β-catenin promotes human EB differentiation. (A) Percentage of GFP⁺ EBs after Δ45β-catenin transduction in each of the 6 donors shown in Figure 6. (B) Western blot analysis of Δ45β-catenin expression in human EBs 24 hours after doxycycline (Dox) treatment. (C) Quantification of erythroid differentiation in response to Dox treatment compared with PBS-treated control cells, as determined by flow cytometry, revealing treatment with Dox (gray bars) not to alter the differentiation of EBs (n = 6; two-way ANOVA). (D) Representative flow cytometry plots of erythroid differentiation, using CD71 and CD235a cell surface staining, gated for GFP⁺ marked lentiviral transduced cells, in response to Dox-inducible Δ45β-catenin expression, as compared with untreated control conditions. (E) Quantification of flow cytometry data, representing fold change of GFP⁺ cells in CD71⁻CD235a⁻-, CD71⁺CD235a⁻-, CD71⁺CD235a⁺-, and CD71⁻CD235a⁺-defined cell stages of EB development, in untreated conditions (pink bars) or stimulated with Dox (gray bars) (n = 6; two-way ANOVA, ∗∗∗∗P < .0001). ns, not significant. (F) Graphical model displaying conserved expression of E-cadherin and β-catenin in human and rat erythroblatst, which upregulation in response to anemia may stimulate erythropoiesis.