Sequence and gene transfer analyses of HLA-CwBL18 (HLA-C blank) and HLA-Cw5 genes. Implications for the control of expression and immunogenicity of HLA-C antigens.
Our previous studies suggested that a serologically undetectable HLA-C blank allele (HLA-CwBL18) is either a variant Cw5 allele or a novel HLA-C Ag. To examine these possibilities, the CwBL18 and Cw5 genes from the TCC (HLA-A1, -A2, -B52, -B18, -Cw-, -Cw-) and QBL (HLA-A26, -B18, -Cw5) EBV-transformed B lymphoblastoid cell lines (LCL) were cloned, sequenced, and transferred into HLA-A, -B, -C null LCL mutant .221 cells. The CwBL18 Ag was detected on the cell surface of CwBL18 transferents by flow cytometry with the anti-class I mAb W6/32 but not by complement-mediated cytotoxicity with currently available HLA-C specific antisera. Sequence analysis of the Cw-BL18 gene indicated that the CwBL18 Ag is "C"-like because it contains all C-locus-specific residues and amino acid replacements commonly found in HLA-C alleles. However, the amino acid sequence of the CwBL18 Ag is unusual; CwBL18 lacks unique allele-specific residues when compared with the sequences of other HLA-C alleles. Moreover, apart from the C-locus-specific differences, the sequence of CwBL18 is identical to the HLA class I consensus sequence. This striking homology of CwBL18 to other HLA class I alleles suggests that CwBL18 may be a weak Ag. Taken together, these data demonstrate that CwBL18 is not a variant Cw5 Ag but is a newly described HLA-C Ag. In contrast to CwBL18, the Cw5 Ag is serologically detectable on the cell surface of Cw5 transferents with HLA-specific allo-antisera. Rather unexpectedly, Cw5 was usually expressed at a lower level than CwBL18 on the surface of .221 transferents as evaluated by W6/32 mAb binding analyses. The sequence of Cw5 revealed several unique amino acid replacements. Two of these substitutions, at residue 35 of the alpha 1 domain and residue 275 of the transmembrane domain, may be responsible for the reduced cell surface expression of Cw5. Additional unique replacements at residues 138 and 177 of the alpha 2 domain suggest that these amino acids may be important in the formation of an epitope recognized by a Cw5-specific antibody.
Tibensky
D
D
Banting and Best Department of Medical Research, University of Toronto, Ontario, Canada.
DeMars
R
R
Holowachuk
E W
EW
Delovitch
T L
TL
D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D005786:Gene Expression Regulation; D005805:Genes, MHC Class I; D015236:HLA-C Antigens / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification; D006650:Histocompatibility Testing; D006801:Humans; D008969:Molecular Sequence Data; D010957:Plasmids; D011487:Protein Conformation; D012689:Sequence Homology, Nucleic Acid; D014162:Transfection
D015236:HLA-C Antigens; C037505:HLA-C*05 antigen