Catenin, beta, phosphorylated (Ser33)

Cell adhesion is important during development, as well as in sorting of cells, induction of cellular morphogenesis and maintenance of tissue integrity.1-3 Ca2+-dependent adhesion of cells involves a multifunctional family of transmembrane glycoproteins termed cadherins that regulate homophilic interactions in cells. These interactions initiate a cascade of events that leads to the structural and functional reorganizations of cells. Cadherins function is mediated by both specific binding of extracellular domains at the cell surface and interaction with components of the cytoplasm. These components include alpha-, beta- and gamma-catenin (plakoglobin) that bind to the cytoplasmic domain of cadherins.4 Beta-catenin (92-97kD) shares 70% sequence identity to a protein encoded by Drosophila armadillo, a segment polarity gene.5-8 Beta-catenin binds to a diverse set of proteins including the presenilins, epidermal growth factor receptor (EGF-R), the actin-binding protein fascin and the transcription factor Teashirt.9-10 Beta-catenin is composed of a series of proteinprotein interaction motifs that allow it to function as a scaffold. The N-terminus domain, containing the binding site for alpha-catenin and the phosphorylation sites, is recognized by GSK3Beta. The C-terminus contains the transcriptional activation domain and the binding site for Teashirt.9-10 Beta-catenin translocates into the nucleus, where it complexes with transcription factors of the LEF-1 family and thus regulates the expression of specific genes. By playing such a dual function, i.e. a structural role in cell junctions and a regulatory role in the nucleus, beta-catenin can transduce changes in cell adhesion and junction formation to control transmembrane signaling and gene expression.1, 11 Beta-catenin-mediated signaling depends on its accumulation and subsequent translocation into the nucleus. The level of beta-catenin is regulated by its association with the tumor suppressor molecule Adenomatous Polyposis Coli (APC), axin, and GSK3Beta. In this complex, GSK3Beta phosphorylates beta-catenin at multiple serine or threonine residues present in the amino terminal region of beta-catenin,1 thereby marking beta-catenin for degradation by the proteasome pathway. The significance of beta-catenin phosphorylation for its stability is most clearly manifested in several types of human cancers. In cells expressing mutant APC, common in human colon cancer and melanoma, beta-catenin accumulates due to the failure of its degradation.1 Moreover, a single amino acid mutation at one of the four critical serine or threonine residues (Ser33, -37 and -45 and Thr-41) at the amino terminal region in the consensus GSK3Beta phosphorylation site, results in deregulated accumulation of beta-catenin and thereby increased signaling through the TCF/beta-catenin transcriptional complex, which contributes to tumorigenesis. Immunofluorescence studies indicate that phosphorylated beta-catenin accumulates in the nuclei of Madin-Darby Canine Kidney (MDCK) and BCAP (human breast cancer) cells and is transiently associated with adherens junctions.12 It was recently shown that phosphorylation of Ser45 and Thr41 precedes the phosphorylation of Ser33 and Ser37. Phosphorylation is mediated by casein kinase IAlph (CKIAlpha), an axin associated kinase.13 Monoclonal antibodies reacting specifically with beta-catenin phosphorylated at Ser33 are essential tools for defining the role of the phosphorylated form in the distributions, interactions, and regulation/deregulation of beta-catenin in signal transduction. Applications: Suitable for use in ELISA and Western Blot. Other applications not tested. Recommended Dilution: ELISA: 5ug/ml. Western Blot: 5ug/ml. Detects a band of ~ 94kD. Optimal dilutions to be determined by the researcher. Cellular Localization: Cytoplasmic when it is unstabilized (high level of phosphorylation) or bound to CDH1. Translocates to the nucleus when it is stabilized (low level of phosphorylation). Storage and Stability: May be stored at 4 degrees C for short-term only. For long-term storage and to avoid repeated freezing and thawing, aliquot Store at -20 degrees C. Aliquots are stable for at least 12 months at -20 degrees C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Further dilutions can be made in assay buffer.