Protein partners of C/EBPε

CCAAT-enhancer binding protein-ε (C/EBPε) is a nuclear transcription factor implicated in the regulation of terminal myeloid differentiation. Using a yeast two-hybrid screen, potential interaction partners of C/EBPε involved in myeloid development were identified. C/EBPε was found to associate with other C/EBP family members, including C/EBPε and CHOP as well as other proteins that are known to contain a leucine-zipper protein interaction motif including CREB2, LDOC1, E6TP1, and AF-17. In addition, C/EBPε demonstrated the potential for interaction with proteins that do not possess a leucine-zipper motif, including proteins that may be involved in sumoylation (protein inhibitor of activated STAT1 [PIAS1] and ubiquitin-conjugating enzyme E2I). As expected, the association of C/EBPε with other C/EBP family members depends on the presence of a functional leucine-zipper motif. Mapping studies of C/EBPε with PIAS1 (as an example of a nonleucine-zipper–containing protein) showed that C/EBPε interacts with the amino-terminal domain of PIAS1. The function of C/EBPε interacting proteins was further investigated. Co-expression of C/EBPε with C/EBPδ resulted in potent transactivation in a lactoferrin reporter system. A gel mobility shift assay suggests that C/EBPε, C/EBPα, and C/EBPδ proteins can bind as heterodimers to a C/EBP consensus DNA-binding site. As CHOP is known to represent a transcriptional repressor, the functional interaction between C/EBPε and CHOP was investigated. Co-expression of C/EBPε and c-Myb with CHOP caused marked transcriptional repression of target reporter genes. Our results suggest heterodimeric partners of C/EBPε modulate the function of C/EBPε in mediating gene transcription during myelopoiesis.