Progression from the Common Lymphoid Progenitor to B/Myeloid PreproB and ProB Precursors during B Lymphopoiesis Requires C/EBPα

Abstract

The C/EBPα transcription factor is required for myelopoiesis, with prior observations suggesting additional contributions to B lymphopoiesis. <i>Cebpa</i> expression is evident in common lymphoid progenitor (CLP) and preproB cells but is absent in proB and preB cells. We previously observed that marrow lacking the <i>Cebpa</i> +37 kb enhancer is impaired in producing B cells upon competitive transplantation. Additionally, a <i>Cebpa</i> enhancer/promoter-hCD4 transgene is expressed in B/myeloid CFU. Extending these findings, pan-hematopoietic murine <i>Cebpa</i> enhancer deletion using Mx1-Cre leads to expanded CLP, fewer preproB cells, markedly reduced proB and preB cells, and reduced mature B cells, without affecting T cell numbers. In contrast, enhancer deletion at the proB stage using Mb1-Cre does not impair B cell maturation. Further evaluation of CLP reveals that the <i>Cebpa</i> transgene is expressed almost exclusively in Flt3⁺ multipotent CLP versus B cell-restricted Flt3^- CLP. In vitro, hCD4⁺ preproB cells produce both B and myeloid cells, whereas hCD4^- preproB cells only produce B cells. Additionally, a subset of hCD4^- preproB cells express high levels of RAG1-GFP, as seen also in proB cells. Global gene expression analysis indicates that hCD4⁺ preproB cells express proliferative pathways, whereas B cell development and signal transduction pathways predominate in hCD4^- preproB cells. Consistent with these changes, <i>Cebpa</i> enhancer-deleted preproB cells downmodulate cell cycle pathways while upregulating B cell signaling pathways. Collectively, these findings indicate that C/EBPα is required for Flt3⁺ CLP maturation into preproB cells and then for proliferative <i>Cebpa</i>^int B/myeloid preproB cells to progress to <i>Cebpa</i>^lo B cell-restricted preproB cells and finally to <i>Cebpa</i>^neg proB cells.