Single amino acid substitution in human platelet glycoprotein Ibbeta is responsible for the formation of the platelet-specific alloantigen Iy(a).
We recently described a new low-frequency platelet alloantigen on the human platelet glycoprotein (GP) Ib-IX complex, termed Iy(a), which was implicated in a severe case of neonatal alloimmune thrombocytopenia. Immunoprecipitation studies with trypsin-treated platelets indicated that the Iy(a) alloantigenic determinants are formed by the membrane-associated remnant moiety of GP Ibalpha (GP Ibalpha(r)) together with GP Ibbeta and GP IX. To elucidate the molecular basis underlying the Iy(a) alloantigen, we amplified GPIbalpha(r), GPIbbeta, and GPIX genes by polymerase chain reaction (PCR). Nucleotide-sequence analysis of these 3 genes showed a G to A transition at position 141 on GPIbbeta gene in a subject positive for Iy(a). This transition resulted in a Gly(15)Glu dimorphism on the N-terminal domain of GPIbbeta. This finding was confirmed by genotyping analysis of 6 Iy(a)-positive subjects by restriction fragment length polymorphism (RFLP) studies using NarI endonuclease. In 300 randomly selected healthy blood donors, one Iy(a)-positive individual was found. Phenotypes determined by monoclonal antibody-specific immobilization of platelet antigens assay and genotypes determined by RFLP were identical in this population. Analysis of Iy(a)-positive platelets showed that the point mutation affected neither the degree of surface expression nor the function of the GP Ibalpha-GP Ibbeta-IX complex on the platelet surface. Transient expression of the GP Ib-IX complex in CHO cells using wild-type GP Ibbeta (Gly(15)) or mutant GP Ibbeta (Glu(15)) allowed us to demonstrate that this single amino acid substitution is sufficient to induce Iy(a) epitope(s). (Blood. 2000;95:1849-1855)
Sachs
U J
UJ
Institute for Clinical Immunology and Transfusion Medicine, Justus Liebig-University, Giessen, Germany.
Kiefel
V
V
Böhringer
M
M
Afshar-Kharghan
V
V
Kroll
H
H
Santoso
S
S
D019943:Amino Acid Substitution; D000818:Animals; D016824:Antigens, Human Platelet / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D001327:Autoimmune Diseases / Q000151:congenital / Q000235:genetics* / Q000276:immunology; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D004252:DNA Mutational Analysis; D000939:Epitopes / Q000235:genetics / Q000276:immunology; D006801:Humans; D007231:Infant, Newborn; D046911:Macromolecular Substances; D010375:Pedigree; D010641:Phenotype; D019038:Platelet Glycoprotein GPIb-IX Complex / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D017354:Point Mutation; D016133:Polymerase Chain Reaction; D012150:Polymorphism, Restriction Fragment Length; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D013921:Thrombocytopenia / Q000151:congenital / Q000235:genetics* / Q000276:immunology
C110429:12bw alloantigen, human; D016824:Antigens, Human Platelet; D000939:Epitopes; D046911:Macromolecular Substances; D019038:Platelet Glycoprotein GPIb-IX Complex; D011993:Recombinant Fusion Proteins