Diversity of peripheral blood human NK cells identified by single cell RNA sequencing

Human Natural Killer (NK) cells in peripheral blood perform many functions but classification of specific subsets has been a long-standing problem. Here, we report single-cell RNA sequencing of NK cells from healthy CMV-negative donors, comparing gene expression in unstimulated and IL-2 activated cells. Unsupervised clustering identified seven NK cell subsets. Three resembled well-described populations, CD56brightCD16-, CD56dimCD16+CD57- and CD56dimCD16+CD57+. CD56dimCD16+CD57- cells sub-divided to include a population with higher chemokine mRNA and increased frequency of KIR expression. Three novel human blood NK cell populations were identified as: a population of type I interferon responding NK cells which were CD56neg; a population exhibiting an in-vivo cytokine-induced memory-like phenotype, including increased granzyme B mRNA in response to IL-2, and finally, a small population, with low ribosomal expression, down-regulation of oxidative phosphorylation and high levels of immediate early response genes indicative of cellular activation. Human cytomegalovirus positive donors also included a higher frequency of adaptive NK cells. Together, these data establish an unexpected diversity in blood NK cells and provide a new framework for analysing NK cell responses in health and disease. Overall design: scRNAseq of sorted NK cells from one healthy individual