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CLIC4 is a chloride ion channel protein that is upregulated and translocates to the nucleus under conditions of cell stress and induces growth arrest and cellular apoptosis upon translocation. CLIC4 is excluded from the nucleus in cancer cell lines and most human tumors, suggesting that regulation of CLIC4 translocation is an important component of tumor development. Despite nuclear residence, CLIC4 has not been shown to modify a signaling pathway. Here we show that TGF-β enhances CLIC4 expression and nuclear translocation in skin keratinocytes. Furthermore, CLIC4 upregulates TGF-β signaling in keratinocytes, analyzed by TGF-β dependent Smad phosphorylation, inhibition of DNA synthesis, activation of TGF-β dependent reporter activity and regulation of TGF-β target gene expression. A yeast two-hybrid screen using CLIC4 sequences as bait revealed Schnurri-2 as a CLIC4 binding protein. Schnurri-2 is a zinc finger protein and is a gene-specific transcription factor as well as an adaptor molecule in the TGF-β signaling pathway in Drosophila. TGF-β increases the expression of Schnurri-2 and enhances the association of CLIC4 and Schnurri-2 in keratinocytes. Our results suggest that Schnurri-2 and CLIC4 together modify the TGF-β signaling pathway and inhibition of CLIC4 nuclear translocation in tumors may be associated with defective TGF-β signaling commonly detected during malignant transformation.

98th AACR Annual Meeting-- Apr 14-18, 2007; Los Angeles, CA