title: Molecular cloning and characterization of NcROP2Fam-1, a member of the ROP2 family of rhoptry proteins in Neospora caninum that is targeted by antibodies neutralizing host cell invasion in vitro.
abstract: Recent publications demonstrated that a fragment of a Neospora caninum ROP2 family member antigen represents a promising vaccine candidate. We here report on the cloning of the cDNA encoding this protein, N. caninum ROP2 family member 1 (NcROP2Fam-1), its molecular characterization and localization. The protein possesses the hallmarks of ROP2 family members and is apparently devoid of catalytic activity. NcROP2Fam-1 is synthesized as a pre-pro-protein that is matured to 2 proteins of 49 and 55 kDa that localize to rhoptry bulbs. Upon invasion the protein is associated with the nascent parasitophorous vacuole membrane (PVM), evacuoles surrounding the host cell nucleus and, in some instances, the surface of intracellular parasites. Staining was also observed within the cyst wall of 'cysts' produced in vitro. Interestingly, NcROP2Fam-1 was also detected on the surface of extracellular parasites entering the host cells and antibodies directed against NcROP2Fam-1-specific peptides partially neutralized invasion in vitro. We conclude that, in spite of the general belief that ROP2 family proteins are intracellular antigens, NcROP2Fam-1 can also be considered as an extracellular antigen, a property that should be taken into account in further experiments employing ROP2 family proteins as vaccines.
authors.affiliation: Institute of Parasitology, Vetsuisse Faculty, University of Bern, Länggass-Strasse 122, CH-3012 Bern, Switzerland.
authors.forename: Ferial
authors.initials: F
authors.lastname: Alaeddine
authors.forename: Andrew
authors.initials: A
authors.lastname: Hemphill
authors.forename: Karim
authors.initials: K
authors.lastname: Debache
authors.forename: Christophe
authors.initials: C
authors.lastname: Guionaud
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000913:Antibodies, Protozoan / Q000276:immunology*; D000953:Antigens, Protozoan / Q000276:immunology*; D001483:Base Sequence; D002522:Chlorocebus aethiops; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D016054:DNA, Protozoan / Q000737:chemistry / Q000235:genetics; D005347:Fibroblasts / Q000469:parasitology; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D008969:Molecular Sequence Data; D018688:Neospora / Q000166:cytology / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D015800:Protozoan Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D016053:RNA, Protozoan / Q000235:genetics; D017422:Sequence Analysis, DNA; D014617:Vacuoles / Q000378:metabolism / Q000469:parasitology; D014709:Vero Cells
chemical_list: D000913:Antibodies, Protozoan; D000953:Antigens, Protozoan; D018076:DNA, Complementary; D016054:DNA, Protozoan; D008565:Membrane Proteins; D015800:Protozoan Proteins; D016053:RNA, Protozoan; C090555:ROP 2 protein, Toxoplasma gondii