Molecular cloning and analysis of stage and tissue-specific expression of cathepsin B encoding genes from Fasciola gigantica

Meemon, Krai; Grams, Rudi; Vichasri-Grams, Suksiri; Hofmann, Annemarie; Korge, Günter; Viyanant, Vithoon; Upatham, E. S.; Habe, Shigehisa; Sobhon, Prasert

doi:10.1016/j.molbiopara.2004.02.010

Cited by 73 publications

(48 citation statements)

References 30 publications

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“…Previous studies have detected significant levels of transcript for cathepsin B cysteine proteases in F. hepatica and F. gigantica infective juveniles and immature liver stages, but the expression of these in adult parasites was low (32)(33)(34). Additionally immunoblotting experiments using antisera raised against a recombinant F. hepatica cathepsin B detected this protease in proteins secreted by immature parasites but not by adult parasites (35).…”

Section: Discussionmentioning

confidence: 92%

Proteomics and Phylogenetic Analysis of the Cathepsin L Protease Family of the Helminth Pathogen Fasciola hepatica

Robinson

Tort

Lowther

et al. 2008

Molecular & Cellular Proteomics

120

131

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Cathepsin L proteases secreted by the helminth pathogen Fasciola hepatica have functions in parasite virulence including tissue invasion and suppression of host immune responses. Using proteomics methods alongside phylogenetic studies we characterized the profile of cathepsin L proteases secreted by adult F. hepatica and hence identified those involved in host-pathogen interaction. Phylogenetic analyses showed that the Fasciola cathepsin L gene family expanded by a series of gene duplications followed by divergence that gave rise to three clades associated with mature adult worms (Clades 1, 2, and 5) and two clades specific to infective juvenile stages (Clades 3 and 4). Consistent with these observations our proteomics studies identified representatives from Clades 1, 2, and 5 but not from Clades 3 and 4 in adult F. hepatica secretory products. Clades 1 and 2 account for 67.39 and 27.63% of total secreted cathepsin Ls, respectively, suggesting that their expansion was positively driven and that these proteases are most critical for parasite survival and adaptation. Sequence comparison studies revealed that the expansion of cathepsin Ls by gene duplication was followed by residue changes in the S2 pocket of the active site. Our biochemical studies showed that these changes result in alterations in substrate binding and suggested that the divergence of the cathepsin L family produced a repertoire of enzymes with overlapping and complementary substrate specificities that could cleave host macromolecules more efficiently. Although the cathepsin Ls are produced as zymogens containing a prosegment and mature domain, all secreted enzymes identified by MS were processed to mature active enzymes. The prosegment region was highly conserved between the clades except at the boundary of prosegment and mature enzyme. Despite the lack of conservation at this section, sites for exogenous cleavage by asparaginyl endopeptidases and a Leu-Ser2His motif for autocatalytic cleavage by cathepsin Ls were preserved.

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Section: Discussionmentioning

confidence: 92%

Proteomics and Phylogenetic Analysis of the Cathepsin L Protease Family of the Helminth Pathogen Fasciola hepatica

Robinson

Tort

Lowther

et al. 2008

Molecular & Cellular Proteomics

120

131

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“…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. Alternatively or additionally, because immunocytochemistry and in situ hybridization have localized cysteine proteases and transcripts to the reproductive structures of other trematodes (44,45) these proteases may be involved in the process of egg production. Other proteins identified by EST2Secretome analysis but not by proteomics, including the previously uncharacterized proteins such as deoxyribonucleases, carboxypeptidase, and trypsin-like peptidases that have homologues in Schistosoma japonicum and C. sinensis, may also be confined to internal tissues of the parasite or be expressed and secreted at levels below the detection capacity of the proteomics methods so far used (cDNAs of these were poorly represented in the transcriptome of the adult parasite).…”

Section: Discussionmentioning

confidence: 99%

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

Robinson

Menon

Donnelly

et al. 2009

Molecular & Cellular Proteomics

250

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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“…Cathepsin B-like proteases have been demonstrated to be one of the major proteases synthesized by newly excysted juvenile (NEJ) and immature Fasciola parasites (Tkalcevic et al 1995;Wilson et al 1998;Law et al 2003;Meemon et al 2004;Cancela et al 2008). The first cathepsin B of Fasciola spp.…”

Section: Cathepsin Bmentioning

confidence: 99%

“…namely FhCatB1 (also known as CB2) was identified in the somatic and excretory-secretory (ES) extracts of F. hepatica NEJs and 5-week-old immature parasites (Tkalcevic et al 1995;Wilson et al 1998;Law et al 2003). Later, three cathepsin B isotypes were identified from stage-specific cDNA libraries of F. gigantica and named FgCatB1, FgCatB2, and FgCatB3 (Meemon et al 2004). The orthologous cathepsin B isotypes (CB1, CB2, CB3) were then identified in juvenile F. hepatica (Cancela et al 2008(Cancela et al , 2010.…”

Section: Cathepsin Bmentioning

confidence: 99%

“…Cathepsin Ls were the major proteases expressed in adult Fasciola spp. (Grams et al 2001;Jefferies et al 2001;Morphew et al 2007;Robinson et al 2009), whereas cathepsin Bs were expressed predominantly in juvenile parasites (Wilson et al 1998;Meemon et al 2004). However, several cathepsin L and B isoforms were also found to differentially express during developmental stages of Fasciola spp.…”

Section: Introductionmentioning

confidence: 99%

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Juvenile-specific cathepsin proteases in Fasciola spp.: their characteristics and vaccine efficacies

Meemon

Sobhon

2015

Parasitol Res

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Fasciolosis, caused by Fasciola hepatica and Fasciola gigantica, is one of the most neglected tropical zoonotic diseases. One sustainable control strategy against these infections is the employment of vaccines that target proteins essential for parasites' invasion and nutrition acquiring processes. Cathepsin proteases are the most abundantly expressed proteins in Fasciola spp. that have been tested successfully as vaccines against fasciolosis in experimental as well as large animals because of their important roles in digestion of nutrients, invasion, and migration. Specifically, juvenile-specific cathepsin proteases are the more effective vaccines because they could block the invasion and migration of juvenile parasites whose immune evasion mechanism has not yet been fully developed. Moreover, because of high sequence similarity and identity of cathepsins from juveniles with those of adults, the vaccines can attack both the juvenile and adult stages. In this article, the characteristics and vaccine potentials of juvenile-specific cathepsins, i.e., cathepsins L and B, of Fasciola spp. were reviewed.

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Molecular cloning and analysis of stage and tissue-specific expression of cathepsin B encoding genes from Fasciola gigantica

Cited by 73 publications

References 30 publications

Proteomics and Phylogenetic Analysis of the Cathepsin L Protease Family of the Helminth Pathogen Fasciola hepatica

Proteomics and Phylogenetic Analysis of the Cathepsin L Protease Family of the Helminth Pathogen Fasciola hepatica

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

Juvenile-specific cathepsin proteases in Fasciola spp.: their characteristics and vaccine efficacies

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