Startseite First identification of a phosphorylcholine-substituted protein from Caenorhabditis elegans: isolation and characterization of the aspartyl protease ASP-6
Artikel
Lizenziert
Nicht lizenziert Erfordert eine Authentifizierung

First identification of a phosphorylcholine-substituted protein from Caenorhabditis elegans: isolation and characterization of the aspartyl protease ASP-6

  • Günter Lochnit , Julia Grabitzki , Björn Henkel , Nektarios Tavernarakis und Rudolf Geyer
Veröffentlicht/Copyright: 2. November 2006
Biological Chemistry
Aus der Zeitschrift Band 387 Heft 10_11

Abstract

Caenorhabditis elegans is a widely accepted model system for parasitic nematodes, drug screening and developmental studies. Similar to parasitic worms, C. elegans expresses glycosphingolipids and glycoproteins carrying, in part, phosphorylcholine (PCho) substitutions, which might play important roles in nematode development, fertility and, at least in the case of parasites, survival within the host. With the exception of a major secretory/excretory product from Acanthocheilonema viteae (ES-62), no protein carrying this epitope has been studied in detail yet. Here we report on the identification, characterization and localization of the aspartyl protease ASP-6 of C. elegans, which is excreted by the nematode in a PCho-substituted form. Within the worm, most prominent expression of the protein is observed in the intestine, while muscle and epithelial cells express asp-6 to a lesser extent. In animals harboring an ASP-6::GFP fusion protein, diffuse fluorescence throughout the body cavity of adult worms indicates that the chimeric protein is secreted.

:

Corresponding author

References

Baumeister, S., Dennis, R.D., Klünder, R., Schares, G., Zahner, H., and Geyer, E. (1994). Litomosoides carinii: macrofilariae-derived glycolipids-chromatography, serology and potential in the evaluation of anthelmintic efficacy. Parasite Immunol.16, 629–641.10.1111/j.1365-3024.1994.tb00319.xSuche in Google Scholar

Becker, M.M., Harrop, S.A., Dalton, J.P., Kalinna, B.H., McManus, D.P., and Brindley, P.J. (1995). Cloning and characterization of the Schistosoma japonicum aspartic proteinase involved in hemoglobin degradation. J. Biol. Chem.270, 24496–24501.10.1074/jbc.270.41.24496Suche in Google Scholar

Brown, A., Girod, N., Billett, E.E., and Pritchard, D.I. (1999). Necator americanus (human hookworm) aspartyl proteinases and digestion of skin macromolecules during skin penetration. Am. J. Trop. Med. Hyg.60, 840–847.10.4269/ajtmh.1999.60.840Suche in Google Scholar

Cipollo, J.F., Costello, C.E., and Hirschberg, C.B. (2002). The fine structure of Caenorhabditis elegans N-glycans. J. Biol. Chem.277, 49143–49157.10.1074/jbc.M208020200Suche in Google Scholar

Cipollo, J.F., Awad, A., Costello, C.E., Robbins, P.W., and Hirschberg, C.B. (2004). Biosynthesis in vitro of Caenorhabditis elegans phosphorylcholine oligosaccharides. Proc. Natl. Acad. Sci. USA101, 3404–3408.10.1073/pnas.0400384101Suche in Google Scholar

Cipollo, J.F., Awad, A.M., Costello, C.E., and Hirschberg, C.B. (2005). N-Glycans of Caenorhabditis elegans are specific to developmental stages. J. Biol. Chem.280, 26063–26072.10.1074/jbc.M503828200Suche in Google Scholar

Eckerskorn, C., 1999. Electroblotting. In: Microcharacterization of proteins, R. Kellner, F. Lottspeich, and H.E. Meyer, eds. (Weinheim, Germany: Wiley-VCH), pp. 35–48.Suche in Google Scholar

Finkelman, F.D. and Urban, J.F. (1992). Cytokines: making the right choice. Parasitol. Today8, 311–314.10.1016/0169-4758(92)90105-BSuche in Google Scholar

Fraser, A.G., Kamath, R.S., Zipperlen, P., Martinez-Campos, M., Sohrmann, M., and Ahringer, J. (2000). Functional genomic analysis of C. elegans chromosome I by systematic RNA interference. Nature408, 325–330.Suche in Google Scholar

Friedl, C.H., Lochnit, G., Zähringer, U., Bahr, U., and Geyer, R. (2003). Structural elucidation of zwitterionic carbohydrates derived from glycosphingolipids of the porcine parasitic nematode Ascaris suum. Biochem. J.369, 89–102.10.1042/bj20021074Suche in Google Scholar

Gallego, S.G., Slade, R.W., and Brindley, P.J. (1998). A cDNA encoding a pepsinogen-like, aspartic protease from the human roundworm parasite Strongyloides stercoralis. Acta Trop.71, 17–26.10.1016/S0001-706X(98)00050-3Suche in Google Scholar

Geier, G., Banaj, H.J., Heid, H., Bini, L., Pallini, V., and Zwilling, R. (1999). Aspartyl proteases in Caenorhabditis elegans. Isolation, identification and characterization by a combined use of affinity chromatography, two-dimensional gel electrophoresis, microsequencing and databank analysis. Eur. J. Biochem.264, 872–879.Suche in Google Scholar

Geldhof, P., Claerebout, E., Knox, D.P., Jagneessens, J., and Vercruysse, J. (2000). Proteinases released in vitro by the parasitic stages of the bovine abomasal nematode Ostertagia ostertagi. Parasitology121, 639–647.10.1017/S0031182000006806Suche in Google Scholar

Gerdt, S., Lochnit, G., Dennis, R.D., and Geyer, R. (1997). Isolation and structural analysis of three neutral glycosphingolipids from a mixed population of Caenorhabditis elegans (Nematoda: Rhabditida). Glycobiology7, 265–275.10.1093/glycob/7.2.265Suche in Google Scholar

Gerdt, S., Dennis, R.D., Borgonie, G., Schnabel, R., and Geyer, R. (1999). Isolation, characterization and immunolocalization of phosphocholine-substituted glycolipids in developmental stages of Caenorhabditis elegans. Eur. J. Biochem.266, 952–963.10.1046/j.1432-1327.1999.00937.xSuche in Google Scholar

Gimenez-Pardo, C., Vazquez-Lopez, C., Armas-Serra, C.D., and Rodriguez-Caaeiro, F. (1999). Proteolytic activity in Caenorhabditis elegans: soluble and insoluble fractions. J. Helminthol.73, 123–127.10.1017/S0022149X99000189Suche in Google Scholar

Goodridge, H.S., Harnett, W., Liew, F.Y., and Harnett, M.M. (2003). Differential regulation of interleukin-12 p40 and p35 induction via Erk mitogen-activated protein kinase-dependent and -independent mechanisms and the implications for bioactive IL-12 and IL-23 responses. Immunology109, 415–425.10.1046/j.1365-2567.2003.01689.xSuche in Google Scholar

Görg, A. and Westermeier, R., 1999. High resolution gel-electrophoretic techniques: qualitative, quantitative and micropreparative applications. In: Microcharacterization of Proteins, R. Kellner, F. Lottspeich, and H.E. Meyer, eds. (Weinheim, Germany: Wiley-VCH), pp. 13–24.Suche in Google Scholar

Grabitzki, J., Sauerland, V., Geyer, R., and Lochnit, G. (2005). Identification of phosphorylcholine substituted peptides by their characteristic mass spectrometric fragmentation. Eur. J. Mass Spectrom.11, 335–344.10.1255/ejms.728Suche in Google Scholar

Griffitts, J.S., Haslam, S.M., Yang, T., Garczynski, S.F., Mulloy, B., Morris, H., Cremer, P.S., Dell, A., Adang, M.J., and Aroian, R.V. (2005). Glycolipids as receptors for Bacillus thuringiensis crystal toxin. Science307, 922–925.10.1126/science.1104444Suche in Google Scholar

Harnett, W. and Harnett, M.M. (2000). Phosphorylcholine: an immunomodulator present on glycoproteins secreted by filarial nematodes. Mod. Asp. Immunobiol.1, 40–42.Suche in Google Scholar

Harnett, W. and Harnett, M.M. (2001). Modulation of the host immune system by phosphorylcholine-containing glycoproteins secreted by parasitic filarial nematodes. Biochim. Biophys. Acta1539, 7–15.10.1016/S0167-4889(01)00101-XSuche in Google Scholar

Harnett, W., Houston, K.M., Amess, R., and Worms, M.J. (1993). Acanthocheilonema viteae: phosphorylcholine is attached to the major excretory-secretory product via an N-linked glycan. Exp. Parasitol.77, 498–502.10.1006/expr.1993.1113Suche in Google Scholar PubMed

Harnett, W., Frame, M.J., Nor, Z.M., MacDonald, M., and Houston, K.M. (1994). Some preliminary data on the nature/structure of the PC-glycan of the major excretory-secretory product of Acanthocheilonema viteae (ES-62). Parasite1, 179–181.10.1051/parasite/1994012179Suche in Google Scholar PubMed

Harnett, W., McInnes, I.B., and Harnett, M.M. (2004). ES-62, a filarial nematode-derived immunomodulator with anti-inflammatory potential. Immunol. Lett.94, 27–33.10.1016/j.imlet.2004.04.008Suche in Google Scholar PubMed

Harrop, S.A., Prociv, P., and Brindley, P.J. (1996). Acasp, a gene encoding a cathepsin D-like aspartic protease from the hookworm Ancylostoma caninum. Biochem. Biophys. Res. Commun.227, 294–302.10.1006/bbrc.1996.1503Suche in Google Scholar

Haslam, S.M. and Dell, A. (2003). Hallmarks of Caenorhabditis elegans N-glycosylation: complexity and controversy. Biochimie 85, 25–32.10.1016/S0300-9084(03)00041-5Suche in Google Scholar

Haslam, S.M., Khoo, K.H., Houston, K.M., Harnett, W., Morris, H.R., and Dell, A. (1997). Characterisation of the phosphorylcholine-containing N-linked oligosaccharides in the excretory-secretory 62 kDa glycoprotein of Acanthocheilonema viteae. Mol. Biochem. Parasitol.85, 53–66.10.1016/S0166-6851(96)02807-1Suche in Google Scholar

Haslam, S.M., Houston, K.M., Harnett, W., Reason, A.J., Morris, H.R., and Dell, A. (1999). Structural studies of N-glycans of filarial parasites. Conservation of phosphorylcholine-substituted glycans among species and discovery of novel chito-oligomers. J. Biol. Chem.274, 20953–20960.10.1074/jbc.274.30.20953Suche in Google Scholar

Haslam, S.M., Gems, D., Morris, H.R., and Dell, A. (2002). The glycomes of Caenorhabditis elegans and other model organisms. Biochem. Soc. Symp.69, 117–134.10.1042/bst030a004aSuche in Google Scholar

Hobert, O. (2002). PCR fusion-based approach to create reporter gene constructs for expression analysis in transgenic C. elegans. Biotechniques32, 728–730.10.2144/02324bm01Suche in Google Scholar

Jolodar, A. and Miller, D.J. (1998). Identification of a novel family of non-lysosomal aspartic proteases in nematodes. Biochim. Biophys. Acta1382, 13–16.10.1016/S0167-4838(97)00141-6Suche in Google Scholar

Kamath, R.S., and Ahringer, J. (2003). Genome-wide RNAi screening in Caenorhabditis elegans. Methods30, 313–321.10.1016/S1046-2023(03)00050-1Suche in Google Scholar

Knox, D.P. and Kennedy, M.W. (1988). Proteinases released by the parasitic larval stages of Ascaris suum, and their inhibition by antibody. Mol. Biochem. Parasitol.28, 207–216.10.1016/0166-6851(88)90005-9Suche in Google Scholar

Knox, D.P. and Jones, D.G. (1990). Studies on the presence and release of proteolytic enzymes (proteinases) in gastro-intestinal nematodes of ruminants. Int. J. Parasitol.20, 243–249.10.1016/0020-7519(90)90106-WSuche in Google Scholar

Kurokawa, T., Wuhrer, M., Lochnit, G., Geyer, H., Markl, J., and Geyer, R. (2002). Hemocyanin from the keyhole limpet Megathura crenulata (KLH) carries a novel type of N-glycans with Gal(β1-6)Man-motifs. Eur. J. Biochem.269, 5459–5473.10.1046/j.1432-1033.2002.03244.xSuche in Google Scholar

Laemmli, U.K. (1970). Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature227, 680–685.10.1038/227680a0Suche in Google Scholar

Lochnit, G., Dennis, R.D., Ulmer, A.J., and Geyer, R. (1998). Structural elucidation and monokine-inducing activity of two biologically active zwitterionic glycosphingolipids derived from the porcine parasitic nematode Ascaris suum. J. Biol. Chem.278, 466–474.10.1074/jbc.273.1.466Suche in Google Scholar

Lochnit, G., Dennis, R.D., and Geyer, R. (2000). Phosphorylcholine substituents in nematodes: structures, occurrence and biological implications. Biol. Chem.381, 839–847.10.1515/BC.2000.106Suche in Google Scholar

Lochnit, G., Bongaarts, R., and Geyer, R. (2005). Searching new targets for anthelmintic strategies: interference with glycosphingolipid biosynthesis and phosphorylcholine metabolism affects development of Caenorhabditis elegans. Int. J. Parasitol.35, 911–923.10.1016/j.ijpara.2005.02.015Suche in Google Scholar

Marshall, F.A., Grierson, A.M., Garside, P., Harnett, W., and Harnett, M.M. (2005). ES-62, an immunomodulator secreted by filarial nematodes, suppresses clonal expansion and modifies effector function of heterologous antigen-specific T cells in vivo. J. Immunol.175, 5817–5826.10.4049/jimmunol.175.9.5817Suche in Google Scholar

Sarkis, G.J., Kurpiewski, M.R., Ashcom, J.D., Jen-Jacobson, L., and Jacobson, L.A. (1988). Proteases of the nematode Caenorhabditis elegans. Arch. Biochem. Biophys.261, 80–90.10.1016/0003-9861(88)90106-3Suche in Google Scholar

Sulston, J. and Hodgkin, J., 1988. Methods. In: The Nematode Caenorhabditis elegans, W.B. Wood, ed. (Plainview, NY, USA: Cold Spring Harbor Laboratory Press), pp. 587–606.Suche in Google Scholar

Syntichaki, P., Xu, K., Driscoll, M., and Tavernarakis, N. (2002). Specific aspartyl and calpain proteases are required for neurodegeneration in C. elegans. Nature419, 939–944.10.1038/nature01108Suche in Google Scholar

Tcherepanova, I., Bhattacharyya, L., Rubin, C.S., and Freedman, J.H. (2000). Aspartic proteases from the nematode Caenorhabditis elegans. Structural organization and developmental and cell-specific expression of asp-1. J. Biol. Chem.275, 26359–26369.10.1074/jbc.M000956200Suche in Google Scholar

Tort, J., Brindley, P.J., Knox, D., Wolfe, K.H., and Dalton, J.P. (1999). Proteinases and associated genes of parasitic helminths. Adv. Parasitol.43, 161–266.10.1016/S0065-308X(08)60243-2Suche in Google Scholar

Vanfleteren, J.R. (1978). Axenic culture of free-living, plant-parasitic, and insect-parasitic nematodes. Annu. Rev. Phytopathol.16, 131–157.10.1146/annurev.py.16.090178.001023Suche in Google Scholar

Wuhrer, M., Rickhoff, S., Dennis, R.D., Lochnit, G., Soboslay, P.T., Baumeister, S., and Geyer, R. (2000). Phosphocholine-containing, zwitterionic glycosphingolipids of adult Onchocerca volvulus as highly conserved, antigenic structures of parasitic nematodes. Biochem. J.348, 417–423.10.1042/bj3480417Suche in Google Scholar

Yan, R., Bienkowski, M.J., Shuck, M.E., Miao, H., Tory, M.C., Pauley, A.M., Brashier, J.R., Stratman, N.C., Mathews, W.R., Buhl, A.E., et al. (1999). Membrane-anchored aspartyl protease with Alzheimer's disease β-secretase activity. Nature 402, 533–537.10.1038/990107Suche in Google Scholar PubMed

Published Online: 2006-11-02
Published in Print: 2006-10-01

©2006 by Walter de Gruyter Berlin New York

Artikel in diesem Heft

  1. Highlight: Redox signaling – mechanisms and biological impact
  2. Paper of the Year 2005: Award to Vanessa Ferreira Merino
  3. Two-site substrate recognition model for the Keap1-Nrf2 system: a hinge and latch mechanism
  4. Hypoxia and lipid signaling
  5. Glutathione peroxidases and redox-regulated transcription factors
  6. Redox regulation of the hypoxia-inducible factor
  7. The l-arginine nitric oxide pathway: avenue for a multiple-level approach to assess vascular function
  8. Protein oxidation and proteolysis
  9. Mitochondrial signaling, TOR, and life span
  10. Pathogenetic interplay between osmotic and oxidative stress: the hepatic encephalopathy paradigm
  11. Regulation of redox-sensitive exofacial protein thiols in CHO cells
  12. N-Ethylmaleimide-sensitive factor: a redox sensor in exocytosis
  13. Aspects of the biological redox chemistry of cysteine: from simple redox responses to sophisticated signalling pathways
  14. Singlet oxygen inactivates protein tyrosine phosphatase-1B by oxidation of the active site cysteine
  15. Regulatory effects of the mitochondrial energetic status on mitochondrial p66Shc
  16. Air pollution-associated fly ash particles induce fibrotic mechanisms in primary fibroblasts
  17. Incinerator fly ash provokes alteration of redox equilibrium and liberation of arachidonic acid in vitro
  18. Unique neuronal functions of cathepsin L and cathepsin B in secretory vesicles: biosynthesis of peptides in neurotransmission and neurodegenerative disease
  19. Two novel mitochondrial and chloroplastic targeting-peptide-degrading peptidasomes in A. thaliana, AtPreP1 and AtPreP2
  20. Switch from actin α1 to α2 expression and upregulation of biomarkers for pressure overload and cardiac hypertrophy in taurine-deficient mouse heart
  21. Human RBM28 protein is a specific nucleolar component of the spliceosomal snRNPs
  22. The β12-β13 loop is a key regulatory element for the activity and properties of the catalytic domain of protein phosphatase 1 and 2B
  23. DNA-binding properties of the recombinant high-mobility-group-like AT-hook-containing region from human BRG1 protein
  24. Papaya glutamine cyclotransferase shows a singular five-fold β-propeller architecture that suggests a novel reaction mechanism
  25. First identification of a phosphorylcholine-substituted protein from Caenorhabditis elegans: isolation and characterization of the aspartyl protease ASP-6
  26. The human cathelicidin peptide LL-37 and truncated variants induce segregation of lipids and proteins in the plasma membrane of Candida albicans
  27. Specificity of human cathepsin S determined by processing of peptide substrates and MHC class II-associated invariant chain
  28. Mast cell-dependent activation of pro matrix metalloprotease 2: a role for serglycin proteoglycan-dependent mast cell proteases
Heruntergeladen am 17.9.2025 von https://www.degruyterbrill.com/document/doi/10.1515/BC.2006.186/html
Button zum nach oben scrollen