Characterization of an expressible nonclassical class I HLA gene.
Screening of a human cosmid library representing genomic DNA from an individual homozygous for the HLA-DR2 B7 A2 haplotype yielded 109 class I HLA-specific clones. One cosmid clone, Ice 6.23, had a full-length nonclassical class I gene within a 5.4-kb HindIII fragment. The Ice 6.23-5.4H gene was cloned into the unique NotI site of an expression vector pSV2.Not, a derivative of pSV2neo, which was constructed to contain a second SV40 early region promoter adjacent to an introduced NotI site. The resulting construct was transfected into the P815-B2M cell line, a derivative of the mouse mastocytoma P815 (HTR) line which expressed human beta2-microglobulin following stable transfection with a cloned human beta2-microglobulin gene. Following transfection the Ice 6.23-5.4 H gene was found to be expressed at both the mRNA and cell surface product levels. DNA sequencing of this gene suggests that it is allelic to the HLA-6.0 gene clone (HLA-G) of Geraghty et al. (Proceedings of the National Academy of Sciences USA, 84:9145, 1987); thereby revealing a HindIII restriction fragment length polymorphism at the HLA-G locus. An extraordinarily high degree of sequence similarity (99.92%) between these two genes, which derive from unrelated HLA haplotypes, suggests strong conservative selection pressure at the HLA-G locus. A flanking single copy sequence probe 4 kb distant from the Ice 6.23-5.4H gene was used to generate long-range restriction mapping at the HLA-G locus.
Pook
M A
MA
University Department of Medical Genetics, St. Mary's Hospital, Manchester, United Kingdom.
Woodcock
V
V
Tassabehji
M
M
Campbell
R D
RD
Summers
C W
CW
Taylor
M
M
Strachan
T
T
D000595:Amino Acid Sequence; D001483:Base Sequence; D004247:DNA / Q000032:analysis*; D015870:Gene Expression; D015723:Gene Library; D005805:Genes, MHC Class I / Q000235:genetics*; D006680:HLA Antigens / Q000235:genetics*; D059951:HLA-G Antigens; D015395:Histocompatibility Antigens Class I / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes; D015183:Restriction Mapping
D006680:HLA Antigens; D059951:HLA-G Antigens; D015395:Histocompatibility Antigens Class I; D015345:Oligonucleotide Probes; D004247:DNA