Reconstitution of the Cytoplasmic Regulation of the Wnt Signaling Pathway Using Xenopus Egg Extracts
The regulation of β-catenin turnover is the central mechanism governing activation of the Wnt signaling pathway. All components of the pathway are present in the early embryo of Xenopus laevis, and Xenopus egg extracts have been used to recapitulate complex biological reactions such as microtubule dynamics, DNA replication, chromatin assembly, and phases of the cell cycle. Herein, we describe a biochemical method for analyzing β-catenin degradation using radiolabeled and luciferase-fusion proteins in Xenopus egg extracts. We show that in such a biochemical system, cytoplasmic β-catenin degradation is regulated by soluble components of the Wnt pathway as well as small molecules.
Key wordsXenopus laevis Ubiquitin Proteasome Egg extract Axin β-Catenin destruction complex
We thank Laurie Lee for critical reading of the manuscript. ASH is supported by a fellowship from the NIH Molecular Endocrinology Training Program. EL is supported by the NIH grants R01GM081635 and R01GM103926.
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