Identification and characterization of GRA6/GRA7 of Neospora caninum in MDBK cells.
Neospora caninum, an apicomplexan parasite, is recognized as a major bovine abortifacient. Dense granule antigens (GRAs) play important roles in the formation and modification of parasitophorous vacuoles (PVs) in Toxoplasma gondii. However, a few studies investigating GRAs have been reported in N. caninum. The aim of the present study was to characterize the dense GRA6/GRA7 of N. caninum in PVs using MDBK cells as a host cell model. Neospora caninum was inoculated into MDBK cells, and changes were observed using a transmission electron microscope (TEM). Neospora caninum GRA6/GRA7 were identified and characterized using bioinformatics, cell fractionation, and immunofluorescence. The TEM results revealed that integrated PVs were present in MDBK cells after N. caninum infection. Bioinformatics analysis showed that NcGRA6/NcGRA7 shared 28.76% and 29.66% homology with T. gondii GRA6/GRA7 (TgGRA6/TgGRA7) but had similar signal peptides, transmembrane domains, and motifs. Cell fractionation and subcellular localization analyses both showed that NcGRA6 was distributed in the lumen and intravacuolar network in soluble and transmembrane forms. The transmembrane form of NcGRA7 was observed in the PV membrane. These data lay a foundation for further study on bovine neosporosis and NcGRA6/NcGRA7 function during PV formation.
College of Veterinary Medicine, Jilin University, Changchun 130062, China.
Jingquan
J
Dong
College of Veterinary Medicine, Jilin University, Changchun 130062, China.
Jianhua
J
Li
College of Veterinary Medicine, Jilin University, Changchun 130062, China.
Jinpeng
J
Wang
College of Veterinary Medicine, Jilin University, Changchun 130062, China.
Fei
F
Li
College of Veterinary Medicine, Jilin University, Changchun 130062, China.
Ju
J
Yang
College of Veterinary Medicine, Jilin University, Changchun 130062, China.
Pengtao
P
Gong
College of Veterinary Medicine, Jilin University, Changchun 130062, China.
He
H
Li
College of Veterinary Medicine, Jilin University, Changchun 130062, China.
Xichen
X
Zhang
D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D002417:Cattle; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D004847:Epithelial Cells / Q000469:parasitology* / Q000648:ultrastructure; D006790:Host-Parasite Interactions; D002479:Inclusion Bodies / Q000469:parasitology* / Q000648:ultrastructure; D007668:Kidney / Q000166:cytology; D018613:Microscopy, Confocal; D046529:Microscopy, Electron, Transmission; D018688:Neospora / Q000378:metabolism / Q000502:physiology* / Q000648:ultrastructure; D015800:Protozoan Proteins / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid
D015800:Protozoan Proteins
MDBK cells; Neospora caninum; dense granule antigen; subcellular localization