More purified CD49fHSC subpopulations also use asymmetric cell division to regulate daughter fates. (A) Experimental design. (B) CD49fHSC real-time differentiation landscapes by Uniform Manifold Approximation and Projection (UMAP). Localization of CD49fHSCs and daughters in landscape including CD71 and CD49c levels before divisions. n = 5 independent experiments; 1093 cells analyzed. (C) Identification of high-dimensional cell states (= clusters) in real-time differentiation landscape. Frequency of CD49f subpopulations and their potential to give rise of daughter cells in different parts of the depicted landscape. (D) CD49c expression of cluster C1 and C5 CD49fHSCs. (E) Both CD49c(high) and (low) CD49fHSCs show a- and symmetric lysosome inheritance. (F) Average frequency of cell fates (= clusters) in daughter cells after a-/symmetric LysoBrite inheritance in CD49c(high) cluster1 HSCs. Sister cell fates (= clusters, colors as in panel C) are comparable after symmetric inheritance, but differ after asymmetric inheritance. Asymmetric Lyso^High sisters more frequently acquire fates (clusters) with higher levels of CD49c than their Lyso^Low sisters. Two-way analysis of variance (ANOVA), Sidak multiple comparison corrected. n = 5 independent experiments; 1093 cells analyzed. (G) Quantification of overall a-/symmetric paired daughter cell fates (= clusters) after a-/symmetric LysoBrite inheritance. Five theoretically possible combinations of mother and daughter clusters (“division modes”) are shown on the left. Lysosome inheritance predicts division modes in CD49c^high and CD49c^low CD49fHSC subpopulations (clusters C1 and C5, respectively). In these subpopulations, after a-/symmetric LysoBrite inheritance, daughter cells a-/symmetrically change mother clusters (yellow/white). Two-way ANOVA, Sidak multiple comparison corrected. n = 5 independent experiments mean ± SEM.