Human C/EBP-ϵ activator and repressor isoforms differentially reprogram myeloid lineage commitment and differentiation

CCAAT enhancer-binding protein-epsilon (C/EBP-ϵ) is required for the terminal differentiation of neutrophils and eosinophils. Human C/EBP-ϵ is expressed as 4 isoforms (32, 30, 27, and 14 kDa) through differential RNA splicing, and alternative promoters and translational start sites. The C/EBP-ϵ32/30 isoforms are transcriptional activators, whereas C/EBP-ϵ27 interacts with and represses GATA-1 transactivation of eosinophil promoters. C/EBP-ϵ14 contains only DNA-binding and -dimerization domains and may function as a dominant-negative regulator. To define functional activities for these C/EBP-ϵ isoforms in myelopoiesis, human CD34+ progenitors were transduced with internal ribosomal entry site–enhanced green fluorescent protein retroviral vectors encoding the 32/30, 27, and 14-kDa isoforms, purified by fluorescence-activated cell sorter, and analyzed in colony-forming assays and suspension cultures. Progenitors transduced with C/EBP-ϵ32/30 default exclusively to eosinophil differentiation and gene expression, independent of interleukin-5, and regardless of inclusion of cytokines to induce other lineages. In contrast, the putative repressor C/EBP-ϵ27 isoform strongly inhibits eosinophil differentiation and gene expression, including GATA-1, promoting granulocyte (neutrophil)-macrophage differen-tiation. The C/EBP-ϵ14 repressor isoform strongly inhibits eosinophil development and gene expression, promoting erythroid differentiation, an effect enhanced by erythropoietin. Thus, C/EBP-ϵ isoforms can reprogram myeloid lineage commitment and differentiation consistent with their predicted activities based on activator and repressor domains and in vitro functional activities.