Single-cell transcriptomics in bone marrow delineates CD56dimGranzymeK+ subset as intermediate stage in NK cell differentiation

In this study we performed single-cell RNA sequencing on 7000 negatively enriched fresh NK and progenitor cells from one healthy bone marrow donor, that were pre-labeled with oligonucleotide-conjugated antibodies against CD34, CD117, CD56 and CXCR6. A new CD56dimGzmK+ subset was identified that shared features with CD56bright and CD56dimGzmK- NK cells based on transcriptome, phenotype (NKG2AhighCD16lowKLRG1highTIGIThigh) and functional analysis in bone marrow and blood, supportive for an intermediate subset. Pseudotime analysis positioned CD56bright, CD56dimGzmK+ and CD56dimGzmK- cells in one differentiation trajectory, while ltNK cells were separated in a distinct lineage. Integrative analysis with bone marrow cells from the human cell atlas did not demonstrate a developmental connection between CD34+ progenitor and NK cells, implicating that bone marrow is not a site of active NK cell development. Overall design: Single-cell RNA sequencing on negatively enriched NK and progenitor cells from one healthy bone marrow donor, combined with oligonucleotide-conjugated antibodies against CD34, CD117, CD56 and CXCR6.