Figure 3.

Ddit3 Is Required in Early Erythroid Specification and Blocks Myeloid Lineage Progression

(A–C) Lineage potential of multipotent mouse BM cells upon the loss of Ddit3 expression. CFC assays of FDCPmix cells (n = 3) (A) and primary KLS cells (n = 3) (B) upon Ddit3 knockdown and of Ddit3 knockout KLS cells (n = 4) (C) are shown. Error bars, SD.

(D) Re-plating capacity of primary BM GMPs upon enforced expression of Ddit3 read in CFC assays (CSIem, empty vector; n = 4). Colonies were scored 7–10 days after plating of transduced cells (plate 1). The cellular content of the colonies obtained was re-seeded into successive CFC assays (plates 2–4) until the exhaustion of colony production.

(E) Distribution of colony types in CFC plate 1. Most GM colonies obtained upon Ddit3-enforced expression have a blast-like appearance. Error bars, SEM.

(F) Representative images of GM colonies in (E) are shown.

(G) PCA plot of the transcriptional profiles of individual GMPs, either untransduced (WT) or transduced with CSIem- or Ddit3-expressing lentiviral vectors, analyzed for the expression of 44 genes. The first two PC explain 24% of the data variance; n = 114 (CSIem), 84 (Ddit3), and 118 (WT).

(H) Gene loadings of PC1 and PC2 in (G). Genes with the most extreme positions along each axis contribute the most to cell separation along the respective PC.