Single-cell RNA-seq reveals new types of human blood dendritic cells, monocytes, and progenitors

What's in a drop of blood? Blood contains many types of cells, including many immune system components. Immune cells used to be characterized by marker-based assays, but now classification relies on the genes that cells express. Villani et al. used deep sequencing at the single-cell level and unbiased clustering to define six dendritic cell and four monocyte populations. This refined analysis has identified, among others, a previously unknown dendritic cell population that potently activates T cells. Further cell culture revealed possible differentiation progenitors within the different cell populations. Science, this issue p. eaah4573 Discovery of additional immune cell subtypes will help identify functions and immune monitoring during disease. INTRODUCTION Dendritic cells (DCs) and monocytes consist of multiple specialized subtypes that play a central role in pathogen sensing, phagocytosis, and antigen presentation. However, their identities and interrelationships are not fully understood, as these populations have historically been defined by a combination of morphology, physical properties, localization, functions, developmental origins, and expression of a restricted set of surface markers. RATIONALE To overcome this inherently biased strategy for cell identification, we performed single-cell RNA sequencing of ~2400 cells isolated from healthy blood donors and enriched for HLA-DR+ lineage− cells. This single-cell profiling strategy and unbiased genomic classification, together with follow-up profiling and functional and phenotypic characterization of prospectively isolated subsets, led us to identify and validate six DC subtypes and four monocyte subtypes, and thus revise the taxonomy of these cells. RESULTS Our study reveals: 1) A new DC subset, representing 2 to 3% of the DC populations across all 10 donors tested, characterized by the expression of AXL, SIGLEC1, and SIGLEC6 antigens, named AS DCs. The AS DC population further divides into two populations captured in the traditionally defined plasmacytoid DC (pDC) and CD1C+ conventional DC (cDC) gates. This split is further reflected through AS DC gene expression signatures spanning a spectrum between cDC-like and pDC-like gene sets. Although AS DCs share properties with pDCs, they more potently activate T cells. This discovery led us to reclassify pDCs as the originally described “natural interferon-producing cells (IPCs)” with weaker T cell proliferation induction ability. 2) A new subdivision within the CD1C+ DC subset: one defined by a major histocompatibility complex class II–like gene set and one by a CD14+ monocyte–like prominent gene set. These CD1C+ DC subsets, which can be enriched by combining CD1C with CD32B, CD36, and CD163 antigens, can both potently induce T cell proliferation. 3) The existence of a circulating and dividing cDC progenitor giving rise to CD1C+ and CLEC9A+ DCs through in vitro differentiation assays. This blood precursor is defined by the expression of CD100+CD34int and observed at a frequency of ~0.02% of the LIN–HLA-DR+ fraction. 4) Two additional monocyte populations: one expressing classical monocyte genes and cytotoxic genes, and the other with unknown functions. 5) Evidence for a relationship between blastic plasmacytoid DC neoplasia (BPDCN) cells and healthy DCs. CONCLUSION Our revised taxonomy will enable more accurate functional and developmental analyses as well as immune monitoring in health and disease. The discovery of AS DCs within the traditionally defined pDC population explains many of the cDC properties previously assigned to pDCs, highlighting the need to revisit the definition of pDCs. Furthermore, the discovery of blood cDC progenitors represents a new therapeutic target readily accessible in the bloodstream for manipulation, as well as a new source for better in vitro DC generation. Although the current results focus on DCs and monocytes, a similar strategy can be applied to build a comprehensive human immune cell atlas. Establishing a human blood monocyte and dendritic cell atlas. We isolated ~2400 cells enriched from the healthy human blood lineage− HLA-DR+ compartment and subjected them to single-cell RNA sequencing. This strategy, together with follow-up profiling and functional and phenotypic characterization, led us to update the original cell classification to include six DCs, four monocyte subtypes, and one conventional DC progenitor. Dendritic cells (DCs) and monocytes play a central role in pathogen sensing, phagocytosis, and antigen presentation and consist of multiple specialized subtypes. However, their identities and interrelationships are not fully understood. Using unbiased single-cell RNA sequencing (RNA-seq) of ~2400 cells, we identified six human DCs and four monocyte subtypes in human blood. Our study reveals a new DC subset that shares properties with plasmacytoid DCs (pDCs) but potently activates T cells, thus redefining pDCs; a new subdivision within the CD1C+ subset of DCs; the relationship between blastic plasmacytoid DC neoplasia cells and healthy DCs; and circulating progenitor of conventional DCs (cDCs). Our revised taxonomy will enable more accurate functional and developmental analyses as well as immune monitoring in health and disease.