An exonic missense mutation c.28G>A is associated with weak B blood group by affecting RNA splicing of the ABO gene

Abstract

Background: The amino acid substitutions caused by ABO gene mutations are usually predicted to impact glycosyltransferase's function or its biosynthesis. Here we report an ABO exonic missense mutation that affects B-antigen expression by decreasing the mRNA level of the ABO gene rather than the amino acid change.

Study design and methods: Serologic studies including plasma total GTB transfer capacity were performed. The exon sequences of the ABO gene were analyzed by Sanger sequencing. B³¹⁰ cDNA with c.28G>A (p.G10R) mutation was expressed in HeLa cells and total GTB transfer capacity in cell supernatant was measured. Flow cytometry was performed on these HeLa cells after transfection, and agglutination of Hela-B_weak cells was also examined. The mRNA of the ABO gene was analyzed by direct sequencing and real-time reverse transcriptase-polymerase chain reaction. A minigene construct was prepared to evaluate the potential of splicing.

Results: While plasma total GTB transfer capacity was undetectable in this B₃ -like individual, the relative percentage of antigen-expressing cells and mean fluorescence index of the B_weak red blood cells (RBCs) were 19 and 14% of normal B RBCs, respectively. There was no significant difference of total GTB transfer capacity in cell supernatant and B-antigen expression on cell surfaces between HeLa cells transfected with B³¹⁰ cDNA and B cDNA. The mRNA expression level of B³¹⁰ in peripheral whole blood was significantly reduced. The amount of splicing is significantly lower in c.28G>A construct compared to that in wild-type construct after transfection in K562 cells.

Conclusion: ABO c.28G>A mutation may cause B₃ -like subgroup by affecting RNA splicing of the ABO gene.