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. Author manuscript; available in PMC: 2012 Jun 29.
Published in final edited form as: Cancer. 2010 Dec 29;117(11):10.1002/cncr.25803. doi: 10.1002/cncr.25803

Figure 6. EGF downregulates NGAL expression by inhibition of NF-κB binding activity.

Figure 6

Figure 6

Figure 6

A. AsPC-1 cells were cotransfected with pGL3-luc/NGAL [−900] or pGL3-luc/NGAL [ NF-κB- mut] and pGL4.74 [hRluc/TK] (control for transfection efficiency) for 24 hours after being treated with 25 ng/ml EGF. Cell lysate was then used for the luciferase activity assay. * represents p<0.05. B. AsPC-1 cells were infected with lentivirus carrying shE-cadherin or its controls for 16 hours and followed by cotransfection with pGL3-luc/NF-κB and pGL4.74[hRluc/TK] for 24 hours. Thereafter, the cell lysate was used for a luciferase activity assay. * represents p<0.05. C. Nuclear extract from AsPC-1 cells treated with EGF for 24 hours was analyzed by EMSA as described in the methods section. The p.c. stands for positive control (nuclear extracts from KBM-5 cells stimulated with TNF-α). D. Model illustrating EGF-mediated downregulation of NGAL. EGF activates ERK signaling by activating EGFR, leading to increased Zeb1 expression and reduced E-cadherin expression, which results in decreased NGAL expression by blocking NF-κB binding to the NGAL promoter.