Single-cell RNASeq analysis to unravel molecular networks driving leukemia in Ebf1+/-Pax5+/- (dHet) B-ALL mice

Here, we report that EBF1 and Pax5 collaborate in a dose-dependent manner to regulate the IL-7-STAT5 signaling pathway and one-carbon metabolism, whereby we found both diminished and enhanced binding of EBF1 and Pax5 to target genes in compound heterozygous mutant mice. Moreover, single-cell RNA sequencing analysis identified a small subset of wild-type pro-B cells on the trajectory to pre-B cells that share gene expression signatures with leukemic Ebf1+/−Pax5+/− pro-B cells. Thus, a normal expression level of EBF1 and Pax5 is required for safeguarding a potentially vulnerable pro-Bcell subset from a transformation to B-ALL. Single cells from the bone marrow, spleen or blood were sorted into 384-well plates and processed using a modified version of the CEL-Seq2 protocol (Hasimshony et al. 2016, Genome Biology, DOI: 10.1186/s13059-016-0938-8).