Fig. 2.

FOXO1 acts to enforce B-cell fate (A) Graph shows the distribution of D_H-J_H rearrangements in LY6D⁺ CLPs. A total of 192 cells/genotype from two independent sorts were assayed. Cells were scored as follows: GL only, germ-line band only; DJ + GL, cells producing one D_HJ_H and one GL DNA fragments; DJ, cells producing one or two D_HJ_H DNA fragments and no GL band; no read out, cells failing to produce detectable PCR products. (B) Graph displays cloning frequency from OP9-coculture experiments using single cell–sorted LY6D⁺ CLPs derived from WT and FOXO1^−/− mice. Cells were cultured in B/NK cell promoting culture conditions. Indicated are percentages of clones containing cells expressing CD19, NK1.1, and/or CD11C. A total of 264 cells per genotype were sorted. (C) Graphs display cloning frequency from OP9-DL1 coculture experiments using single-sorted LY6D⁻ (Left) and LY6D⁺ (Right) CLPs isolated from WT and FOXO1^−/− mice. Shown are the mean ± SEM. A total of 192 cells/genotype were sorted in two independent experiments. (D) Result from microarrays analysis displaying genes that are changed by a factor of twofold between WT and FOXO1^−/− LY6D⁺ CLPs. Displayed data are derived from two microarray replicas using cells from independent sorts. (E) LY6D⁺ CLPs from WT and FOXO1^−/− mice were analyzed by real-time PCR for the abundance of the indicated transcripts. Values were normalized for HPRT expression and are shown as mean ± SEM using mRNA from two independent sorts. (F) Multiplex single cell RT-PCR from LY6D⁺ CLPs. Graph indicates the percentages of cells expressing the indicated genes. A total of 96 cells were assayed for each genotype.