Full-length recombinant choline transporter-like protein 2 containing arginine 154 reconstitutes the epitope recognized by HNA-3a antibodies

摘要

BACKGROUND: Recent reports have shown that the HNA-3a leukocyte antigen, a target for antibodies that cause severe transfusion-related acute lung injury, correlates with an arginine 154 (rather than glutamine) polymorphism in choline transporter-like protein 2 (CTL2) but did not show directly that R154 determines HNA-3a. CTL2 peptides containing R154 are recognized by only half of HNA-3a antibodies studied to date. Constructs that react with all HNA-3a antibodies are needed to fully define the HNA-3a epitope. STUDY DESIGN AND METHODS: HEK293 cells were transfected with cDNA encoding full-length CTL2 linked to green fluorescent protein (GFP). Transfectants were selected for GFP expression and tested with antibodies specific for HNA-3a and -3b. RESULTS: Each of 20 HNA-3a antibodies reacted preferentially with HEK293 cells expressing the R154 CTL2 construct. An HNA-3b antibody reacted only with CTL2 (Q154). CONCLUSIONS: These findings provide direct evidence that R154 in the context of full-length CTL2 is both necessary and sufficient to create the HNA-3a epitope but suggest that posttranslational modifications of the protein, for example, S-S bonds or addition of glycans, are necessary for recognition of HNA-3a by many antibodies. This could complicate development of an assay for large-scale screening of blood donors to detect anti-HNA-3a.