Figure 3.

c-Abl acts on p53 to regulate Mdm2, but not p21 or Noxa, expression. (a) HepG2 cells were transfected with plasmid carrying the luciferase reporter gene controlled by the Mdm2, p21^WAF1 (p21), or Noxa promoter alone or along with Abl^PP constructs. After 24 h, cells were treated or not with HGF for 24 h, and luciferase activity was assayed on cell lysates. (b) HepG2 cells stably transfected with Abl shRNA or control plasmid were serum starved, then stimulated with HGF, and Mdm2 protein levels were analyzed by immunoblotting. (c) HGF-induced Mdm2 upregulation is impaired in HepG2 cells transfected with p53 shRNA plasmid. (d) Imatinib impairs HGF-induced upregulation of Mdm2 mRNA levels in HepG2 cells. Cells were serum starved for 3 h and then stimulated with HGF in the presence or not of Imatinib (10 μM). Total RNA was extracted at the indicated times with Trizol and reverse transcription was performed. Levels of Mdm2 mRNA were measured by quantitative real-time PCR (RU, relative unit). (e) HepG2 cells transiently transfected with dominant negative (Abl^Kin−) or constitutive active (Abl^PP) forms of c-Abl were serum starved, stimulated with HGF as in (c) and Mdm2 protein levels were analyzed by immunoblotting. (f) Upregulation of Mdm2 by HGF does not occur in p53-null Hep3B cells. (g) Constitutive active Abl^PP does not upregulate Mdm2 levels in the absence of p53