(a) t-SNE map with a principal curve fitted to all cells within the progenitor clusters (1,2,3,7). (b) Heatmap of z-score transformed pseudo-temporal expression profiles of a number of multipotency and lineage marker genes. Cells were ordered along the principal curve in (a) and profiles were smoothened by a local regression. The bottom panel depicts a local regression of the fate bias using the same temporal ordering. Successive expression of multipotency markers Kit, Ly6a, Ifitm1, Cd34, Cd48, and Flt3 is consistent with the ordering of multipotent progenitor (MPP) stages MPP1 to MPP4 as previously inferred by bulk measurements1,2. Lineage markers comprise Gata2, Car2, Gata1, Pf4 for the erythrocyte/megakaryocyte lineage, Cebpa, Csf1r, Mpo for the granulocyte and monocyte lineage, Itgax for the conventional dendritic cell lineage, and Irf8, Tcf4 for the plasmacytoid dendritic cell lineage, and Il7r, Rag1, Ebf1, Dntt for the B cell lineage. (c) Pictorial representation of the derived lineage tree. In (a) and (c) data are shown for 1,802 cells from 3 independent experiments with n = 4 mice. In (b) profiles are shown for 1,416 cells from three independent experiments with n = 4 mice. (d) Hematopoietic lineage tree inferred by StemID2. Only significant links are shown (P<0.01). The color of the link indicates the -log10p-value. The color of the vertices indicates the entropy. The thickness indicates the link score reflecting how densely a link is covered with cells. The lineage tree is consistent with independently derived fate bias estimates: erythrocytes (cluster 9) branch off from cluster 1, while cluster 2 and 3 give rise to granulocytes/monocytes (cluster 17), and cluster 2 and 7 comprise progenitors of pDCs (cluster 12) and B cells (cluster 10). (e) Barplot of StemID2 scores for hematopoietic clusters. Cluster 1, which shows highest expression of HSC markers, such as Ifitm12, receives the highest StemID2 score. In (a) and (b), only clusters with >5 cells were included and a link score cut-off of 0.5 was applied. The StemID2 in (d) and (e) computation has been performed on 1,802 cells from three independent experiments with n = 4 mice.
1. Wilson, A. et al. Hematopoietic Stem Cells Reversibly Switch from Dormancy to Self-Renewal during Homeostasis and Repair. Cell 135, 1118–1129 (2008).
2. Cabezas-Wallscheid, N. et al. Identification of Regulatory Networks in HSCs and Their Immediate Progeny via Integrated Proteome, Transcriptome, and DNA Methylome Analysis. Cell Stem Cell 15, 507–522 (2014).