Poster presentation, P173

Official XXIst International Pigment Cell Conference website - 21-24 Sept 2011, Bordeaux - France | updated: September 04 2011

ICAT overexpression increases motility of melanoma cells in culture

ICAT (Inhibitor of β-catenin and TCF) is a small acidic protein of 81 amino-acids that negatively regulates β-catenin co-transcriptional activity by competing with TCF/LEF factors for binding to armadillo repeats of β-catenin. Given that β-catenin is involved in proliferation of melanoma cells through the regulation of M-MITF and BRN2 transcription factors, we have evaluated the incidence of ICAT overexpression on melanoma cells in culture. At the cellular level, stable overexpression of wild-type ICAT-GFP in Mel501 melanoma cells did not affect their proliferation. However, in a wound scratch assay, ! Mel501 cells overexpressing ICAT-GFP were found to migrate more rapidly than GFP-positive control cells. At the molecular level, mRNA transcripts of M-MITF and BRN-2 were reduced. Luciferase assays with the M-MITF and BRN-2 promoters showed that ICAT competes with LEF1 for binding to β-catenin thus reducing promoter activities. Site-directed mutagenesis of ICAT residues revealed that Asp66 and Glu75 in the carboxy terminal domain are critical for competition with LEF1. However, their absence does not prevent the formation of a stable ICAT/β-catenin protein complex, whereas Tyr15, Lys19 and Val22 residues in the amino terminal domain of ICAT are essential for its formation. These results indicate that ICAT strongly binds to β-catenin through its N-terminal end and mediates competition with LEF1 through its C-terminal region. Down-regulation of β-catenin targets could increase melanoma cells motility in vitro by altering expression of focal adhesion molecules.

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