Regulation of β-Catenin Levels and Localization by Overexpression of Plakoglobin and Inhibition of the Ubiquitin-Proteasome System

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Abstract
β-Catenin and plakoglobin (γ-catenin) are closely related molecules of the armadillo family of proteins. They are localized at the submembrane plaques of cell–cell adherens junctions where they form independent complexes with classical cadherins and α-catenin to establish the link with the actin cytoskeleton. Plakoglobin is also found in a complex with desmosomal cadherins and is involved in anchoring intermediate filaments to desmosomal plaques. In addition to their role in junctional assembly, β-catenin has been shown to play an essential role in signal transduction by the Wnt pathway that results in its translocation into the nucleus. To study the relationship between plakoglobin expression and the level of β-catenin, and the localization of these proteins in the same cell, we employed two different tumor cell lines that express N-cadherin, and α- and β-catenin, but no plakoglobin or desmosomal components. Individual clones expressing various levels of plakoglobin were established by stable transfection. Plakoglobin overexpression resulted in a dose-dependent decrease in the level of β-catenin in each clone. Induction of plakoglobin expression increased the turnover of β-catenin without affecting RNA levels, suggesting posttranslational regulation of β-catenin. In plakoglobin overexpressing cells, both β-catenin and plakoglobin were localized at cell– cell junctions. Stable transfection of mutant plakoglobin molecules showed that deletion of the N-cadherin binding domain, but not the α-catenin binding domain, abolished β-catenin downregulation. Inhibition of the ubiquitin-proteasome pathway in plakoglobin overexpressing cells blocked the decrease in β-catenin levels and resulted in accumulation of both β-catenin and plakoglobin in the nucleus. These results suggest that (a) plakoglobin substitutes effectively with β-catenin for association with N-cadherin in adherens junctions, (b) extrajunctional β-catenin is rapidly degraded by the proteasome-ubiquitin system but, (c) excess β-catenin and plakoglobin translocate into the nucleus.