Identification of the C/EBP伪 C-terminal tail residues involved in the protein interaction with GABP and their potency in myeloid differentiation of K562 cells
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- 作者：Toshibumi Shimokawa ; Satoshi Nunomura ; Daisuke Fujisawa ; Chisei Ra
- 关键词：AML ; acute myeloid leukemia ; BR ; basic region ; bZIP ; basic leucine zipper ; C/EBP伪 ; CCAAT/enhancer-binding protein 伪 ; ChIP ; chromatin immunoprecipitation ; DBD ; DNA-binding domain ; EGFP ; enhanced green fluorescent protein ; EMSA ; electrophoretic mobility shift assay ; GST ; glutathione S-transferase ; HSV-1 ; herpes simplex virus type 1 ; IRES ; internal ribosome entry site ; LZ ; leucine zipper ; mtFKBP ; mutated FK506-binding protein-12 ; ODN ; oligodeoxyribonucleotides ; PCR ; polymerase chain reaction ; RT-
- 刊名：Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms
- 出版年：November, 2013
- 年：2013
- 卷：1829
- 期：11
- 页码：1207-1217
- 全文大小：1048 K
文摘
The CCAAT/enhancer-binding protein 伪 (C/EBP伪) is the member of a family of related basic leucine zipper (bZIP) transcription factors and is critical for granulopoiesis. We previously demonstrated that C/EBP伪 interacts with the ETS domain of widely expressed GABP伪, which leads to cooperative transcriptional activation of the myeloid-specific promoter for human FCAR encoding the Fc receptor for IgA (Fc伪R, CD89) in part by facilitating recruitment of C/EBP伪 to the promoter. The C/EBP伪 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 GABP伪 interacts with the last 18 residues of the C/EBP伪 C-terminus beyond the bZIP DNA-binding and dimerizing region. Deletion of this C-terminus resulted in loss of GABP伪 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/EBP伪 mutant incapable of interacting with GABP伪 does not induce expression of granulocytic differentiation markers including CD15, CD11b, GCSF-R and C/EBP蔚, and does not inhibit proliferation, whereas wild type does. These results demonstrate the functional importance of the C/EBP伪 C-terminus beyond the bZIP DNA-binding and dimerization region, which may mediate cooperative activation by C/EBP伪 and GABP of myeloid-specific genes involved in C/EBP伪-dependent granulopoiesis.