title: A Simple Method to Assess Abundance of the β-Catenin Signaling Pool in Cells.
abstract: β-catenin (CTNNB1) is a dual-function cell-cell adhesion/transcriptional co-activator protein and an essential transducer of canonical Wnt signals. Although a number of established techniques and reagents are available to quantify the nuclear signaling activity of β-catenin (e.g., TCF-dependent reporter assays, nuclear accumulation of β-catenin, and generation of N-terminally hypophosphorylated β-catenin), there are cell-type and context-dependent limitations of these methods. Since the posttranscriptional stabilization of β-catenin outside of the cadherin complex appears universally required for β-catenin signaling, the following method allows for simple assessment of the cadherin-free fraction of β-catenin in cells, using a GST-tagged form of ICAT (Inhibitor of β-Catenin and Tcf) as an affinity matrix. This method is more sensitive and quantitative than immunofluorescence and may be useful in studies that implicate TCF-independent signaling events.
authors: Department of Pulmonary Medicine, Feinberg School of Medicine, Northwestern University, 240 East Huron St., McGaw Pavilion, M-323, Chicago, IL, 60611, USA.
authors: Annette S
authors: AS
authors: Flozak
authors: Department of Pulmonary Medicine, Feinberg School of Medicine, Northwestern University, 240 East Huron St., McGaw Pavilion, M-323, Chicago, IL, 60611, USA. a-lam4@northwestern.edu.
authors: Anna P
authors: AP
authors: Lam
authors: Department of Pulmonary Medicine, Feinberg School of Medicine, Northwestern University, 240 East Huron St., McGaw Pavilion, M-323, Chicago, IL, 60611, USA. c-gottardi@northwestern.edu.|Department of Cellular and Molecular Biology, Feinberg School of Medicine, Northwestern University, 303 East Chicago Ave., Chicago, IL, 60611, USA. c-gottardi@northwestern.edu.
authors: Cara J
authors: CJ
authors: Gottardi
mesh_terms: D015820:Cadherins / Q000737:chemistry / Q000235:genetics; D002448:Cell Adhesion / Q000235:genetics; D002467:Cell Nucleus / Q000737:chemistry*; D008967:Molecular Biology / Q000379:methods*; D014158:Transcription, Genetic; D051153:Wnt Proteins / Q000737:chemistry / Q000302:isolation & purification; D060449:Wnt Signaling Pathway; D051176:beta Catenin / Q000235:genetics / Q000302:isolation & purification*
chemical_list: D015820:Cadherins; D051153:Wnt Proteins; D051176:beta Catenin
keywords: Affinity precipitation; Fusion proteins; GST (glutathione S transferase); ICAT; TCF (T-cell factor); Western blot; Wnt; β-catenin