RESUMO
The molecular basis of the HNA-3a/b (5b/a) leukocyte antigen system has not yet been defined despite evidence that HNA-3a-specific antibodies are particularly prone to cause severe, often fatal, transfusion-related lung injury. We used genome-wide single nucleotide polymorphism scanning and sequencing of DNA from persons of different HNA-3a/b phenotypes to identify a single single nucleotide polymorphism in exon 7 of the CLT2 gene (SLC44A2) that predicts an amino acid substitution in the first extracellular loop of choline transporter-like protein 2, a member of the choline transporter-like protein family of membrane glycoproteins, and correlates perfectly with HNA-3a/b phenotypes (R154 encodes HNA-3a; Q154 encodes HNA-3b). Mass spectrometric analysis of proteins immunoprecipitated from leukocytes by anti-HNA-3a provided direct evidence that anti-HNA-3a recognizes choline transporter-like protein 2. These findings will enable large-scale genotyping for HNA-3a/b to identify blood donors at risk to have HNA-3a-specific antibodies and should facilitate development of practical methods to detect such antibodies and prevent transfusion-related lung injury.
Assuntos
Isoantígenos/genética , Glicoproteínas de Membrana/genética , Proteínas de Membrana Transportadoras/genética , Neutrófilos/imunologia , Neutrófilos/metabolismo , Sequência de Aminoácidos , Substituição de Aminoácidos/genética , Substituição de Aminoácidos/fisiologia , Arginina/genética , Sequência de Bases , Análise Mutacional de DNA , Estudo de Associação Genômica Ampla , Ácido Glutâmico/genética , Humanos , Isoantígenos/imunologia , Isoantígenos/metabolismo , Espectrometria de Massas , Glicoproteínas de Membrana/imunologia , Glicoproteínas de Membrana/metabolismo , Proteínas de Membrana Transportadoras/imunologia , Proteínas de Membrana Transportadoras/metabolismo , Dados de Sequência Molecular , Proteínas Mutantes/imunologia , Proteínas Mutantes/metabolismo , Polimorfismo de Nucleotídeo Único/fisiologiaRESUMO
BACKGROUND: People with the human neutrophil antigen (HNA)-3b/3b type can make HNA-3a antibodies, which have been reported to cause immune neutropenia disorders and are especially prone to cause severe cases of transfusion-related acute lung injury. However, knowledge of HNA-3 allele frequencies outside Caucasian populations is limited. We developed a high-throughput genotyping assay and determined the HNA-3a/3b genotype frequencies in six different racial and ethnic groups. STUDY DESIGN AND METHODS: Genotyping utilized TaqMan 5' exonuclease chemistry and real-time polymerase chain reaction. A total of 742 DNA samples from six different racial and ethnic groups were genotyped for HNA-3a and HNA-3b. RESULTS: The genotyping assay showed 100% sensitivity and specificity compared to sequencing and phenotyping and had high throughput. A significant percentage of Caucasians (6.5%), Han Chinese (16%), and Asian Indians (6%) typed HNA-3b/3b, but only a small percentage of Hispanics (1%) and no African or Native Americans. CONCLUSIONS: The HNA-3 genotyping assay had high sensitivity, specificity, and sample throughput. HNA-3b/b genotype results determined for 742 individuals representing six different racial and ethnic groups showed that there could be a significant risk of producing anti-HNA-3a in Chinese, as well as in Caucasian and Asian Indian blood donor populations, but a very low risk in Hispanic, African, or Native American populations.