Expression of South East Asian Ovalocytic Band 3 Disrupts Erythroblast Cytokinesis and Reticulocyte Maturation
Expression of South East Asian Ovalocytic Band 3 Disrupts Erythroblast Cytokinesis and Reticulocyte Maturation
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DOI：
10.3389/fphys.2020.00357
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发表时间：
2020-04-28
影响因子：
4
通讯作者：
Bruce, Lesley J.
中科院分区：
文献类型：
--
作者：
Flatt, Joanna F.;Stevens-Hernandez, Christian J.;Bruce, Lesley J.
东南亚人卵圆细胞增多症是由于红细胞阴离子交换蛋白AE 1（带3）中9个氨基酸的杂合缺失所致。第一个成功出生的个体纯合子的报告显示与严重的红细胞生成不良性贫血。先证者红细胞成像显示其表面存在带3，Wr（B）抗原表达减少，血型糖蛋白C、CD 44和CD 147免疫反应性增加。来自杂合子东南亚卵圆细胞增多症红细胞的膜的免疫印迹显示CD 44、CD 147和钙网蛋白的定量增加，表明网织红细胞成熟的缺陷，以及带3的残基Tyr 359和膜上的过氧化物氧还蛋白-2的磷酸化增加，分别表明带3的运输和氧化应激改变。纯合和杂合东南亚卵圆细胞增多症红系祖细胞的体外培养产生双核和多核细胞。去核在纯合细胞中严重受损，在杂合细胞中减少。形成了带3的大的内部囊泡积累，其与其他质膜蛋白和自噬体标记物LC 3共定位，但不与ER、高尔基体或再循环内体标记物共定位。带3从成红细胞裂解物在正染阶段的免疫沉淀显示增加的相互作用的突变体带3与热休克蛋白，泛素和细胞骨架蛋白，锚蛋白，血影蛋白和肌动蛋白。我们还发现突变带3与非肌肌球蛋白IIA和IIB形成强烈的相互作用，而这种相互作用在野生型成红细胞中不能检测到。与此一致，非肌肉肌球蛋白IIA和肌动蛋白的本地化在一些东南亚卵圆细胞增多症成红细胞受到干扰。这些发现为理解突变膜蛋白表达引起的体内红细胞生成不良提供了新的见解。
Southeast Asian Ovalocytosis results from a heterozygous deletion of 9 amino acids in the erythrocyte anion exchange protein AE1 (band 3). The report of the first successful birth of an individual homozygous for this mutation showed an association with severe dyserythropoietic anemia. Imaging of the proband's erythrocytes revealed the presence of band 3 at their surface, a reduction in Wr(b) antigen expression, and increases in glycophorin C, CD44, and CD147 immunoreactivity. Immunoblotting of membranes from heterozygous Southeast Asian Ovalocytosis red cells showed a quantitative increase in CD44, CD147, and calreticulin suggesting a defect in reticulocyte maturation, as well as an increase in phosphorylation at residue Tyr359 of band 3, and peroxiredoxin-2 at the membrane, suggesting altered band 3 trafficking and oxidative stress, respectively. In vitro culture of homozygous and heterozygous Southeast Asian Ovalocytosis erythroid progenitor cells produced bi- and multi-nucleated cells. Enucleation was severely impaired in the homozygous cells and reduced in the heterozygous cells. Large internal vesicular accumulations of band 3 formed, which co-localized with other plasma membrane proteins and with the autophagosome marker, LC3, but not with ER, Golgi or recycling endosome markers. Immunoprecipitation of band 3 from erythroblast cell lysates at the orthochromatic stage showed increased interaction of the mutant band 3 with heat shock proteins, ubiquitin and cytoskeleton proteins, ankyrin, spectrin and actin. We also found that the mutant band 3 forms a strong interaction with non-muscle myosins IIA and IIB, while this interaction could not be detected in wild type erythroblasts. Consistent with this, the localization of non-muscle myosin IIA and actin was perturbed in some Southeast Asian Ovalocytosis erythroblasts. These findings provide new insights toward understanding in vivo dyserythropoiesis caused by the expression of mutant membrane proteins.