A novel functional mechanism of Rac1 GTPase mediated regulation of Wnt signalling pathway
Aberrant expression of Rac1 GTPase (Rac1) occurs in many tumors while its splice variant, Rac1b, is specifically expressed in breast and colon tumors, and shows only limited functional similarities to Rac1. We recently discovered a novel cross-talk between Rac1 and the canonical Wnt signaling pathway in cancer. We showed that active Rac1 stimulates the transcriptional co-activator function of ß-catenin, resulting in enhanced activation of Wnt signaling. The polybasic region of Rac1/Rac1b acts as a nuclear localization signal and mutation of this region prevents nuclear translocation of Rac1 alongwith abolishing its ability for Wnt activation. This study was aimed at characterization of the role of Rac1b in the regulation of Wnt pathway and investigation of functional mechanisms underlying Rac1 mediated modulation of ß-catenin/TCF complex activation in cancer. In co-immunoprecipitation studies, both Rac1b and Rac1 functionally interact with Dishevelled-3 (Dvl3) or ß-catenin to mediate synergistic activation of wnt target genes and GSK-3ß is able to block this activation. We observed that physical association of Rac1b/Rac1 with Dvl3, ß-catenin and TCF-4 predominantly occurres in the nucleus. We next investigated whether Rac1 is present in the transcription factor complex comprised of ß-catenin and TCF-4 at the promoters of Wnt target genes. To address this, we performed chromatin immunoprecipitation (ChIP) assays and show for the first time, that Rac1 indeed associates with Wnt responsive promoters in vivo. The association of Rac1 with TCF binding elements was also confirmed in vitro via electrophoretic mobility shift assay (EMSA). Furthermore, by quantitative RT-PCR analysis, active Rac1 expression was observed to cause increased transcription of Wnt target genes, cyclin D1 and c-myc. These findings provide exciting insights into the novel functional mechanism underlying regulation of canonical Wnt signaling in cancer.