Discuss the dual functions of beta catenin PDF

Preview

Summary

Discuss the dual-functions of Beta-catenin and why formation of cadherin mediated cell-cell junctions supresses the Wnt signalling pathway.β-catenin is an 88kDa protein which has multiple different functions dependent upon its cellular location. The protein can act as a transcription factor, promoti...

Description

Discuss the dual-functions of Beta-catenin and why formation of cadherin mediated cell-cell junctions supresses the Wnt signalling pathway. β-catenin is an 88kDa protein which has multiple different functions dependent upon its cellular location. The protein can act as a transcription factor, promoting the upregulation of genes necessary for cell proliferation and determining cellular polarity. This function is important in epithelial to mesenchymal transition (EMT), in which polarised immobile epithelial cells change their morphology to highly motile fibroblastoid mesenchymal cells. EMT is important for embryonic development but is also drives cancer invasion and metastases. β-catenin also functions as an important adaptor protein in cell-cell adherens junctions acting as a bridge between cadherins and the actin cytoskeleton. When β-catenin is present within these cell-cell junctions it cannot function as a transcription factor due to be sequestered in the junction and unable to move to the nucleus. β-catenin possess an N-terminal domain of around 130 amino acids. Within this domain resides four regulatory serine/threonine phosphorylation sites, these are important in regulating β-catenin as a transcription factor. The middle region of β-catenin consists of around 535 amino acids and contains 13 tri-helical armadillo repeats which twist into a rigid superhelix conformation. The C-terminal of the protein is the transactivational domain which recruits transcriptional co-activators necessary for β-catenins function to upregulate genes. For β-catenin to function as a transcription factor, the Wnt signalling pathway must be induced. Without this signalling pathway any free cytosolic β-catenin is targeted for degredation by the formation of a degredation complex. Free cytosolic β-catenin is recognised and bound by the scaffold molecules AXIN and APC. This binding creates a platform for the kinases casein kinase 1α (CK1α) and glycogen synthase kinase 3β (GSK3β) to phosphorylate β-catenin. AXIN acts to increase the activity of GSK3β on β-catenin, whilst APC stabilises the phosphorylated state of β-catenin, preventing its dephosphorylation my protein phosphotase 2A (PP2A). CK1α phosphorylates β-catenin at Ser45, priming β-catenin for subsequent phosphorylation by GSK3β at Thr41, Ser37 and Ser33. The phosphorylation of Serine’s 37 and 33 creates a binding site for the F-box protein β-Trcp, which is one of four subunits of the SCF ubiquitin ligase complex. β-catenin is subsequently ubiquitinated, targeting it for degredation by the 26S proteosome. This prevents β-catenin from translocating to the nucleus and upregulating its target genes. In the absence of the signalling pathway Wnt target genes are repressed by the DNA bound T-cell factor/lymphoid enhancer factor (TCF/LEF) family of proteins. Gene expression is repressed by TCF/LEF interacting with the repressor Groucho (TLE1 in humans), which promotes histone deacetylation and chromatin compaction. The Wnt signalling pathway is promoted by Wnt proteins which bind to frizzled (Fz) receptors. These seven transmembrane domain receptors show high topological homology with G-protein coupled receptors. Wnt binds to the N-terminal extracellular cysteine-rich domain of the Fz receptor. Binding initiates recruitment and phosphorylation of the co-receptor low-density-lipoprotein-receptor related protein 5/6 (LRP5/6). LRP 5 and 6 both possess 5 repeated PPPSPxS motifs where P represents proline, S represents serine/threonine and x is a variable residue. This motif is phosphorylated by GSK3 and CK1 with GSK3 phosphorylating PPPSP, priming the phosphorylation of xS by CK1. The phosphorylation provides a binding site for the scaffold protein AXIN. The recruitment of AXIN to the membrane prevents it from forming the β-catenin degredation complex mentioned above, allowing free cytosolic β-catenin to persist. The translocation of AXIN to the plasma membrane is not completely understood, however it is believed to involve the microtubule actin cross-linking factor 1 (MACF1). Binding of Wnt is also triggers activation of the activity of the cytoplasmic phosphoprotein Dishevelled (Dsh). Dsh acts to inhibit the phosphorylation of β-catenin by GSK3β, this increases the stability of β-catenin, allowing its accumulation in the cytoplasm. The stabilised β-catenin moves into the nucleus, where it can act as a transcription factor. The nuclear

accumulation of β-catenin results in a complex forming with TCF with β-catenin displacing Groucho removing its repressing activity on target genes. The forming of the complex results in recruitment of co-activators for gene expression. TCF proteins are high mobility group DNA-binding factors and bind to the consensus sequence CCTTTGWW called the Wnt responsive element (WRE) where W represents either Thymine of Adenine. Binding to the consensus sequence causes significant DNA bending that may alter local chromatin structure. The recruitment of co-activators such as BCL9 and Pygo in Drosphilia is essential for β-catenin-dependent transcription. Wnt target genes are diverse and cell and context specific with Wnt/β-catenin signalling regulating proliferation, fate specification and differentiation in numerous developmental stages and adult tissue homeostasis. Mutations in βcatenin have been linked to cancer with as much as 10% of cancers being linked to such mutations. Mutations in Thr41, Ser37 and Ser33 are frequently observed in cancers with the mutations generating β-catenin that escapes phosphorylation and degredation allowing translocation to the nucleus in the absence of Wnt signalling, leading to uncontrolled cell proliferation. β-catenin is an important feature of adherens junctions, acting to link cadherins to the actin cytoskeleton to form strong cell-cell adhesions. The cadherin family is a group of functionally related glycoproteins and is comprised of a variety of different subclasses which are distinct in specificity and tissue distribution. We will focus on Ecadherin which is expresses in preimplantation embryos and in non-neuronal epithelial tissue. Cell adhesion is promoted via a homophilic mechanism with cadherins only being able to bind to the same subtype e.g. E-cadherin to E-cadherin. The N-terminal domain of E-cadherin is comprised of 5 repeat domains (EC1-5) with the N-terminal 113 amino acids containing a highly conserved HAV sequence which is critical for ligand binding and signalling. It is this domain which forms the trans-cadherin interactions between neighbouring cells. The domain also contains the Ca2+ binding motifs. Ca2+ binding is essential for maintaining the correct conformation to allow cadherin binding. Calcium promotes an elongated, rigid conformation. Without calcium the domain becomes disordered and not elongated, resembling a globular structure that cannot interact with neighbouring cadherin molecules. The trans-cadherin interaction involves a partial swapping of an N-terminal β-strand present in the EC1 domains of two cadherin molecules. The swapped strands are stabilised by a tryptophan residue from one monomer docking into a pocket in the EC1 domain of another monomer. Stability is also provided by salt-bridge interactions formed by positively charged N-terminal residues. The cytoplasmic domain of E-cadherin has 2 relatively well-defined catenin binding domains. A 94 amino acid juxtamembrane domain which binds P120 catenin and an extended region to the C-terminus which binds β-catenin via a highly conserved LSSL motif. When β-catenin is bound it is unable to act as a transcription factor and is protected from the degredation complex. β-catenin can therefore not promote cell proliferation when cadherin-cadherin junctions are present. This is known as contact inhibition. The ratio of bound to free cytosolic β-catenin is therefore an important control of EMT. Increased EMT is caused by increased free cytosolic β-catenin this can be the result of decreased E-cadherin expression, meaning less β-catenin is sequestered in cell junctions. Decreased α-catenin is also related to increased EMT. α-catenin acts as another adaptor protein in cell-cell junctions, linking βcatenin to the actin cytoskeleton, decreased levels of α-catenin result in increased levels of cytoplasmic β-catenin resulting in increased EMT. Post-translational modification are also important in regulating the activity of β-catenin. Phosphorylation of tyrosine 654 in β-catenin by Src results in a decreased affinity for E-cadherin due to an inability to form a hydrogen bond with aspartic acid 665 in E-cadherin. This modification therefore promotes the Wnt signalling pathway. The phosphorylation of tyrosine 142 on β-catenin

reduces affinity for α-catenin, increasing accumulation of cytosolic β-catenin. This phosphorylation also promotes the binding of the transcriptional co-activator BCL9, promoting transcription of Wnt target genes and preventing protein-protein interactions. The overlapping of the binding site for αcatenin and the transcriptional co-activator allow for efficient regulation of the activity of β-catenin....