Department of Blood Group Serology and Transfusion Medicine, Medical University of Graz, Graz, Austria.
Institute of Biochemistry, Technical University of Graz, Graz, Austria.
Transfusion. 2020 Jul;60(7):1564-1572. doi: 10.1111/trf.15907. Epub 2020 Jun 4.
Blood group A and B antigens are synthesized by glycosyltransferases regulated by a complex molecular genetic background. A multibase deletion in the ABO gene was identified in two related blood donors. To define its hereditary character and to evaluate genotype-phenotype associations, a detailed study including 30 family members was conducted.
ABO phenotyping was performed with agglutination techniques and adsorption-elution tests. The secretor status was determined. Allele-specific sequencing of ABO and genotyping of family members by a mutation-specific polymerase chain reaction were carried out. Functional analysis included cloning of complementary DNA and transfection experiments in HeLa cells. The antigen expression was investigated by flow cytometry and adsorption-elution method.
Sequencing analysis revealed a 24-bp deletion in Exon 5 and the adjacent intronic region of ABO. The alteration was inherited by 16 family members. Nine of them being heterozygous for the mutated allele failed to express A antigen on their erythrocytes as found by routine typing. In particular samples, however, adsorption-elution studies indicated inconclusive results. HeLa cells transfected with aberrant gene transcripts did not express blood group antigen A.
The variation causes defects in messenger RNA splicing, most likely inactivating the transferase as observed by serological typing and in vitro expression analysis. These data suggest a novel mechanism associated with blood group O and extend the knowledge of exceptionally rare ABO splice site mutations and deletions. With increased understanding of the molecular bases of ABO, the diagnostics may be further enhanced to ensure the safest possible use of the blood supply.
血型 A 和 B 抗原是由受复杂分子遗传背景调控的糖基转移酶合成的。在两位相关献血者的 ABO 基因中发现了一个多碱基缺失。为了确定其遗传特征并评估基因型-表型相关性，进行了一项包括 30 名家庭成员的详细研究。
采用凝集技术和吸附洗脱试验进行 ABO 表型分析。确定分泌状态。对 ABO 进行等位基因特异性测序，并通过突变特异性聚合酶链反应对家庭成员进行基因分型。功能分析包括 cDNA 克隆和 HeLa 细胞中的转染实验。通过流式细胞术和吸附洗脱法研究抗原表达。
测序分析显示，ABO 的外显子 5 和相邻内含子区域发生了 24 个碱基缺失。该改变通过 16 名家庭成员遗传。其中 9 名杂合子突变等位基因未能在其红细胞上表达 A 抗原，如常规分型所示。然而，在特定样本中，吸附洗脱研究表明结果不确定。转染异常基因转录本的 HeLa 细胞不表达血型抗原 A。
该变异导致信使 RNA 剪接缺陷，最可能使转移酶失活，如血清学分型和体外表达分析所示。这些数据表明与血型 O 相关的新机制，并扩展了异常罕见的 ABO 剪接位点突变和缺失的知识。随着对 ABO 分子基础的深入了解，诊断可能会进一步增强，以确保血液供应的安全使用。
