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. 2015 Apr 23;33(5):1419–1433. doi: 10.1002/stem.1944

Figure 4.

Figure 4

E‐cadherin associates with PI3K/Akt activation in response to short‐term Wnt/β‐catenin signaling‐enhanced human embryonic stem cell (hESC) self‐renewal. (A): Decreased phosphorylated AKT (pAKTS473) in hESCs after E‐cadherin knockdown for 48 hours (Western blot). (B): Increased phosphorylated AKT (pAKTS473) after E‐cadherin upregulation. E‐cadherin‐Tet‐On hESCs were treated with DOX (2 µg/ml) for 48 hours, followed by Western blot analysis. (C): Inhibition of PI3/Akt with LY294002 abrogates the enhanced clonogenic capacity induced by E‐cadherin upregulation. E‐cadherin‐Tet‐On hESCs were treated with or without DOX (2 µg/ml) in the presence or absence of LY294002 (5 µM) for 24 hours, followed by clonogenic assays (n = 3). (D): E‐cadherin knockdown abolishes short‐term GSK3 inhibition‐induced upregulation of E‐cadherin and phosphorylated Akt, while specific Akt activator SC79 counteracts the effect of E‐cadherin knockdown. After siRNA knockdown for 44 hours, hESCs (H9 and H1 lines) were treated with either CHIR99021 (6 µM) or vehicle (DMSO) for 6 hours, followed by Akt inhibitor VIII (6 µM), or Akt activator II SC79 (6 µg/ml), or vehicle for an additional 30 minutes. All data in this figure are mean ± SD and were generated from H1 and H9 lines. **, p < .01.