C/EBPβ isoforms sequentially regulate regenerating mouse hematopoietic stem/progenitor cells

The transcription factor CCAAT enhancer-binding protein β (C/EBPβ) is required for stress-induced granulopoiesis at the level of hematopoietic stem/progenitor cells (HSPCs); however, its role and mechanisms of action in HSPCs are unknown. In this study, we assessed the regulation and functions of C/EBPβ in HSPCs, especially under stress conditions. After 5-fluorouracil treatment or bone marrow transplantation, Cebpb-/- HSPCs exhibited impaired cell-cycle activation and myeloid differentiation at the early and late phases of regeneration, respectively, whereas at steady state, Cebpb deficiency did not affect HSPCs. C/EBPβ was upregulated in response to hematopoietic stress, especially in CD150high long term-hematopoietic stem cells (LT-HSCs). Intracellular flow cytometric analysis that detected distinct domains of C/EBPβ revealed that, among the 3 isoforms of C/EBPβ, liver-enriched inhibitory protein (LIP) was upregulated in LT-HSCs prior to liver-enriched activating protein (LAP)/LAP* during regeneration. Early upregulation of LIP promoted cell-cycle entry of LT-HSCs by positively regulating Myc and expanded the HSPCs pool. Subsequent myeloid differentiation of amplified HSPCs was mediated by LAP/LAP*, which were upregulated at a later phase of regeneration. Collectively, our findings show that stress-induced sequential upregulation of C/EBPβ isoforms is critical for fine-tuning the proliferation and differentiation of regenerating HSPCs.