The clonal and molecular aetiology of Flt3L-mediated 'emergency' dendritic cell development

Flt3 ligand (Flt3L) promotes an increased generation of type 1 conventional dendritic cells (cDC1s), resulting in enhanced immunity against infections and cancer. Here, we employ cellular barcoding to understand how Flt3L regulates single haematopoietic stem and progenitor cell (HSPC) fate. Our results demonstrate that although Flt3L stimulation can recruit some additional cDC1-generating HSPCs, the major contributing factor to higher cDC1 numbers is through enhanced clonal expansion. This selective cDC1 expansion occurs primarily via multi-/oligo-potent clones, without compromising their clonal output to other lineages. We then develop Divi-Seq to simultaneously profile division history, surface phenotype and the transcriptional state of single HSPCs during the early phase of the response. We discover that Flt3L-responsive HSPCs maintain a proliferative 'early progenitor'-like state, which leads to a selective emergence of CD11c+cKit+ transitional precursors with high cellular output to cDC1s. These findings inform the mechanistic action of Flt3L in natural immunity and immunotherapy. Overall design: Mouse HSPCs were CTV labelled and transplanted into non-irradiated recipients, which received PBS or Flt3L treatment for 3 days. Donor-derived cells (CTV+) on day 4 were isolated from recipient bone marrow and index sorted into 384 PCR plate, with library generated using the CEL-seq2 protocol. Please note that the cells were pooled per plate in the library preparation and the cell barcode annotation is provided.