Inappropriate activation of downstream target genes by the oncoprotein beta-catenin is implicated in development of numerous human cancers. beta-catenin and its fruitfly counterpart Armadillo act as a coactivator in the canonical Wnt/Wingless pathway by binding to Tcf/Lef transcription factors. Here we report a conserved nuclear protein, named Chibby, which was identified in a screen for proteins that directly interact with the C-terminal region of beta-catenin. In mammalian cultured cells we demonstrate that Chibby inhibits beta-catenin-mediated transcriptional activation by competing with Lef-1 to bind to beta-catenin. Inhibition of Drosophila Chibby by RNA interference results in segment polarity defects that mimick a wingless gain-of-function phenotype, and overexpression of the wingless target genes engrailed and Ultrabithorax. In addition, epistasis experiments indicate that chibby acts downstream of wingless and upstream of armadillo.

beta-catenin plays essential roles in cell adhesion and Wnt signaling, while deregulation of beta-catenin is associated with multiple diseases including cancers. Here, we report the crystal structures of full-length zebrafish beta-catenin and a human beta-catenin fragment that contains both the armadillo repeat and the C-terminal domains. Our structures reveal that the N-terminal region of the C-terminal domain, a key component of the C-terminal transactivation domain, forms a long alpha helix that packs on the C-terminal end of the armadillo repeat domain, and thus forms part of the beta-catenin superhelical core. The existence of this helix redefines our view of interactions of beta-catenin with some of its critical partners, including ICAT and Chibby, which may form extensive interactions with this C-terminal domain alpha helix. Our crystallographic and NMR studies also suggest that the unstructured N-terminal and C-terminal tails interact with the ordered armadillo repeat domain in a dynamic and variable manner.

Transcriptional coactivators play a crucial role in gene expression by communicating between regulatory factors and the basal transcription machinery. The coactivator multiprotein bridging factor 1 (MBF1) was originally identified as a bridging molecule that connects the Drosophila nuclear receptor FTZ-F1 and TATAbinding protein (TBP). The MBF1 sequence is highly conserved across species from Saccharomyces cerevisiae to human. Here we provide evidence acquired in vitro and in vivo that yeast MBF1 mediates GCN4-dependent transcriptional activation by bridging the DNA-binding region of GCN4 and TBP. These findings indicate that the coactivator MBF1 functions by recruiting TBP to promoters where DNA-binding regulators are bound.Studies on transcriptional control in eukaryotes have revealed many regulatory proteins that bind to control elements on DNA (17,25). Typical regulators such as GAL4 and GCN4 consist of two domains (29), a DNA-binding domain that binds to a control element in a sequence-specific fashion and a transcriptional activation domain that somehow stimulates the basal transcription machinery. A variety of observations have led to the proposal that some transactivation domains may facilitate binding of TATA-binding protein (TBP) to a promoter (32, 37 see reference 35 for a review). In many cases, a member of another class of transcription factors, termed coactivators, adapters, or mediators, is necessary to connect a regulatory protein and a component of the basal transcription machinery (see references 23 and 30 for reviews). Recent studies demonstrated the importance of these non-DNA-binding transcription factors in gene expression (1,4,5,11,14,19,20,27,31).In in vitro transcription studies, it was found that an insect coactivator, multiprotein bridging factor 1 (MBF1), can recruit TBP to a promoter carrying the FTZ-F1-binding site by interconnecting 34). A homology search of the databases showed that the MBF1 sequence is highly conserved across species from Saccharomyces cerevisiae to human (34). A key question concerning MBF1, and its close relatives, is its in vivo function. To address this issue, we have started genetic studies of MBF1. We report here that MBF1 serves as a coactivator of GCN4 in the yeast S. cerevisiae. MATERIALS AND METHODSYeast strains and plasmids. The GCN4 disruptant used in this study was KY803 (trp1-⌬1 ura3-52 leu2-P1 gcn4-⌬1) (13). Wild-type strain KT130 (trp1-⌬1 ura3-52 leu2-P1) was constructed by homologous recombination after introduction of GCN4 genomic DNA into KY803. KT130 was used to generate the ⌬mbf1 strain by replacement of the sequence encoding amino acid residues 64 to 131 of yeast MBF1 (yMBF1) (34) with a LEU2 selectable marker.To construct pMBF1, the 3-kb EcoRI genomic fragment encompassing the entire MBF1 regulatory and coding regions was subcloned into YCplac33 (9). YCp88 (13), a centromeric vector containing the constitutive DED1 promoter, was used to express GCN4 or its mutants in KY803. pYCGCN4, pYCGCN4⌬bZIP, and pYCGCN4bZIP carry the entire coding regi...

Chibby (Cby) is a conserved component of the Wnt–β-catenin pathway. Cby physically interacts with β-catenin to repress its activation of transcription. To elucidate the function of Cby in vertebrates, we generated Cby−/− mice and found that after 2–3 d of weight loss, the majority of mice die before or around weaning. All Cby−/− mice develop rhinitis and sinusitis. When challenged with Pseudomonas aeruginosa isolates, Cby−/− mice are unable to clear the bacteria from the nasal cavity. Notably, Cby−/− mice exhibit a complete absence of mucociliary transport caused by a marked paucity of motile cilia in the nasal epithelium. Moreover, ultrastructural experiments reveal impaired basal body docking to the apical surface of multiciliated cells. In support of these phenotypes, endogenous Cby protein is localized at the base of cilia. As the phenotypes of Cby−/− mice bear striking similarities to primary ciliary dyskinesia, Cby−/− mice may prove to be a useful model for this condition.