In adherens junctions, alpha-catenin links the cadherin-beta-catenin complex to the actin-based cytoskeleton. alpha-catenin is a homodimer in solution, but forms a 1:1 heterodimer with beta-catenin. ...The crystal structure of the alpha-catenin dimerization domain, residues 82-279, shows that alpha-catenin dimerizes through formation of a four-helix bundle in which two antiparallel helices are contributed by each protomer. A slightly larger fragment, comprising residues 57-264, binds to beta-catenin. A chimera consisting of the alpha-catenin-binding region of beta-catenin linked to the amino terminus of alpha-catenin 57-264 behaves as a monomer in solution, as expected, since beta-catenin binding disrupts the alpha-catenin dimer. The crystal structure of this chimera reveals the interaction between alpha- and beta-catenin, and provides a basis for understanding adherens junction assembly.
Spatial and functional organization of cells in tissues is determined by cell-cell adhesion, thought to be initiated through trans-interactions between extracellular domains of the cadherin family of ...adhesion proteins, and strengthened by linkage to the actin cytoskeleton. Prevailing dogma is that cadherins are linked to the actin cytoskeleton through β-catenin and α-catenin, although the quaternary complex has never been demonstrated. We test this hypothesis and find that α-catenin does not interact with actin filaments and the E-cadherin-β-catenin complex simultaneously, even in the presence of the actin binding proteins vinculin and α-actinin, either in solution or on isolated cadherin-containing membranes. Direct analysis in polarized cells shows that mobilities of E-cadherin, β-catenin, and α-catenin are similar, regardless of the dynamic state of actin assembly, whereas actin and several actin binding proteins have higher mobilities. These results suggest that the linkage between the cadherin-catenin complex and actin filaments is more dynamic than previously appreciated.
In the canonical Wnt signaling pathway, β-catenin activates target genes through its interactions with Tcf/Lef-family transcription factors and additional transcriptional coactivators. The crystal ...structure of ICAT, an inhibitor of β-catenin-mediated transcription, bound to the armadillo repeat domain of β-catenin, has been determined. ICAT contains an N-terminal helilical domain that binds to repeats 11 and 12 of β-catenin, and an extended C-terminal region that binds to repeats 5–10 in a manner similar to that of Tcfs and other β-catenin ligands. Full-length ICAT dissociates complexes of β-catenin, Lef-1, and the transcriptional coactivator p300, whereas the helical domain alone selectively blocks binding to p300. The C-terminal armadillo repeats of β-catenin may be an attractive target for compounds designed to disrupt aberrant β-catenin-mediated transcription associated with various cancers.
β-catenin is essential for cadherin-based cell adhesion and Wnt/Wingless growth factor signaling. In these roles, it binds to cadherins, Tcf-family transcription factors, and the tumor suppressor ...gene product Adenomatous Polyposis Coli (APC). A core region of β-catenin, composed of 12 copies of a 42 amino acid sequence motif known as an armadillo repeat, mediates these interactions. The three-dimensional structure of a protease-resistant fragment of β-catenin containing the armadillo repeat region has been determined. The 12 repeats form a superhelix of helices that features a long, positively charged groove. Although unrelated in sequence, the β-catenin binding regions of cadherins, Tcfs, and APC are acidic and are proposed to interact with this groove.
C-type (Ca(2+)-dependent) animal lectins such as mannose-binding proteins mediate many cell-surface carbohydrate-recognition events. The crystal structure at 1.7 A resolution of the ...carbohydrate-recognition domain of rat mannose-binding protein complexed with an oligomannose asparaginyl-oligosaccharide reveals that Ca2+ forms coordination bonds with the carbohydrate ligand. Carbohydrate specificity is determined by a network of coordination and hydrogen bonds that stabilizes the ternary complex of protein, Ca2+ and sugar. Two branches of the oligosaccharide crosslink neighbouring carbohydrate-recognition domains in the crystal, enabling multivalent binding to a single oligosaccharide chain to be visualized directly.
Epithelial cell-cell junctions, organized by adhesion proteins and the underlying actin cytoskeleton, are considered to be stable structures maintaining the structural integrity of tissues. Contrary ...to the idea that α-catenin links the adhesion protein E-cadherin through β-catenin to the actin cytoskeleton, in the accompanying paper we report that α-catenin does not bind simultaneously to both E-cadherin-β-catenin and actin filaments. Here we demonstrate that α-catenin exists as a monomer or a homodimer with different binding properties. Monomeric α-catenin binds more strongly to E-cadherin-β-catenin, whereas the dimer preferentially binds actin filaments. Different molecular conformations are associated with these different binding states, indicating that α-catenin is an allosteric protein. Significantly, α-catenin directly regulates actin-filament organization by suppressing Arp2/3-mediated actin polymerization, likely by competing with the Arp2/3 complex for binding to actin filaments. These results indicate a new role for α-catenin in local regulation of actin assembly and organization at sites of cadherin-mediated cell-cell adhesion.
Wnt growth factors mediate cell fate determination during embryogenesis and in the renewal of tissues in the adult. Wnts act by stabilizing cellular levels of the transcriptional coactivator ...beta-catenin, which forms complexes with sequence-specific DNA-binding Tcf/Lef transcription factors. In the absence of nuclear beta-catenin, Tcf/Lefs act as transcriptional repressors by binding to Groucho/TLE proteins. The molecular basis of the switch from transcriptional repression to activation during Wnt signaling has not been clear, in particular whether factors other than beta-catenin are required to disrupt the interaction between Groucho/TLE and Tcf/Lef. Using highly purified proteins, we demonstrate that beta-catenin displaces Groucho/TLE from Tcf/Lef by binding to a previously unidentified second, low-affinity binding site on Lef-1 that includes sequences just N-terminal to the DNA-binding domain, and that overlaps the Groucho/TLE-binding site.
Celotno besedilo
Dostopno za:
DOBA, IJS, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
N-ethylmaleimide–sensitive fusion protein (NSF) is a cytosolic ATPase required for many intracellular vesicle fusion reactions. NSF consists of an amino-terminal region that interacts with other ...components of the vesicle trafficking machinery, followed by two homologous ATP-binding cassettes, designated D1 and D2, that possess essential ATPase and hexamerization activities, respectively. The crystal structure of D2 bound to Mg
2PLUSPUSSIGN-AMPPNP has been determined at 1.75 Å resolution. The structure consists of a nucleotide-binding and a helical domain, and it is unexpectedly similar to the first two domains of the clamp-loading subunit δ′ of
E. coli DNA polymerase III. The structure suggests several regions responsible for coupling of ATP hydrolysis to structural changes in full-length NSF.
Cadherins are single pass transmembrane proteins that mediate Ca(2+)-dependent homophilic cell-cell adhesion by linking the cytoskeletons of adjacent cells. In adherens junctions, the cytoplasmic ...domain of cadherins bind to beta-catenin, which in turn binds to the actin-associated protein alpha-catenin. The physical properties of the E-cadherin cytoplasmic domain and its interactions with beta-catenin have been investigated. Proteolytic sensitivity, tryptophan fluorescence, circular dichroism, and (1)H NMR measurements indicate that murine E-cadherin cytoplasmic domain is unstructured. Upon binding to beta-catenin, the domain becomes resistant to proteolysis, suggesting that it structures upon binding. Cadherin-beta-catenin complex stability is modestly dependent on ionic strength, indicating that, contrary to previous proposals, the interaction is not dominated by electrostatics. Comparison of 18 cadherin sequences indicates that their cytoplasmic domains are unlikely to be structured in isolation. This analysis also reveals the presence of PEST sequences, motifs associated with ubiquitin/proteosome degradation, that overlap the previously identified beta-catenin-binding site. It is proposed that binding of cadherins to beta-catenin prevents recognition of degradation signals that are exposed in the unstructured cadherin cytoplasmic domain, favoring a cell surface population of catenin-bound cadherins capable of participating in cell adhesion.
Axin and the adenomatous polyposis coli (APC) tumor suppressor protein are components of the Wnt/Wingless growth factor signaling pathway. In the absence of Wnt signal, Axin and APC regulate ...cytoplasmic levels of the proto‐oncogene β‐catenin through the formation of a large complex containing these three proteins, glycogen synthase kinase 3β (GSK3β) and several other proteins. Both Axin and APC are known to be critical for β‐catenin regulation, and truncations in APC that eliminate the Axin‐binding site result in human cancers. A protease‐resistant domain of Axin that contains the APC‐binding site is a member of the regulators of G‐protein signaling (RGS) superfamily. The crystal structures of this domain alone and in complex with an Axin‐binding sequence from APC reveal that the Axin–APC interaction occurs at a conserved groove on a face of the protein that is distinct from the G‐protein interface of classical RGS proteins. The molecular interactions observed in the Axin–APC complex provide a rationale for the evolutionary conservation seen in both proteins.
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