An Ikaros-driven in vitro system captures key features of in vivo B cell progenitor differentiation.

(A) Schematic representation of self-renewing (left) and differentiating (right) B cell progenitors and the nomenclature used to refer to these populations in the literature. (B) Experimental design of Ikaros-induced B3 cell differentiation. RNA-seq was performed at the indicated time points. The experiment was designed so that sources of variability could be traced to library preparation and sequencing biases and corrected (see Methods). Effects of 4-OHT were monitored by addition of 4-OHT to control vector cells and found to be negligible. (C) Pseudotemporal ordering of cells as a function of their scRNA-seq transcriptomes. The pseudotime trajectory is shown in black. (D) Gene set analysis of gene expression trends in B3 differentiation. Each box contains the total number of genes and the major functional terms (q S1 Data. (F) Comparison of genes differentially expressed in IKZF1 mutated human B-ALL and the B3 differentiation. Numbers denote common genes that are differentially expressed in IKZF1-mutated versus IKZF1 wild-type human B-ALL and the top 200 genes regulated at the indicated times after Ikaros induction in B3 cells. The color scale indicates the odds ratio for the enrichment of Ikaros-regulated gene during B3 cell differentiation among differentially expressed genes in human B-ALL with and without IKZF1 mutations. B-ALL, B cell progenitor acute lymphoblastic leukemia; ERt2; Fr.C, proliferating B cell progenitor; Fr.D, differentiating B cell progenitor; HA, haemagglutinin; JAK-STAT, Janus kinase/signal transducers and activators of transcription; RNA-seq, RNA sequencing; scRNA-seq, single cell RNA sequencing; TCA, tricarboxylic acid cycle; 4-OHT, 4-hydroxy-tamoxifen.