Molecular cloning and expression of feline CD3epsilon.
The cDNA of feline CD3epsilon, one of the T-cell receptor components, was cloned from a feline T-lymphoblastoid cell line (MYA-1 cells) and peripheral blood mononuclear cells and thymocytes of cats by polymerase chain reaction. Sequencing analysis revealed that the open reading frame of feline CD3epsilon consists of 606 base pairs encoding a predicted molecular mass of 25 kDa transmembrane protein which lacks N-glycosylation site. Comparison of the predicted amino acid sequence of feline CD3epsilon with those of other mammalians' homologues revealed that a relatively low homology was present in the extracellular domain. However, the cytoplasmic domain contained several characteristic motifs highly conserved across the species. These motifs were known to be important for signal transduction upon T-cell activation or endoplasmic reticulum retention. In addition, the feline CD3epsilon protein was expressed in an insect cell line (Sf9) by a baculovirus expression system. The expression was confirmed by indirect immunofluorescence assay and immunoblotting analysis using an anti-human CD3epsilon polyclonal antibody. These results will provide additional information for understanding the feline immune system.
Nishimura
Y
Y
Department of Veterinary Microbiology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Japan.
Miyazawa
T
T
Ikeda
Y
Y
Izumiya
Y
Y
Nakamura
K
K
Cai
J S
JS
Sato
E
E
Kohmoto
M
M
Mikami
T
T
D000595:Amino Acid Sequence; D000818:Animals; D016367:Baculoviridae / Q000737:chemistry; D001483:Base Sequence; D015153:Blotting, Western / Q000662:veterinary; D017252:CD3 Complex / Q000737:chemistry / Q000235:genetics*; D002415:Cats / Q000276:immunology*; D003001:Cloning, Molecular; D017931:DNA Primers / Q000737:chemistry; D019084:Fluorescent Antibody Technique, Indirect / Q000662:veterinary; D005786:Gene Expression Regulation; D006801:Humans; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016366:Open Reading Frames; D012333:RNA, Messenger / Q000737:chemistry; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000662:veterinary; D016415:Sequence Alignment / Q000662:veterinary; D017422:Sequence Analysis, DNA / Q000662:veterinary; D017386:Sequence Homology, Amino Acid; D014162:Transfection / Q000235:genetics
D017252:CD3 Complex; D017931:DNA Primers; D012333:RNA, Messenger