Skip to main content
NCBI home page
Infection and Immunity logoLink to Infection and Immunity
. 1996 Oct;64(10):4279–4287. doi: 10.1128/iai.64.10.4279-4287.1996

Subcellular localization and functional characterization of Nc-p43, a major Neospora caninum tachyzoite surface protein.

A Hemphill 1
PMCID: PMC174368  PMID: 8926100

Abstract

Neospora caninum is a recently identified coccidian parasite which shares many features with, but is clearly distinct from, Toxoplasma gondii. N. caninum tachyzoites infect a wide range of mammalian cells both in vivo and in vitro. The mechanisms by which infection is achieved are largely unknown. Recent evidence has suggested that a receptor-ligand system in which one or several host cell receptors bind to one or several parasite ligands is involved. Parasite cell surface-associated molecules such as the recently identified Nc-p43 antigen are prime suspects for being implicated in this physical interaction. In this study it is shown that invasion of Vero cell monolayers by N. caninum tachyzoites in vitro is impaired on incubation of parasites with subagglutinating amounts of affinity-purified antibodies directed against Nc-p43. Postembedding immunogold labeling with anti-Nc-p43 antibodies demonstrated that Nc-p43 is localized not only on the parasite cell surface but also within dense granules and rhoptries. The fate of Nc-p43 during intracellular proliferation of N. caninum tachyzoites and subsequent maturation of the parasitophorous vacuole was also studied.

Full Text

The Full Text of this article is available as a PDF (4.5 MB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Achbarou A., Mercereau-Puijalon O., Sadak A., Fortier B., Leriche M. A., Camus D., Dubremetz J. F. Differential targeting of dense granule proteins in the parasitophorous vacuole of Toxoplasma gondii. Parasitology. 1991 Dec;103(Pt 3):321–329. doi: 10.1017/s0031182000059837. [DOI] [PubMed] [Google Scholar]
  2. Adams J. H., Hudson D. E., Torii M., Ward G. E., Wellems T. E., Aikawa M., Miller L. H. The Duffy receptor family of Plasmodium knowlesi is located within the micronemes of invasive malaria merozoites. Cell. 1990 Oct 5;63(1):141–153. doi: 10.1016/0092-8674(90)90295-p. [DOI] [PubMed] [Google Scholar]
  3. Aikawa M., Torii M., Sjölander A., Berzins K., Perlmann P., Miller L. H. Pf155/RESA antigen is localized in dense granules of Plasmodium falciparum merozoites. Exp Parasitol. 1990 Oct;71(3):326–329. doi: 10.1016/0014-4894(90)90037-d. [DOI] [PubMed] [Google Scholar]
  4. Barr B. C., Rowe J. D., Sverlow K. W., BonDurant R. H., Ardans A. A., Oliver M. N., Conrad P. A. Experimental reproduction of bovine fetal Neospora infection and death with a bovine Neospora isolate. J Vet Diagn Invest. 1994 Apr;6(2):207–215. doi: 10.1177/104063879400600212. [DOI] [PubMed] [Google Scholar]
  5. Bouvier J., Etges R. J., Bordier C. Identification and purification of membrane and soluble forms of the major surface protein of Leishmania promastigotes. J Biol Chem. 1985 Dec 15;260(29):15504–15509. [PubMed] [Google Scholar]
  6. Charif H., Darcy F., Torpier G., Cesbron-Delauw M. F., Capron A. Toxoplasma gondii: characterization and localization of antigens secreted from tachyzoites. Exp Parasitol. 1990 Jul;71(1):114–124. doi: 10.1016/0014-4894(90)90014-4. [DOI] [PubMed] [Google Scholar]
  7. Cheng Q., Saul A. Sequence analysis of the apical membrane antigen I (AMA-1) of Plasmodium vivax. Mol Biochem Parasitol. 1994 May;65(1):183–187. doi: 10.1016/0166-6851(94)90127-9. [DOI] [PubMed] [Google Scholar]
  8. Couvreur G., Sadak A., Fortier B., Dubremetz J. F. Surface antigens of Toxoplasma gondii. Parasitology. 1988 Aug;97(Pt 1):1–10. doi: 10.1017/s0031182000066695. [DOI] [PubMed] [Google Scholar]
  9. Donelson J. E., Fulton A. B. Parasitology. Skirmishes on the border. Nature. 1992 Apr 9;356(6369):480–481. doi: 10.1038/356480a0. [DOI] [PubMed] [Google Scholar]
  10. Dubey J. P., Carpenter J. L., Speer C. A., Topper M. J., Uggla A. Newly recognized fatal protozoan disease of dogs. J Am Vet Med Assoc. 1988 May 1;192(9):1269–1285. [PubMed] [Google Scholar]
  11. Dubey J. P., Lindsay D. S. Neosporosis. Parasitol Today. 1993 Dec;9(12):452–458. doi: 10.1016/0169-4758(93)90099-2. [DOI] [PubMed] [Google Scholar]
  12. Entzeroth R., Dubremetz J. F., Hodick D., Ferreira E. Immunoelectron microscopic demonstration of the exocytosis of dense granule contents into the secondary parasitophorous vacuole of Sarcocystis muris (Protozoa, Apicomplexa). Eur J Cell Biol. 1986 Aug;41(2):182–188. [PubMed] [Google Scholar]
  13. Furtado G. C., Cao Y., Joiner K. A. Laminin on Toxoplasma gondii mediates parasite binding to the beta 1 integrin receptor alpha 6 beta 1 on human foreskin fibroblasts and Chinese hamster ovary cells. Infect Immun. 1992 Nov;60(11):4925–4931. doi: 10.1128/iai.60.11.4925-4931.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Furtado G. C., Slowik M., Kleinman H. K., Joiner K. A. Laminin enhances binding of Toxoplasma gondii tachyzoites to J774 murine macrophage cells. Infect Immun. 1992 Jun;60(6):2337–2342. doi: 10.1128/iai.60.6.2337-2342.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Galinski M. R., Barnwell J. W. Plasmodium vivax: Merozoites, invasion of reticulocytes and considerations for malaria vaccine development. Parasitol Today. 1996 Jan;12(1):20–29. doi: 10.1016/0169-4758(96)80641-7. [DOI] [PubMed] [Google Scholar]
  16. Grimwood J., Smith J. E. Toxoplasma gondii: redistribution of tachyzoite surface protein during host cell invasion and intracellular development. Parasitol Res. 1995;81(8):657–661. doi: 10.1007/BF00931842. [DOI] [PubMed] [Google Scholar]
  17. Hemphill A., Affolter M., Seebeck T. A novel microtubule-binding motif identified in a high molecular weight microtubule-associated protein from Trypanosoma brucei. J Cell Biol. 1992 Apr;117(1):95–103. doi: 10.1083/jcb.117.1.95. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Hemphill A., Gottstein B. Identification of a major surface protein on Neospora caninum tachyzoites. Parasitol Res. 1996;82(6):497–504. doi: 10.1007/s004360050152. [DOI] [PubMed] [Google Scholar]
  19. Hemphill A., Gottstein B., Kaufmann H. Adhesion and invasion of bovine endothelial cells by Neospora caninum. Parasitology. 1996 Feb;112(Pt 2):183–197. doi: 10.1017/s0031182000084754. [DOI] [PubMed] [Google Scholar]
  20. Holder A. A., Freeman R. R. Biosynthesis and processing of a Plasmodium falciparum schizont antigen recognized by immune serum and a monoclonal antibody. J Exp Med. 1982 Nov 1;156(5):1528–1538. doi: 10.1084/jem.156.5.1528. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Holder A. A., Freeman R. R. Immunization against blood-stage rodent malaria using purified parasite antigens. Nature. 1981 Nov 26;294(5839):361–364. doi: 10.1038/294361a0. [DOI] [PubMed] [Google Scholar]
  22. Kasper L. H., Mineo J. R. Attachment and invasion of host cells by Toxoplasma gondii. Parasitol Today. 1994 May;10(5):184–188. doi: 10.1016/0169-4758(94)90026-4. [DOI] [PubMed] [Google Scholar]
  23. Kimata I., Tanabe K. Secretion by Toxoplasma gondii of an antigen that appears to become associated with the parasitophorous vacuole membrane upon invasion of the host cell. J Cell Sci. 1987 Sep;88(Pt 2):231–239. doi: 10.1242/jcs.88.2.231. [DOI] [PubMed] [Google Scholar]
  24. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  25. Leriche M. A., Dubremetz J. F. Characterization of the protein contents of rhoptries and dense granules of Toxoplasma gondii tachyzoites by subcellular fractionation and monoclonal antibodies. Mol Biochem Parasitol. 1991 Apr;45(2):249–259. doi: 10.1016/0166-6851(91)90092-k. [DOI] [PubMed] [Google Scholar]
  26. Lindsay D. S., Speer C. A., Toivio-Kinnucan M. A., Dubey J. P., Blagburn B. L. Use of infected cultured cells to compare ultrastructural features of Neospora caninum from dogs and Toxoplasma gondii. Am J Vet Res. 1993 Jan;54(1):103–106. [PubMed] [Google Scholar]
  27. Mineo J. R., Kasper L. H. Attachment of Toxoplasma gondii to host cells involves major surface protein, SAG-1 (P30). Exp Parasitol. 1994 Aug;79(1):11–20. doi: 10.1006/expr.1994.1054. [DOI] [PubMed] [Google Scholar]
  28. Nutman T. B., Zimmerman P. A., Kubofcik J., Kostyu D. D. A universally applicable diagnostic approach to filarial and other infections. Parasitol Today. 1994 Jun;10(6):239–243. doi: 10.1016/0169-4758(94)90127-9. [DOI] [PubMed] [Google Scholar]
  29. Ossorio P. N., Schwartzman J. D., Boothroyd J. C. A Toxoplasma gondii rhoptry protein associated with host cell penetration has unusual charge asymmetry. Mol Biochem Parasitol. 1992 Jan;50(1):1–15. doi: 10.1016/0166-6851(92)90239-g. [DOI] [PubMed] [Google Scholar]
  30. Perkins M. E. Rhoptry organelles of apicomplexan parasites. Parasitol Today. 1992 Jan;8(1):28–32. doi: 10.1016/0169-4758(92)90308-o. [DOI] [PubMed] [Google Scholar]
  31. Saffer L. D., Mercereau-Puijalon O., Dubremetz J. F., Schwartzman J. D. Localization of a Toxoplasma gondii rhoptry protein by immunoelectron microscopy during and after host cell penetration. J Protozool. 1992 Jul-Aug;39(4):526–530. doi: 10.1111/j.1550-7408.1992.tb04844.x. [DOI] [PubMed] [Google Scholar]
  32. Smith J. E. A ubiquitous intracellular parasite: the cellular biology of Toxoplasma gondii. Int J Parasitol. 1995 Nov;25(11):1301–1309. doi: 10.1016/0020-7519(95)00067-c. [DOI] [PubMed] [Google Scholar]
  33. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Werk R. How does Toxoplasma gondii enter host cells? Rev Infect Dis. 1985 Jul-Aug;7(4):449–457. doi: 10.1093/clinids/7.4.449. [DOI] [PubMed] [Google Scholar]
  35. Wessel D., Flügge U. I. A method for the quantitative recovery of protein in dilute solution in the presence of detergents and lipids. Anal Biochem. 1984 Apr;138(1):141–143. doi: 10.1016/0003-2697(84)90782-6. [DOI] [PubMed] [Google Scholar]
  36. de Carvalho L., Souto-Padrón T., de Souza W. Localization of lectin-binding sites and sugar-binding proteins in tachyzoites of Toxoplasma gondii. J Parasitol. 1991 Feb;77(1):156–161. [PubMed] [Google Scholar]

Articles from Infection and Immunity are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES