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. Author manuscript; available in PMC: 2012 Apr 1.
Published in final edited form as: Nat Cell Biol. 2011 Aug 14;13(10):1244–1251. doi: 10.1038/ncb2313

Figure 4.

Figure 4

Notch-Mediated Degradation of β-Catenin Requires Numb and Lysosomal Activity. a, Human colon cancer cells (SW480) transfected with pcDNA-Myc or tethered Notch (V1774L)-Myc constructs, immunoprecipitated (IP) with anti-Myc antibody and immunoblotted (IB) with anti-Numb antibody. Expression of tethered Notch was detected with anti-Myc antibody; expression of pcDNA-myc was confirmed by PCR. b, Relative β-Catenin/TCF luciferase activity of ESCs transfected with control, N1ICD or tethered Notch (V1774L) in the presence or absence of Numb/Numbl siRNA and cultured in BIO for 72 h. c, Western analysis of active β-Catenin in ESCs transfected with control or tethered Notch (V1774L) in the presence or absence of Numb/Numbl siRNA. d, Western analysis of active β-Catenin in ESCs treated with control or DAPT in the presence or absence of Bafilomycin A1, which inhibits lysosomal activity. All luciferase values were normalized to Renilla activity (mean± s. d.; n = 4; *P < 0.01; NS, not significant). P values were determined using two-tailed Student’s t-test, type II (see Methods). Gapdh antibody was used as a loading control. Numbers on Western blots correspond to relative quantification. Con, control.