N- and C-terminal Transactivation Domains of GATA1 Protein Coordinate Hematopoietic Program
N- and C-terminal Transactivation Domains of GATA1 Protein Coordinate Hematopoietic Program
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DOI：
10.1074/jbc.m112.370437
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发表时间：
2012-06-15
影响因子：
4.8
通讯作者：
Shimizu, Ritsuko
中科院分区：
文献类型：
--
作者：
Kaneko, Hiroshi;Kobayashi, Eri;Shimizu, Ritsuko
转录因子 GATA1 调节一系列对于造血细胞向红细胞和巨核细胞谱系分化很重要的基因的表达。 GATA1 中已鉴定出三个功能域，一个位于 N 末端 (N-TAD) 的反式激活域和两个位于分子中部的锌指域。尽管 N-TAD 被认为是 GATA1 的孤立反式激活结构域，但唐氏综合症白血病的临床观察表明可能还存在其他反式激活结构域。在本研究中，我们在报告基因共转染测定中发现，通过删除 C 端 95 个氨基酸，GATA1 的反式激活活性显着降低，而 DNA 结合活性或自缔合潜力没有显着减弱。因此，我们生成了表达缺乏 C 末端区域的 GATA1 的转基因小鼠品系 (GATA1-Delta CT)。当我们将这些转基因小鼠系与 Gata1 缺陷小鼠杂交时，我们发现 GATA1-Delta CT 转基因使 Gata1 缺陷小鼠免于胚胎致死。使用与内源性 GATA1 几乎相似水平的 GATA1 Delta CT 挽救的胚胎在胚胎发育超过 13.5 天后，显示出严重贫血并伴有未成熟红系细胞的积累，与通过缺乏 N-TAD 的内源性水平的 GATA1 (GATA1-Delta NT) 挽救的胚胎的情况一样。 GATA1-Delta CT 和 GATA1-Delta NT 挽救的胚胎中，不同的靶基因组受到影响。我们还发现，在两个获救胚胎系中，GATA1 在未成熟巨核细胞的细胞周期控制中的功能减弱。因此，这些结果表明 GATA1 具有两个独立的反式激活结构域：N-TAD 和 C-TAD。 N-TAD 和 C-TAD 都保留了体内正常造血的冗余和特定活性。
Transcription factor GATA1 regulates the expression of a cluster of genes important for hematopoietic cell differentiation toward erythroid and megakaryocytic lineages. Three functional domains have been identified in GATA1, a transactivation domain located in the N terminus (N-TAD) and two zinc finger domains located in the middle of the molecule. Although N-TAD is known as a solitary transactivation domain for GATA1, clinical observations in Down syndrome leukemia suggest that there may be additional transactivation domains. In this study, we found in reporter co-transfection assays that transactivation activity of GATA1 was markedly reduced by deletion of the C-terminal 95 amino acids without significant attenuation of the DNA binding activity or self-association potential. We therefore generated transgenic mouse lines that expressed GATA1 lacking the C-terminal region (GATA1-Delta CT). When we crossed these transgenic mouse lines to the Gata1-deficient mouse, we found that the GATA1-Delta CT transgene rescued Gata1-deficient mice from embryonic lethality. The embryos rescued with an almost similar level of GATA1 Delta CT to endogenous GATA1 developed beyond embryonic 13.5 days, showing severe anemia with accumulation of immature erythroid cells, as was the case for the embryos rescued by endogenous levels of GATA1 lacking N-TAD (GATA1-Delta NT). Distinct sets of target genes were affected in the embryos rescued by GATA1-Delta CT and GATA1-Delta NT. We also found attenuated GATA1 function in cell cycle control of immature megakaryocytes in both lines of rescued embryos. These results thus demonstrate that GATA1 has two independent transactivation domains, N-TAD and C-TAD. Both N-TAD and C-TAD retain redundant as well as specific activities for proper hematopoiesis in vivo.