Fig. 4.

Regulation of MT1–MMP and LAMC2 by ZEB1 in CRC cells with mutant APC. (A) ZEB1 activates MT1–MMP expression in CRC cells. (Left) ZEB1 activates MT1–MMP transcription. SW480 cells were transfected with 0.8 μg of the human MT1–MMP promoter along with 1.0 μg of empty expression vector or equal molar amounts of ZEB1 expression vector. (Center) ZEB1 activates endogenous MT1–MMP mRNA. SW480 cells transfected with a siRNA control or specific siRNAs for ZEB1, β-catenin, and TCF4 were assessed for relative levels of MT1–MMP mRNA by qRT-PCR. (Right) ZEB1 regulates endogenous MT1–MMP protein. SW480 cells were transiently transfected with 50 nM of a siRNA for ZEB1 or a control siRNA. Protein expression was assessed by Western blot with Abs for MT1–MMP (LEM2/15.8), ZEB1 (H-102), and α-tubulin (B5-1-2). (B) ZEB1 activates LAMC2 expression in CRC cells. (Left) ZEB1 activates LAMC2 transcription. Same as in A, but using both wild-type LAMC2 promoter (LAMC2) or a mutant version where two ZEB1 binding sites had been mutated (LAMC2-MUT) (Center) ZEB1 activates endogenous LAMC2 mRNA. Same as in A, but for LAMC2 mRNA. (Right) ZEB1 regulates endogenous LAMC2 protein. Same as in A, but a LAMC2 (D4B5) Ab was used.