Identification of the C/EBPα C-terminal tail residues involved in the protein interaction with GABP and their potency in myeloid differentiation of K562 cells
Identification of the C/EBPα C-terminal tail residues involved in the protein interaction with GABP and their potency in myeloid differentiation of K562 cells
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鉴定参与与 GABP 的蛋白质相互作用的 C/EBPα C 末端残基及其在 K562 细胞骨髓分化中的效力
DOI：
10.1016/j.bbagrm.2013.09.004
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发表时间：
2013
期刊：
影响因子：
--
通讯作者：
and Chisei Ra
中科院分区：
文献类型：
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作者：
Toshibumi Shimokawa;Satoshi Nunomura;Daisuke Fujisawa;and Chisei Ra
CCAAT/增强子结合蛋白α（C/EBPα）是碱性亮氨酸拉链（bZIP）转录因子家族的成员，对粒细胞生成至关重要。我们先前证明C/EBPα与广泛表达的GABPα的ETS结构域相互作用，导致编码伊加Fc受体（FcαR，CD 89）的人FCAR骨髓特异性启动子协同转录激活，部分原因是促进C/EBPα募集到启动子。C/EBPα分子在其N端含有反式激活结构域（TADs），在其C端含有DNA结合和二聚化bZIP结构。我们证明GABPα与bZIP DNA结合和二聚化区域以外的C/EBPα C末端的最后18个残基相互作用。该C末端的缺失导致GABPα相互作用的丧失，但不影响其DNA结合能力，表明其不是同源二聚体形成所必需的。此外，当与VP 16 β的C-末端融合时，C-末端赋予在异源β上与GABP功能协同的能力。我们确定了一个三个氨基酸的延伸（氨基酸341-343），这是重要的功能和蛋白质与GABP的相互作用。不能与GABPα相互作用的C/EBPα突变体在K562细胞中的异位表达不诱导粒细胞分化标志物包括CD 15、CD 11b、GCSF-R和C/EBPε的表达，也不抑制细胞增殖，而野生型则有此作用。这些结果证明了C/EBPα C端在bZIP DNA结合和二聚化区域之外的功能重要性，其可能介导C/EBPα和GABP对参与C/EBPα依赖性粒细胞生成的骨髓特异性基因的协同激活。
The CCAAT/enhancer-binding protein α (C/EBPα) is the member of a family of related basic leucine zipper (bZIP) transcription factors and is critical for granulopoiesis. We previously demonstrated that C/EBPα interacts with the ETS domain of widely expressed GABPα, which leads to cooperative transcriptional activation of the myeloid-specific promoter for humanFCARencoding the Fc receptor for IgA (FcαR, CD89) in part by facilitating recruitment of C/EBPα to the promoter. The C/EBPα molecule contains transactivation domains (TADs) at its N-terminus and a DNA-binding and dimerization bZIP structure at its C-terminus. We demonstrate here that GABPα interacts with the last 18 residues of the C/EBPα C-terminus beyond the bZIP DNA-binding and dimerizing region. Deletion of this C-terminus resulted in loss of GABPα interaction but not affecting its DNA binding ability, indicating that it is not required for homodimer formation. Moreover, the C-terminus confers the ability to functionally synergize with GABP on a heterologous TAD when fused to the C-terminus of the VP16 TAD. We identified a three-amino acid stretch (amino acids 341–343) that is important for both functional and protein interactions with GABP. Ectopic expression in K562 cells of C/EBPα mutant incapable of interacting with GABPα does not induce expression of granulocytic differentiation markers including CD15, CD11b, GCSF-R and C/EBPε, and does not inhibit proliferation, whereas wild type does. These results demonstrate the functional importance of the C/EBPα C-terminus beyond the bZIP DNA-binding and dimerization region, which may mediate cooperative activation by C/EBPα and GABP of myeloid-specific genes involved in C/EBPα-dependent granulopoiesis.