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Biological function of Dictyocaulus viviparus asparaginyl peptidase legumain-1 and its suitability as a vaccine target

Published online by Cambridge University Press:  25 September 2017

JULIA HOLZHAUSEN ,
CLAAS HAAKE ,
SABINE SCHICHT ,
PETRA HINSE ,
DANIELA JORDAN ,
ELISABETH KREMMER  and
CHRISTINA STRUBE
JULIA HOLZHAUSEN
Affiliation:
Institute for Parasitology, Centre for Infection Medicine, University of Veterinary Medicine Hannover, Buenteweg 17, 30559 Hanover, Germany
CLAAS HAAKE
Affiliation:
Institute for Parasitology, Centre for Infection Medicine, University of Veterinary Medicine Hannover, Buenteweg 17, 30559 Hanover, Germany
SABINE SCHICHT
Affiliation:
Institute for Parasitology, Centre for Infection Medicine, University of Veterinary Medicine Hannover, Buenteweg 17, 30559 Hanover, Germany
PETRA HINSE
Affiliation:
Institute for Parasitology, Centre for Infection Medicine, University of Veterinary Medicine Hannover, Buenteweg 17, 30559 Hanover, Germany
DANIELA JORDAN
Affiliation:
Institute for Parasitology, Centre for Infection Medicine, University of Veterinary Medicine Hannover, Buenteweg 17, 30559 Hanover, Germany
ELISABETH KREMMER
Affiliation:
Helmholtz Zentrum München, German Research Center for Environmental Health, Institute of Molecular Immunology, Marchioninistraße 25, 81377 Munich, Germany
CHRISTINA STRUBE*
Affiliation:
Institute for Parasitology, Centre for Infection Medicine, University of Veterinary Medicine Hannover, Buenteweg 17, 30559 Hanover, Germany
*
*Corresponding author: Institute for Parasitology, Centre for Infection Medicine, University of Veterinary Medicine Hannover, Buenteweg 17, 30559 Hanover, Germany; Email: christina.strube@tiho-hannover.de
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Summary

The present study characterized the biological function of the asparaginyl peptidase legumain-1 (LEG-1) of the bovine lungworm Dictyocaulus viviparus and its suitability as a recombinant vaccine against dictyocaulosis. Quantitative real-time PCR and immunoblot analysis revealed LEG-1 to be almost exclusively transcribed and expressed in parasitic lungworm stages. Immunohistochemistry localized the enzyme in the parasite's gut, which was confirmed by immunoblots detecting LEG-1 in the gut as well as male testes. LEG-1 was recombinantly (rLEG-1) expressed in the yeast Pichia pastoris and subsequently analysed in activity assays for its enzyme functions and substrate specificity. For sufficient functionality, rLEG-1 needed trans-activation through D. viviparus cathepsin L-2, indicating a novel mechanism of legumain activation. After trans-activation, rLEG-1 worked best at pH 5·5 and 35–39 °C and cleaved a legumain-specific artificial substrate as well as the natural substrates bovine collagen types I and II. In a clinical vaccination trial, rLEG-1 did not protect against challenge infection. Results of in vitro characterization, transcription pattern and localization enhance the presumption that LEG-1 participates in digestion processes of D. viviparus. Since rLEG-1 needs trans-activation through a cathepsin, it is probably involved in an enzyme cascade and therefore remains interesting as a candidate in a multi-component vaccine.

Keywords

Legumainbovine lungwormnematodeenzyme activityimmunohistochemistryvaccinationimmunizationhidden antigen
Type
Research Article
Information
Parasitology , Volume 145 , Issue 3 , March 2018 , pp. 378 - 392
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Copyright
Copyright © Cambridge University Press 2017 

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Footnotes

†

Present address: Department of Biology II, Ludwig-Maximilians University, Großhaderner Straße 2, 82152 Martinsried, Germany.

References

Adisakwattana, P., Viyanant, V., Chaicumpa, W., Vichasri-Grams, S., Hofmann, A., Korge, G., Sobhon, P. and Grams, R. (2007). Comparative molecular analysis of two asparaginyl endopeptidases and encoding genes from Fasciola gigantica . Molecular and Biochemical Parasitology 156, 102–116.Google Scholar
Caffrey, C. R., Mathieu, M. A., Gaffney, A. M., Salter, J. P., Sajid, M., Lucas, K. D., Franklin, C., Bogyo, M. and McKerrow, J. H. (2000). Identification of a cDNA encoding an active asparaginyl endopeptidase of Schistosoma mansoni and its expression in pichia pastoris . FEBS Letters 466, 244–248.Google Scholar
Chen, J. M., Dando, P. M., Rawlings, N. D., Brown, M. A., Young, N. E., Stevens, R. A., Hewitt, E., Watts, C. and Barrett, A. J. (1997). Cloning, isolation, and characterization of mammalian legumain, an asparaginyl endopeptidase. The Journal of Biological Chemistry 272, 8090–8098.Google Scholar
Chlichlia, K., Bahgat, M., Ruppel, A. and Schirrmacher, V. (2001). DNA vaccination with asparaginyl endopeptidase (Sm32) from the parasite Schistosoma mansoni: anti-fecundity effect induced in mice. Vaccine 20, 439–447.Google Scholar
Cox, J. C. and Coulter, A. R. (1997). Adjuvants – a classification and review of their modes of action. Vaccine 15, 248–256.Google Scholar
Dall, E. and Brandstetter, H. (2016). Structure and function of legumain in health and disease. Biochemie 122, 126–150.CrossRefGoogle ScholarPubMed
Dalton, J. P. and Brindley, P. J. (1996). Schistosome asparaginyl endopeptidase SM32 in hemoglobin digestion. Parasitology Today 12, 125.Google Scholar
Dalton, J. P., Hola-Jamriska, L. and Brindley, P. J. (1995). Asparaginyl endopeptidase activity in adult Schistosoma mansoni . Parasitology 111, 575–580.Google Scholar
Dalton, J. P., Brindley, P. J., Donnelly, S. and Robinson, M. W. (2009). The enigmatic asparaginyl endopeptidase of helminth parasites. Trends in Parasitology 25, 59–61.CrossRefGoogle ScholarPubMed
Horn, M., Nussbaumerová, M., Šanda, M., Kovářová, Z., Srba, J., Franta, Z., Sojka, D., Bogyo, M., Caffrey, C. R., Kopáček, P. and Mareš, M. (2009). Hemoglobin digestion in blood-feeding ticks: mapping a multipeptidase pathway by functional proteomics. Chemistry & Biology 16, 1053–1063.Google Scholar
Jarrett, W. F. H., McIntyre, W. I. M., Jennings, F. W. and Mulligan, S. T. (1957). The natural history of parasitic bronchitis with notes on prophylaxis and treatment. Veterinary Record 69, 1329–1339.Google Scholar
Joekel, D., Hinse, P., Raulf, M. K., Schicht, S., Baumer, W., Werling, D., Kremmer, E. and Strube, C. (2015). Vaccination of calves with yeast- and bacterial-expressed paramyosin from the bovine lungworm Dictyocaulus viviparus . Parasite Immunology 37, 614–623.Google Scholar
Kembhavi, A. A., Buttle, D. J., Knight, C. G. and Barrett, A. J. (1993). The two cysteine endopeptidases of legume seeds: purification and characterization by use of specific fluorometric assays. Archives of Biochemistry and Biophysics 303, 208–213.Google Scholar
Knox, D. P., Smith, S. K., Redmond, D. L. and Smith, W. D. (2005). Protection induced by vaccinating sheep with a thiol-binding extract of Haemonchus contortus membranes is associated with its protease components. Parasite Immunology 27, 121–126.Google Scholar
Krautz-Peterson, G. and Skelly, P. J. (2008). Schistosome asparaginyl endopeptidase (legumain) is not essential for cathepsin B1 activation in vivo . Molecular and Biochemical Parasitology 159, 54–58.Google Scholar
Kuerpick, B., Schnieder, T. and Strube, C. (2013). Evaluation of a recombinant cathepsin L1 ELISA and comparison with the Pourquier and ES ELISA for the detection of antibodies against Fasciola hepatica . Veterinary Parasitology 193, 206–213.Google Scholar
Laabs, E. M., Schnieder, T. and Strube, C. (2011). Transcriptional differences between hypobiotic and non-hypobiotic preadult larvae of the bovine lungworm Dictyocaulus viviparus . Parasitology Research 110, 151–159.Google Scholar
Laabs, E. M., Schnieder, T. and Strube, C. (2012). In vitro studies on the sexual maturation of the bovine lungworm Dictyocaulus viviparus during the development of preadult larvae to adult worms. Parasitology Research 110, 1249–1259.Google Scholar
McKeand, J. B. (2000). Vaccine development and diagnostics of Dictyocaulus viviparus . Parasitology 120(Suppl.), S17–S23.CrossRefGoogle ScholarPubMed
Murray, J. and Smith, W. D. (1994). Ingestion of host immunoglobulin by three non-blood-feeding nematode parasites of ruminants. Research in Veterinary Science 57, 387–389.CrossRefGoogle ScholarPubMed
Newton, S. E. and Munn, E. A. (1999). The development of vaccines against gastrointestinal nematode parasites, particularly Haemonchus contortus . Parasitology Today 15, 116–122.Google Scholar
Nguyen, P. V., Srihari, S. and Leong, H. W. (2013). Identifying conserved protein complexes between species by constructing interolog networks. BMC Bioinformatics 14(Suppl. 16), S8.Google Scholar
Oliver, E. M., Skuce, P. J., McNair, C. M. and Knox, D. P. (2006). Identification and characterization of an asparaginyl proteinase (legumain) from the parasitic nematode, Haemonchus contortus . Parasitology 133, 237–244.Google Scholar
Porter, D. A. (1936). The ingestion of the inflammatory exudate by swine lungworms. Journal of Parasitology 22, 411–412.CrossRefGoogle Scholar
Redmond, D. L. and Knox, D. P. (2004). Protection studies in sheep using affinity-purified and recombinant cysteine proteinases of adult haemonchus contortus . Vaccine 22, 4252–4261.Google Scholar
Sajid, M., McKerrow, J. H., Hansell, E., Mathieu, M. A., Lucas, K. D., Hsieh, I., Greenbaum, D., Bogyo, M., Salter, J. P., Lim, K. C., Franklin, C., Kim, J. H. and Caffrey, C. R. (2003). Functional expression and characterization of Schistosoma mansoni cathepsin B and its trans-activation by an endogenous asparaginyl endopeptidase. Molecular and Biochemical Parasitology 131, 65–75.Google Scholar
Schnieder, T., Epe, C., von Samson-Himmelstjerna, G. and Kohlmetz, C. (1996). The development of protective immunity against gastrointestinal nematode and lungworm infections after use of an ivermectin bolus in first-year grazing calves. Veterinary Parasitology 64, 239–250.CrossRefGoogle ScholarPubMed
Scott, C. A., McKeand, J. B. and Devaney, E. (1996). A longitudinal study of local and peripheral isotype/subclass antibodies in Dictyocaulus viviparus-infected calves. Veterinary Immunology and Immunopathology 53, 235–247.Google Scholar
Strube, C., Buschbaum, S., Wolken, S. and Schnieder, T. (2008). Evaluation of reference genes for quantitative real-time PCR to investigate protein disulfide isomerase transcription pattern in the bovine lungworm Dictyocaulus viviparus . Gene 425, 36–43.CrossRefGoogle ScholarPubMed
Strube, C., Buschbaum, S., von Samson-Himmelstjerna, G. and Schnieder, T. (2009). Stage-dependent transcriptional changes and characterization of paramyosin of the bovine lungworm Dictyocaulus viviparus . Parasitology International 58, 334–340.CrossRefGoogle ScholarPubMed
Strube, C., Buschbaum, S. and Schnieder, T. (2012). Genes of the bovine lungworm Dictyocaulus viviparus associated with transition from pasture to parasitism. Infection, Genetics and Evolution 12, 1178–1188.Google Scholar
Strube, C., Haake, C., Sager, H., Schorderet Weber, S., Kaminsky, R., Buschbaum, S., Joekel, D., Schicht, S., Kremmer, E., Korrell, J., Schnieder, T. and von Samson-Himmelstjerna, G. (2015). Vaccination with recombinant paramyosin against the bovine lungworm Dictyocaulus viviparus considerably reduces worm burden and larvae shedding. Parasites & Vectors 8, 733.Google Scholar
Tamura, K., Stecher, G., Peterson, D., Filipski, A. and Kumar, S. (2013). MEGA6: molecular evolutionary genetics analysis version 6·0. Molecular Biology and Evolution 30, 2725–2729.Google Scholar
Urquhart, G. M., Jarrett, W. F., Bairden, K. and Bonazzi, E. F. (1981). Control of parasitic bronchitis in calves: vaccination or treatment? The Veterinary Record 108, 180–182.Google Scholar
Wood, I. B., Amaral, N. K., Bairden, K., Duncan, J. L., Kassai, T., Malone, J. B. Jr., Pankavich, J. A., Reinecke, R. K., Slocombe, O., Taylor, S. M. and Vercruysse, J. (1995). World Association for the Advancement of Veterinary Parasitology (W.A.A.V.P.) second edition of guidelines for evaluating the efficacy of anthelmintics in ruminants (bovine, ovine, caprine). Veterinary Parasitology 58, 181–213.Google Scholar
Yamane, T., Takeuchi, K., Yamamoto, Y., Li, Y. H., Fujiwara, M., Nishi, K., Takahashi, S. and Ohkubo, I. (2002). Legumain from bovine kidney: its purification, molecular cloning, immunohistochemical localization and degradation of annexin II and vitamin D-binding protein. Biochimica et Biophysica Acta 1596, 108–120.Google Scholar