosomes between different T. cruzi strains. Despite these studies, the extent of diversity in CCNV among T. cruzi strains based on a read depth coverage analysis has not been determined.We identify the CCNV in T. cruzi strains from the TcI, TcII and TcIII DTUs, by analyzing the depth coverage of short reads from these strains using the 41 CL Brener chromosomes as reference. This study led to the identification of a broader extent of CCNV in T. cruzi than was previously speculated. The TcI DTU strains have very few aneuploidies, while the strains from TcII and TcIII DTUs present a high degree of chromosomal expansions. Chromosome 31, which is the only chromosome that is supernumerary in all six T. cruzi samples evaluated in this study, is enriched with genes related to glycosylation pathways, highlighting the importance of glycosylation to parasite survival.Increased gene copy number due to chromosome amplification may contribute to alterations in gene expression, which represents a strategy that may be crucial for parasites that mainly depend on post-transcriptional mechanisms to control gene expression.
| S-EPMC4491234 | biostudies-literature
Project description:The regulation of transcription in trypanosomes is unusual. To modulate protein synthesis during their complex developmental stages, these unicellular microorganisms rely largely on post-transcriptional gene expression pathways. These pathways include a plethora of RNA-binding proteins (RBPs) that modulate all steps of the mRNA life cycle in trypanosomes and help organize transcriptomes into clusters of post-transcriptional regulons. The aim of this work was to characterize an RNA regulon comprising numerous transcripts of trypomastigote-associated cell-surface glycoproteins that are preferentially expressed in the infective stages of the human parasite Trypanosoma cruzi. In vitro and in vivo RNA-binding assays disclosed that these glycoprotein mRNAs are targeted by the small trypanosomatid-exclusive RBP in T. cruzi, U-rich RBP 1 (TcUBP1). Overexpression of a GFP-tagged TcUBP1 in replicative parasites resulted in >10 times up-regulated expression of transcripts encoding surface proteins and in changes in their subcellular localization from the posterior region to the perinuclear region of the cytoplasm, as is typically observed in the infective parasite stages. Moreover, RT-quantitative PCR analysis of actively translated mRNAs by sucrose cushion fractionation revealed an increased abundance of these target transcripts in the polysome fraction of TcUBP1-induced samples. Because these surface proteins are involved in cell adherence or invasion during host infection, we also carried out in vitro infections with TcUBP1-transgenic trypomastigotes and observed that TcUBP1 overexpression significantly increases parasite infectivity. Our findings provide evidence for a role of TcUBP1 in trypomastigote stage-specific gene regulation important for T. cruzi virulence.
| S-EPMC6664183 | biostudies-literature