Abstract
Backgroundβ-catenin is a key mediator of the canonical Wnt pathway as it associates with members of the T-cell factor (TCF) family at Wnt-responsive promoters to drive the transcription of Wnt target genes. Recently, we showed that Rac1 GTPase synergizes with β-catenin to increase the activity of a TCF-responsive reporter. This synergy was dependent on the nuclear presence of Rac1, since inhibition of its nuclear localization effectively abolished the stimulatory effect of Rac1 on TCF-responsive reporter activity. We hypothesised that Rac1 plays a direct role in enhancing the transcription of endogenous Wnt target genes by modulating the β-catenin/TCF transcription factor complex.ResultsWe employed chromatin immunoprecipitation studies to demonstrate that Rac1 associates with the β-catenin/TCF complex at Wnt-responsive promoters of target genes. This association served to facilitate transcription, since overexpression of active Rac1 augmented Wnt target gene activation, whereas depletion of endogenous Rac1 by RNA interference abrogated this effect. In addition, the Rac1-specific exchange factor, Tiam1, potentiated the stimulatory effects of Rac1 on the canonical Wnt pathway. Tiam1 promoted the formation of a complex containing Rac1 and β-catenin. Furthermore, endogenous Tiam1 associated with endogenous β-catenin, and this interaction was enhanced in response to Wnt3a stimulation. Intriguingly, Tiam1 was recruited to Wnt-responsive promoters upon Wnt3a stimulation, whereas Rac1 was tethered to TCF binding elements in a Wnt-independent manner.ConclusionTaken together, our results suggest that Rac1 and the Rac1-specific activator Tiam1 are components of transcriptionally active β-catenin/TCF complexes at Wnt-responsive promoters, and the presence of Rac1 and Tiam1 within these complexes serves to enhance target gene transcription. Our results demonstrate a novel functional mechanism underlying the cross-talk between Rac1 and the canonical Wnt signalling pathway.
Highlights
Rac1 GTPase is a member of the Rho family of small GTPases, which play critical roles in the regulation of various cellular processes that include reorganization of the actin cytoskeleton, cell-cycle progression, intercellular adhesion, and gene expression [reviewed in [1]]
Rho family proteins act as molecular switches that cycle between an inactive GDP-bound state and an active GTP-bound state. This cycling is regulated by proteins such as guanine nucleotide exchange factors (GEFs) that activate Rho GTPases by accelerating GDP to GTP exchange, and GTPase activating proteins (GAPs) that deactivate Rho GTPases by increasing their intrinsic rate of GTP hydrolysis
We have previously shown that active GTP-bound Rac1, but not the inactive GDP-bound form, synergizes with βcatenin to induce the activity of a T-cell factor (TCF)-responsive reporter (TOPFlash) [34]
Summary
Rac GTPase is a member of the Rho family of small GTPases, which play critical roles in the regulation of various cellular processes that include reorganization of the actin cytoskeleton, cell-cycle progression, intercellular adhesion, and gene expression [reviewed in [1]]. Rho family proteins act as molecular switches that cycle between an inactive GDP-bound state and an active GTP-bound state. This cycling is regulated by proteins such as guanine nucleotide exchange factors (GEFs) that activate Rho GTPases by accelerating GDP to GTP exchange, and GTPase activating proteins (GAPs) that deactivate Rho GTPases by increasing their intrinsic rate of GTP hydrolysis. Overexpression of the Rac1-specific GEF, T-cell lymphoma invasion and metastasis 1 (Tiam1), has been reported in highly invasive breast tumours [4] and colon carcinomas [5,6,7,8], and may contribute to elevated Rac signalling in these cancers. One mechanism by which dysregulated Rac signalling may promote tumourigenesis is by modulating the activities of various transcription factors, including nuclear factor-kappa B (NF-κB), activator protein-1 (AP-1), and signal transducer and activator of transcription 3 (STAT3), which regulate the transcription of genes involved in tumourigenic events such as cell proliferation, tumour angiogenesis, and cell survival [reviewed in [9]]
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