ENU mutagenesis identifies the first mouse mutants reproducing human β-thalassemia at the genomic level
ENU mutagenesis identifies the first mouse mutants reproducing human β-thalassemia at the genomic level
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DOI：
10.1016/j.bcmd.2012.09.004
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发表时间：
2013-02-01
影响因子：
2.3
通讯作者：
Curtis, David J.
中科院分区：
文献类型：
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作者：
Brown, Fiona C.;Scott, Nicholas;Curtis, David J.
正向遗传筛选已在许多物种中进行，以确定特定器官系统中的表型。我们已经进行了大规模的N-乙基-N-亚硝基脲（ENU）诱变筛选，以确定显性突变，扰乱小鼠红细胞生成。突变小鼠显示红细胞平均细胞体积（MCV）大于三个标准偏差的群体平均值进行了鉴定。其中两个细胞系，RBC 13和RBC 14，表现为低色素性小细胞性贫血，伴有明显的网织红细胞增多症、脾肿大和红细胞存活减少。从杂合子杂交的定时妊娠显示，四分之一的胚胎表现出严重的贫血，不能存活超过胚胎日（E）18.5，与纯合子β-地中海贫血一致。用β-地中海贫血小鼠系进行的遗传互补研究再现了复合杂合子中的胚胎致死性，基因组定制捕获阵列和β-珠蛋白基因座的大规模平行测序鉴定了致病突变。RBC 13细胞系在β-主要基因外显子2的密码子40处显示无义突变，这与人类常见的β（0）39地中海贫血突变相似。RBC 14细胞系在β-主要基因的多聚腺苷酸化信号处表现出突变，精确复制了人类β-地中海贫血突变。RBC 13和RBC 14系是第一个在基因组水平复制人β-地中海贫血的β-地中海贫血小鼠模型，因此突出了ENU诱变筛选在产生人类疾病小鼠模型中的作用。(C)2012 Elsevier Inc. All rights reserved.
Forward genetic screens have been performed in many species to identify phenotypes in specific organ systems. We have undertaken a large-scale N-ethyl-N-nitrosourea (ENU) mutagenesis screen to identify dominant mutations that perturb erythropoiesis in mice. Mutant mice that displayed an erythrocyte mean cell volume (MCV) greater than three standard deviations from the population mean were identified. Two of these lines, RBC13 and RBC14, displayed a hypochromic, microcytic anemia, accompanied by a marked reticulocytosis, splenomegaly and diminished red cell survival. Timed pregnancies from heterozygous intercrosses revealed that a quarter of the embryos displayed severe anemia and did not survive beyond embryonic day (E) 18.5, consistent with homozygous beta-thalassemia. Genetic complementation studies with a beta-thalassemia mouse line reproduced the embryonic lethality in compound heterozygotes and a genomic custom capture array and massively parallel sequencing of the beta-globin locus identified the causative mutations. The RBC13 line displayed a nonsense mutation at codon 40 in exon 2 of the beta-major gene, invoking parallels with the common beta(0)39 thalassemia mutation seen in humans. The RBC14 line exhibited a mutation at the polyadenylation signal of the beta-major gene, exactly replicating a human beta-thalassemia mutation. The RBC13 and RBC14 lines are the first beta-thalassemia mouse models that reproduce human beta-thalassemia at the genomic level, and as such highlight the power of ENU mutagenesis screens in generating mouse models of human disease. (C) 2012 Elsevier Inc. All rights reserved.