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. Author manuscript; available in PMC: 2013 Oct 1.
Published in final edited form as: Nat Struct Mol Biol. 2013 Feb 24;20(4):447–453. doi: 10.1038/nsmb.2505

Figure 4. Disruption of the SIX1–EYA Transcriptional Complex Inhibits SIX1-mediated TGF-β Signaling and Characteristics of EMT.

Figure 4

(A) MCF7-V17E and delE133 cells fail to induce p-SMAD3 protein to the same levels as SIX1 WT, as demonstrated by Western blot performed on whole cell lysates. β–actin was used as a loading control.

(B) MCF7-V17E and delE133 cells fail to induce TGF-β responsive 3TP-luciferase reporter activity to levels observed in SIX1 WT cells. Values represent biological replicates with SEM error bars (n=3) and are normalized to renilla luciferase levels.

(C) MCF7-V17E and delE133 cells fail to reduce cytokeratin 18 levels and induce fibronectin levels, as is observed in MCF7-SIX1 WT cells. Expression of cytokeratin 18 and fibronectin were determined using qRT-PCR.

(D) Expression of SIX1 V17E and delE133 in MCF7 cells fails to decrease the adhesion of the cells to the matrix proteins Collagen I, Collagen IV and Laminin, whereas expression of SIX1 WT leads to decreased adhesion to these matrices. Relative adherence measured by crystal violet staining. P-values represent statistical analysis using a paired t-test.