Toward a more precise immunophenotypic and molecular definition of human cMoPs.

Monocytes arise from hematopoietic progenitors in the bone marrow and play crucial roles in both homeostasis and inflammation. Human common monocyte progenitors (cMoPs) were originally identified as CLEC12A+CD64hi cells but have recently been redefined as CD115+CD64hi/int cells. We therefore evaluated whether CLEC12A+CD64hi cMoPs and CD115+CD64hi/int cMoPs represent the same cell type using flow cytometry, scRNA-seq, morphological, and functional analyses. We demonstrate that including CD115 in the gating strategy, rather than CLEC12A, yields a more accurate definition of cMoPs. Differentiation assays demonstrate the transition of CD115+CD64int cMoPs into CD115+CD64hi cMoPs and a comparable capacity for the two cMoPs to generate monocyte progeny. scRNA-seq clustering of the CD115+CD64hi/int cMoPs identified five different transcriptional features including maturation-associated programs, IFN response and stress-related genes. ATAC-seq analysis of CD115+CD64hi/int cMoPs identified IRF8, PU.1, and also C/EBPδ as transcription factors putatively involved in monocyte development. Based on a more accurate definition and analysis, our data demonstrate the molecular complexity of human cMoPs.