Flatworm Schistosoma Mansoni Research Articles

Descriptive study on risk of increased morbidity of schistosomiasis and graft loss after liver transplantation.

Solid-organ transplantation procedures have witnessed a surge in frequency. Consequently, increased attention to associated infections and their impact on graft success is warranted. The liver is the principal target for infection by the flatworm Schistosoma mansoni. Hence, rigorous screening protocols for this parasite should be implemented for liver transplantation donors and recipients. This study investigated the risks posed by schistosomiasis-infected liver tissues for successful liver transplantation (LT), considering donors and recipients, by analyzing reported cases. Among the 43 patients undergoing LT (donors = 19; recipients = 24), 32 were infected with S. mansoni, five were infected with other Schistosoma species, and no identification was made in four patients. Reported follow-up periods ranged from 1 to 132 months, and all patients achieved successful recovery. As these helminths do not replicate in their vertebrate hosts, immunosuppressive treatment is not expected to promote increased morbidity or reactivation. Moreover, suspected or confirmed schistosomiasis infections often have a benign course, and generally, should not prevent LT. The available literature was reviewed and a provisional screening protocol has been proposed.

The stage- and sex-specific transcriptome of the human parasite Schistosoma mansoni

The flatworm Schistosoma mansoni is an important but neglected pathogen that causes the disease schistosomiasis in millions of people worldwide. The parasite has a complex life cycle, undergoing sexual reproduction in a mammalian host and asexual replication in a snail host. Understanding the molecular mechanisms that the parasite uses to transition between hosts and develop into dimorphic reproductively competent adults may reveal new strategies for control. We present the first comprehensive transcriptomic analysis of S. mansoni, from eggs to sexually naïve worms. Focusing on eight life stages spanning free-living water-borne and parasitic stages from both intermediate and definitive hosts, we have generated deep RNA-seq data for five replicates per group for a total of 75 data sets. The data were produced using a single approach to increase the accuracy of stage-to-stage comparisons and made accessible via a user-friendly tool to visualise and explore gene expression (https://lifecycle.schisto.xyz/). These data are valuable for understanding the biology and sex-specific development of schistosomes and the interpretation of complementary genomic and functional genetics studies.

Codon usage in the flatworm Schistosoma mansoni is shaped by the mutational bias towards A+T and translational selection, which increases GC-ending codons in highly expressed genes

Schistosoma mansoni is a trematode flatworm that parasitizes humans and produces a disease called bilharzia. At the genomic level, it is characterized by a low genomic GC content and an "isochore-like" structure, where GC-richest regions, mainly placed at the extremes of the chromosomes, are interspersed with low GC-regions. Furthermore, the GC-richest regions are at the same time the gene-richest, and where the most heavily expressed genes are placed. Taking these features into account, we decided to reanalyze the codon usage of this flatworm. Our results show that a) when all genes are considered together, the strong mutational bias towards A + T leads to a predominance of A/T-ending codons, b) a multivariate analysis discriminates between highly and lowly expressed genes, c) the sequences expressed at highest levels display a significant increase in G/C-ending codons, d) when comparing the molecular distances with a closely related species the synonymous distance in highly expressed genes is significantly lower than in lowly expressed sequences. Therefore, we conclude that despite previous results, which were performed with a small sample of genes, codon usage in S. mansoni is the result of two forces that operate in opposite directions: while mutational bias leads to a predominance of A/T codons, translational selection, working at the level of speed, increment G/C ending triplets.

The anthelminthic drug praziquantel is a selective agonist of the sensory transient receptor potential melastatin type 8 channel

Praziquantel is the most effective anthelminthic drug for the treatment of schistosomiasis, an infectious disease caused by the platyhelminth Schistosoma mansoni. While praziquantel is known to trigger calcium influx into schisostomes, followed by spastic paralysis of the worms and tegumental disruption, the mechanism of action of the drug is not completely understood. Although relatively well tolerated, praziquantel has been reported to cause mild adverse effects, including nausea, abdominal pain and headaches. As a number of putative Transient Receptor Potential (TRP) channel genes have recently been predicted in S. mansoni, we sought to investigate the effect of praziquantel on three mammalian TRP channels, TRP melastatin type 8 (TRPM8), TRP vanilloid type 1 (TRPV1) and TRP ankyrin type 1 (TRPA1). Using calcium microfluorimetry and the patch clamp technique, we recorded the effect of praziquantel on HEK293T cells expressing recombinant TRPM8, TRPV1 or TRPA1, as well as on cultured dorsal root ganglion (DRG) neurons from wild type and TRPM8 null mutant mice. We discovered that praziquantel is a relatively potent and selective partial agonist of the mammalian and avian cold and menthol receptor TRPM8. The activation of cultured DRG neurons by clinically relevant concentrations of praziquantel is predominantly mediated by TRPM8. Our results may provide clues to a better understanding of praziquantel's mechanism of action and its adverse effects.

A gene expression atlas of adult Schistosoma mansoni and their gonads

RNA-Seq has proven excellence in providing information about the regulation and transcript levels of genes. We used this method for profiling genes in the flatworm Schistosoma mansoni. This parasite causes schistosomiasis, an infectious disease of global importance for human and animals. The pathology of schistosomiasis is associated with the eggs, which are synthesized as a final consequence of male and female adults pairing. The male induces processes in the female that lead to the full development of its gonads as a prerequisite for egg production. Unpaired females remain sexually immature. Based on an organ-isolation method we obtained gonad tissue for RNA extraction from paired and unpaired schistosomes, with whole adults included as controls. From a total of 23 samples, we used high-throughput cDNA sequencing (RNA-Seq) on the Illumina platform to profile gene expression between genders and tissues, with and without pairing influence. The data obtained provide a wealth of information on the reproduction biology of schistosomes and a rich resource for exploitation through basic and applied research activities.

Sm10.3, a member of the micro-exon gene 4 (MEG-4) family, induces erythrocyte agglutination in vitro and partially protects vaccinated mice against Schistosoma mansoni infection.

BackgroundThe parasitic flatworm Schistosoma mansoni is a blood fluke that causes schistosomiasis. Current schistosomiasis control strategies are mainly based on chemotherapy, but many researchers believe that the best long-term strategy to control disease is a combination of drug treatment and immunization with an anti-schistosome vaccine. Numerous antigens that are expressed at the interface between the parasite and the mammalian host have been assessed. Among the most promising molecules are the proteins present in the tegument and digestive tract of the parasite.Methodology/Principal FindingsIn this study, we evaluated the potential of Sm10.3, a member of the micro-exon gene 4 (MEG-4) family, for use as part of a recombinant vaccine. We confirmed by real-time PCR that Sm10.3 was expressed at all stages of the parasite life cycle. The localization of Sm10.3 on the surface and lumen of the esophageal and intestinal tract in adult worms and lung-stage schistosomula was confirmed by confocal microscopy. We also show preliminary evidence that rSm10.3 induces erythrocyte agglutination in vitro. Immunization of mice with rSm10.3 induced a mixed Th1/Th2-type response, as IFN-γ, TNF-α, and low levels of IL-5 were detected in the supernatant of cultured splenocytes. The protective effect conferred by vaccination with rSm10.3 was demonstrated by 25.5–32% reduction in the worm burden, 32.9–43.6% reduction in the number of eggs per gram of hepatic tissue, a 23.8% reduction in the number of granulomas, an 11.8% reduction in the area of the granulomas and a 39.8% reduction in granuloma fibrosis.Conclusions/SignificanceOur data suggest that Sm10.3 is a potential candidate for use in developing a multi-antigen vaccine to control schistosomiasis and provide the first evidence for a possible role for Sm10.3 in the blood feeding process.

Why do we need three dynamins? (Comment on DOI 10.1002/bies.201200033)

All living organisms must exchange matter with the environment. Clathrin-mediated endocytosis (CME) is a fundamental process by which eukaryotic cells engulf und internalize substances from the surrounding. After cargo-containing invaginated clathrin-coated pits are formed, dynamin forms collars around the necks of these nascent vesicles and, by acting as a mechanochemical enzyme, uses the energy from GTP hydrolysis to mediate fission from the plasma membrane. Dynamins are large GTPases build up of five domains: the G domain (responsible for GTP substrate binding and hydrolysis), the middle domain, the pleckstrin-homology domain (PHD, which mediates binding to membrane lipids), the GTPase effector domain and the proline-rich domain. Vertebrates encode three dynamins. The neuron-specific dynamin 1 and the ubiquitously expressed dynamin 2 participate in CME. The function of dynamin 3 is poorly understood; it is found in testis and brain. In this issue, Liu et al. 1 discuss the evolution of the dynamins by looking primarily at the PHD. A counterintuitive order of appearance was suggested, in which the neuronal dynamin 1 evolved first, and the ubiquitous dynamin 2 was a later invention. Further, the evolution of dynamin 1 was suggested to have been driven by the development of the central nervous system (CNS). The discussion is based on a phylogenetic analysis presented in the paper and earlier biochemical findings made by the authors 2. They suggest that dynamin 1 is a curvature generator and that dynamin 2 is a curvature sensor, and that these distinct properties are mainly due to Tyr600 and Leu600, respectively. The key residue is part of the variable loop 3 (VL3), one of the three VLs of the PHD forming the phospholipid interaction pocket. Indeed, exchanging the VL3 leucine for tyrosine turned dynamin 2 into a potent curvature generator. In agreement, the curvature-generating potency of dynamin 1 was reduced by a Y600L mutation. Leu600 is only found in dynamin 2; dynamin 1, and dynamin 3 encode a Tyr600. Drosophila and Caenorhabditis elegans, fly and worm, possess only one classical dynamin, which is primarily expressed in the nervous system and has a tyrosine in VL3. Interestingly, in the 1970s, the importance of dynamin for neuronal signal transduction was demonstrated for the first time through a mutation, shibire, in its gene, which caused a reversible temperature-dependent paralysis in fly. Liu et al. identified dynamin-coding sequences in the protists Monosiga brevicollis and Capsaspora owczarzaki, and early metazoans that lacked CNS. In the sequences, a histidine was found in VL3. Remarkably, a flatworm (Schistosoma mansoni) owning a CNS was shown to encode two dynamins, one containing histidine and the other tyrosine in the VL3. It appears likely that the powerful curvature generator, dynamin 1, with a VL3 tyrosine, evolved for the special needs of neuronal signal transmission, where rapid recycling of synaptic vesicles is required. Dynamin 2 could have evolved in vertebrates as a sensor that controls fidelity of CME and releases only mature vesicles from plasma membrane. Could the VL3 residue also contribute to lipid composition specificity, and control dynamin isoform-specific functions in distinct cell types and subcellular compartments? What is dynamin 3 needed for? The dynamin superfamily encompasses proteins – classical dynamins and dynamin-like proteins – participating in deformation of biological membranes, and involved in multiple cellular processes from endocytosis and vesicle trafficking to division of organelles and cells. It is believed that the proteins originally evolved as components of cell and organelle division machineries. Is the existence of an additional dynamin in testis, the place of spermatogenesis and a multitude of cell division processes, just a coincident? Intriguingly, there is a further unusual large GTPase expressed only in testis. Irgc belongs to the immunity-related GTPases (IRGs), a family that consists of more than 20 members in the mouse, but is almost lost in man 3. IRGs participate in the destruction of the membrane surrounding the protozoan parasite Toxoplasma gondii in infected cells. Irgc is highly conserved between mouse and human, and is the only full-length remnant of the IRG family in humans. Furthermore, Irgc is probably the only IRG protein not induced by interferons and without a function in immunity.

Combination of de novo assembly of massive sequencing reads with classical repeat prediction improves identification of repetitive sequences in Schistosoma mansoni

The genome of the parasitic platyhelminth Schistosoma mansoni is composed of approximately 40% of repetitive sequences of which roughly 20% correspond to transposable elements. When the genome sequence became available, conventional repeat prediction programs were used to find these repeats, but only a fraction could be identified. To exhaustively characterize the repeats we applied a new massive sequencing based strategy: we re-sequenced the genome by next generation sequencing, aligned the sequencing reads to the genome and assembled all multiple-hit reads into contigs corresponding to the repetitive part of the genome. We present here, for the first time, this de novo repeat assembly strategy and we confirm that such assembly is feasible. We identified and annotated 4,143 new repeats in the S. mansoni genome. At least one third of the repeats are transcribed. This strategy allowed us also to identify 14 new microsatellite markers, which can be used for pedigree studies. Annotations and the combined (previously known and new) 5,420 repeat sequences (corresponding to 47% of the genome) are available for download (http://methdb.univ-perp.fr/downloads/).

Computational Vaccinology: An Important Strategy to Discover New Potential S. mansoni Vaccine Candidates

The flatworm Schistosoma mansoni is a blood fluke parasite that causes schistosomiasis, a debilitating disease that occurs throughout the developing world. Current schistosomiasis control strategies are mainly based on chemotherapy, but many researchers believe that the best long-term strategy to control schistosomiasis is through immunization with an antischistosomiasis vaccine combined with drug treatment. Several papers on Schistosoma mansoni vaccine and drug development have been published in the past few years, representing an important field of study. The advent of technologies that allow large-scale studies of genes and proteins had a remarkable impact on the screening of new and potential vaccine candidates in schistosomiasis. In this postgenomic scenario, bioinformatic technologies have emerged as important tools to mine transcriptomic, genomic, and proteomic databases. These new perspectives are leading to a new round of rational vaccine development. Herein, we discuss different strategies to identify potential S. mansoni vaccine candidates using computational vaccinology.

A P2X receptor from the tardigrade species Hypsibius dujardini with fast kinetics and sensitivity to zinc and copper.

BackgroundOrthologs of the vertebrate ATP gated P2X channels have been identified in Dictyostelium and green algae, demonstrating that the emergence of ionotropic purinergic signalling was an early event in eukaryotic evolution. However, the genomes of a number of animals including Drosophila melanogaster and Caenorhabditis elegans, both members of the Ecdysozoa superphylum, lack P2X-like proteins, whilst other species such as the flatworm Schistosoma mansoni have P2X proteins making it unclear as to what stages in evolution P2X receptors were lost. Here we describe the functional characterisation of a P2X receptor (HdP2X) from the tardigrade Hypsibius dujardini demonstrating that purinergic signalling is preserved in some ecdysozoa.ResultsATP (EC50 ~44.5 μM) evoked transient inward currents in HdP2X with millisecond rates of activation and desensitisation. HdP2X is antagonised by pyridoxal-phosphate-6-azophenyl-2',4' disulfonic acid (IC50 15.0 μM) and suramin (IC50 22.6 μM) and zinc and copper inhibit ATP-evoked currents with IC50 values of 62.8 μM and 19.9 μM respectively. Site-directed mutagenesis showed that unlike vertebrate P2X receptors, extracellular histidines do not play a major role in coordinating metal binding in HdP2X. However, H306 was identified as playing a minor role in the actions of copper but not zinc. Ivermectin potentiated responses to ATP with no effect on the rates of current activation or decay.ConclusionThe presence of a P2X receptor in a tardigrade species suggests that both nematodes and arthropods lost their P2X genes independently, as both traditional and molecular phylogenies place the divergence between Nematoda and Arthropoda before their divergence from Tardigrada. The phylogenetic analysis performed in our study also clearly demonstrates that the emergence of the family of seven P2X channels in human and other mammalian species was a relatively recent evolutionary event that occurred subsequent to the split between vertebrates and invertebrates. Furthermore, several characteristics of HdP2X including fast kinetics with low ATP sensitivity, potentiation by ivermectin in a channel with fast kinetics and distinct copper and zinc binding sites not dependent on histidines make HdP2X a useful model for comparative structure-function studies allowing a better understanding of P2X receptors in higher organisms.

Crystallization and X-ray analysis of theSchistosoma mansoniguanidino kinase

The 716-amino-acid guanidino kinase from the parasitic flatworm Schistosoma mansoni results from the fusion of two guanidino kinase subunits. Crystals of this 80 kDa protein have been obtained in the monoclinic space group P2(1), with unit-cell parameters a = 52.7, b = 122.1, c = 63.2 A, beta = 108.5 degrees . Synchrotron data were collected to 2.8 A resolution on ESRF beamline ID29. The structure was solved by the molecular-replacement method, using the 357-amino-acid structure of the arginine kinase from Trypanosoma cruzi as the search model.

A bone morphogenetic protein homologue in the parasitic flatworm, Schistosoma mansoni

Members of the bone morphogenetic protein (BMP) subfamily of cytokines control many aspects of metazoan development including patterning and organogenesis. Despite the recognition that schistosomes possess key components of a BMP signaling pathway, a BMP-like ligand in the parasitic flatworm Schistosoma mansoni remained elusive. Here, we describe the cloning and characterisation of an S. mansoni BMP ( SmBMP). SmBMP is most closely related to BMP homologues from the free-living flatworms Schmidtea mediterranea and Dugesia japonica, with 51% and 47% identity at the amino acid level, respectively. Based on reverse transcription-PCR, SmBMP is expressed throughout the mammalian life-cycle of the parasite in both male and female schistosomes. In support of these results, antibodies to SmBMP successfully immunoprecipitated the protein in adult male and female antigen preparations with more protein detected in male parasites. Immunofluorescent studies localised SmBMP to the protonephridia of adult parasites, and SmBMP was identified in the excretory/secretory products of adult male parasites via immunoprecipitation. With the previous description of a TGF-β subfamily homologue in S. mansoni, ligands representing both arms of the TGF-β superfamily have now been described in this trematode.

Aza-peptidyl Michael Acceptors. A New Class of Potent and Selective Inhibitors of Asparaginyl Endopeptidases (Legumains) from Evolutionarily Diverse Pathogens

Aza-peptide Michael acceptors with the general structure of Cbz-Ala-Ala-AAsn- trans-CH=CHCOR are a new class of inhibitors specific for the asparaginyl endopeptidases (AE) (legumains). Structure-activity relationships (SARs) were characterized for a set of 31 aza-peptide Michael acceptors with AEs derived from three medically important parasites: the protist Trichomonas vaginalis, the hard tick Ixodes ricinus, and the flatworm Schistosoma mansoni. Despite arising from phylogenetically disparate organisms, all three AEs shared a remarkably similar SAR with lowest IC50 values extending into the picomolar range. The results suggest an evolutionary constraint on the topography of the prime side of the active site. SAR also revealed that esters in the P1' position are more potent than disubstituted amides and that monosubstituted amides and alkyl derivatives show little or no inhibition. The preferred P1' residues have aromatic substituents. Aza-asparaginyl Michael acceptors react with thiols, which provides insight into the mechanism of their inhibition of asparaginyl endopeptidases.

Schistosoma mansoni: SmE78, a nuclear receptor orthologue of Drosophila ecdysone-induced protein 78

Drosophila ecdysone-induced protein 78 (E78) belongs to the nuclear receptor (NR) superfamily, it plays a role directly related to ecdysone signaling. We isolated a cDNA of Drosophila E78 orthologue from the Platyhelminth Schistosoma mansoni (SmE78). It is the first E78 orthologue known outside of the molting animals—the Ecdysozoa. The SmE78 cDNA is 3471 base pairs long and contains an entire open reading frame (ORF) encoding a 1087 amino acid protein. Phylogenetic analysis of the ligand-binding domain (LBD) demonstrates that the LBD of SmE78 is phylogenetically related to the Drosophila E78. Gene structure of SmE78 was determined showing it to consist of six exons spanning more than 32 kbp. Quantitative real-time reverse-transcriptase polymerase chain reaction (RT-PCR) demonstrated that SmE78 was expressed throughout schistosome development but with the highest levels of expression in miracidia and egg stage. The result is consistent with the previous studies that Ecdysterone was effective in stimulating host location activities in miracidia. The data suggest that SmE78 may be involved in transduction of an ecdysone signal in S. mansoni.

Atypical properties of a conventional calcium channel β subunit from the platyhelminth Schistosoma mansoni

BackgroundThe function of voltage-gated calcium (Cav) channels greatly depends on coupling to cytoplasmic accessory β subunits, which not only promote surface expression, but also modulate gating and kinetic properties of the α1 subunit. Schistosomes, parasitic platyhelminths that cause schistosomiasis, express two β subunit subtypes: a structurally conventional β subunit and a variant β subunit with unusual functional properties. We have previously characterized the functional properties of the variant Cavβ subunit. Here, we focus on the modulatory phenotype of the conventional Cavβ subunit (SmCavβ) using the human Cav2.3 channel as the substrate for SmCavβ and the whole-cell patch-clamp technique.ResultsThe conventional Schistosoma mansoni Cavβ subunit markedly increases Cav2.3 currents, slows macroscopic inactivation and shifts steady state inactivation in the hyperpolarizing direction. However, currents produced by Cav2.3 in the presence of SmCavβ run-down to approximately 75% of their initial amplitudes within two minutes of establishing the whole-cell configuration. This suppressive effect was independent of Ca2+, but dependent on intracellular Mg2+-ATP. Additional experiments revealed that SmCavβ lends the Cav2.3/SmCavβ complex sensitivity to Na+ ions. A mutant version of the Cavβ subunit lacking the first forty-six amino acids, including a string of twenty-two acidic residues, no longer conferred sensitivity to intracellular Mg2+-ATP and Na+ ions, while continuing to show wild type modulation of current amplitude and inactivation of Cav2.3.ConclusionThe data presented in this article provide insights into novel mechanisms employed by platyhelminth Cavβ subunits to modulate voltage-gated Ca2+ currents that indicate interactions between the Ca2+ channel complex and chelated forms of ATP as well as Na+ ions. These results have potentially important implications for understanding previously unknown mechanisms by which platyhelminths and perhaps other organisms modulate Ca2+ currents in excitable cells.

Evolution of a novel subfamily of nuclear receptors with members that each contain two DNA binding domains

BackgroundNuclear receptors (NRs) are important transcriptional modulators in metazoans which regulate transcription through binding to the promoter region of their target gene by the DNA binding domain (DBD) and activation or repression of mRNA synthesis through co-regulators bound to the ligand binding domain (LBD). NRs typically have a single DBD with a LBD.ResultsThree nuclear receptors named 2DBD-NRs, were identified from the flatworm Schistosoma mansoni that each possess a novel set of two DBDs in tandem with a LBD. They represent a novel NR modular structure: A/B-DBD-DBD-hinge-LBD. The 2DBD-NRs form a new subfamily of NRs, VII. By database mining, 2DBD-NR genes from other flatworm species (Schmidtea mediterranea and Dugesia japonica), from Mollusks (Lottia gigantean) and from arthropods (Daphnia pulex) were also identified. All 2DBD-NRs possess a P-box sequence of CEACKK in the first DBD, which is unique to 2DBD-NRs, and a P-box sequence of CEGCKG in the second DBD. Phylogenetic analyses of both DBD and ligand binding domain sequences showed that 2DBD-NR genes originate from a common two DBD-containing ancestor gene. A single 2DBD-NR orthologue was found in Arthropoda, Platyhelminths and Mollusca. Subsequent 2DBD-NR gene evolution in Mollusks and Platyhelminths involved gene duplication. Chromosome localization of S. mansoni 2DBD-NR genes by Fluorescent in situ hybridization (FISH) suggests that 2DBD-NR genes duplicated on different chromosomes in the Platyhelminths. Dimerization of Sm2DBDα indicates that 2DBD-NRs may act as homodimers, suggesting either that two repeats of a half-site are necessary for each DBD of 2DBD-NRs to bind to its target gene, or that each 2DBD-NR can recognize multiple sites.Conclusion2DBD-NRs share a common ancestor gene which possessed an extra DBD that likely resulted from a recombination event. After the split of the Arthropods, Mollusks and Platyhelminths, 2DBD-NR underwent a recent duplication in a common ancestor of Mollusks, while two rounds of duplication occurred in a common ancestor of the Platyhelminths. This demonstrates that certain NR gene underwent recent duplication in Prostostome lineages after the split of the Prostostomia and Deuterostomia.

Identification and characterization of a nuclear receptor subfamily I member in the Platyhelminth Schistosoma mansoni (SmNR1)

A cDNA encoding a nuclear receptor subfamily I member in the platyhelminth Schistosoma mansoni (SmNR1) was identified and characterized. SmNR1 cDNA is 2406 bp long and contains an open reading frame encoding a 715 residue protein. Phylogenetic analysis demonstrates that SmNR1 is a divergent member of nuclear receptor subfamily I with no known orthologue. SmNR1 was localized to S. mansoni chromosome 1 by fluorescent in situ hybridization. Gene structure of SmNR1 was determined showing it to consist of eight exons spanning more than 14 kb. Quantitative real-time RT-PCR showed that SmNR1 was expressed throughout schistosome development with a higher expression in eggs, sporocysts and 21-day worms. SmNR1 contains an autonomous transactivation function (AF1) in the A/B domain as demonstrated in a yeast one-hybrid assay; it interacts with SmRXR1 in a yeast two-hybrid assay and in a glutathione S-transferase pull-down assay. Electrophoretic mobility shift assay showed that SmNR1 could form a heterodimer with SmRXR1 to bind to DNA elements containing the half-site AGGTCA, a direct repeat of the half-site separated by 0-5 nucleotides (DR1-DR5) and a palindrome repeat of the half-site not separated by nucleic acids (Pal0). Transient transfection in mammalian COS-7 cells showed that SmNR1/SmRXR1 could enhance the transcriptional activation of a DR2-dependent reporter gene. Our results demonstrate that SmNR1 is a partner of SmRXR1.

Nucleic acid binding properties of SmZF1, a zinc finger protein of Schistosoma mansoni

During its life cycle, the flat worm Schistosoma mansoni is exposed to diverse environmental conditions and changes its morphological form. Each change calls for distinct patterns of gene expression. In order to understand the regulation of gene expression, it is necessary to identify regulatory elements in the promoter region of genes, and DNA transacting factors that control transcription. Zinc finger protein domains are responsible for transcription regulation of diverse genes in a wide range of organisms and are also involved in the promotion of protein–protein interactions. A transcript homologous to zinc finger gene sequences was isolated from a S. mansoni adult worm cDNA library and named SmZF1. It codes for a protein of 164 amino acids presenting three C 2H 2 type zinc finger motifs. The recombinant SmZF1 protein was expressed and used on electrophoretic mobility shift assays to investigate the binding specificity of SmZF1 for DNA and RNA oligonucleotides. Our results demonstrated that SmZF1 binds both ds and ss DNA oligonucleotides, with an apparent preference for the specific D1-3DNA oligonucleotide, and also binds RNA oligonucleotides with lower affinity. Although we found that SmZF1 recognises DNA and RNA oligonucleotides not containing putative target sites, SmZF1 binds preferentially to sequence specific sites. Furthermore, unrelated oligonucleotides are not able to abolish this interaction. In silico studies identified putative SmZF1 binding sites in the complete genome of three model organisms and in partial genome sequences of S. mansoni. Six Drosophila genes presented these binding sites in their promoter region, indicating that they might be controlled by transcription factors containing zinc fingers motifs. Taken together, these results suggest that SmZF1 acts as a putative transcription factor of S. mansoni.

Gene linkage and steady state RNAs suggest trans-splicing may be associated with a polycistronic transcript in Schistosoma mansoni

Spliced leader (SL) trans-splicing generates the 5′ end of mature mRNAs through the addition of a small exon to pre-mRNAs in some flagellates (kinetoplastida and euglenoids) and metazoans (nematodes and flatworms). Although SL addition in the kinetoplastida and a subset of nematode genes serves to resolve multicistronic mRNAs into monocistronic, capped mRNAs, information regarding the functional significance of trans-splicing in flatworms is limited. We describe here the identification and characterization of a closely linked gene upstream from the trans-spliced enolase gene in the flatworm Schistosoma mansoni. This gene produces a non- trans-spliced mRNA encoding a ubiquinol binding protein, UbCRBP, that is a component of the ubiquinol-cytochrome C reductase complex. The distance between the UbCRBP polyadenylation site and the enolase trans-splice acceptor site is exceptionally short, only 54 nucleotides. Primer extension (5′ RACE), RT-PCR, and RNase mapping have identified steady state, cis-spliced RNAs which significantly overlap both the UbCRBP and enolase genes. These transcripts contain the 5′ ends of mature UbCRBP mRNAs, extend through UbCRBP, across the intergenic region, and a significant distance 3′ into the enolase gene. Interestingly, the close linkage between the UbCRBP and enolase genes is conserved in a second flatworm, Fasciola hepatica, which also trans-splices the downstream enolase gene. Taken together, the role of SL addition in resolving multicistronic transcripts in both C. elegans and the kinetoplastida, the conservation of UbCRBP/enolase gene linkage in two divergent trematodes, and the multicistronic organization of schistosome UbCRBP/enolase RNAs are consistent with the suggestion that these two genes are likely to be cotranscribed and that trans-splicing in flatworms may be associated with polycistronic transcripts.

A selenium-containing phospholipid-hydroperoxide glutathione peroxidase in Schistosoma mansoni.

The 100000Xg supernatant parasite platyhelminth Schistosoma mansoni exhibits a glutathione peroxidase activity with the substrate phosphatidylcholine hydroperoxide. Purification yielded a protein of 20 kDa molecular mass both on gel filtration column chromatography and SDS/PAGE, thus suggesting that S. mansoni expresses a protein similar to the mammalian selenoenzynic phospholipid-hydroperoxide glutathione peroxidase. Kinetic analysis and substrate specificity corroborated this assumption, the second-order rate constants for the oxidation of the ground-state enzyme (k+1) being higher with phosphatidylcholine hydroperoxide than with other peroxide substrates, such as cumene liydroperoxide or H2O2, and quantitatively similar to those of mammalian phospholipid-hydroperoxide glutathione peroxidase. Partial sequencing of the protein and selenium measurement by neutron activation analysis established that the purified peroxidase corresponded to the product of the S. mansoni gene previously reported and supposed to encode a selenium-containing glutathione peroxidase [Roche, C., Williams, D. L., Khalife, J., LePresle, T., Capron, A. & Pierce, R. J. (1994) Cloning and characterization of gene encoding Schistosoma mansoni glutathione peroxidase, Gene 138, 149 - 152]. S. mansoni thus contains a scienoperoxidase sharing molecular mass, catalytic efficiency and substrate specificity with phospholipid-hydroperoxide glutathione peroxidase, dismantling the concept that those enzymes are unique to vertebrate organisms.