A nonsense mutation in zebrafish gata1 causes the bloodless phenotype in vlad tepes
A nonsense mutation in zebrafish gata1 causes the bloodless phenotype in vlad tepes
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DOI：
10.1073/pnas.082695299
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发表时间：
2002-04-16
影响因子：
11.1
通讯作者：
Liu, PP
中科院分区：
文献类型：
--
作者：
Lyons, SE;Lawson, ND;Liu, PP
Wad Tepes(VLT(M651))是通过大规模化学诱变筛选分离到的仅有的5个“无血”斑马鱼突变体之一。它的特点是血细胞前体细胞严重减少，循环开始时血细胞很少或没有。我们现在报告突变表型的特征和VLT(M651)突变基因的鉴定。VLT(M651)突变纯合子胚胎受精后24小时造血干细胞标志物表达正常，髓系和淋巴系标志物表达正常。对红系发育的分析表明，红系标志的表达是不同的。通过定位和候选基因克隆的方法，我们在VLT(M651)中发现了GATA1基因1015C->；T(Arg-339->；Stop)的无义突变。无义突变位于两个锌指的C末端，导致截断的蛋白质无法结合DNA或介导GATA特异性反式激活。含有斑马鱼GATA1基因的BAC克隆能够挽救VLT(M651)的无血表型。这些结果表明，VLT(M651)突变是斑马鱼中一种未知的GATA I等位基因。VLT(M651)突变揭示了GATA1在体内的结构和功能，表明GATA1在斑马鱼的造血中起着重要的作用，在哺乳动物和斑马鱼之间的功能具有显著的保守性，并为未来的造血途径的研究提供了一个强有力的工具。
Wad tepes (vlt(m651)) is one of only five "bloodless" zebrafish mutants isolated through large-scale chemical mutagenesis screening. It is characterized by a severe reduction in blood cell progenitors and few or no blood cells at the onset of circulation. We now report characterization of the mutant phenotype and the identification of the gene mutated in vlt(m651). Embryos homozygous for the vlt(m651) mutation had normal expression of hematopoietic stem cell markers through 24 h postfertilization, as well as normal expression of myeloid and lymphoid markers. Analysis of erythroid development revealed variable expression of erythroid markers. Through positional and candidate gene cloning approaches we identified a nonsense mutation in the gata1 gene, 1015C --> T (Arg-339 --> Stop), in vlt(m651). The nonsense mutation was located C-terminal to the two zinc fingers and resulted in a truncated protein that was unable to bind DNA or mediate GATA-specific transactivation. A BAC clone containing the zebrafish gata1 gene was able to rescue the bloodless phenotype in vlt(m651). These results show that the vlt(m651) mutation is a previously uncharacterized gata I allele in the zebrafish. The vlt(m651) mutation sheds new light on Gata1 structure and function in vivo, demonstrates that Gata1 plays an essential role in zebrafish hematopoiesis with significant conservation of function between mammals and zebrafish, and offers a powerful tool for future studies of the hematopoietic pathway.