Cloning and Expression of the Major SecretedCathepsin B-Like Protein from Juvenile
            <i>Fasciola hepatica</i>
            andAnalysis of Immunogenicity following Liver FlukeInfection

Law, Ruby H. P.; Smooker, Peter M.; Irving, James A.; Piedrafita, David; Ponting, Rebecca C.; Kennedy, N.; Whisstock, James C.; Pike, Robert N.; Spithill, Terry W.

doi:10.1128/iai.71.12.6921-6932.2003

Cited by 85 publications

(51 citation statements)

References 53 publications

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“…Other examples are the transcripts encoding six novel cathepsin B cysteine proteases and five new asparaginyl endopeptidases. Although some isotypes of these proteases are secreted by the infective larvae and immature liver stage flukes (see below), they are absent from the secretome of adult F. hepatica or F. gigantica as previously reported using biochemical, proteomics, and immunoblotting methods (8,40,41). Considering these observations and the fact that cathepsin B and asparaginyl endopeptidases also function internally within cells (42,43), it is likely that these newly discovered proteases have functions within the internal tissues of this complex multicellular parasite and are involved in generalized protein turnover, remodeling, and/or catabolism.…”

Section: Discussionmentioning

confidence: 77%

An Integrated Transcriptomics and Proteomics Analysis of the Secretome of the Helminth Pathogen Fasciola hepatica

Robinson

Menon

Donnelly

et al. 2009

Molecular & Cellular Proteomics

251

249

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To infect their mammalian hosts, Fasciola hepatica larvae must penetrate and traverse the intestinal wall of the duodenum, move through the peritoneum, and penetrate the liver. After migrating through and feeding on the liver, causing extensive tissue damage, the parasites move to their final niche in the bile ducts where they mature and produce eggs. Here we integrated a transcriptomics and proteomics approach to profile Fasciola secretory proteins that are involved in host-pathogen interactions and to correlate changes in their expression with the migration of the parasite. Prediction of F. hepatica secretory proteins from 14,031 expressed sequence tags (ESTs) available from the Wellcome Trust Sanger Centre using the semiautomated EST2Secretome pipeline showed that the major components of adult parasite secretions are proteolytic enzymes including cathepsin L, cathepsin B, and asparaginyl endopeptidase cysteine proteases as well as novel trypsinlike serine proteases and carboxypeptidases. Proteomics analysis of proteins secreted by infective larvae, immature flukes, and adult F. hepatica showed that these proteases are developmentally regulated and correlate with the passage of the parasite through host tissues and its encounters with different host macromolecules. Proteases such as FhCL3 and cathepsin B have specific functions in larvae activation and intestinal wall penetration, whereas FhCL1, FhCL2, and FhCL5 are required for liver penetration and tissue and blood feeding. Besides proteases, the parasites secrete an array of antioxidants that are also highly regulated according to their migration through host tissues. However, whereas the proteases of F. hepatica are secreted into the parasite gut via a classical endoplasmic reticulum/Golgi pathway, we speculate that the antioxidants, which all lack a signal sequence, are released via a non-classical trans-tegumental pathway. Fasciola hepatica is a helminth (worm) parasite with a worldwide distribution. Although traditionally regarded as a parasite of livestock, particularly sheep and cattle, that results in a large economic loss to the agricultural community it has recently emerged as an important human infection in many regions of the world, including South America, Iran, Egypt, and mainland South-East Asia (1). Dormant larvae contained within cysts adhere to vegetation and emerge as infective juveniles (newly excysted juveniles (NEJs)) 1 in the duodenum following ingestion by animals or humans. They infect their hosts by rapidly penetrating the intestinal wall and entering the peritoneal cavity where they break through the liver capsule. After 8 -12 weeks of consistent burrowing, feeding, and growth within the liver parenchyma they move to their final destination within the bile ducts where they mature and produce enormous numbers of eggs (2). The two distinct clinical phases of fasciolosis are directly related to the migration of the parasites: acute fasciolosis, which manifests as fever, abdominal pain, weight loss, and hepatomegaly, is associated with...

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Section: Discussionmentioning

confidence: 77%

An Integrated Transcriptomics and Proteomics Analysis of the Secretome of the Helminth Pathogen Fasciola hepatica

Robinson

Menon

Donnelly

et al. 2009

Molecular & Cellular Proteomics

251

249

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“…Proteolytic enzymes are important for digestion of the host tissue and are used in the parasite's defense mechanism. The proteolytic enzymes and antioxidants have been proposed as vaccine candidates against fascioliasis (5)(6)(7)(8)(9)(10)(11)(12)(13).…”

mentioning

confidence: 99%

Fasciola hepatica Fatty Acid Binding Protein Induces the Alternative Activation of Human Macrophages

Figueroa-Santiago

Espino

2014

Infect Immun

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The liver fluke Fasciola hepatica is a highly evolved parasite that uses sophisticated mechanisms to evade the host immune response. The immunosuppressive capabilities of the parasite have been associated with antigens secreted through the parasite's tegument, called excretory-secretory products (ESPs). Proteomic studies have identified the fatty acid binding protein (FABP) as one of molecules present in the parasite ESPs. Although FABP has been investigated for potential use in the development of vaccines against fascioliasis, its direct interaction with cells of immune system has not been studied. In this study, FABP was purified in native form from soluble extracts of F. hepatica adult flukes using a combination of molecular sieving chromatography and preparative isoelectric focusing. The immunological effect of the purified protein, termed Fh12, was assayed in vitro using monocyte-derived macrophages (MDM) obtained from healthy human donors. Results from the assay indicate that Fh12 produced a significantly increased arginase expression and activity and induced the expression of chitinase-3-like protein (CHI3L1). The assay also showed that Fh12 downregulated the production of nitric oxide (NO) and the expression of nitric oxide synthase (NOS2). This indicates that Fh12 induced the production of alternatively activated macrophages (AAM). The results also demonstrated the ability of Fh12 to downregulate the secretion of the proinflammatory and inflammatory cytokines tumor necrosis factor alpha (TNF-␣), interleukin-12 (IL-12), and IL-1␤B, even after stimulation with lipopolysaccharide (LPS), as well as its ability to stimulate the overexpression of IL-10. These results suggest a potent anti-inflammatory role for Fh12, which could occur via targeting of Toll-like receptor 4 (TLR4).

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“…The migration of the parasite in host tissues is mediated by the release of proteolytic enzymes that can degrade the tissue barriers. Potent proteolytic enzymes of cysteine, serine, and metalloprotease classes have been identified in secretory products of many of the parasites (8,19,26,27,30). Serine proteases have been reported to have strong hydrolytic activity against a wide range of extracellular matrix (ECM) components and human plasma proteins; hence, they are thought to play vital roles in the host tissue invasion process (17).…”

mentioning

confidence: 99%

In SituImmunolocalization and Stage-Dependent Expression of a Secretory Serine Protease inLeishmania donovaniand Its Role as a Vaccine Candidate

Choudhury

Das

Bhaumik

et al. 2010

Clin Vaccine Immunol

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Proteases have been found to play essential roles in many biological processes, including the pathogenesis of leishmaniasis. Most parasites rely on their intracellular and extracellular protease repertoire to invade and multiply in mammalian host cells. However, few studies have addressed serine proteases in Leishmania and their role in host pathogenesis. Here we report the intracellular distribution of a novel L. donovani secretory serine protease in the flagellar pocket, as determined by immunogold labeling. Flow cytometry and confocal immunofluorescence analysis revealed that the expression of the protease diminishes sequentially from virulent to attenuated strains of this species and is also highly associated with the metacyclic stage of L. donovani promastigotes. The level of internalization of parasites treated with the anti-115-kDa antibody into host macrophages was significantly reduced from that of non-antibody-treated parasites, suggesting that this serine protease probably plays a role in the infection process. In vivo studies confirmed that this serine protease is a potential vaccine candidate. Altogether, the 115-kDa serine protease might play vital roles in L. donovani pathogenesis and hence could be recognized as a potential candidate for drug design.Leishmania species, belonging to the family Trypanosomatidae, are a group of protozoan pathogens that cause a spectrum of chronic diseases ranging from self-healing cutaneous lesions to a lethal visceral disorder. They represent a major health problem in tropical and subtropical areas around the world. Leishmania species have a relatively simple life cycle, with parasite stage differentiation regulated by environmental signals encountered in their different hosts. They are digenetic parasites: the flagellated promastigote forms, which can be derived in axenic culture, differentiate within the alimentary tracts of sandfly vectors from a replicating procyclic to a nonreplicating, infectious metacyclic stage, whereas obligately intracellular amastigotes live and replicate in mammalian mononuclear phagocytes, such as macrophages (1).Recent advances in genomic analysis of several of the major global parasites have revealed key factors involved in the pathogenesis of parasite diseases. Among the major virulence factors identified are parasite-derived proteases. Parasite proteases play significant roles in the pathogenesis of parasitic diseases; the proteases are involved in the invasion of the host via parasite migration through tissue barriers, degradation of host proteins for their nutrition, immune evasion, and activation of inflammation (21). The migration of the parasite in host tissues is mediated by the release of proteolytic enzymes that can degrade the tissue barriers. Potent proteolytic enzymes of cysteine, serine, and metalloprotease classes have been identified in secretory products of many of the parasites (8,19,26,27,30). Serine proteases have been reported to have strong hydrolytic activity against a wide range of extracellular matrix (ECM) c...

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Cloning and Expression of the Major SecretedCathepsin B-Like Protein from Juvenile Fasciola hepatica andAnalysis of Immunogenicity following Liver FlukeInfection

Cited by 85 publications

References 53 publications

An Integrated Transcriptomics and Proteomics Analysis of the Secretome of the Helminth Pathogen Fasciola hepatica

An Integrated Transcriptomics and Proteomics Analysis of the Secretome of the Helminth Pathogen Fasciola hepatica

Fasciola hepatica Fatty Acid Binding Protein Induces the Alternative Activation of Human Macrophages

In SituImmunolocalization and Stage-Dependent Expression of a Secretory Serine Protease inLeishmania donovaniand Its Role as a Vaccine Candidate

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