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. Author manuscript; available in PMC: 2012 Aug 15.
Published in final edited form as: Int J Cancer. 2010 Dec 9;129(4):799–809. doi: 10.1002/ijc.25750

Figure 5. Gal-4 mechanism.

Figure 5

(A). Effect of gal-4 transient transfection in ATRFLOX, HCT-116 and HT-29-2 cells. Cells were transiently transfected with either gal-4 plasmid (Gal-4) or vector plasmid (C). Cell lysates (10 μg each) prepared after 24 h were analyzed for expression of gal-4, cyclin D1, p21, p15 and β-actin. (B). Effect of gal-4 RNAi3 transfection in HT-29 cells. HT-29 cells were transfected either with gal-4 RNAi3 (RNAi3) or Stealth RNAi negative control (C) using electroporation and cell lysates at 96 hrs were analyzed for the expression of gal-4, p15, β-catenin, Phospho-p53 and β-actin. (C). Immunoprecipitation of gal-4 with Wnt signaling proteins. Antibodies against gal-4 (rabbit), β-catenin (rabbit), axin (rabbit) and APC (mouse), mouse IgG (m) and rabbit IgG (r) (controls) were used in the immunoprecipitation from HT-29 cell lysates. The IP's were probed with anti-mouse-gal-4 and anti-mouse-β-catenin antibodies. Prominent bands observed were 37 kDa gal-4 band and 92 kDa β-Catenin band. All the IPs could pull down both gal-4 and β-catenin. (D). Gal-4 effect on Wnt signaling pathway. HCT-116 cells transfected either with gal-4 plasmid (Gal-4) or vector plasmid (C) and cell lysates at 48 h were analyzed for β-catenin, Naked 1, Dvl2, Dvl3, GSK-3β, axin1, TCF1, TCF4 and β-actin. (E). Gal-4 effect on Wnt target genes. HCT-116 cells transfected either with gal-4 plasmid (Gal-4) or vector plasmid (C) for 48 h and cell lysates were analyzed for the expression of gal-4, c-Myc, LRP6, Ephrin B1 and β-actin.