Fig. 4. Slug represses the activity of the mouse E-cadherin promoter both in transient and stable MDCK transfectants. (A) MDCK cells were transiently cotransfected with 200 ng of the -178 wild-type E-cadherin promoter construct fused to the luciferase reporter gene in the presence of the indicated amounts of pcDNA3 (Mock), pcDNA3-Slug or pcDNA3-Snail vectors. Luciferase and renilla activities were determined 24 hours after transfection. The activity of the promoter is expressed relative to that obtained in the mock-transfected cells. Results represent the mean±s.d. of at least two independent results. (B) The activity of the -178 wild-type E-cadherin promoter is strongly reduced or completely silenced in Slug- and Snail-expressing cells. MDCK-mock, two independent MDCK-Slug clones and MDCK-Snail cells were transiently transfected with the -178 wild-type (white bars) or mE-pal (grey bars) E-cadherin constructs fused to the CAT reporter gene. Luciferase and CAT activities were determined 24 hours after transfection. The activity of the promoter constructs is represented relative to that of the -178 wt construct detected in the mock-transfected clone. Results represent the mean±s.d. of two independent experiments. The relative levels of activity for both constructs in each cell line are also indicated at the bottom. Slu1I represents one subclone isolated from an original Slu1 clone; Slu3 represents an independently isolated clone.

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