Parasite Cell Surface Research Articles

Role of a novel uropod-like cell membrane protrusion in the pathogenesis of the parasite Trichomonas vaginalis.

Trichomonas vaginalis causes trichomoniasis, the most common non-viral sexually transmitted disease worldwide. As an extracellular parasite, adhesion to host cells is essential for the development of infection. During attachment, the parasite changes its tear ovoid shape to a flat ameboid form, expanding the contact surface and migrating through tissues. Here, we have identified a novel structure formed at the posterior pole of adherent parasite strains, resembling the previously described uropod, which appears to play a pivotal role as an anchor during the attachment process. Moreover, our research demonstrates that the overexpression of the tetraspanin T. vaginalis TSP5 protein (TvTSP5), which is localized on the cell surface of the parasite, notably enhances the formation of this posterior anchor structure in adherent strains. Finally, we demonstrate that parasites that overexpress TvTSP5 possess an increased ability to adhere to host cells, enhanced aggregation and reduced migration on agar plates. Overall, these findings unveil novel proteins and structures involved in the intricate mechanisms of T. vaginalis interactions with host cells.

Silver(I) and Copper(II) 1,10-Phenanthroline-5,6-dione Complexes as Promising Antivirulence Strategy against Leishmania: Focus on Gp63 (Leishmanolysin).

Leishmaniasis, caused by protozoa of the genus Leishmania, encompasses a group of neglected diseases with diverse clinical and epidemiological manifestations that can be fatal if not adequately and promptly managed/treated. The current chemotherapy options for this disease are expensive, require invasive administration and often lead to severe side effects. In this regard, our research group has previously reported the potent anti-Leishmania activity of two coordination compounds (complexes) derived from 1,10-phenanthroline-5,6-dione (phendione): [Cu(phendione)3].(ClO4)2.4H2O and [Ag(phendione)2].ClO4. The present study aimed to evaluate the effects of these complexes on leishmanolysin (gp63), a virulence factor produced by all Leishmania species that plays multiple functions and is recognized as a potential target for antiparasitic drugs. The results showed that both Ag-phendione (-74.82 kcal/mol) and Cu-phendione (-68.16 kcal/mol) were capable of interacting with the amino acids comprising the active site of the gp63 protein, exhibiting more favorable interaction energies compared to phendione alone (-39.75 kcal/mol) or 1,10-phenanthroline (-45.83 kcal/mol; a classical gp63 inhibitor) as judged by molecular docking assay. The analysis of kinetic parameters using the fluorogenic substrate Z-Phe-Arg-AMC indicated Vmax and apparent Km values of 0.064 µM/s and 14.18 µM, respectively, for the released gp63. The effects of both complexes on gp63 proteolytic activity were consistent with the in silico assay, where Ag-phendione exhibited the highest gp63 inhibition capacity against gp63, with an IC50 value of 2.16 µM and the lowest inhibitory constant value (Ki = 5.13 µM), followed by Cu-phendione (IC50 = 163 µM and Ki = 27.05 µM). Notably, pretreatment of live L. amazonensis promastigotes with the complexes resulted in a significant reduction in the expression of gp63 protein, including the isoforms located on the parasite cell surface. Both complexes markedly decreased the in vitro association indexes between L. amazonensis promastigotes and THP-1 human macrophages; however, this effect was reversed by the addition of soluble gp63 molecules to the interaction medium. Collectively, our findings highlight the potential use of these potent complexes in antivirulence therapy against Leishmania, offering new insights for the development of effective treatments for leishmaniasis.

Apical Secretory Glycoprotein Complex Contributes to Cell Attachment and Entry by Cryptosporidium parvum.

Cryptosporidium parvum is an enteric pathogen that invades epithelial cells in the intestine, where it resides at the apical surface in a unique epicellular location. Compared with those of related apicomplexan parasites, the processes of host cell attachment and invasion by C. parvum are poorly understood. The streamlined C. parvum genome contains numerous mucin-like glycoproteins, several of which have previously been shown to mediate cell attachment, although the majority are unstudied. Here, we identified the antigens recognized by monoclonal antibody (MAb) 1A5, which stains the apical end of sporozoites and mature merozoites. Immunoprecipitation with MAb 1A5 followed by mass spectrometry identified a heterodimer comprised of paralogous proteins which are related to additional orthologs in the genome of C. parvum and related species. Paralogous glycoproteins recognized by MAb 1A5 heterodimerize as a complex displayed on the parasite surface, and they also interact with lectins that suggest that they contain mucin-like, O-linked oligosaccharides. Although the gene encoding one of the paralogs was readily disrupted by CRISPR/Cas9 gene editing, its partner, which contains a mucin-like domain related to GP900, was refractory to deletion. Combined with the ability of MAb 1A5 to partially neutralize host cell attachment by sporozoites, these findings define a new family of secretory glycoproteins that participate in cell invasion by Cryptosporidium spp. IMPORTANCE Although Cryptosporidium is extremely efficient at penetrating mucus and invading epithelial cells in the intestine, the mechanism of cell attachment is poorly understood. To expand our understanding of this process, we characterized the antigens recognized by a monoclonal antibody that stains the apical end of invasive stages called sporozoites and merozoites. Our studies identify a family of glycoproteins that form heterodimers on the parasite cell surface to facilitate host cell attachment and entry. By further defining the role of mucin-like glycoproteins in host cell attachment, our studies may lead to strategies to disrupt cell adhesion and thereby decrease infection.

Molecular Assessment of Domain I of Apical Membrane Antigen I Gene in Plasmodium falciparum: Implications in Plasmodium Invasion, Taxonomy, Vaccine Development, and Drug Discovery.

Given its global morbidity and mortality rates, malaria continues to be a major public health concern. Despite significant progress in the fight against malaria, efforts to control and eradicate the disease globally are in jeopardy due to lack of a universal vaccine. The conserved short peptide sequences found in Domain I of Plasmodium falciparum apical membrane antigen 1 (PfAMA1), which are exposed on the parasite cell surface and in charge of Plasmodium falciparum invasion of host cells, make PfAMA1 a promising vaccine candidate antigen. The precise amino acids that make up these conserved short peptides are still unknown, and it is still difficult to pinpoint the molecular processes by which PfAMA1 interacts with the human host cell during invasion. The creation of a universal malaria vaccine based on the AMA1 antigen is challenging due to these knowledge limitations. This study used genome mining techniques to look for these particular short peptides in PfAMA1. Thirty individuals with Plasmodium falciparum malaria had blood samples taken using Whatman's filter papers. DNA from the parasite was taken out using the Chelex technique. Domain I of the Plasmodium falciparum AMA1 gene was amplified using nested polymerase chain reactions, and the amplified products were removed, purified, and sequenced. The DNA sequence generated was converted into the matching amino acid sequence using bioinformatic techniques. These amino acid sequences were utilized to search for antigenic epitopes, therapeutic targets, and conserved short peptides in Domain I of PfAMA1. The results of this investigation shed important light on the molecular mechanisms behind Plasmodium invasion of host cells, a potential PfAMA1 vaccine antigen sequence, and prospective malaria treatment options in the future. Our work offers fresh information on malaria medication and vaccine research that has not been previously discussed.

Global overview of Leishmania virulence factors, and the role of GP63 in promastigotes

Protozoa parasites of the genus Leishmania have a great ability to avoid damage in the hostile environments they encounter throughout their life cycle within the host’s body. Parasites have developed many virulence factors to ensure their persistence and replication within the host, and the first role of these factors is to attenuate the host’s defenses against them through innate and adaptive immunity, as evidence indicates that the determinants of parasite virulence are responsible for evading the host’s defenses, allowing these organisms to survive on Alive in the host’s hostile immune environment. Understanding the molecular details of how these pathogens persist with impunity under extreme conditions is beginning to begin. The fact that Leishmania parasites have adapted not only to survive but are likely to reproduce is due to the protection afforded by specialized molecules on the parasite’s cell surface. Although recent years have seen clear and significant progress in the research on Leishmania in different directions, many issues have yet to be clarified. The surface of all parasites, especially protozoa, usually undergoes pronounced changes during their life cycle. It is of particular interest in the case of protozoan parasites of the genus Leishmania whose surface is exposed to different and hostile environments within vertebrate and invertebrate organisms. Because of the importance that the cell surface of pathogenic parasites plays in their interaction with the host for survival, some efforts have been devoted to their characterization. This review aims to give an overview of the main virulence factors that contribute to parasite survival and survival. We have attempted to provide a brief picture of the factors that influence the interaction of the parasite in its host, further highlighting GP63 as a critical virulence factor affecting macrophage physiology as well as the functioning of the immune system.

Screening of a Library of Recombinant Schistosoma mansoni Proteins With Sera From Murine and Human Controlled Infections Identifies Early Serological Markers.

BackgroundSchistosomiasis is a major global health problem caused by blood-dwelling parasitic worms, which is currently tackled primarily by mass administration of the drug praziquantel. Appropriate drug treatment strategies are informed by diagnostics that establish the prevalence and intensity of infection, which, in regions of low transmission, should be highly sensitive.MethodsTo identify sensitive new serological markers of Schistosoma mansoni infections, we have compiled a recombinant protein library of parasite cell-surface and secreted proteins expressed in mammalian cells.ResultsTogether with a time series of sera samples from volunteers experimentally infected with a defined number of male parasites, we probed this protein library to identify several markers that can detect primary infections with as low as 10 parasites and as early as 5 weeks postinfection.ConclusionsThese new markers could be further explored as valuable tools to detect ongoing and previous S mansoni infections, including in endemic regions where transmission is low.

Reduced expression of a rhomboid protease, EhROM1, correlates with changes in the submembrane distribution and size of the Gal/GalNAc lectin subunits in the human protozoan parasite, Entamoeba histolytica.

Entamoeba histolytica is a food- and waterborne parasite that causes amebic dysentery and amoebic liver abscesses. Adhesion is one of the most important virulence functions as it facilitates motility, colonization of host, destruction of host tissue, and uptake of nutrients by the parasite. The parasite cell surface adhesin, the Gal/GalNAc lectin, facilitates parasite-host interaction by binding to galactose or N-acetylgalactosamine residues on host components. It is composed of heavy (Hgl), intermediate (Igl), and light (Lgl) subunits. Igl is constitutively localized to lipid rafts (cholesterol-rich membrane domains), whereas Hgl and Lgl transiently associate with rafts. When all three subunits are localized to rafts, galactose-sensitive adhesion is enhanced. Thus, submembrane location may regulate the function of this adhesion. Rhomboid proteases are a conserved family of intramembrane proteases that also participate in the regulation of parasite-host interactions. In E. histolytica, one rhomboid protease, EhROM1, cleaves Hgl as a substrate, and knockdown of its expression inhibits parasite-host interactions. Since rhomboid proteases are found within membranes, it is not surprising that lipid composition regulates their activity and enzyme-substrate binding. Given the importance of the lipid environment for both rhomboid proteases and the Gal/GalNAc lectin, we sought to gain insight into the relationship between rhomboid proteases and submembrane location of the lectin in E. histolytica. We demonstrated that EhROM1, itself, is enriched in highly buoyant triton-insoluble membranes reminiscent of rafts. Reducing rhomboid protease activity, either pharmacologically or genetically, correlated with an enrichment of Hgl and Lgl in rafts. In a mutant cell line with reduced EhROM1 expression, there was also a significant augmentation of the level of all three Gal/GalNAc subunits on the cell surface and an increase in the molecular weight of Hgl and Lgl. Overall, the study provides insight into the molecular mechanisms governing parasite-host adhesion for this pathogen.

Signal peptide recognition in Trypanosoma cruzi GP82 adhesin relies on its localization at protein N-terminus

Trypanosoma cruzi, the causative agent of Chagas disease, has a dense coat of GPI-anchored virulence factors. T. cruzi GPI-anchored adhesin GP82 is encoded by a repertoire of transcripts containing several in-frame initiation codons located up-stream from that adjacent to the predicted signal peptide (SP). Transfection of T. cruzi epimastigotes with constructs encoding GP82 starting at the SP or from the farthest up-stream methionine confirmed protein expression on the parasite cell surface, comparable to the native GP82. Proteins were fully functional, inducing parasite adhesion to HeLa cells and lysosome mobilization, events required for parasite invasion. Transgenic and native GP82 proteins showed indistinguishable electrophoretic mobility, suggesting similar processing of the SP. Deletion of SP generated a ~72 kDa protein devoid of N-linked oligosaccharides allowing irrefutable identification of GP82 precursor. SP transposition to an internal region of GP82 rendered the signal unrecognizable by the signal peptidase and incapable to direct the nascent protein for ER-membrane association. Altogether our data strongly suggests that GP82 SP fails to function as transmembrane domain and its recognition by the signal peptidase shows strict dependence on the signal localization at protein N-terminus. This report presents the first experimental characterization of the full-length GP82 and its signal peptide.

Genomic Cloning and Sequence Analysis of Trypanosoma brucei rhodesiense gene Encoding Putative N-glycosylation Enzyme

Background: Trypanosoma brucei rhodesiense is a haemoflagellate parasite of zoonotic significance. Aside from its public health importance, this parasite subspecies gained notoriety because of their effective system to circumvent the immune response of vertebrate host. The parasite cell surface is covered with millions of VSG dimers, which serve as an almost infinite repertoire of biomolecules needed for evasion of host immune system. Around two decades ago, it was resolved that all trypanosome VSG is associated with one or more N-linked oligosaccharides, with a range of structures including high mannose and complex types. This complex process of protein modification known as N-linked glycosylation is catalyzed by oligosaccharyl transferase (OST). In general, the incorporation of glycan structures can alter protein’s antigenic properties and recently it was established that glycan molecules associated with VSG were found to be important in several aspects of trypanosome-host interaction, especially during parasite evasion of the host defense mechanisms. Therefore, our major interest is to clone and characterize the trypanosome OST. Materials, Methods & Results: The template genomic DNA for PCR amplification was extracted as described previously. In an attempt to clone Trypanosoma brucei rhodesiense putative oligosaccharyl transferase, an amplicon of ~2000 bp was obtained having an open reading frame of 2057 bp and deduced primary structure composed of 685 amino acid residues (TbrOST II). Comparison of TbrOST II ORF with annotated putative oligosaccharyl transferase in the genome of other organisms revealed sequence identity to other kinetoplastid. TbrOST II had high nucleotide (Ns) and amino acid (As) sequence similarity with the genomes of T. brucei gambiense (Ns:99%; As:78%) and T. brucei (Ns:95-98%; As:77%-98%). There was also significant nucleotide and amino acid sequence identity in the genomes of T. cruzi (Ns:74%; As:63%), Leishmania infantum (Ns:70-83%; As:46-57%), L. braziliensis (Ns:69-81%; As:46-55%) and L. major (Ns:69-80%; As:46-57%). Sequence similarity (71-77%) from other origins was also exhibited. The nucleotide sequence alignments and analysis were performed using the Oxford University Mac Vector 6.5 sequence analysis software and CLC Workbench 5.6 software. Discussion: The nucleotide BLAST results indicate that sequence identity is higher between species of the same genus rather than of the same family. It is known that T. brucei, T. gambiense and T. rhodesiense are members of the Brucei-complex or Brucei group. Although T. brucei brucei has more similarities with T. brucei rhodesiense than T. brucei gambiense, these parasites are morphologically indistinguishable. This is the probable reason why high sequence identity was displayed by other subspecies of the Brucei group. In addition, the high percent identity possessed by TbrOST II with other trypansomatids agrees with the evolutionarily conserved characteristics of the established OST. The DNA sequence data of TbrOST II showing similar sequences in the genome of other organisms further corroborate the previous reports regarding the ubiquitous nature of OST in other life forms. Based on the size of the amplicon and significant percentage of nucleotide and amino acid sequence identity to homologues within the genome of related species and various organisms, the results strongly indicate that TbrOST II is a trypanosome oligosaccharyl transferase gene candidate that should be fully characterized and subjected to functional genomic studies. The study reports the molecular cloning and sequencing of a potential oligosaccharyl transferase gene in T. brucei rhodesiense (TbrOST II). The sequence data has been deposited in the GenBank with accession number GU475126.

Alpha-v–containing integrins are host receptors for the Plasmodium falciparum sporozoite surface protein, TRAP

Malaria-causing Plasmodium sporozoites are deposited in the dermis by the bite of an infected mosquito and move by gliding motility to the liver where they invade and develop within host hepatocytes. Although extracellular interactions between Plasmodium sporozoite ligands and host receptors provide important guidance cues for productive infection and are good vaccine targets, these interactions remain largely uncharacterized. Thrombospondin-related anonymous protein (TRAP) is a parasite cell surface ligand that is essential for both gliding motility and invasion because it couples the extracellular binding of host receptors to the parasite cytoplasmic actinomyosin motor; however, the molecular nature of the host TRAP receptors is poorly defined. Here, we use a systematic extracellular protein interaction screening approach to identify the integrin αvβ3 as a directly interacting host receptor for Plasmodium falciparum TRAP. Biochemical characterization of the interaction suggests a two-site binding model, requiring contributions from both the von Willebrand factor A domain and the RGD motif of TRAP for integrin binding. We show that TRAP binding to cells is promoted in the presence of integrin-activating proadhesive Mn2+ ions, and that cells genetically targeted so that they lack cell surface expression of the integrin αv-subunit are no longer able to bind TRAP. P. falciparum sporozoites moved with greater speed in the dermis of Itgb3-deficient mice, suggesting that the interaction has a role in sporozoite migration. The identification of the integrin αvβ3 as the host receptor for TRAP provides an important demonstration of a sporozoite surface ligand that directly interacts with host receptors.

Life Stage-Specific Cargo Receptors Facilitate Glycosylphosphatidylinositol-Anchored Surface Coat Protein Transport in Trypanosoma brucei.

The critical virulence factor of bloodstream-form Trypanosoma brucei is the glycosylphosphatidylinositol (GPI)-anchored variant surface glycoprotein (VSG). Endoplasmic reticulum (ER) exit of VSG is GPI dependent and relies on a discrete subset of COPII machinery (TbSec23.2/TbSec24.1). In other systems, p24 transmembrane adaptor proteins selectively recruit GPI-anchored cargo into nascent COPII vesicles. Trypanosomes have eight putative p24s (TbERP1 to TbERP8) that are constitutively expressed at the mRNA level. However, only four TbERP proteins (TbERP1, -2, -3, and -8) are detectable in bloodstream-form parasites. All four colocalize to ER exit sites, are required for efficient GPI-dependent ER exit, and are interdependent for steady-state stability. These results suggest shared function as an oligomeric ER GPI-cargo receptor. This cohort also mediates rapid forward trafficking of the soluble lysosomal hydrolase TbCatL. Procyclic insect-stage trypanosomes have a distinct surface protein, procyclin, bearing a different GPI anchor structure. A separate cohort of TbERP proteins (TbERP1, -2, -4, and -8) are expressed in procyclic parasites and also function in GPI-dependent ER exit. Collectively, these results suggest developmentally regulated TbERP cohorts, likely in obligate assemblies, that may recognize stage-specific GPI anchors to facilitate GPI-cargo trafficking throughout the parasite life cycle. IMPORTANCE African trypanosomes are protozoan parasites that cause African sleeping sickness. Critical to the success of the parasite is the variant surface glycoprotein (VSG), which covers the parasite cell surface and which is essential for evasion of the host immune system. VSG is membrane bound by a glycolipid (GPI) anchor that is attached in the earliest compartment of the secretory pathway, the endoplasmic reticulum (ER). We have previously shown that the anchor acts as a positive forward trafficking signal for ER exit, implying a cognate receptor mechanism for GPI recognition and loading in coated cargo vesicles leaving the ER. Here, we characterize a family of small transmembrane proteins that act at adaptors for this process. This work adds to our understanding of general GPI function in eukaryotic cells and specifically in the synthesis and transport of the critical virulence factor of pathogenic African trypanosomes.

Trypanosoma cruzi Invasion into Host Cells: A Complex Molecular Targets Interplay.

Chagas' disease is still a worldwide threat, with estimated from 6 to 7 million infected people, mainly in Latin America. Despite all efforts, especially from international consortia (DNDi, NMTrypI), to develop an innovative therapeutic strategy against this disease, no candidate has achieved full requirements for clinical use yet. In this review, we point out the general molecular and cellular mechanisms involved in T. cruzi cell invasion and elucidate the roles of specific parasite and host targets in the progress of Chagas' disease. Among these molecular targets are Gp85/transsialidase, mucins, cruzipain and oligopeptidase B, found in parasite cell surface, and Galectin-3 and Toll-like receptors present in host cells. Thus, the deep understanding of their interplay and involvement on T. cruzi host cell adhesion, invasion and evasion from host immune may expand the chances for discovering new therapeutic agents against this neglected disease. Additionally, these targets may represent a remarkable strategy to block parasite invasion in the early stages of infection.

Modulation of Cell Sialoglycophenotype: A Stylish Mechanism Adopted by Trypanosoma cruzi to Ensure Its Persistence in the Infected Host.

Trypanosoma cruzi, the etiological agent of Chagas disease exhibits multiple mechanisms to guarantee its establishment and persistence in the infected host. It has been well demonstrated that T. cruzi is not able to synthesize sialic acids (Sia). To acquire the monosaccharide, the parasite makes use of a multifunctional enzyme called trans-sialidase (Tc-TS). Since this enzyme has no analogous in the vertebrate host, it has been used as a target in drug therapy development. Tc-TS preferentially catalyzes the transfer of Sia from the host glycoconjugates to the terminal β-galactopyranosyl residues of mucin-like molecules present on the parasite’s cell surface. Alternatively, the enzyme can sialylate/re-sialylate glycoconjugates expressed on the surface of host cells. Since its discovery, several studies have shown that T. cruzi employs the Tc-TS activity to modulate the host cell sialoglycophenotype, thus favoring its perpetuation in the infected vertebrate. In this review, we summarize the dynamic of host/parasite sialoglycophenotype modulation, highlighting its role in the subversion of host immune response in order to promote the establishment of persistent chronic infection.

Gene family phylogeny and the evolution of parasite cell surfaces

Parasite genomes typically contain unique contingency gene families encoding multi-copy effector proteins that are often expressed abundantly on the parasite cell surface and beyond. The functions of these gene families are incompletely understood but it is clear that they perform fundamental roles at the host-parasite interface. Over evolutionary timescales, the evolution of these gene families is likely to have decisive effects on the pathology and virulence of parasitic infections. In this review, I will compare the evolutionary dynamics of multiple examples from trypanosomatids and apicomplexan parasites to demonstrate how their inherent mutability makes their phylogeny very different to ‘normal’ gene families. I will argue that phylogenetic analyses could help to understand the functions of these enigmatic genes.

Global Gene Expression Profiling through the Complete Life Cycle of Trypanosoma vivax.

The parasitic flagellate Trypanosoma vivax is a cause of animal trypanosomiasis across Africa and South America. The parasite has a digenetic life cycle, passing between mammalian hosts and insect vectors, and a series of developmental forms adapted to each life cycle stage. Each point in the life cycle presents radically different challenges to parasite metabolism and physiology and distinct host interactions requiring remodeling of the parasite cell surface. Transcriptomic and proteomic studies of the related parasites T. brucei and T. congolense have shown how gene expression is regulated during their development. New methods for in vitro culture of the T. vivax insect stages have allowed us to describe global gene expression throughout the complete T. vivax life cycle for the first time. We combined transcriptomic and proteomic analysis of each life stage using RNA-seq and mass spectrometry respectively, to identify genes with patterns of preferential transcription or expression. While T. vivax conforms to a pattern of highly conserved gene expression found in other African trypanosomes, (e.g. developmental regulation of energy metabolism, restricted expression of a dominant variant antigen, and expression of ‘Fam50’ proteins in the insect mouthparts), we identified significant differences in gene expression affecting metabolism in the fly and a suite of T. vivax-specific genes with predicted cell-surface expression that are preferentially expressed in the mammal (‘Fam29, 30, 42’) or the vector (‘Fam34, 35, 43’). T. vivax differs significantly from other African trypanosomes in the developmentally-regulated proteins likely to be expressed on its cell surface and thus, in the structure of the host-parasite interface. These unique features may yet explain the species differences in life cycle and could, in the form of bloodstream-stage proteins that do not undergo antigenic variation, provide targets for therapy.

The trans-sialidase, the major Trypanosoma cruzi virulence factor: Three decades of studies.

Chagas' disease is a potentially life-threatening disease caused by the protozoan parasite Trypanosoma cruzi. Since the description of Chagas'disease in 1909 extensive research has identified important events in the disease in order to understand the biochemical mechanism that modulates T. cruzi-host cell interactions and the ability of the parasite to ensure its survival in the infected host. Exactly 30 years ago, we presented evidence for the first time of a trans-sialidase activity in T. cruzi (T. cruzi-TS). This enzyme transfers sialic acid from the host glycoconjugates to the terminal β-galactopyranosyl residues of mucin-like molecules on the parasite's cell surface. Thenceforth, many articles have provided convincing data showing that T. cruzi-TS is able to govern relevant mechanisms involved in the parasite's survival in the mammalian host, such as invasion, escape from the phagolysosomal vacuole, differentiation, down-modulation of host immune responses, among others. The aim of this review is to cover the history of the discovery of T. cruzi-TS, as well as some well-documented biological effects encompassed by this parasite's virulence factor, an enzyme with potential attributes to become a drug target against Chagas disease.

Structural model of a putrescine-cadaverine permease from Trypanosoma cruzi predicts residues vital for transport and ligand binding

The TcPOT1.1 gene from Trypanosoma cruzi encodes a high affinity putrescine-cadaverine transporter belonging to the APC (amino acid/polyamine/organocation) transporter superfamily. No experimental three-dimensional structure exists for any eukaryotic member of the APC family, and thus the structural determinants critical for function of these permeases are unknown. To elucidate the key residues involved in putrescine translocation and recognition by this APC family member, a homology model of TcPOT1.1 was constructed on the basis of the atomic co-ordinates of the Escherichia coli AdiC arginine/agmatine antiporter crystal structure. The TcPOT1.1 homology model consisted of 12 transmembrane helices with the first ten helices organized in two V-shaped antiparallel domains with discontinuities in the helical structures of transmembrane spans 1 and 6. The model suggests that Trp241 and a Glu247-Arg403 salt bridge participate in a gating system and that Asn245, Tyr148 and Tyr400 contribute to the putrescine-binding pocket. To test the validity of the model, 26 site-directed mutants were created and tested for their ability to transport putrescine and to localize to the parasite cell surface. These results support the robustness of the TcPOT1.1 homology model and reveal the importance of specific aromatic residues in the TcPOT1.1 putrescine-binding pocket.

(Not) helping Plasmodium break in

An integral step in the establishment of malarial infection is invasion of the host erythrocyte by the parasite. Previous work had reported that presenilin-like signal peptide peptidase (SPP) has a role during invasion by Plasmodium falciparum, targeting the cytoskeletal protein Band 3 on the erythrocyte surface; this, surprisingly, implied that, although SPP is an ER protein, it is secreted during invasion. However, a separate study found that SPP has a role in growth, not invasion, of Plasmodium berghei. Here, Marapana et al. sought to resolve the precise contribution of SPP during the establishment of infection. Using fluorescence imaging, they found that SPP resides in the ER and does not relocalize to the parasite cell surface during invasion. Importantly, blocking SPP with a range of inhibitors had no effect on invasion; however, it did interfere with parasite development, which suggests that the protein could be targeted for therapeutic purposes.

Exposure to Host Ligands Correlates with Colocalization of Gal/GalNAc Lectin Subunits in Lipid Rafts and Phosphatidylinositol (4,5)-Bisphosphate Signaling in Entamoeba histolytica

Entamoeba histolytica is an intestinal parasite that causes dysentery and liver abscess. Parasite cell surface receptors, such as the Gal/GalNAc lectin, facilitate attachment to host cells and extracellular matrix. The Gal/GalNAc lectin binds to galactose or N-acetylgalactosamine residues on host components and is composed of heavy (Hgl), intermediate (Igl), and light (Lgl) subunits. Although Igl is constitutively localized to lipid rafts (cholesterol-rich membrane domains), Hgl and Lgl transiently associate with this compartment in a cholesterol-dependent fashion. In this study, trophozoites were exposed to biologically relevant ligands to determine if ligand binding influences the submembrane distribution of the subunits. Exposure to human red blood cells (hRBCs) or collagen, which are bona fide Gal/GalNAc lectin ligands, was correlated with enrichment of Hgl and Lgl in rafts. This enrichment was abrogated in the presence of galactose, suggesting that direct lectin-ligand interactions are necessary to influence subunit location. Using a cell line that is able to attach to, but not phagocytose, hRBCs, it was shown that physical attachment to ligands was not sufficient to induce the enrichment of lectin subunits in rafts. Additionally, the mutant had lower levels of phosphatidylinositol (4,5)-bisphosphate (PIP(2)); PIP(2) loading restored the ability of this mutant to respond to ligands with enrichment of subunits in rafts. Finally, intracellular calcium levels increased upon attachment to collagen; this increase was essential for the enrichment of lectin subunits in rafts. Together, these data provide evidence that ligand-induced enrichment of lectin subunits in rafts may be the first step in a signaling pathway that involves both PIP(2) and calcium signaling.

Host cell invasion by apicomplexans: what do we know?

Apicomplexan zoites enter host cells by forming and actively moving through a tight junction (TJ) formed between the parasite and host cell surfaces. Although the TJ was first described decades ago, its molecular characterization has proved difficult mainly because of its transient existence during an internalization process that lasts only seconds. In the past 7 years, work has led to a model of the TJ in which the association between AMA1 and RON proteins structures the TJ and bridges the cytoskeletons of the two cells. However, more recent work questions this view. Here, we critically discuss the current model and speculate on alternative models of the AMA1-RON association and of the apicomplexan TJ.