Biolistic transformation of Schistosoma mansoni with 5′ flanking regions of two peptidase genes promotes tissue-specific expression

Wippersteg, Volker; Sajid, Mohammed; Walshe, Deirdre; Khiem, Dustin; Salter, Jason P.; McKerrow, James H.; Grevelding, Christoph G.; Caffrey, Conor R.

doi:10.1016/j.ijpara.2005.02.002

Cited by 37 publications

(22 citation statements)

References 19 publications

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“…In addition, heritable transmission of plasmid-based transgenes has been reported (123,124). Similar findings have been described in schistosomes, and stage- and tissue-specific expression of schistosome genes has been investigated using reporter transgenes driven by the promoters of schistosome genes, including cathepsin L (125). Recently, Brindley and coworkers have described somatic transgenesis of S. mansoni facilitated by either the piggyBac transposon (126) or the murine leukemia retrovirus (127), including the first demonstration of integration of reporter genes into the chromosomes of a parasitic worm.…”

Section: Helminth Genomics Postgenomics and Gene Manipulationmentioning

confidence: 57%

Helminth infections: the great neglected tropical diseases

et al. 2008

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Helminths are parasitic worms. They are the most common infectious agents of humans in developing countries and produce a global burden of disease that exceeds better-known conditions, including malaria and tuberculosis. As we discuss here, new insights into fundamental helminth biology are accumulating through newly completed genome projects and the nascent application of transgenesis and RNA interference technologies. At the same time, our understanding of the dynamics of the transmission of helminths and the mechanisms of the Th2-type immune responses that are induced by infection with these parasitic worms has increased markedly. Ultimately, these advances in molecular and medical helminth biology should one day translate into a new and robust pipeline of drugs, diagnostics, and vaccines for targeting parasitic worms that infect humans.

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Section: Helminth Genomics Postgenomics and Gene Manipulationmentioning

confidence: 57%

Helminth infections: the great neglected tropical diseases

et al. 2008

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“…SmCB1 has been immunolocalized to both the lumen and gastrodermis (37). Biolistic transformation of adult worms with a green fluorescent protein construct containing a 5Ј-flanking region of SmCL1 showed expression of reporter gene product in the gut of adult schistosome worms (53), and SmCL1 has been immunolocalized to the gastrodermal cells (14). It is therefore possible that cysteine proteases produce some degradation at a less acidic pH, in the parasite gut lumen at pH 6 -6.5.…”

Section: Discussionmentioning

confidence: 99%

A Multienzyme Network Functions in Intestinal Protein Digestion by a Platyhelminth Parasite

Delcroix

Sajid

Caffrey

et al. 2006

Journal of Biological Chemistry

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Proteases frequently function not only as individual enzymes but also in cascades or networks. A notable evolutionary switch occurred in one such protease network that is involved in protein digestion in the intestine. In vertebrates, this is largely the work of trypsin family serine proteases, whereas in invertebrates, cysteine proteases of the papain family and aspartic proteases assume the role. Utilizing a combination of protease class-specific inhibitors and RNA interference, we deconvoluted such a network of major endopeptidases functioning in invertebrate intestinal protein digestion, using the parasitic helminth, Schistosoma mansoni as an experimental model. We show that initial degradation of host blood proteins is ordered, occasionally redundant, and substrate-specific. Although inhibition of parasite cathepsin D had a greater effect on primary cleavage of hemoglobin, inhibition of cathepsin B predominated in albumin degradation. Nevertheless, in both cases, inhibitor combinations were synergistic. An asparaginyl endopeptidase (legumain) also synergized with cathepsin B and L in protein digestion, either by zymogen activation or facilitating substrate cleavage. This protease network operates optimally in acidic pH compartments either in the gut lumen or in vacuoles of the intestinal lining cells. Defining the role of each of these major enzymes now provides a clearer understanding of the function of a complex protease network that is conserved throughout invertebrate evolution. It also provides insights into which of these proteases are logical targets for development of chemotherapy for schistosomiasis, a major global health problem.

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“…Cytochemical studies have shown that the cathepsin L and cathepsin F proteases of schistosomes localize to the gastrodermis of the gut and participate in extracellular digestion of ingested host tissues [40]. Caffrey and colleagues showed that promoter regions of the SmCF gene could drive reporter transgene expression in the gut of adult S. mansoni [45], thus providing strong support for a specific role for this protease in digestion of host blood [40]. CsCF-6 is also predominantly expressed in the gut of the adult C. sinensis where it is likely has a nutritional role [13].…”

Section: Discussionmentioning

confidence: 99%

Cathepsin F Cysteine Protease of the Human Liver Fluke, Opisthorchis viverrini

et al. 2009

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BackgroundThe liver fluke Opisthorchis viverrini is classified as a class I carcinogen due to the association between cholangiocarcinoma and chronic O. viverrini infection. During its feeding activity within the bile duct, the parasite secretes several cathepsin F cysteine proteases that may induce or contribute to the pathologies associated with hepatobiliary abnormalities.Methodology/Principal FindingsHere, we describe the cDNA, gene organization, phylogenetic relationships, immunolocalization, and functional characterization of the cathepsin F cysteine protease gene, here termed Ov-cf-1, from O. viverrini. The full length mRNA of 1020 nucleotides (nt) encoded a 326 amino acid zymogen consisting of a predicted signal peptide (18 amino acids, aa), prosegment (95 aa), and mature protease (213 aa). BLAST analysis using the Ov-CF-1 protein as the query revealed that the protease shared identity with cathepsin F-like cysteine proteases of other trematodes, including Clonorchis sinensis (81%), Paragonimus westermani (58%), Schistosoma mansoni and S. japonicum (52%), and with vertebrate cathepsin F (51%). Transcripts encoding the protease were detected in all developmental stages that parasitize the mammalian host. The Ov-cf-1 gene, of ∼3 kb in length, included seven exons interrupted by six introns; the exons ranged from 69 to 267 bp in length, the introns from 43 to 1,060 bp. The six intron/exon boundaries of Ov-cf-1 were conserved with intron/exon boundaries in the human cathepsin F gene, although the gene structure of human cathepsin F is more complex. Unlike Ov-CF-1, human cathepsin F zymogen includes a cystatin domain in the prosegment region. Phylogenetic analysis revealed that the fluke, human, and other cathepsin Fs branched together in a clade discrete from the cathepsin L cysteine proteases. A recombinant Ov-CF-1 zymogen that displayed low-level activity was expressed in the yeast Pichia pastoris. Although the recombinant protease did not autocatalytically process and activate to a mature enzyme, trans-processing by Fasciola hepatica cathepsin L cleaved the prosegment of Ov-CF-1, releasing a mature cathepsin F with activity against the peptide Z-Phe-Arg-NHMec >50 times that of the zymogen. Immunocytochemistry using antibodies raised against the recombinant enzyme showed that Ov-CF-1 is expressed in the gut of the mature hermaphroditic fluke and also in the reproductive structures, including vitelline glands, egg, and testis. Ov-CF-1 was detected in bile duct epithelial cells surrounding the flukes several weeks after infection of hamsters with O. viverrini and, in addition, had accumulated in the secondary (small) bile ducts where flukes cannot reach due to their large size.Conclusions/SignificanceA cathepsin F cysteine protease of the human liver fluke O. viverrini has been characterized at the gene and protein level. Secretion of this protease may contribute to the hepatobiliary abnormalities, including cholangiocarcinogenesis, observed in individuals infected with this parasite.

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Biolistic transformation of Schistosoma mansoni with 5′ flanking regions of two peptidase genes promotes tissue-specific expression

Cited by 37 publications

References 19 publications

Helminth infections: the great neglected tropical diseases

Helminth infections: the great neglected tropical diseases

A Multienzyme Network Functions in Intestinal Protein Digestion by a Platyhelminth Parasite

Cathepsin F Cysteine Protease of the Human Liver Fluke, Opisthorchis viverrini

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