title: Mechanistic Evaluation for Mixed-field Agglutination in the K562 Cell Study Model with Exon 3 Deletion of A1 Gene.
abstract: In the case of blood type B3 with typical mixed-field agglutination of RBCs in the presence of anti-B or anti-AB antibody, a number of genetic alternations have been reported. It is well known that the IVS3+5G→A mutation in the B gene destroys the consensus of the splice donor site leading to exon 3 skipping during mRNA splicing. The lack of exon 3 likely causes a short stem region, producing an unstable B3 protein, and is concomitant with a decrease in B3 protein expression. Whether the phenomenon also appears in the type A blood group is of question. In this study, we evaluate whether exon 3 deletion in the blood type A gene also results in mixed-field phenotype. Site-directed mutagenesis was used to generate cDNA encoding A1 gene with exon 3 deletion. The cDNA was stably expressed in K562 cells. The expression of A antigen was compared with expression in parental K562 cells that did not express A antigen and in the stable K562 cell line expressing A(1) cDNA by flow cytometry analyses. The expression of A antigen in A1 stable cells and parental K562 cells was set as 100% and 0%, respectively. The mean relative percentage of A antigen expression for the cells of A1 with exon 3 deletion was 59.9% of A1 stable cells. Consistent with the observations of B3, which is B gene with exon 3 deletion, mixed field agglutination was observed for the cells expressing A1 with exon 3 deletion. Exon 3 deletion results in mixed field phenotype in both type A and B RBCs. However, the degree of antigen expression change for exon 3 deletion in A gene was less severe when compared with the deletion occurred in B gene.
authors.affiliation: Department of Laboratory Medicine, Chang Gung Memorial Hospital, Taoyuan County, Taiwan Department of Medical Biotechnology and Laboratory Science, College of Medicine, Chang Gung University, Taoyuan County, Taiwan a12048@adm.cgmh.org.tw.
authors.forename: Ding-Ping
authors.initials: DP
authors.lastname: Chen
authors.affiliation: Department of Medical Biotechnology and Laboratory Science, College of Medicine, Chang Gung University, Taoyuan County, Taiwan Molecular Medicine Research Center, Chang Gung University, Taoyuan County, Taiwan.
authors.forename: Ching-Ping
authors.initials: CP
authors.lastname: Tseng
authors.affiliation: Department of Laboratory Medicine, Chang Gung Memorial Hospital, Taoyuan County, Taiwan.
authors.forename: Chi-Jui
authors.initials: CJ
authors.lastname: Lin
authors.affiliation: Department of Laboratory Medicine, Chang Gung Memorial Hospital, Taoyuan County, Taiwan.
authors.forename: Wei-Ting
authors.initials: WT
authors.lastname: Wang
authors.affiliation: Department of Laboratory Medicine, Chang Gung Memorial Hospital, Taoyuan County, Taiwan Department of Pathology, School of Medicine, Chang Gung University, Taoyuan County, Taiwan.
authors.forename: Chien-Feng
authors.initials: CF
authors.lastname: Sun
mesh_terms: D000017:ABO Blood-Group System / Q000235:genetics* / Q000378:metabolism; D000371:Agglutination; D000483:Alleles; D005091:Exons; D006801:Humans; D020014:K562 Cells / Q000502:physiology*; D017384:Sequence Deletion
chemical_list: D000017:ABO Blood-Group System
keywords: RNA splicing; exon 3 skipping; mixed field; point mutation; site-directed mutagenesis