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Thanks to the increasing availability of the parasitic Platyhelminthes genomes in recent years, several studies have been directed to the identification of the nuclear receptors set expressed by these organisms. Nevertheless, important gaps in our knowledge remain to be addressed, concerning their mechanism of action, ligands, co-regulator proteins, and DNA binding sequences on target genes. The proposed review chapter will be an account of research into the nuclear receptors field of parasitic Platyhelminthes. Several in vitro effects of host steroid hormones on Taenia and Echinococcus species were observed, however, the classical mammalian estrogen, androgen, or progesterone receptors could not be identified in databases. Nonetheless, novel nuclear receptors and related proteins and genes, are being identified and characterized. The elucidation of their target genes as well as ligands in parasitic Platyhelminthes could allow discovery of new and specific pathways differing from those of their hosts. In this sense, these parasitic proteins seem to be good putative targets of new drugs.

Nuclear receptors are ligand-activated transcription factors capable of regulating the expression of complex gene networks. The family includes seven subfamilies of protein with a wide phylogenetic distribution. A novel subfamily with two DNA-binding domains (2DBDs) has been first reported in

Two fatty acid binding proteins, EgFABP1 and EgFABP2, were isolated from the parasitic platyhelminth Echinococcus granulosus. These proteins bind fatty acids and have particular relevance in flatworms since de novo fatty acids synthesis is absent. Therefore platyhelminthes depend on the capture and intracellular distribution of host's lipids and fatty acid binding proteins could participate in lipid distribution. To elucidate EgFABP's roles, we investigated their intracellular distribution in the larval stage by a proteomic approach. Our results demonstrated the presence of EgFABP1 isoforms in cytosolic, nuclear, mitochondrial and microsomal fractions, suggesting that these molecules could be involved in several cellular processes.

This work describes two new fatty acid binding proteins (FABPs) identified in the parasite platyhelminth Mesocestoides vogae (syn. corti). The corresponding polypeptide chains share 62% identical residues and overall 90% similarity according to clustalx default conditions. Compared with Cestoda FABPs, these proteins share the highest similarity score with the Taenia solium protein. M. vogae FABPs are also phylogenetically related to the FABP3/FABP4 mammalian FABP subfamilies. The native proteins were purified by chromatographical procedures, and apparent molecular mass and isoelectric point were determined. Immunolocalization studies determined the localization of the expression of these proteins in the larval form of the parasite. The genomic exon–intron organization of both genes is also reported, and supports new insights on intron evolution. Consensus motifs involved in splicing were identified.

We describe the 1.6 A crystal structure of the fatty-acid-binding protein EgFABP1 from the parasitic platyhelminth Echinococcus granulosus. E. granulosus causes hydatid disease, which is a major zoonosis. EgFABP1 has been implicated in the acquisition, storage, and transport of lipids, and may be important to the organism since it is incapable of synthesising most of its lipids de novo. Moreover, EgFABP1 is a promising candidate for a vaccine against hydatid disease. The crystal structure reveals that EgFABP1 has the expected 10-stranded beta-barrel fold typical of the family of intracellular lipid-binding proteins, and that it is structurally most similar to P2 myelin protein. We describe the comparison of the crystal structure of EgFABP1 with these proteins and with an older homology model for EgFABP1. The electron density reveals the presence of a bound ligand inside the cavity, which we have interpreted as palmitic acid. The carboxylate group of the fatty acid interacts with the protein's P2 motif, consisting of a conserved triad R em leader R-x-Y. The hydrophobic tail of the ligand assumes a fairly flat, U-shaped conformation and has relatively few interactions with the protein.We discuss some of the structural implications of the crystal structure of EgFABP1 for related platyhelminthic FABPs.

Tropomyosins are a family of actin-binding proteins with diverse roles in actin filament function. One of the best characterized roles is the regulation of muscle contraction. Tropomyosin isoforms can be generated from different genes, and from alternative promoters and alternative splicing from the same gene. In this work, we have isolated sequences for tropomyosin isoforms from the cestode Mesocestoides corti, and searched for tropomyosin genes and isoforms in other flatworms. Two genes are conserved in the cestodes M. corti and Echinococcus multilocularis, and in the trematode Schistosoma mansoni. Both genes have the same structure, and each gene gives rise to at least two different isoforms, a high molecular weight (HMW) and a low molecular weight (LMW) one. Because most exons are duplicated and spliced in a mutually exclusive fashion, isoforms from one gene only share one exon and are highly divergent. The gene duplication preceded the divergence of neodermatans and the planarian Schmidtea mediterranea. Further duplications occurred in Schmidtea, coupled to the selective loss of duplicated exons, resulting in genes that only code for HMW or LMW isoforms. A polyclonal antibody raised against a HMW tropomyosin from Echinococcus granulosus was demonstrated to specifically recognize HMW tropomyosin isoforms of M. corti, and used to study their expression during segmentation. HMW tropomyosins are expressed in muscle layers, with very low or absent levels in other tissues. No expression of HMW tropomyosins is present in early or late genital primordia, and expression only begins once muscle fibers develop in the genital ducts. Therefore, HMW tropomyosins are markers for the development of muscles during the final differentiation of genital primordia.

Tropomyosins (Trps) constitute a family of actin filament-binding proteins found in all eukaryotic cells. In muscle cells, they play a central role in contraction by regulating calcium-sensitive interaction of actin and myosin. In non-muscle cells, tropomyosins regulate actin filament organization and dynamics. Trps genes exhibit extensive cell type-specific isoform diversity generated by alternative splicing. Here, we report the characterization of tropomyosin gene transcribed sequences from the parasitic platyhelminth Echinococcus granulosus. Using RT-PCR approach we isolated three isoforms (egtrpA, egtrpB and egtrpC), which display significant homologies to know tropomyosins of different phylogenetic origin. The corresponding gene, egtrp (5656 bp), contains eight introns and nine exons. Southern blot hybridization studies showed that egtrp is present as single copy locus in E. granulosus. We demonstrated that egtrp expresses three different transcripts which differ in alternatively spliced exon 4 and intron VI. Interestingly, intron VI suffers intron retention and contains an internal stop codon in frame. Three major bands are also detected by Western blot analysis using a specific anti-rEgTrp antiserum. Immune-localization and in situ hybridization studies showed that egtrp transcription and translation is mostly localized at the protoscoleces suckers. This is the first report of alternative splicing in this parasite.

This work describes two new fatty acid binding proteins (FABPs) identified in the parasite platyhelminth Mesocestoides vogae (syn. corti). The corresponding polypeptide chains share 62% identical residues and overall 90% similarity according to CLUSTALX default conditions. Compared with Cestoda FABPs, these proteins share the highest similarity score with the Taenia solium protein. M. vogae FABPs are also phylogenetically related to the FABP3/FABP4 mammalian FABP subfamilies. The native proteins were purified by chromatographical procedures, and apparent molecular mass and isoelectric point were determined. Immunolocalization studies determined the localization of the expression of these proteins in the larval form of the parasite. The genomic exon-intron organization of both genes is also reported, and supports new insights on intron evolution. Consensus motifs involved in splicing were identified.

EgFABP1 is a developmentally regulated intracellular fatty acid binding protein characterized in the larval stage of parasitic platyhelminth Echinococcus granulosus. It is structurally related to the heart group of fatty acid binding proteins (H-FABPs). Binding properties and ligand affinity of recombinant EgFABP1 were determined by fluorescence spectroscopy using cis- and trans-parinaric acid. Two binding sites for cis- and trans-parinaric acid were found (K(d(1)) 24+/-4 nM, K(d(2)) 510+/-60 nM for cis-parinaric acid and K(d(1)) 32+/-4 nM, K(d(2)) 364+/-75 nM for trans-parinaric). A putative third site for both fatty acids is discussed. Binding preferences were determined using displacement assays. Arachidonic and oleic acids presented the highest displacement percentages for EgFABP1. The Echinococcus FABP is the unique member of the H-FABP group able to bind two long chain fatty acid molecules with high affinity. Structure-function relationships and putative roles for EgFABP1 in E. granulosus metabolism are discussed.

Dogs are the main source of human cystic echinococcosis. An oral vaccine would be an important contribution to control programs in endemic countries. We conducted two parallel experimental trials in Morocco and Tunisia of a new oral vaccine candidate against Echinococcus granulosus in 28 dogs. The vaccine was prepared using two recombinant proteins from adult worms, a tropomyosin (EgTrp) and a fibrillar protein similar to paramyosin (EgA31), cloned and expressed in a live attenuated strain of Salmonella enterica serovar typhimurium. In each country, five dogs were vaccinated with the associated EgA31 and EgTrp; three dogs received only the vector Salmonella; and six dogs were used as different controls. The vaccinated dogs received two oral doses of the vaccine 21 d apart, and were challenged 20 d later with 75,000 living protoscoleces. The controls were challenged under the same conditions. All dogs were sacrificed 26–29 d postchallenge, before the appearance of eggs, for safety reasons. We studied the histological responses to both the vaccine and control at the level of the duodenum, the natural localization of the cestode. Here we show a significant decrease of parasite burden in vaccinated dogs (70% to 80%) and a slower development rate in all remaining worms. The Salmonella vaccine EgA31-EgTrp demonstrated a high efficacy against E. granulosus promoting its potential role in reducing transmission to humans and animals., Author Summary In many countries in the world, livestock and humans are affected with hydatid disease, which is caused by the development, in the viscera, of the larval stage of the cestode Echinococcus granulosus. They become infected by ingesting the eggs of this parasite, which are passed in the feces of the dog—the host of the adult worm. Domestic dogs are key in the transmission to livestock and humans. This disease remains a major economic and public health problem in affected countries. Because dogs are quickly reinfected, control programs in these locations include monthly anthelmintic deworming. These control measures, however, are burdensome for the owner, so they often fail. In contrast, vaccination can take place in control programs at different stages of the parasite life cycle. For example, currently an effective recombinant vaccine for sheep has been developed that should work indirectly to reduce infection in dogs, which tend to eat sheep offal. However, we propose that a recombinant oral vaccine given to the small number of dogs keeping the herd would decrease the number of Echinococcus granulosus adult worms and, consequently, the number of infective eggs. This measure would help reduce the contamination risk factors for humans and livestock, and would be cost-effective for the owners of the dogs.