Identification of potential CTL epitopes of bovine RSV using allele-specific peptide motifs from bovine MHC class I molecules.
Respiratory syncytial virus (RSV) is a major cause of lower respiratory tract infection in young infants and housed calves. Depletion of CD8+ lymphocytes from calves inhibited their ability to clear the virus from the nasopharynx and lungs. To study these cells further, a cytotoxic T lymphocyte (CTL) assay was established. CTL could be demonstrated in the peripheral blood of gnotobiotic calves 7-10 days post infection (p.i.) with RSV and in lungs 10 days p.i. This response was both MHC-restricted and virus-specific. Following separation of the lung lymphocytes by magnetic activated cell sorting, it was shown that the cytolytic activity was mediated by cells of the CD8+ phenotype. To identify epitopes recognised by bovine CTL, the consensus motifs from MHC class I alleles found in the herd at Compton were identified. cDNA libraries were constructed and screened for full length class I sequences. The isolated cDNA clones were then transfected into mouse P815 cells and the expressed product immunoprecipitated and matched with a serological specificity. The bovine MHC class I molecules were isolated from lysed transfected cells by affinity chromatography, using a monoclonal antibody specific for bovine MHC class I, and bound peptides were separated by reverse-phase HPLC. Analysis of the protein sequences of bovine RSV for the defined motifs has identified potential CTL epitopes.
Gaddum
R M
RM
Institute for Animal Health, Compton, Newbury, UK.
Ellis
S A
SA
Willis
A C
AC
Cook
R S
RS
Staines
K A
KA
Thomas
L H
LH
Taylor
G
G
D000483:Alleles; D000818:Animals; D002417:Cattle; D018604:Epitope Mapping; D000939:Epitopes / Q000276:immunology*; D015395:Histocompatibility Antigens Class I / Q000276:immunology*; D010446:Peptide Fragments / Q000276:immunology*; D018112:Respiratory Syncytial Virus, Bovine / Q000276:immunology*; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology* / Q000821:virology*; D014407:Tumor Cells, Cultured
D000939:Epitopes; D015395:Histocompatibility Antigens Class I; D010446:Peptide Fragments