cDNA cloning and eukaryotic expression of feline CD9.
A monoclonal antibody (vpg15) has been described which can block infection with feline immunodeficiency virus (FIV) and which recognizes the feline homologue of CD9. In order to study the role of feline CD9 in infection with FIV we have molecularly cloned a cDNA encoding feline CD9 by R.A.C.E (rapid amplification of cDNA ends). The amino acid sequence of feline CD9 displays 95.1, 93.8 and 90.7% homology to human, murine and bovine CD9, respectively. Although feline CD9 appears most homologous to human CD9, it has two important features in common with bovine and murine CD9: the presence of a histidine residue at position 192 which is absent from the corresponding position (194) in human CD9; and the absence of two asparagine residues which are found at positions 51 and 52 of human CD9. Feline CD9 is unique in that it lacks a potential N-linked glycosylation site in the first extracellular loop, a feature common to CD9 of other species. Despite the high degree of sequence homology, significant cross-species variation occurred in the two predicted extracellular loops, notably between amino acids 169 to 180 of the second loop. When feline CD9 was expressed on human and murine cells, it was recognized by both the conformation-dependent feline CD9-specific antibody, vpg15, and the cross-species reactive anti-human CD9 antibody, FMC56, confirming that the feline CD9 clone encoded a protein which was synthesized, transported to the cell surface and expressed in a similar conformation to native feline CD9. However, although the vpg15 antibody did not recognize human CD9 when expressed on human epithelial cells, it reacted with human CD9 when expressed on murine fibroblast cells. It is possible therefore, that the conformational epitope recognized by the vpg15 epitope is sensitive to either species- or tissue-specific post-translational modification.
Willett
B J
BJ
Department of Veterinary Pathology, University of Glasgow Veterinary School, U.K.
Neil
J C
JC
D000595:Amino Acid Sequence; D000818:Animals; D015703:Antigens, CD / Q000235:genetics*; D001483:Base Sequence; D002415:Cats; D003001:Cloning, Molecular; D017931:DNA Primers / Q000737:chemistry; D018076:DNA, Complementary / Q000235:genetics; D005434:Flow Cytometry; D006801:Humans; D008562:Membrane Glycoproteins / Q000235:genetics; D051379:Mice; D008969:Molecular Sequence Data; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D060245:Tetraspanin 29
D015703:Antigens, CD; C554869:CD9 protein, human; C554870:Cd9 protein, mouse; D017931:DNA Primers; D018076:DNA, Complementary; D008562:Membrane Glycoproteins; D060245:Tetraspanin 29