2023 - SIS-SEQ IDENTIFIES THE EARLIEST LINEAGE PRIMING REGULATORS OF DENDRITIC CELL FATE

Dendritic cells (DCs) are immune cells important for the detection and immunity against pathogens, self-antigens and cancer. They include 3 subtypes, conventional DC type 1 (cDC1), cDC type 2 (cDC2) and plasmacytoid DC (pDC) which all derive from a common hematopoietic stem cell progenitor population (HSPCs). Recent evidence has shown that this population is heterogenous for fate and not all HSPCs produce every DC subtype (Naik et al., 2007). To understand the earliest lineage priming regulators of DC fate, one would have to overcome the problem of the destructive nature of scRNA-seq, which makes it impossible for a single cell to be tested for both fate and its transcription profile. Here, we achieve this with a novel approach where a single progenitor cell is allowed to divide into a few cells, some of which are tested for fate while others tested for gene expression. By correlating clonal fate with gene expression, clone-by-clone, for 109 clones, we identified 490 genes that correlated with cDC1, cDC2 and/or pDC fate bias. All genes were tested in a pooled CRISPR/Cas9 screen in DC cultures where several novel genes were identified that regulated DC fate – the knockout of some genes enhanced numbers of particular DC subtypes, whereas others reduced numbers. This system could be used to study heterogeneity of fate at clonal level in other populations, including stem cells and cancer cells to reveal the transcriptional origin of fate decisions.