Differential Contribution of the Gata1 Gene Hematopoietic Enhancer to Erythroid Differentiation
Differential Contribution of the Gata1 Gene Hematopoietic Enhancer to Erythroid Differentiation
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DOI：
10.1128/mcb.01572-08
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发表时间：
2009-03-01
影响因子：
5.3
通讯作者：
Yamamoto, Masayuki
中科院分区：
文献类型：
--
作者：
Suzuki, Mikiko;Moriguchi, Takashi;Yamamoto, Masayuki
GATA1 是红细胞分化的关键调节因子。为了研究 Gata1 基因表达如何以阶段特异性方式受到调节，制备了表达来自细菌人工染色体 (G1BAC-GFP) 中 Gata1 基因座的绿色荧光蛋白 (GFP) 报告基因的转基因小鼠系。我们发现 GFP 报告基因的表达忠实地再现了 Gata1 基因的表达。使用 GFP 荧光与造血表面标记相结合，我们建立了两种红系祖细胞级分的纯化方案，称为突发形成单位-红系细胞相关红系祖细胞 (BREP) 和 CFU-红系细胞相关红系祖细胞 (CREP) 级分。我们检查了Gata1基因造血增强子（G1HE）和增强子核心中高度保守的GATA盒的功能。 G1HE 的缺失和 GATA 盒的取代突变均导致 BREP 级分中 GFP 表达几乎完全丧失，但 CREP 阶段的表达仅部分受到抑制，表明 GATA 盒对 Gata1 的 BREP 阶段表达的关键贡献。一致地，从染色体 Gata1 位点定向删除 G1HE 会引起 BREP 部分中 Gata1 基因的表达抑制，从而导致 BREP 阶段造血祖细胞的异常积累。这些结果证明了 Gata1 基因表达以分化阶段特异性方式动态调节的生理意义。
GATA1 is a key regulator of erythroid cell differentiation. To examine how Gata1 gene expression is regulated in a stage-specific manner, transgenic mouse lines expressing green fluorescent protein (GFP) reporter from the Gata1 locus in a bacterial artificial chromosome (G1BAC-GFP) were prepared. We found that the GFP reporter expression faithfully recapitulated Gata1 gene expression. Using GFP fluorescence in combination with hematopoietic surface markers, we established a purification protocol for two erythroid progenitor fractions, referred to as burst-forming units-erythroid cell-related erythroid progenitor (BREP) and CFU-erythroid cell-related erythroid progenitor (CREP) fractions. We examined the functions of the Gata1 gene hematopoietic enhancer (G1HE) and the highly conserved GATA box in the enhancer core. Both deletion of the G1HE and substitution mutation of the GATA box caused almost complete loss of GFP expression in the BREP fraction, but the CREP stage expression was suppressed only partially, indicating the critical contribution of the GATA box to the BREP stage expression of Gata1. Consistently, targeted deletion of G1HE from the chromosomal Gata1 locus provoked suppressed expression of the Gata1 gene in the BREP fraction, which led to aberrant accumulation of BREP stage hematopoietic progenitor cells. These results demonstrate the physiological significance of the dynamic regulation of Gata1 gene expression in a differentiation stage-specific manner.