Reciprocal Negative Regulation between the Guanine Nucleotide Exchange Factor C3G and β-Catenin

Abstract

The guanine nucleotide exchange factor C3G (RAPGEF1) regulates proliferation, migration, and differentiation of cells and is essential for mammalian embryonic development. The molecular effectors of C3G dependent functions are poorly understood. Here we report that C3G functions as a negative regulator of β-catenin, a major player in pathways known to be deregulated in human cancers. In mammalian cells, C3G is present in a complex with cellular β-catenin. The proline rich Crk binding region of C3G and residues 90-525 of β-catenin are sufficient for the interaction. Knockdown of cellular C3G stimulated, and its overexpression repressed, β-catenin/TCF transcription activity. C3G acts by destabilizing β-catenin protein and inhibiting its nuclear accumulation. Nuclear extracts of C3G overexpressing cells showed reduced binding to TCF consensus oligos. C3G exerts its effects independent of its function as an exchange factor. It also inhibits stability and activity of an N-terminal deletion construct of β-catenin that is not subject to GSK3β dependent phosphorylation, suggesting that C3G exerts its effect independent of GSK3β. β-catenin repression by C3G was not significantly altered in the presence of proteasome inhibitors, MG132 or lactacystin, suggesting that alternate mechanisms are engaged by C3G to cause β-catenin turnover. C3G expression represses β-catenin target gene expression, and stable clones of MCF-7 breast cancer cells expressing C3G showed reduced migration. Activation of cellular β-catenin or expression of constitutively active β-catenin resulted in reduced C3G expression, indicating that C3G gene expression is negatively regulated by β-catenin. Our results identify a novel property of C3G in functioning as a negative regulator of β-catenin signaling by promoting its degradation. In addition, we show that β-catenin inhibits C3G expression, forming a feedback loop. © The Author(s) 2013.