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Dynamic and Static Interactions between p120 Catenin and E-Cadherin Regulate the Stability of Cell-Cell Adhesion
Abstract
The association of p120 catenin (p120) with the juxtamembrane domain (JMD) of the cadherin cytoplasmic tail is critical for the surface stability of cadherin-catenin cell-cell adhesion complexes. Here, we present the crystal structure of p120 isoform 4A in complex with the JMD core region (JMD(core)) of E-cadherin. The p120 armadillo repeat domain contains modular binding pockets that are complementary to electrostatic and hydrophobic properties of the JMD(core). Single-residue mutations within the JMD(core)-binding site of p120 abolished its interaction with E- and N-cadherins in vitro and in cultured cells. These mutations of p120 enabled us to clearly differentiate between N-cadherin-dependent and -independent steps of neuronal dendritic spine morphogenesis crucial for synapse development. NMR studies revealed that p120 regulates the stability of cadherin-mediated cell-cell adhesion by associating with the majority of the JMD, including residues implicated in clathrin-mediated endocytosis and Hakai-dependent ubiquitination of E-cadherin, through its discrete "dynamic" and "static" binding sites.
... d Detachment time-course assay. Data represent the percentage of remaining attached cells after being treated with trypsin at different timepoints (n = 9, mean ± SD; Welch's t-test, * P < 0.05) ◂ machinery [21][22][23]. Since CTNND1-depleted cells showed both morphological and E-cadherin alterations, hampered cell-to-cell interaction was suspected. ...
... Its protein product, p120ctn, has a crucial role in maintaining cell-to-cell adhesion at adherens junctions, regulating the activation of small RhoGTPases in the cytoplasm and modulating nuclear transcription. Specifically, it directly interacts within the JMD of E-cadherin and controls E-cadherin turnover by hampering Hakai and endocytic machinery interaction [21][22][23]. ...
... Of particular note is the different effects observed either over E-cadherin expression or localization. It has been reported that the stability of E-cadherin at the adherens junctions is dependent on an equilibrium of cadherin retention at the membrane, driven by p120ctn, and cadherin internalization, induced by either the clathrin endocytic machinery or the Hakai-dependent ubiquitination and subsequent degradation [23]. Since the endocytic or ubiquitination fate of E-cadherin seems to be associated with different p120ctn binding regions, the specific location of p120ctn variants may influence or favor one mechanism over another. ...
Background
CDH1 and CTNNA1 remain as the main genes for hereditary gastric cancer. However, they only explain a small fraction of gastric cancer cases with suspected inherited basis. In this study, we aimed to identify new hereditary genes for early-onset gastric cancer patients (EOGC; < 50 years old).
Methods
After germline exome sequencing in 20 EOGC patients and replication of relevant findings by gene-panel sequencing in an independent cohort of 152 patients, CTNND1 stood out as an interesting candidate gene, since its protein product (p120ctn) directly interacts with E-cadherin. We proceeded with functional characterization by generating two knockout CTNND1 cellular models by gene editing and introducing the detected genetic variants using a lentiviral delivery system. We assessed β-catenin and E-cadherin levels, cell detachment, as well as E-cadherin localization and cell-to-cell interaction by spheroid modeling.
Results
Three CTNND1 germline variants [c.28_29delinsCT, p.(Ala10Leu); c.1105C > T, p.(Pro369Ser); c.1537A > G, p.(Asn513Asp)] were identified in our EOGC cohorts. Cells encoding CTNND1 variants displayed altered E-cadherin levels and intercellular interactions. In addition, the p.(Pro369Ser) variant, located in a key region in the E-cadherin/p120ctn binding domain, showed E-cadherin mislocalization.
Conclusions
Defects in CTNND1 could be involved in germline predisposition to gastric cancer by altering E-cadherin and, consequently, cell-to-cell interactions. In the present study, CTNND1 germline variants explained 2% (3/172) of the cases, although further studies in larger external cohorts are needed.
... Given that both substitutions are solventfacing in the N-cadherin/p120 complex structure, they do not significantly alter any binding interactions upon model equilibration. Analysis of our models indicated that the NH2-terminal region of the N-cadherin peptide binds p120-catenin through a series of electrostatic interactions, as described in (41). These remained unperturbed between the WT and phosphomimic-SE or phospho-Ser-788 N-cadherin/p120-catenin simulations. ...
... Previous work has demonstrated that p120-catenin binds to the juxtamembrane region (aa774-aa790) of N-cadherin (39) that contains Ser-788. This region of N-cadherin, which is normally disordered, is anchored to p120-catenin via electrostatic interactions involving the Asp774-Glu775-Glu776 (DEE) motif and hydrophobic interactions encompassing residues Leu-787, Ser-788, Gln-789 and Leu-790 (41). Asp-774 and Leu-790 are separated by more than 3 nm in the N-cadherin-bound p120-catenin form, and there is no obvious mechanism through which phosphorylation of Ser-788 would directly influence the structure, dynamics, and binding capacity of the DEE motif. ...
... Molecular dynamics simulations using the mouse N-cadherin/p120 complex model (based off the human E-cadherin/p120-catenin structure; pdb accession number 3L6X) were run on WT, the S787E phosphomimic variant and the phospho-S788 form of N-cadherin, introduced by the 'swap' command in YASARA (41,65). These simulations were run at 310°K, 150 mM NaCl, pH 7.4 using the Amber ff14 forcefield in a simulation cell with periodic boundaries for 100 ns. ...
Obscurins are giant cytoskeletal proteins with structural and regulatory roles. Obscurin-B (~870 kDa), the largest known isoform, contains two enzymatically active Ser/Thr kinase (kin) domains, kin1 and kin2, which belong to the myosin light chain kinase (MLCK) family. Kin1 binds to and phosphorylates N-cadherin, a major component of the intercalated disc (ICD), the unique sarcolemmal microdomain that mediates the mechanochemical coupling of adjacent cardiomyocytes. Obscurin-B containing kin1 and N-cadherin co-localize at cell junctions in embryonic rat ventricular myocytes (ERVM), and their co-distribution is regulated by Ca2+. Phosphoproteomics analysis revealed that obscurin-kin1 phosphorylates N-cadherin at Ser-788 located within the juxtamembrane region of its cytoplasmic domain with an apparent Kcat of ~5.05 min-1. Overexpression of obscurin-kin1 or phosphomimic-Ser-788-Glu N-cadherin in ERVM markedly increases cell adhesion and chemical coupling. Importantly, phosphomimic-Ser-788-Glu N-cadherin exhibits significantly reduced binding to p120-catenin, while overexpression of phosphoablated-Ser-788-Ala N-cadherin increases RhoA activity. Consistent with an essential role of the obscurin-kin1/N-cadherin axis in cardiomyocyte coupling, it is deregulated in end-stage human heart failure. Given the nearly ubiquitous expression of obscurin and N-cadherin, our findings may have broad applicability in deciphering the obscurin-kin1/N-cadherin axis that likely mediates cell coupling in diverse tissues and organs.
... Most cadherins are bound to p120: some 82.6 ± 3.6% of VE-cadherin and 75.5 ± 7.7% of N-cadherins 68 . X-ray crystallography shows that it likely forms oligomers with filaments organised in a crystallographic screw, with a pitch of 17.2nm and 3 residues per turn 69 . This naturally would allow cadherin-catenin complex interactions to be stabilised with a spacing of 17.2nm. ...
... Allowing for sub-optimal packing within a punctate adhesion and the fluorescent labels on the cadherins making the adhesions appear slightly larger, this separation between neighbouring cadherins is consistent with punctate adhesions with 6 cadherins and ≈ 50 nm wide as seen by Wu et al. 24 . The strength of cadherin tail-p120 bonds can be very accurately found to be J = 11k B T from the detailed isothermal titration calorimetry measurements conducted by Ishiyama et al. 69 (see their Figure 6A). This value is stronger than the J ≤ 7k B T suggested for cis bond 24 , suggesting that p120-cadherin bonds form preferentially prior to cadherin-cadherin cis bonds. ...
... This value is stronger than the J ≤ 7k B T suggested for cis bond 24 , suggesting that p120-cadherin bonds form preferentially prior to cadherin-cadherin cis bonds. (Note that there is a possibility that the oligomerization of p120 might have been a crystallisation artifact, as discussed in 69 . However, because recent work by? supports a role of p120 in E-cadherin clustering, we consider the geometry described in this paragraph to be the best model for nearest-neighbour cadherin-cadherin interactions in the remainder of this article.). ...
Cadherins mediate cell-cell adhesion and help the cell determine its shape and function. Here we study collective cadherin organization and interactions within cell-cell contact areas, and find the cadherin density at which a ‘gas-liquid’ phase transition occurs, when cadherin monomers begin to aggregate into dense clusters. We use a 2D lattice model of a cell-cell contact area, and coarse-grain to the continuous number density of cadherin to map the model onto the Cahn-Hilliard coarsening theory. This predicts the density required for nucleation, the characteristic length scale of the process, and the number density of clusters. The analytical predictions of the model are in good agreement with experimental observations of cadherin clustering in epithelial tissues.
... The variant c.2281G > A occurs in the cytoplasmic tail of E-cadherin whose role is to regulate downstream cell-cell adhesion signaling ( Figure 5A). The corresponding amino acid was solved as part of the juxta-membrane domain core region (JMD core) [31], which interacts with p120 catenin (p120) ( Figure 5B). In the co-crystal structure, it corresponds to an 18-amino-acid peptide (residues 756-773) that interacts with the Armadillo (ARM) domain of p120. ...
... Such a property would have a significant consequence by restricting the plasticity of the mutant form to conformations other than that of the WT form. Moreover, G761 has been shown to be highly conserved in the JMD core, and the GGG motif (residues 759-763) is crucial for the formation of a rotational structure that interacts with residues F437, W477, and N478 of p120 (ACMG-PP2-PP3) [31]. For JI-007 with the c.2281G > A CDH1 variant, we observed a loss of E-cadherin protein expression by IHC in GC FFPE tissue. ...
... This is a major hallmark of tumor malignancy, which is induced by a variety of factors, including transcriptional regulation, mutation, and aberrant cadherin internalization ( Figure 6) [40]. The ubiquitin-dependent endocytosis of E-cadherin [41] was associated with the depletion of E-cadherin from the cell surface [31], highlighted by the loss of membranous staining of E-cadherin in tumor cells in our results ( Figure 3C,D). ...
Mutational screening of the CDH1 gene is a standard treatment for patients who fulfill Hereditary Diffuse Gastric Cancer (HDGC) testing criteria. In this framework, the classification of variants found in this gene is a crucial step for the clinical management of patients at high risk for HDGC. The aim of our study was to identify CDH1 as well as CTNNA1 mutational profiles predisposing to HDGC in Tunisia. Thirty-four cases were included for this purpose. We performed Sanger sequencing for the entire coding region of both genes and MLPA (Multiplex Ligation Probe Amplification) assays to investigate large rearrangements of the CDH1 gene. As a result, three cases, all with the HDGC inclusion criteria (8.82% of the entire cohort), carried pathogenic and likely pathogenic variants of the CDH1 gene. These variants involve a novel splicing alteration, a missense c.2281G > A detected by Sanger sequencing, and a large rearrangement detected by MLPA. No pathogenic CTNNA1 variants were found. The large rearrangement is clearly pathogenic, implicating a large deletion of two exons. The novel splicing variant creates a cryptic site. The missense variant is a VUS (Variant with Uncertain Significance). With ACMG (American College of Medical Genetics and Genomics) classification and the evidence available, we thus suggest a revision of its status to likely pathogenic. Further functional studies or cosegregation analysis should be performed to confirm its pathogenicity. In addition, molecular exploration will be needed to understand the etiology of the other CDH1-and CTNNA1-negative cases fulfilling the HDGC inclusion criteria. Citation: Ben Aissa-Haj, J.; Kabbage, M.; Othmen, H.; Saulnier, P.; Kettiti Tounsi, H..; Jaballah-Gabteni, A.; Ferah, A.; Medhioub, M.; Khsiba, A.; Mahmoudi, M.; et al. CDH1
... ;https://doi.org/10.1101https://doi.org/10. /2024 catenin levels results in a significant increase in E-cadherin internalization (Davis et al., 2003;Ireton et al., 2002;Ishiyama et al., 2010). ...
... Once internalized, E-cadherin can be recycled back to the plasma membrane, sequestered in internal compartments, or sent for degradation (Bryant and Stow, 2004). Endocytosis of E-cadherin is triggered by the disassembly of the E-cadherin/p120-catenin complex (Davis et al., 2003;Ireton et al., 2002;Ishiyama et al., 2010). Our results suggest that the downregulation of E-cadherin levels in SGEF KD cells is initiated by the dissociation of p120-catenin from the E-cadherin complex, followed by internalization of E-cadherin through endocytosis and degradation by the proteasome. ...
SGEF, a RhoG specific GEF, can form a ternary complex with the Scribble polarity complex proteins Scribble and Dlg1, which regulates the formation and maintenance of adherens junctions and barrier function of epithelial cells. Notably, silencing SGEF results in a dramatic downregulation of the expression of both E-cadherin and ZO-1. However, the molecular mechanisms involved in the regulation of this pathway are not known. Here, we describe a novel signaling pathway governed by the Scribble/SGEF/Dlg1 complex. Our results show that an intact ternary complex is required to maintain the stability of the apical junctions, the expression of ZO-1, and TJ permeability. In contrast, only SGEF is necessary to regulate E-cadherin expression. The absence of SGEF destabilizes the E-cadherin/catenin complex at the membrane, triggering a positive feedback loop that exacerbates the phenotype through the repression of E-cadherin transcription in a process that involves the internalization of E-cadherin by endocytosis, β-catenin signaling and the transcriptional repressor Slug.
... Plakophilins are members of a p120-catenin subfamily of armadillo repeat motif proteins [16][17][18]. This distinctive feature, as determined by crystallography of p120-catenin and plakophilin-1 [19,20], is the nine armadillo repeat motif organization of the armadillo domain interrupted between repeats four and five by an insert of approximately 60 amino acids. The p120-catenin subfamily is subdivided into two groups. ...
... We present the cryoEM structure plakophilin-3, consisting of the entire armadillo repeat motifs domain, although the loop region lacks continuous density in the Coulomb map. As a member of the p120-related armadillo repeat subfamily proteins, the overall arrangement of the armadillo repeat motifs domain of plakophilin-3 resembles the crystal structures of plakophilin-1 [19] (PDB entry 1xm9) and p120-catenin [20] (PDB entry 3l6x). Plakophilin-3 and plakophilin-1 do not have an additional loop between the fourth and fifth armadillo repeats e and f, as compared to p120-catenin, which retains this loop. ...
Plakophilin-3 is a ubiquitously expressed protein found widely in epithelial cells and is a critical component of desmosomes. The plakophilin-3 carboxy-terminal domain harbors nine armadillo repeat motifs with largely unknown functions. Here, we report the 5 Å cryogenic electron microscopy (cryoEM) structure of the armadillo repeat motif domain of plakophilin-3, one of the smaller cryoEM structures reported to date. We find that this domain is a monomer or homodimer in solution. In addition, using an in vitro actin co-sedimentation assay, we show that the armadillo repeat domain of plakophilin-3 directly interacts with F-actin. This feature, through direct interactions with actin filaments, could be responsible for the observed association of extra-desmosomal plakophilin-3 with the actin cytoskeleton directly attached to the adherens junctions in A431 epithelial cells. Further, we demonstrate, through lipid binding analyses, that plakophilin-3 can effectively be recruited to the plasma membrane through phosphatidylinositol-4,5-bisphosphate-mediated interactions. Collectively, we report on novel properties of plakophilin-3, which may be conserved throughout the plakophilin protein family and may be behind the roles of these proteins in cell-cell adhesion.
... A remarkable example of the interplay between static and dynamic PPI is the mechanism p120 catenin uses to regulate the stability of cell-cell adhesion. This mechanism relies on the binding of the intrinsically disordered cytoplasmic tail of cadherin through both "static" and "dynamic" interfaces [10]. The central amino acid sequence of the cadherin cytoplasmic tail strongly binds p120 through a well-structured "static" interface. ...
... On the other hand, the unfolded region of the cadherin tail flanking the protein N-terminus also weakly but dynamically interacts with p120. Fluctuations of this second "dynamic" binding site provide a marginal contribution to the binding affinity, but play a crucial function in determining cadherin cell fate by masking a Leucine-Leucine motif which hinders internalization via cadherin-mediated endocytosis [10]. Given the fundamental role played by protein partnerships in cells, networks of PPI are tightly regulated to deliver specific signaling over time and space. ...
Protein–protein interfaces play fundamental roles in the molecular mechanisms underlying pathophysiological pathways and are important targets for the design of compounds of therapeutic interest. However, the identification of binding sites on protein surfaces and the development of modulators of protein–protein interactions still represent a major challenge due to their highly dynamic and extensive interfacial areas. Over the years, multiple strategies including structural, computational, and combinatorial approaches have been developed to characterize PPI and to date, several successful examples of small molecules, antibodies, peptides, and aptamers able to modulate these interfaces have been determined. Notably, peptides are a particularly useful tool for inhibiting PPIs due to their exquisite potency, specificity, and selectivity. Here, after an overview of PPIs and of the commonly used approaches to identify and characterize them, we describe and evaluate the impact of chemical peptide libraries in medicinal chemistry with a special focus on the results achieved through recent applications of this methodology. Finally, we also discuss the role that this methodology can have in the framework of the opportunities, and challenges that the application of new predictive approaches based on artificial intelligence is generating in structural biology.
... Cell-cell adhesion junctions (AJs) initiate and maintain intercellular adhesion, regulate the organization of the underlying actin cytoskeleton, and establish hubs for cell signaling and gene transcription regulation (Garcia et al., 2018) AJs play important roles in maintaining tissue structure and cell polarity, which limit the cell movement and proliferation. Catenin delta 1 (CTNND1), also known as p120, is a member of the cadherin-series protein complex and directly binds to the cytoplasmic tail of E-cadherin through a pre-conserved juxtamembrane domain (Ishiyama et al., 2010) Also, γ-catenin binds to the catenin-binding domain of Ecadherin. Therefore, E-cadherin, CTNND1, α-catenin, and γ-catenin are the major proteins involved in cellcell AJ formation and maintenance. ...
... These results suggest that CTNND1 regulates EMT, proliferation, migration, and invasion in pancreatic cancer cells by regulating the Wnt/β-catenin signaling pathway. CTNND1 and β-catenin jointly bind to E-cadherin (Ishiyama et al., 2010) The different degrees of loss of E-cadherin and CTNND1 leads to different roles of CTNND1 in tumors ( Thoreson and Reynolds, 2002) The initial loss of CTNND1 leads to the instability of the E-cadherin complex, resulting in the reduction of E-cadherin expression level. However, the rst loss of E-cadherin leads to an increase in CTNND1 in the cytoplasm (Soto et al., 2008;Thoreson and Reynolds, 2002) CTNND1 binds the transcription repressor kaiso in the cytoplasm (Daniel and Reynolds, 1999) resulting in the inhibited translocation of kaiso to the nucleus and promoted β-catenin expression (Ogden et al., 2008;van Roy and McCrea, 2005;Xue et al., 2017) Therefore, CTNND1 accumulation leads to an increased level of β-catenin in the cytoplasm (Jiang et al., 2012;Liu et al., 2014;Spring et al., 2005) and activated Wnt/β-catenin signaling pathway, which plays a key role in the apoptosis, proliferation, and metastasis of cancer cells EMT can regulate a variety of cancers, including pancreatic cancer (Liu et al., 2019) EMT is a key step in metastasis in patients with pancreatic cancer and associated with poor prognosis (Nachiyappan et al., 2022, p. 1) Our study showed that CTNND1 knockdown signi cantly inhibited the Wnt/β-catenin pathway, as well as the proliferation, migration, invasion, and EMT phenotype of pancreatic cancer cells. ...
Background: Catenin delta 1 (CTNND1) is upregulated in many tumors and is closely associated with poor prognosis. However, the role of CTNND1 in pancreatic cancer and its underlying mechanisms remain unclear
Methods: The expression of CTNND1 in pancreatic cancer and normal tissues in the TCGA and GTEX databases was preliminarily screened and further verified by immunohistochemistry (IHC) and qPCR. Transwell, wound healing, and cell proliferation assays were used to study the effect of CTNND1 on epithelial-mesenchymal transition (EMT), proliferation, migration, and invasion of pancreatic cancer cells. Western blot experiments verified the signaling pathway mediating the effect of CTNND1 on pancreatic cancer progression. The expression of CTNND1 in the TCGA database, clinical pancreatic cancer samples, and pancreatic cancer cells was significantly upregulated.
Results: We found that the silent CTNND1 in pancreatic cancer cells significantly inhibited the EMT, proliferation, migration, and invasion of pancreatic cancer cells. In addition, the silencing of CTNND1 in pancreatic cancer cells inhibited the Wnt/β-catenin signaling pathway. LiCl (a Wnt/β-catenin-specific activator) treatment partially restored the EMT, proliferation, migration, and invasion abilities of CTNND1-silenced pancreatic cancer cells.
Conclusion: Our research confirmed that CTNND1 can regulate the EMT, proliferation, migration, and invasion of pancreatic cancer through the WNT/β-catenin signaling pathway.
... We next sought to determine the basis for cadherin selectivity by MARCH family ligases. Classical cadherins contain endocytic motifs nestled within the p120-catenin binding site on the cadherin juxtamembrane domain [21,36,37]. Previous studies have implicated transmembrane domains in MARCH ligase substrate selectivity and found that the ligases typically target membrane proximal lysine residues [38][39][40]. ...
Cadherin family proteins play a central role in epithelial and endothelial cell-cell adhesion. The dynamic regulation of cell adhesion is achieved in part through endocytic membrane trafficking pathways that modulate cadherin cell surface levels. Here, we define the role for various MARCH family ubiquitin ligases in the regulation of cadherin degradation. We find that MARCH2 selectively downregulates VE-cadherin, resulting in loss of adherens junction proteins at cell borders and a loss of endothelial barrier function. Interestingly, N-cadherin is refractory to MARCH ligase expression, demonstrating that different classical cadherin family proteins are differentially regulated by MARCH family ligases. Using chimeric cadherins, we find that the specificity of different MARCH family ligases for different cadherins is conferred by the cadherin transmembrane domain. Further, juxta-membrane lysine residues are required for cadherin degradation by MARCH proteins. These findings expand our understanding of cadherin regulation and highlight a new role for mammalian MARCH family ubiquitin ligases in differentially regulating cadherin turnover.
... MiR-409-3 inhibited GC invasion through targeting RDX [28]. Catenin-δ1 (CTNND1) is a key regulator of cell-cell adhesion [114]. Catenin-δ1 binds to juxtamembrane domain of E-cadherin, which subsequently stabilizes E-cadherin. ...
Late diagnosis is one of the main reasons for high mortality rates in cancer patients. Therefore, investigating the molecular mechanisms involved in tumor progression can improve the cancer diagnosis in the early stages of the tumor progression. MicroRNAs (miRNAs) have important roles in regulation of cell growth, proliferation, metabolism, and migration. Since, deregulation of miR-409 has been reported in a wide range of cancers, in the present review, we investigated the molecular mechanisms of miR-409 during tumor progression and invasion. It has been shown that miR-409 functions as a tumor suppressor in different tumor types. MiR-409 can reduce tumor cell proliferation, growth, and migration by regulation of signaling pathways, cellular metabolism, transcription factors, and cellular adhesion. This review can be an effective step in introducing miR-409 as a non-invasive marker in cancer patients.
... При этом гены катенина дельта 1 (CTNND1) и L-селектина (SELL), связанные с межклеточным взаимодействием, напротив, увеличивали экспрессию. Катенин дельта 1 является одним из ключевых регуляторов межклеточной адгезии, а также контролирует активность ГТФаз семейства Rho и динамику цитоскелета [18]. L-селектин относится к семейству рецепторов адгезии/хоминга и обнаруживается на поверхности лейкоцитов. ...
Hibernating myocardium, potentially capable of restoring its function, is of clinical and scientific
interest. Histological and molecular analysis of the hibernating myocardium in the zones of hypokinesis and normokinesis in a patient with ischemic heart disease was performed.
Material and methods. A molecular and immunohistochemical study of left ventricular myocardial
biopsies was performed in a patient after surgical reconstruction of the left ventricle in combination with surgical revascularization.
Results. Morphological examination revealed a violation of the structure of cardiomyocytes, which
correlates with the accumulation of matrix metalloproteinase type 9 in the cytoplasm of cardiomyocytes in these zones against the background of partial or complete destruction of their basement
membranes formed by type IV collagen.
Analysis of the differential expression of genes encoding extracellular matrix proteins and matrixassociated molecules revealed a shift in the transcription profile due to a decrease in the expression
of collagen genes, matrix proteases, and cell-matrix interaction molecules.
Conclusion. Based on the analysis, it can be concluded that the hibernating myocardium, as a
result of the destruction of the structure of the sarcormers and basal membranes of cardiomyocytes,
a decrease in the activity of extracellular matrix genes, is not able to further provide contractile
function, and viable cells that are detected during morphological examination probably function
only as a protective mechanism during early scar formation.
... This feature is suggested to favor binding towards hydrophobic grooves and promote stabilization less stringently (40,41,47). This type of dynamic interface is proposed to be advantageous to conformational fluctuations and facilitate posttranslational modification in a signaling network (41,(48)(49)(50)(51). Similarly, the hydrophobic lower NBD1 interactions with the . ...
Many ATP-binding cassette (ABC) transporters are regulated by phosphorylation on long and disordered loops. which makes their interactions a challenge to visualize. We have trapped an activated state of the regulatory domain (R-domain) of Yeast Cadmium Factor 1 (Ycf1) by enzymatically enriching the phosphorylated state. A 3.2 Å cryo-EM structure reveals an R-domain structure with four phosphorylated residues and a position for the entire R-domain. The structure reveals key R-domain interactions including a bridging interaction between NBD1 and NBD2 as well as an interaction the R-insertion, another regulatory region. We systematically probe these interactions with a linker substitution strategy along the R-domain and find a close match with these interactions and survival under Ycf1-dependent growth conditions. We propose a model where four overlapping phosphorylation sites bridge several regions of Ycf1 to engage in a transport-competent state.
... MDCK cells generally express E-cadherin as the predominant cadherin (18). The binding of p120 catenin to E-cadherin, localized to the cell membrane, is known to inhibit endocytosis, leading to the stability of cadherin-catenin cell-cell adhesion complexes (19,20). RhoA activation is known to decrease Rac1 activity (8) and induce myosin activation through phosphorylation of the myosin II light chain, leading to the assembly of TJ proteins (21). ...
Background: Liquid-liquid interfaces, such as lacrimal fluid of the eye, are pervasive in living organism. While culturing cells in an environment that closely mimics in vivo conditions enhances our understanding of cell characteristics and unveils underlying mechanisms, practical considerations have led to predominant use of solid surfaces, such as polystyrene or glass, for cell culture. In this study, we introduce a novel culture design aimed at promoting uniform maturation of epithelial cells using liquid materials.
Results: Culturing Madin–Darby canine kidney (MDCK) cells at the liquid-liquid interface yielded reduced migration and stimulated active cell growth. Similar to solid-liquid interfaces, cells cultured on a fibronectin-coated liquid-liquid interface exhibited active migration and growth, ultimately reaching a confluent state. These cells exhibited reduced stress fiber formation and adopted a cobblestone-like shape, which led to their even distribution in the culture vessel. Analysis of staining images of the tight junction (TJ) protein ZO-2 revealed that the frequency of ZO-2-positive cells, FZ, in cultures on the fibronectin-coated liquid-liquid interface was 2.2-fold higher than that on the non-coated liquid-liquid interface at t = 72 h. To inhibit stress fiber formation and apoptosis, the exposure of cells to Y27632, a specific inhibitor of the Rho-associated protein kinase (ROCK), facilitated uniform maturation (FZ = 0.73). Additionally, a similar trend was observed in cells cultured on the fibronectin-coated liquid-liquid interface (FZ = 0.80) at t = 72 h. In Y27632-exposed cells on the liquid-liquid interface, the value obtained by subtracting the standard deviation of the ratio of nucleus densities in each region that compartmentalized a culture vessel from 1, denoted as HLN, was 0.93 0.01, indicating even cell distribution in the culture vessel at t = 72 h. Across all culture conditions, the HLN and FZ values exhibited a close correlation (R² = 0.93), suggesting that cell distribution in the culture vessel is related to the formation of TJs in the late phase of maturation.
Conclusions: Our findings suggest that the behaviors of epithelial cells on liquid-liquid interfaces contribute to the promotion of their uniform maturation.
... Third, the function of CTNND1 also supports it as a potential risk gene for anxiety disorders. CTNND1 encodes the p120-catenin (p120) protein, which regulates cell-cell adhesion by interacting with E-cadherin 69 . In addition, p120 regulates RhoA activity 70 and the Wnt signalling pathway 71 , two pathways with important roles in the central nervous system. ...
Anxiety disorders are the most prevalent mental disorders. However, the genetic etiology of anxiety disorders remains largely unknown. Here we conducted a genome-wide meta-analysis on anxiety disorders by including 74,973 (28,392 proxy) cases and 400,243 (146,771 proxy) controls. We identified 14 risk loci, including 10 new associations near CNTNAP5, MAP2, RAB9BP1, BTN1A1, PRR16, PCLO, PTPRD, FARP1, CDH2 and RAB27B. Functional genomics and fine-mapping pinpointed the potential causal variants, and expression quantitative trait loci analysis revealed the potential target genes regulated by the risk variants. Integrative analyses, including transcriptome-wide association study, proteome-wide association study and colocalization analyses, prioritized potential causal genes (including CTNND1 and RAB27B). Evidence from multiple analyses revealed possibly causal genes, including RAB27B, BTN3A2, PCLO and CTNND1. Finally, we showed that Ctnnd1 knockdown affected dendritic spine density and resulted in anxiety-like behaviours in mice, revealing the potential role of CTNND1 in anxiety disorders. Our study identified new risk loci, potential causal variants and genes for anxiety disorders, providing insights into the genetic architecture of anxiety disorders and potential therapeutic targets.
... This protein is another member of the catenin protein family and contains armadillo repeat domains that facilitate its interaction with other proteins including E-Cadherin ( Figure 12). Association or displacement from E-Cadherin regulates the stability of E-Cadherin in adherens junctions [71]. The p120 protein has other interactions and roles that regulate epithelial and endothelial barriers. ...
Familial Exudative Vitreoretinopathy (FEVR), Norrie disease, and persistent fetal vascular syndrome (PFVS) are extremely rare retinopathies that are clinically distinct but are unified by abnormal retinal endothelial cell function, and subsequent irregular retinal vascular development and/or aberrant inner blood-retinal-barrier (iBRB) function. The early angiogenesis of the retina and its iBRB is a delicate process that is mediated by the canonical Norrin Wnt-signaling pathway in retinal endothelial cells. Pathogenic variants in genes that play key roles within this pathway, such as NDP, FZD4, TSPAN12, and LRP5, have been associated with the incidence of these retinal diseases. Recent efforts to further elucidate the etiology of these conditions have not only highlighted their multigenic nature but have also resulted in the discovery of pathological variants in additional genes such as CTNNB1, KIF11, and ZNF408, some of which operate outside of the Norrin Wnt-signaling pathway. Recent discoveries of FEVR-linked variants in two other Catenin genes (CTNND1, CTNNA1) and the Endoplasmic Reticulum Membrane Complex Subunit-1 gene (EMC1) suggest that we will continue to find additional genes that impact the neural retinal vasculature, especially in multi-syndromic conditions. The goal of this review is to briefly highlight the current understanding of the roles of their encoded proteins in retinal endothelial cells to understand the essential functional mechanisms that can be altered to cause these very rare pediatric retinal vascular diseases.
... This protein is another member of the catenin protein family and contains armadillo repeat domains that facilitate its interaction with other proteins including E-Cadherin. Association or displacement from E-Cadherin regulates the stability of E-Cadherin in adherens junctions [75]. The p120 protein has other interactions and roles that regulate epithelial and endothelial barriers. ...
Familial Exudative Vitreoretinopathy (FEVR), Norrie disease, and persistent fetal vascular syndrome (PFVS) are extremely rare retinopathies that are clinically distinct but are unified by abnormal retinal endothelial cell function – and subsequent irregular retinal vascular development and/or aberrant inner blood-retinal-barrier (iBRB) function. The early angiogenesis of the retina and its iBRB is a delicate process that is mediated by the canonical Norrin Wnt-signaling pathway in retinal endothelial cells. Pathogenic variants in genes that play key roles within this pathway such as NDP, FZD4, TSPAN12, and LRP5, have been associated with the incidence of these retinal diseases. Recent efforts to further elucidate the etiology of these conditions have not only highlighted their multigenic nature but have also resulted in the discovery of pathological variants in additional genes such as CTNNB1, KIF11, and ZNF408, some of which operate outside of the Norrin Wnt-signaling pathway. Recent discoveries of FEVR-linked variants in two other Catenin genes (CTNND1, CTNNA1) and the Endoplasmic Reticulum Membrane Complex Subunit-1 gene (EMC1) suggest that we will continue to find additional genes that impact the neural retinal vasculature, especially in multi-syndromic conditions. The goals of this review are to briefly highlight the current understanding of the roles of their encoded proteins in retinal endothelial cells to understand the essential functional mechanisms that can be altered to cause these very rare pediatric retinal vascular diseases.
... Cadherins are a family of calcium-dependent cell adhesion molecules critical throughout neural development with crucial roles ranging from neuroblast migration, axon finding, and synapse formation (Suzuki and Takeichi, 2008;Hirano and Takeichi, 2012). Cadherins are known binding partners of δ-catenin (Silverman et al., 2007;Yuan et al., 2015), but how this interaction occurs has primarily been inferred from data for E-cadherin/p120catenin interaction (Ishiyama et al., 2010) due to the high sequence homology between p120-catenin and δ-catenin (Paffenholz and Franke, 1997). ...
Astrocytes control the formation of specific synaptic circuits via cell adhesion and secreted molecules. Astrocyte synaptogenic functions are dependent on the establishment of their complex morphology. However, it is unknown if distinct neuronal cues differentially regulate astrocyte morphogenesis. δ-Catenin was previously thought to be a neuron-specific protein that regulates dendrite morphology. We found δ-catenin is also highly expressed by astrocytes and required both in astrocytes and neurons for astrocyte morphogenesis. δ-Catenin is hypothesized to mediate transcellular interactions through the cadherin family of cell adhesion proteins. We used structural modeling and biochemical analyses to reveal that δ-catenin interacts with the N-cadherin juxtamembrane domain to promote N-cadherin surface expression. An autism-linked δ-catenin point mutation impaired N-cadherin cell surface expression and reduced astrocyte complexity. In the developing mouse cortex, only lower-layer cortical neurons express N-cadherin. Remarkably, when we silenced astrocytic N-cadherin throughout the cortex, only lower-layer astrocyte morphology was disrupted. These findings show that δ-catenin controls astrocyte–neuron cadherin interactions that regulate layer-specific astrocyte morphogenesis.
... To regulate this essential function, components of the AJ are in a constant state of protein turnover and can be strengthened or weakened as the biochemical or mechanical situation warrants. The regulation necessary for this kind of rapid response partially involves post-translational modifications; phosphorylation of the cadherin tail, p120, and β-catenin influence both protein-protein affinity and the recruitment of peripheral proteins to the AJ [27][28][29]. However, proteolysis of AJ proteins also plays a role in normal AJ maintenance and signaling. ...
The intercalated disk is a cardiac specific structure composed of three main protein complexes—adherens junctions, desmosomes, and gap junctions—that work in concert to provide mechanical stability and electrical synchronization to the heart. Each substructure is regulated through a variety of mechanisms including proteolysis. Calpain proteases, a class of cysteine proteases dependent on calcium for activation, have recently emerged as important regulators of individual intercalated disk components. In this review, we will examine how calcium homeostasis regulates normal calpain function. We will also explore how calpains modulate gap junctions, desmosomes, and adherens junctions activity by targeting specific proteins, and describe the molecular mechanisms of how calpain dysregulation leads to structural and signaling defects within the heart. We will then examine how changes in calpain activity affects cardiomyocytes, and how such changes underlie various heart diseases.
... The cytoplasmic tail of classical cadherins harbors two distinct binding sites for two catenins, p120-and β-catenin, which have distinct functions (Figure 2a,b). For E-cadherin, the best-studied classical cadherin, binding of p120-catenin to the juxtamembrane domain of the cytoplasmic tail promotes its stability by masking residues mediating Hakai-dependent ubiquitination and clathrin-mediated endocytosis (Fujita et al. 2002, Ishiyama et al. 2010. Free cytoplasmic p120catenin, but not cadherin-bound p120-catenin, can inhibit RhoA activity that promotes actin polymerization and myosin contractility (Anastasiadis et al. 2000) (Figure 2b). ...
Multicellular organisms generate tissues of diverse shapes and functions from cells and extracellular matrices. Their adhesion molecules mediate cell-cell and cell-matrix interactions, which not only play crucial roles in maintaining tissue integrity but also serve as key regulators of tissue morphogenesis. Cells constantly probe their environment to make decisions: They integrate chemical and mechanical information from the environment via diffusible ligand- or adhesion-based signaling to decide whether to release specific signaling molecules or enzymes, to divide or differentiate, to move away or stay, or even whether to live or die. These decisions in turn modify their environment, including the chemical nature and mechanical properties of the extracellular matrix. Tissue morphology is the physical manifestation of the remodeling of cells and matrices by their historical biochemical and biophysical landscapes. We review our understanding of matrix and adhesion molecules in tissue morphogenesis, with an emphasis on key physical interactions that drive morphogenesis.
Expected final online publication date for the Annual Review of Cell and Developmental Biology, Volume 39 is October 2023. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.
... It is made available under a preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in The copyright holder for this this version posted January 23, 2023. ; https://doi.org/10.1101/2023.01.23.525158 doi: bioRxiv preprint from the plasma membrane via a superhelical surface formed by 12 tandem ARM repeats (Choi et al, 2009;Huber & Weis, 2001;Ishiyama et al, 2010). To explore this, AlphaFold2, which has recently been shown to perform well for the prediction of protein-peptide complexes (Choi et al., 2009;Jumper et al., 2021), was used to generate a model for the complex of TgGAC1670-2639 with the C-terminal 20 amino acids residues of MIC2. ...
The phylum of Apicomplexa groups intracellular parasites that employ substrate-dependent gliding motility to invade host cells, egress from the infected cells and cross biological barriers. The glideosome associated connector (GAC) is a conserved protein essential to this process. GAC facilitates the association of actin filaments with surface transmembrane adhesins and the efficient transmission of the force generated by myosin translocation of actin to the cell surface substrate. Here, we present the crystal structure of Toxoplasma gondii GAC and reveal a unique, supercoiled armadillo repeat region that adopts a closed ring conformation. Characterisation of the membrane binding interface within the C-terminal PH domain as well as an N-terminal fragment necessary for association with F-actin suggest that GAC adopts multiple conformations. A multi-conformational model for assembly of GAC within the glideosome is proposed.
... The isomers may have completely different effects. By boosting P120's affinity for E-cadherin, P120 isoforms 3 and 4 can help epithelial cells adhere to one another (68). In contrast, p120 isoform 1 promotes RAC1 activity and stimulates cell migration and invasiveness by blocking the RHOA-ROCK signaling pathway (69). ...
Background
As a processing method of RNA precursors, alternative splicing (AS) is critical to normal cellular activities. Aberrant AS events are associated with cancer development and can be promising targets to treat cancer. However, no detailed and unbiased study describes the current state of AS of cancer research. We aim to measure and recognize the current state and trends of AS cancer research in this study.
Methods
The Web of Science Core Collection was used to acquire the articles. Utilizing three bibliometric tools (CiteSpace, VOSviewer, R-bibliometrix), we were able to measure and recognize the influence and collaboration data of individual articles, journals, and co-citations. Analysis of co-occurrence and burst information helped us identify the trending research areas related to AS of cancer.
Results
From 2012 to 2021, the total number of papers on AS of cancer published in 766 academic journals was 3,507, authored by 20,406 researchers in 405 institutions from 80 countries/regions. Research involving AS of cancer genes was primarily conducted in the United States and China; simultaneously, the Chinese Academy of Sciences, Fudan University, and National Cancer Institute were the institutions with strong research capabilities. Scorilas Andreas is the scholar with the most publications, while the most co-citations were generated by Wang, Eric T. Plos One published the most papers on AS of cancer, while J Biol Chem was the most co-cited academic journal in this field. The results of keyword co-occurrence analysis can be divided into three types: molecular (P53, CD44, androgen receptor, srsf3, esrp1), pathological process (apoptosis, EMT, metastasis, angiogenesis, proliferation), and disease (breast cancer, colorectal cancer, prostate cancer, hepatocellular carcinoma, gastric cancer).
Conclusion
Research on AS of cancer has been increasing in intensity over the past decade. Current AS of cancer studies focused on the hallmarks of AS in cancer and AS signatures including diagnostic and therapeutic targets. Among them, the current trends are splicing factors regulating epithelial–mesenchymal transition and other hallmarks, aberrant splicing events in tumors, and further mechanisms. These might give researchers interested in this field a forward-looking perspective and inform further research.
... In normal epithelia, binding of CDH1 to CTNNB1/β-catenin assures maintenance of the epithelial integrity preventing β-catenin from being released from the cytoplasmic signaling protein complex, enter the nucleus and bind to members of the DNA binding protein family LEF/TCF [4][5][6]. TEAD4, a transcription factor driving expression of YAP/TAZ signaling target genes, is known to be regulated by LEF/TCF in organoids derived from different gastrointestinal cell types including those from stomach [7][8][9]. Thus, the selective targeting of LEF/TCF in GC cells lacking CDH1 and having an activated b-catenin pathway may represent a promising therapeutic approach for GC treatment. ...
Loss of CDH1/Cadherin-1 is a common step towards the acquisition of an abnormal epithelial phenotype. In gastric cancer (GC), mutation and/or downregulation of CDH1/Cadherin-1 is recurrent in sporadic and hereditary diffuse GC type. To approach the molecular events downstream of CDH1/Cadherin-1 alterations and their relevance in gastric carcinogenesis, we queried public databases for genetic and DNA methylation data in search of molecular signatures with a still-uncertain role in the pathological mechanism of GC. In all GC subtypes, modulated genes correlating with CDH1/Cadherin-1 aberrations are associated with stem cell and epithelial-to-mesenchymal transition pathways. A higher level of genes upregulated in CDH1-mutated GC cases is associated with reduced overall survival. In the diffuse GC (DGC) subtype, genes downregulated in CDH1-mutated compared to cases with wild type CDH1/Cadherin-1 resulted in being strongly intertwined with the DREAM complex. The inverse correlation between hypermethylated CpGs and CDH1/Cadherin-1 transcription in diverse subtypes implies a common epigenetic program. We identified nonredundant protein-encoding isoforms of 22 genes among those differentially expressed in GC compared to normal stomach. These unique proteins represent potential agents involved in cell transformation and candidate therapeutic targets. Meanwhile, drug-induced and CDH1/Cadherin-1 mutation-related gene expression comparison predicts FIT, GR-127935 hydrochloride, amiodarone hydrochloride in GC and BRD-K55722623, BRD-K13169950, and AY 9944 in DGC as the most effective treatments, providing cues for the design of combined pharmacological treatments. By integrating genetic and epigenetic aspects with their expected functional outcome, we unveiled promising targets for combinatorial pharmacological treatments of GC.
... The isomers may have completely different effects. By boosting P120's affinity for Ecadherin, P120 isoforms 3 and 4 can help epithelial cells adhere to one another (68). In contrast, p120 isoform 1 promotes RAC1 activity and stimulates cell migration and invasiveness by blocking the RHOA-ROCK signaling pathway (69). ...
Background
As a processing method of RNA precursors, alternative splicing plays a critical role in normal cell physiological activities. Aberrant alternative splicing events are associated with cancer development and are promising targets for cancer treatment. However, no detailed and unbiased study describes the current state of alternative splicing of cancer research. This study aims to conduct a bibliometric analysis of alternative splicing of cancer research in the last decade to measure and recognize the current state and trends.
Methods
Web of Science Core Collection was used to acquire the articles. Utilizing three bibliometric tools (CiteSpace, VOSviewer, R-bibliometrix), we were able to measure and recognize the influence and collaboration data of individual articles, journals, and co-citations. Analysis of co-occurrence and bursts information helped us identify the trending research areas related to alternative splicing of cancer.
Results
From 2012 to 2021, the total number of papers on alternative splicing of cancer published in 766 academic journals was 3,507, authored by 20,406 researchers in 405 institutions from 80 countries/regions. Research involving alternative splicing of cancer genes was primarily conducted in the United States and China, with the United States retaining the prime status; simultaneously, the Chinese Academy of Sciences, Fudan University and National Cancer Institute were the institutions with strong research capabilities. Scorilas Andreas is the scholar with the most publications, while the most co-citations were generated by Wang, Eric T. Plos One published the most papers on alternative splicing of cancer, while J biol chem was the most co-cited academic journal in this field. Furthermore, these publications covered a wide range of areas, but were still primarily focused on the fields of molecular, biology, and immunology. The results of keyword co-occurrence analysis can be divided into three types: molecular (P53, CD44, Androgen receptor, SRSF3, ESRP1), pathological process (apoptosis, EMT, metastasis, angiogenesis, proliferation), and disease (breast cancer, colorectal cancer, prostate cancer, hepatocellular carcinoma, gastric cancer).
Conclusion
Research on alternative splicing of cancer has been a hot topic over the past decade, increasing in intensity each year. Current alternative splicing of cancer studies focused on hallmarks of alternative splicing in cancer, and alternative splicing signatures including diagnostic and therapeutic targets. Among them, the current trends are splicing factors regulating epithelial-mesenchymal transition and other hallmarks, aberrant splicing events in tumors, and further mechanisms. These might give researchers interested in this field a forward-looking perspective and inform further research.
... The junctional protein Ecadherin (CDH1) is heavily associated with cell borders in OVCAR3 cells ( Figure 5C) where it likely participates in calcium-dependent cell-cell interactions (Sundfeldt et al., 1997). While CDH1 was not biotinylated in the BioID experiment, its interacting proteins, CTNND1 (Ishiyama et al., 2010), DLG1 (Marziali et al., 2019), and CXADR (Nilchian et al., 2019) were. ...
Claudin-4, a protein with the structure of classic claudins most often found in cell-cell junctions, is frequently overexpressed in epithelial cancers where its localization has not been studied. In this study we aimed to find out where this membrane protein is localized in an ovarian tumor model, OVCAR3 cells, that express high levels of the protein. Immunohistochemical studies showed claudin-4 staining in a perinuclear region, at most plasma membranes and in cytoplasmic puncta. Native claudin-4 did not overlap with phosphorylated claudin-4, which was partially located in focal adhesions. Using claudin-4 BioID technology identified more precisely we confirmed that large amounts of claudin-4 are localized to the Golgi compartment, including in dispersed Golgi in cells where claudin-4 is partially knocked down and in dividing cells. Claudin-4 appears to be present in the vicinity of several types of cell-cell junctions, but there is no evidence that it forms tight junctions in these tumor cells. Both claudin-4, the Golgi marker GM130, and the plasma membrane receptor Notch2 were found in dispersed Golgi in dividing cells. This definition of the cellular architecture of claudin-4 should provide a framework for better understanding of the function of claudin-4 in tumor cells and its molecular interactions.
... E-cadherin is the crucial factor involved in Epithelial-Mesenchymal Transition (EMT) and helps in cell connection. E-cadherin downregulation could be an important factor for cancer cell migration and metastasis (Ishiyama et al., 2010;Wang et al., 2017) and re-expression of this molecule is known to block invasiveness (Birchmeier and Behrens, 1994). In addition, clinical studies report Matrix metalloproteinase (MMP9) expression with progression of gynecological cancers. ...
Cervical cancer is leading cause of cancer death in females worldwide. Vaginal lactobacilli colonizing cervical area are known to play an important role in maintaining cervical physiological conditions to ward away vaginal infections including bacterial vaginosis (BV) and cancer prevention. There are limited studies to study effect of Lactobacilli isolated from different sources on cervical cancer. The objective of the study was to investigate the potential of cell-free culture supernatants (CFCs) or metabolites of twelve well-characterized Lactobacillus species from different microenvironments for their anti-proliferative properties on HPV16 and HPV18 cervical cancer cells and to investigate the mechanisms of anti-proliferative and anti-metastatic activities. Lactobacillus metabolites exerted a dose, strain and cell line-dependent effect on cervical cells as demonstrated by 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide (MTT) assay. The metabolites from vaginalis and L. salivarius exhibited the lowest half-maximal inhibitory concentration (IC50) on HeLa (131 and 167 ng/ml) respectively and SiHa (149 and 205ng /ml) respectively. Lactobacilli demonstrating greater inhibitory effect produced majorly L-lactic acid and hydrogen peroxide (H2O2). Treatment with lactobacilli CFCs significantly upregulated E- cadherin levels in HeLa (p=0.0451) and SiHa (p=0.0051) cells and downregulated matrix metalloproteinase 9 (MMP9) levels in Hela cells (p=0.0465) as measured by ELISA. Lactobacillus-derived metabolites could be explored as biotherapeutics for the control of HPV infections and cervical cancer.
... (Ishiyama et al., 2010), DLG1(Marziali et al., 2019), and CXADR (Nilchian et al., 2019) were. The heavy staining of DLG1 in OVCAR3 cells (Figure 5C) is consistent with the hypothesis that adherens junctions may be important in cell-cell interactions of ovarian cancer cells.Members of the protein families that make up desmosomes (Figure 5A (Green et al., 2019);), the transmembrane cadherin family (desmoglein, DSG2, and desmocollin, DSC2), the armadillo family protein (plakoglobin, JUP) ...
... The p120-catenin subfamily of catenins strengthen the cadherin complex through facilitating the clustering of cadherin monomers into structures called cadherin nanoclusters (Yap et al., 1998;Ishiyama et al., 2010;Yap et al., 2015). These nanoclusters have historically been referred to as spotty or punctate adherens junctions (pAJs) and are observed in lens fiber cells with electron microscopy (Lo, 1988;Niessen and Gottardi, 2008). ...
The etiology of age-related cortical cataracts is not well understood but is speculated to be related to alterations in cell adhesion and/or the changing mechanical stresses occurring in the lens with time. The role of cell adhesion in maintaining lens transparency with age is difficult to assess because of the developmental and physiological roles that well-characterized adhesion proteins have in the lens. This report demonstrates that Arvcf, a member of the p120-catenin subfamily of catenins that bind to the juxtamembrane domain of cadherins, is an essential fiber cell protein that preserves lens transparency with age in mice. No major developmental defects are observed in the absence of Arvcf, however, cortical cataracts emerge in all animals examined older than 6-months of age. While opacities are not obvious in young animals, histological anomalies are observed in lenses at 4-weeks that include fiber cell separations, regions of hexagonal lattice disorganization, and absence of immunolabeled membranes. Compression analysis of whole lenses also revealed that Arvcf is required for their normal biomechanical properties. Immunofluorescent labeling of control and Arvcf-deficient lens fiber cells revealed a reduction in membrane localization of N-cadherin, β-catenin, and αN-catenin. Furthermore, super-resolution imaging demonstrated that the reduction in protein membrane localization is correlated with smaller cadherin nanoclusters. Additional characterization of lens fiber cell morphology with electron microscopy and high resolution fluorescent imaging also showed that the cellular protrusions of fiber cells are abnormally elongated with a reduction and disorganization of cadherin complex protein localization. Together, these data demonstrate that Arvcf is required to maintain transparency with age by mediating the stability of the N-cadherin protein complex in adherens junctions.
... While this variant is expressed throughout the fetal and adult brain, the shorter variant lacking this insert is predominant in most regions of the developing and mature brain (Kawamura et al., 1999). Currently, there are no known functional differences between these two isoforms (Ishiyama et al., 2010), but the location of the 25-residue insert on the longer isoform suggests a potential role in regulating the structure and function of the insert region (Yuan and Arikkath, 2017). Delta-catenin has established roles at adherens junctions where it stabilizes N-cadherin (Fannon and Colman, 1996;Benson and Tanaka, 1998;Israely et al., 2004;Yuan et al., 2015). ...
Small Rho GTPases are molecular switches that are involved in multiple processes including regulation of the actin cytoskeleton. These GTPases are activated (turned on) and inactivated (turned off) through various upstream effector molecules to carry out many cellular functions. One such upstream modulator of small Rho GTPase activity is delta-catenin, which is a protein in the p120-catenin subfamily that is enriched in the central nervous system. Delta-catenin affects small GTPase activity to assist in the developmental formation of dendrites and dendritic spines and to maintain them once they mature. As the dendritic arbor and spine density are crucial for synapse formation and plasticity, delta-catenin’s ability to modulate small Rho GTPases is necessary for proper learning and memory. Accordingly, the misregulation of delta-catenin and small Rho GTPases has been implicated in several neurological and non-neurological pathologies. While links between delta-catenin and small Rho GTPases have yet to be studied in many contexts, known associations include some cancers, Alzheimer’s disease (AD), Cri-du-chat syndrome, and autism spectrum disorder (ASD). Drawing from established studies and recent discoveries, this review explores how delta-catenin modulates small Rho GTPase activity. Future studies will likely elucidate how PDZ proteins that bind delta-catenin further influence small Rho GTPases, how delta-catenin may affect small GTPase activity at adherens junctions when bound to N-cadherin, mechanisms behind delta-catenin’s ability to modulate Rac1 and Cdc42, and delta-catenin’s ability to modulate small Rho GTPases in the context of diseases, such as cancer and AD.
... p120 was an upstream regulator of neurogenesis and cell cycle pathways in glioma patients and a predictor of poor clinical prognosis [36]. p120 could inhibit RhoA and stabilize the cadherin and β-catenin on the plasma membrane [37]. p120 stabilized the membrane association of β-catenin, thereby preventing the accumulation of β-catenin nuclei and the excessive activation of the WNTpathway during EMT [38]. ...
Background:
Oral submucous fibrosis (OSF) is a potentially malignant disease of the oral cavity. New molecular predictors are needed to identify the high risk of malignant transformation in potentially malignant oral lesions. Our purpose is to explore PTPRZ1 and p120/β-catenin pathogenesis in the carcinogenesis of OSF to identify novel drug targets.
Methods:
The expression of PTPRZ1, p120, and β-catenin in clinical tissues was detected. Then, PTPRZ1, p120, β-catenin, RhoA, Rac1, CDC42, cyclin D1, and c-myc expressions were detected by qRT-PCR and western blot. CCK-8 was applied to measure hOMF cells viability. Wound healing and transwell assay were applied to measure cell migration and invasion. Western blot and IF detected the distribution of p-p120 and p-β-catenin. Tumor formation experiment explored PTPRZ1 effects on OSF.
Results:
PTPRZ1, p120, and β-catenin were abnormally expressed in cancer tissues. PTPRZ1 regulated the phosphorylation of p120/β-catenin. Western blot and IF showed that in the oe-NC group, p-p120 and p-β-catenin were expressed in the cell membrane. p-p120 and p-β-catenin were expressed in the cytoplasm and nucleus of the oe-PTPRZ1 group. In vitro experimental results revealed overexpression of PTPRZ1 and β-catenin, and silencing of p120 promoted cell proliferation, migration, and invasion. The tumor volume and weight in the sh-PTPRZ1 group were significantly reduced. IHC revealed the positive rate of PTPRZ1 was also low.
Conclusions:
Overexpression of PTPRZ1 regulated the phosphorylation of p120/β-catenin to promote OSF malignancy.
Plasmodium falciparum, the causative agent of malaria, poses a significant global health challenge, yet much of its biology remains elusive. A third of the genes in the P. falciparum genome lack annotations regarding their function, impeding our understanding of the parasite's biology. In this study, we employ structure predictions and the DALI search algorithm to analyse proteins encoded by uncharacterized genes in the reference strain 3D7 of P. falciparum. By comparing AlphaFold predictions to experimentally determined protein structures in the Protein Data Bank, we found similarities to known domains in 353 proteins of unknown function, shedding light on their potential functions. The lowest-scoring 5% of similarities were additionally validated using the size-independent TM-align algorithm, confirming the detected similarities in 88% of the cases. Notably, in over 70 P. falciparum proteins the presence of domains resembling heptatricopeptide repeats, which are typically involvement in RNA binding and processing, was detected. This suggests this family, which is important in transcription in mitochondria and apicoplasts, is much larger in Plasmodium parasites than previously thought. The results of this domain search provide a resource to the malaria research community that is expected to inform and enable experimental studies.
α-catenin (α-cat) displays force-dependent unfolding and binding to actin filaments through direct and indirect means, but features of adherens junction structure and function most vulnerable to loss of these allosteric mechanisms have not been directly compared. By reconstituting an α-cat F-actin-binding domain unfolding mutant known to exhibit enhanced binding to actin (α-cat-H0-FABD ⁺ ) into α-cat knock-out Madin Darby Canine Kidney (MDCK) cells, we show that partial loss of the α-cat catch bond mechanism (via an altered H0 α-helix) leads to stronger epithelial sheet integrity with greater co-localization between the α-cat-H0-FABD ⁺ mutant and actin. α-cat-H0-FABD ⁺ -expressing cells are less efficient at closing scratch-wounds, suggesting reduced capacity for more dynamic cell-cell coordination. Evidence that α-cat-H0-FABD ⁺ is equally accessible to the conformationally sensitive α18 antibody epitope as WT α-cat and shows similar vinculin recruitment suggests this mutant engages lower tension cortical actin networks, as its M-domain is not persistently open. Conversely, α-cat-M-domain salt-bridge mutants with persistent recruitment of vinculin and phosphorylated myosin light chain show only intermediate monolayer adhesive strengths, but display less directionally coordinated and thereby slower migration speeds during wound-repair. These data show α-cat M- and FABD-unfolding mutants differentially impact cell-cell cohesion and migration properties, and suggest signals favoring α-cat-cortical actin interaction without persistent M-domain opening may improve epithelial monolayer strength through enhanced coupling to lower tension actin networks.
[Media: see text]
Epithelial tissue forms and maintains a critical barrier function in the body. A novel culture design aimed at promoting uniform maturation of epithelial cells using liquid materials is described. Culturing Madin–Darby canine kidney (MDCK) cells at the liquid–liquid interface yielded reduced migration and stimulated active cell growth. Similar to solid–liquid interfaces, cells cultured on a fibronectin-coated liquid–liquid interface exhibited active migration and growth, ultimately reaching a confluent state. These cells exhibited reduced stress fiber formation and adopted a cobblestone-like shape, which led to their even distribution in the culture vessel. To inhibit stress fiber formation and apoptosis, the exposure of cells on liquid–liquid interfaces to Y27632, a specific inhibitor of the Rho-associated protein kinase (ROCK), facilitated tight junction formation (frequency of ZO-2-positive cells, F Z = 0.73). In Y27632-exposed cells on the liquid–liquid interface, the value obtained by subtracting the standard deviation of the ratio of nucleus densities in each region that compartmentalized a culture vessel from 1, denoted as H LN , was 0.93 0.01, indicated even cell distribution in the culture vessel at t = 72 h. The behavior of epithelial cells on liquid–liquid interfaces contributes to the promotion of their uniform maturation.
Epithelial tissue forms and maintains a critical barrier function in the body. A novel culture design aimed at promoting uniform maturation of epithelial cells using liquid materials is described. Culturing Madin–Darby canine kidney (MDCK) cells at the liquid–liquid interface yielded reduced migration and stimulated active cell growth. Similar to solid–liquid interfaces, cells cultured on a fibronectin-coated liquid–liquid interface exhibited active migration and growth, ultimately reaching a confluent state. These cells exhibited reduced stress fiber formation and adopted a cobblestone-like shape, which led to their even distribution in the culture vessel. To inhibit stress fiber formation and apoptosis, the exposure of cells on liquid–liquid interfaces to Y27632, a specific inhibitor of the Rho-associated protein kinase (ROCK), facilitated tight junction formation (frequency of ZO-2-positive cells, F Z = 0.73). In Y27632-exposed cells on the liquid–liquid interface, the value obtained by subtracting the standard deviation of the ratio of nucleus densities in each region that compartmentalized a culture vessel from 1, denoted as H LN , was 0.93 0.01, indicated even cell distribution in the culture vessel at t = 72 h. The behavior of epithelial cells on liquid–liquid interfaces contributes to the promotion of their uniform maturation.
Epithelial tissue forms and maintains a critical barrier function in the body. A novel culture design aimed at promoting uniform maturation of epithelial cells using liquid materials is described. Culturing Madin–Darby canine kidney (MDCK) cells at the liquid–liquid interface yielded reduced migration and stimulated active cell growth. Similar to solid–liquid interfaces, cells cultured on a fibronectin-coated liquid–liquid interface exhibited active migration and growth, ultimately reaching a confluent state. These cells exhibited reduced stress fiber formation and adopted a cobblestone-like shape, which led to their even distribution in the culture vessel. To inhibit stress fiber formation and apoptosis, the exposure of cells on liquid–liquid interfaces to Y27632, a specific inhibitor of the Rho-associated protein kinase (ROCK), facilitated tight junction formation (frequency of ZO-2-positive cells, FZ = 0.73). In Y27632-exposed cells on the liquid–liquid interface, the value obtained by subtracting the standard deviation of the ratio of nucleus densities in each region that compartmentalized a culture vessel from 1, denoted as HLN, was 0.93 ± 0.01, indicated even cell distribution in the culture vessel at t = 72 h. The behavior of epithelial cells on liquid–liquid interfaces contributes to the promotion of their uniform maturation.
COPD causes significant morbidity and mortality worldwide. Epithelial damage is fundamental to disease pathogenesis, although the mechanisms driving disease remain undefined. Published evidence from a COPD cohort (SPIROMICS) and confirmed in a second cohort (COPDgene) demonstrate a polymorphism in Fucosyltransferese-2 (FUT2) is a trans-pQTL for E-cadherin, which is critical in COPD pathogenesis. We found by MALDI-TOF analysis that FUT2 increased terminal fucosylation of E-cadherin. Using atomic force microscopy, we found that FUT2-dependent fucosylation enhanced E-cadherin-E-cadherin bond strength, mediating the improvement in monolayer integrity. Tracheal epithelial cells from Fut2-/- mice have reduced epithelial integrity, which is recovered with reconstitution of Fut2. Overexpression of FUT2 in COPD derived epithelia rescues barrier function. Fut2-/- mice show increased susceptibility in an elastase model of disease developing both emphysema and fibrosis. We propose this is due to the role of FUT2 in proliferation and cell differentiation. Overexpression of FUT2 significantly increased proliferation. Loss of Fut2 results in accumulation of Spc+ cells suggesting a failure of alveolar type 2 cells to undergo transdifferentiation to alveolar type 1. Using a combination of population data, genetically manipulated mouse models, and patient-derived cells, we present a novel mechanism by which post-translational modifications modulate tissue pathology and serve as a proof of concept for the development of a disease-modifying target in COPD.
E-cadherin is an essential cell‒cell adhesion protein that mediates canonical cadherin-catenin complex formation in epithelial lateral membranes. Ankyrin-G (AnkG), a scaffold protein linking membrane proteins to the spectrin-based cytoskeleton, coordinates with E-cadherin to maintain epithelial cell polarity. However, the molecular mechanisms governing this complex formation and its relationships with the cadherin-catenin complex remain elusive. Here, we report that AnkG employs a promiscuous manner to encapsulate three discrete sites of E-cadherin by the same region, a dynamic mechanism that is distinct from the canonical 1:1 molar ratio previously described for other AnkG or E-cadherin-mediated complexes. Moreover, we demonstrate that AnkG-binding-deficient E-cadherin exhibited defective accumulation at the lateral membranes and show that disruption of interactions resulted in cell polarity malfunction. Finally, we demonstrate that E-cadherin is capable of simultaneously anchoring to AnkG and β-catenin, providing mechanistic insights into the functional orchestration of the ankyrin-spectrin complex with the cadherin-catenin complex. Collectively, our results show that complex formation between E-cadherin and AnkG is dynamic, which enables the maintenance of epithelial cell polarity by ensuring faithful targeting of the adhesion molecule-scaffold protein complex, thus providing molecular mechanisms for essential E-cadherin-mediated complex assembly at cell‒cell junctions.
A prototype of cross-membrane signal transduction is that extracellular binding of cell surface receptors to their ligands induces intracellular signaling cascades. However, much less is known about the process in the opposite direction, called inside-out signaling. Recent studies show that it plays a more important role in regulating the functions of many cell surface receptors than we used to think. In particular, in cadherin-mediated cell adhesion, recent experiments indicate that intracellular binding of the scaffold protein p120-catenin can promote extracellular clustering of cadherin and alter its adhesive function. The underlying mechanism, however, is not well understood. To explore possible mechanisms, we designed a new multiscale simulation procedure. Using all-atom molecular dynamics simulations, we found that the conformational dynamics of the cadherin extracellular region can be altered by the intracellular binding of p120-catenin. More intriguingly, by integrating all-atom simulation results into coarse-grained random sampling, we showed that the altered conformational dynamics of cadherin caused by the binding of p120-catenin can increase the probability of lateral interactions between cadherins on the cell surface. These results suggest that p120-catenin could allosterically regulate the cis-dimerization of cadherin through two mechanisms. First, p120-catenin controls the extracellular conformational dynamics of cadherin. Second, p120-catenin oligomerization can further promote cadherin clustering. Our study, therefore, suggests a mechanistic foundation for the inside-out signaling in cadherin-mediated cell adhesion, while the computational framework can be generally applied to other cross-membrane signal transduction systems.
Epithelial cells can become polyploid upon tissue injury, but mechanosensitive cues that trigger this state are poorly understood. Using α-catenin (α-cat) knock-out Madin Darby Canine Kidney (MDCK) cells reconstituted with wild-type and mutant forms of α-cat as a model system, we find that an established α-cat actin-binding domain unfolding mutant designed to reduce force-sensitive binding to F-actin (α-cat-H0-FABD+) can promote cytokinesis failure, particularly along epithelial wound-fronts. Enhanced α-cat coupling to cortical actin is neither sufficient nor mitotic cell-autonomous for cytokinesis failure, but critically requires the mechanosensitive Middle-domain (M1-M2-M3) and neighboring cells. Disease relevant α-cat M-domain missense mutations known to cause a form of retinal pattern dystrophy (α-cat E307K or L436P) are associated with elevated binucleation rates via cytokinesis failure. Similar binucleation rates are seen in cells expressing an α-cat salt-bridge destabilizing mutant (R551A) designed to promote M2-M3 domain unfurling at lower force thresholds. Since binucleation is strongly enhanced by removal of the M1 as opposed to M2-M3 domains, cytokinetic fidelity is most sensitive to α-cat M2-M3 domain opening. To identify α-cat conformation-dependent proximity partners that contribute to cytokinesis, we used a biotin-ligase approach to distinguished proximity partners that show enhanced recruitment upon α-cat M-domain unfurling (R551A). We identified Leucine Zipper Tumor Suppressor 2 (LZTS2), an abscission factor previously implicated in cytokinesis. We confirm that LZTS2 enriches at the midbody, but discover it also localizes to tight and tricellular junctions. LZTS2 knock-down promotes binucleation in both MDCK and Retinal Pigmented Epithelial (RPE) cells. α-cat mutants with persistent M2-M3 domain opening showed elevated junctional enrichment of LZTS2 from the cytosol compared α-cat wild-type cells. These data implicate LZTS2 as a mechanosensitive effector of α-cat that is critical for cytokinetic fidelity. This model rationalizes how persistent mechano-activation of α-cat may drive tension-induced polyploidization of epithelia post-injury and suggests an underlying mechanism for how pathogenic α-cat mutations drive macular dystrophy.
Cadherin family proteins play a central role in epithelial and endothelial cell-cell adhesion. The dynamic regulation of cell adhesion is achieved in part through endocytic membrane trafficking pathways that modulate cadherin cell surface levels. Here, we define the role for various MARCH family ubiquitin ligases in the regulation of cadherin degradation. We find that MARCH2 selectively downregulates VE-cadherin, resulting in loss of adherens junction proteins at cell borders and a loss of endothelial barrier function. Interestingly, N-cadherin is refractory to MARCH ligase expression, demonstrating that different classical cadherin family proteins are differentially regulated by MARCH family ligases. Using chimeric cadherins, we find that the specificity of different MARCH family ligases for different cadherins is conferred by the cadherin transmembrane domain. Further, juxta-membrane lysine residues are required for cadherin degradation by MARCH proteins. These findings expand our understanding of cadherin regulation and highlight a new role for mammalian MARCH family ubiquitin ligases in differentially regulating cadherin turnover.
Plasmodium falciparum , the causative agent of malaria, poses a significant global health challenge, yet much of its biology remains elusive. A third of the genes in the P. falciparum genome lack annotations regarding their function, impeding our understanding of the parasite’s biology. In this study, we employed structure predictions and the DALI search algorithm to analyse proteins encoded by uncharacterized genes in the reference strain 3D7 of P. falciparum .
By comparing Alphafold predictions to experimentally determined protein structures in the Protein Data Bank, we found similarities to known domains in 353 proteins of unknown function, shedding light on their potential functions. The lowest-scoring 5% of similarities were additionally validated using the size-independent TM-align algorithm, confirming the detected similarities in 88% of the cases. Notably, in over 70 P. falciparum proteins the presence of domains resembling heptatricopeptide repeats, which are typically involvement in RNA binding and processing, was detected. This suggests this family, which is important in transcription in mitochondria and apicoplasts, is much larger in Plasmodium parasites than previously thought. The results of this domain search provide a resource to the malaria research community that is expected to inform and enable experimental studies.
Breast cancer is one of the most common invasive cancers among women. The leading cause of difficulty in treating breast cancer patients is metastasis. Because cell migration is closely related to breast cancer metastasis, elucidating the detailed mechanism by which breast cancer cells promote their migration is crucial for improving the prognosis of patients. In this study, we investigated the relationship between breast cancer cell migration and Mind bomb1 (MIB1), an E3 ubiquitin ligase. We found that the downregulation of MIB1 promotes the cell migration of MCF7, a breast cancer-derived cell line. Furthermore, knockdown of MIB1 caused a reduction in CTNND1 and thereby impaired E-cadherin membrane localization in the cell boundary region. Taken together, our data suggest that MIB1 might play a role in suppressing breast cancer cell migration.
The cell-cell adhesion cadherin-catenin complexes recruit vinculin to the adherens junction (AJ) to modulate the mechanical couplings between neighboring cells. However, it is unclear how vinculin influences the AJ structure and function. Here, we identified two patches of salt bridges that lock vinculin in the head-tail autoinhibited conformation and reconstituted the full-length vinculin activation mimetics bound to the cadherin-catenin complex. The cadherin-catenin-vinculin complex contains multiple disordered linkers and is highly dynamic, which poses a challenge for structural studies. We determined the ensemble conformation of this complex using small angle X-ray and selective deuteration/contrast variation small angle neutron scattering. In the complex, both α-catenin and vinculin adopt an ensemble of flexible conformations, but vinculin has fully open conformations with the vinculin head and actin-binding tail domains well separated from each other. F-actin binding experiments show that the cadherin-catenin-vinculin complex binds and bundles F-actin. However, when the vinculin actin-binding domain is removed from the complex, only a minor fraction of the complex binds to F-actin. The results show that the dynamic cadherin-catenin-vinculin complex employs vinculin as the primary F-actin binding mode to strengthen AJ-cytoskeleton interactions.
The cardiac sodium channel NaV1.5 is an essential modulator of cardiac excitability, with decreased NaV1.5 levels at the plasma membrane and consequent reduction in sodium current (INa) leading to potentially lethal cardiac arrhythmias. NaV1.5 is distributed in a specific pattern at the plasma membrane of cardiomyocytes, with localization at the crests, grooves, and T-tubules of the lateral membrane, and particularly high levels at the intercalated disc region. NaV1.5 forms a large macromolecular complex with and is regulated by interacting proteins, some of which are specifically localised at either the lateral membrane or intercalated disc. One of the NaV1.5 trafficking routes is via microtubules (MTs), which are regulated by MT plus-end tracking proteins (+TIPs). In our search for mechanisms involved in targeted delivery of NaV1.5, we here provide an overview of previously demonstrated interactions between NaV1.5 interacting proteins and +TIPs, which potentially (in)directly impact on NaV1.5 trafficking. Strikingly, +TIPs interact extensively with several intercalated disc- and lateral membrane-specific NaV1.5 interacting proteins. Recent work indicates that this interplay of +TIPs and NaV1.5 interacting proteins mediates the targeted delivery of NaV1.5 at specific cardiomyocyte subcellular domains, while also being potentially relevant for the trafficking of other ion channels. These observations are especially relevant for diseases associated with loss of NaV1.5 specifically at the lateral membrane (such as Duchenne muscular dystrophy), or at the intercalated disc (for example, arrhythmogenic cardiomyopathy), and open up potential avenues for development of new anti-arrhythmic therapies.
The phylum of Apicomplexa groups intracellular parasites that employ substrate-dependent gliding motility to invade host cells, egress from the infected cells and cross biological barriers. The glideosome associated connector (GAC) is a conserved protein essential to this process. GAC facilitates the association of actin filaments with surface transmembrane adhesins and the efficient transmission of the force generated by myosin translocation of actin to the cell surface substrate. Here, we present the crystal structure of Toxoplasma gondii GAC and reveal a unique, supercoiled armadillo repeat region that adopts a closed ring conformation. Characterisation of the solution properties together with membrane and F-actin binding interfaces suggest that GAC adopts several conformations from closed to open and extended. A multi-conformational model for assembly and regulation of GAC within the glideosome is proposed.
TBK1-IRF3 complex plays vital roles in antiviral immune responses, its regulatory mechanisms are currently incompletely understood. p120-catenin (p120), an armadillo-repeat protein, mainly regulates the stability of classical cadherins and the development of epithelial-to-mesenchymal transitions (EMTs). Here we report that p120 is a positive regulator of type I IFN production. Ectopic expression of p120 enhanced Vesicular stomatitis virus and Sendai-virus-induced type I IFN production, whereas knockdown of p120 expression suppressed type I IFN production. Mechanistically, p120 promoted phosphorylation of IRF3 via stabilizing the TBK1-IRF3 complex. Consistently, p120 knock down mice are more susceptible to VSV infection as indicated by higher tissue viral titers, less IFN-I production and greater infiltration of immune cells. This study reveals p120 as an important positive regulator in innate immunity and identifies that p120 facilitates host antiviral response through stabilizing TBK1-IRF3 complex.
Metastasis is a pivotal event that accelerates the prognosis of cancer patients towards mortality. Therapies that aim to induce cell death in metastatic cells require a more detailed understanding of the metastasis for better mitigation. Towards this goal, we discuss the details of two distinct but overlapping pathways of metastasis: a classical reversible epithelial-to-mesenchymal transition (hybrid-EMT)-driven transport pathway and an alternative cell death process-driven blebbishield metastatic-witch (BMW) transport pathway involving reversible cell death process. The knowledge about the EMT and BMW pathways is important for the therapy of metastatic cancers as these pathways confer drug resistance coupled to immune evasion/suppression. We initially discuss the EMT pathway and compare it with the BMW pathway in the contexts of coordinated oncogenic, metabolic, immunologic, and cell biological events that drive metastasis. In particular, we discuss how the cell death environment involving apoptosis, ferroptosis, necroptosis, and NETosis in BMW or EMT pathways recruits immune cells, fuses with it, migrates, permeabilizes vasculature, and settles at distant sites to establish metastasis. Finally, we discuss the therapeutic targets that are common to both EMT and BMW pathways.
Background
The cadherin‐associated protein p120 catenin regulates convergent extension through interactions with cadherin proteins, Cdc42, and Rac1, as we previously showed in zebrafish (Danio rerio). Phosphorylation of p120 catenin changes the nature of its activity in vitro but is virtually unexplored in embryos. We used our previously developed antisense RNA splice‐site morpholino targeted to endogenous p120 catenin‐δ1 to cause defects in axis elongation probing the functions of three p120 catenin tyrosine‐phosphorylation sites in gastrulating zebrafish embryos.
Results
The morpholino‐induced defects were rescued by co‐injections with mouse p120 catenin‐δ1‐3A mRNAs mutated at residues Y228 and Y217 to a non‐phosphorylatable phenylalanine (F) or mutated at residue Y335 to a phosphomimetic glutamic acid (E). Co‐injection of the complementary mutations Y228E, Y217E, or Y335F mRNAs partially rescued embryos whereas dual mutation to Y228E‐Y217E blocked rescue. Immunopurification showed Y228F mutant proteins preferentially interacted with Rac1, potentially promoting cell migration. In contrast, the phosphomimetic Y228E preferentially interacted with E‐cadherin increasing adhesion. Both Y228F and Y335F strongly bind VAV2.
Conclusions
p120 catenin serves dual roles during gastrulation of zebrafish. Phosphorylation and dephosphorylation of tyrosine residues Y217, Y228, and Y335 precisely balance cell adhesion and cell migration to facilitate somite compaction and axis elongation.
Familial exudative vitreoretinopathy (FEVR) is a hereditary disorder that can cause vision loss. The CTNND1 gene encodes a cellular adhesion protein p120-catenin (p120), which is essential for vascularization, yet the function of p120 in postnatal physiological angiogenesis remains unclear. Here, we applied whole-exome sequencing (WES) on 140 probands of FEVR families and identified three candidate variants in the human CTNND1 gene. We performed inducible deletion of Ctnnd1 in the postnatal mouse endothelial cells (ECs) and observed typical phenotypes of FEVR. Immunofluorescence of retina flat mounts also revealed immune responses, including reactive astrogliosis and microgliosis accompanied by abnormal Vegfa expression. Using an unbiased proteomics analysis in combination with in vivo or in vitro approaches, we propose that p120 is critical for the integrity of cadherin/catenin complex, and that p120 activates Wnt signaling activity by protecting β-catenin from Gsk3β-ubiquitin-guided degradation. Treatment of CTNND1-depleted HRECs with Gsk3β inhibitors LiCl or CHIR-99021 successfully enhanced cell proliferation by preventing β-catenin from degradation. Moreover, LiCl treatment increased vessel density in Ctnnd1-deficient mouse retinas. Functional analysis also revealed that variants in CTNND1 cause FEVR by compromising the expression of adherens junctions (AJs) and Wnt signaling activity. Additionally, genetic interactions between p120 and β-catenin or α-catenin revealed by double heterozygous deletion in mice further confirmed that p120 regulates vascular development through the Wnt/Cadherin axis. Together, we propose that CTNND1 is a novel candidate gene associated with FEVR, and that variants in CTNND1 can cause FEVR through the Wnt/Cadherin axis.
Publisher Summary X-ray data can be collected with zero-, one-, and two-dimensional detectors, zero-dimensional (single counter) being the simplest and two-dimensional the most efficient in terms of measuring diffracted X-rays in all directions. To analyze the single-crystal diffraction data collected with these detectors, several computer programs have been developed. Two-dimensional detectors and related software are now predominantly used to measure and integrate diffraction from single crystals of biological macromolecules. Macromolecular crystallography is an iterative process. To monitor the progress, the HKL package provides two tools: (1) statistics, both weighted (χ 2 ) and unweighted (R-merge), where the Bayesian reasoning and multicomponent error model helps obtain proper error estimates and (2) visualization of the process, which helps an operator to confirm that the process of data reduction, including the resulting statistics, is correct and allows the evaluation of the problems for which there are no good statistical criteria. Visualization also provides confidence that the point of diminishing returns in data collection and reduction has been reached. At that point, the effort should be directed to solving the structure. The methods presented in the chapter have been applied to solve a large variety of problems, from inorganic molecules with 5 A unit cell to rotavirus of 700 A diameters crystallized in 700 × 1000 × 1400 A cell.
The NMRPipe system is a UNIX software environment of processing, graphics, and analysis tools designed to meet current routine and research-oriented multidimensional processing requirements, and to anticipate and accommodate future demands and developments. The system is based on UNIX pipes, which allow programs running simultaneously to exchange streams of data under user control. In an NMRPipe processing scheme, a stream of spectral data flows through a pipeline of processing programs, each of which performs one component of the overall scheme, such as Fourier transformation or linear prediction. Complete multidimensional processing schemes are constructed as simple UNIX shell scripts. The processing modules themselves maintain and exploit accurate records of data sizes, detection modes, and calibration information in all dimensions, so that schemes can be constructed without the need to explicitly define or anticipate data sizes or storage details of real and imaginary channels during processing. The asynchronous pipeline scheme provides other substantial advantages, including high flexibility, favorable processing speeds, choice of both all-in-memory and disk-bound processing, easy adaptation to different data formats, simpler software development and maintenance, and the ability to distribute processing tasks on multi-CPU computers and computer networks.
Phaser is a program for phasing macromolecular crystal structures by both molecular replacement and experimental phasing methods. The novel phasing algorithms implemented in Phaser have been developed using maximum likelihood and multivariate statistics. For molecular replacement, the new algorithms have proved to be significantly better than traditional methods in discriminating correct solutions from noise, and for single-wavelength anomalous dispersion experimental phasing, the new algorithms, which account for correlations between F
+ and F
−, give better phases (lower mean phase error with respect to the phases given by the refined structure) than those that use mean F and anomalous differences ΔF. One of the design concepts of Phaser was that it be capable of a high degree of automation. To this end, Phaser (written in C++) can be called directly from Python, although it can also be called using traditional CCP4 keyword-style input. Phaser is a platform for future development of improved phasing methods and their release, including source code, to the crystallographic community.
Most transmembrane proteins are selected as transport-vesicle cargo through the recognition of short, linear amino-acid motifs in their cytoplasmic portions by vesicle coat proteins. For clathrin-coated vesicles, the motifs are recognized by clathrin adaptors. The AP2 adaptor complex (subunits alpha, beta2, mu2 and sigma2) recognizes both major endocytic motifs: YxxPhi motifs (where Phi can be F, I, L, M or V) and [ED]xxxL[LI] acidic dileucine motifs. Here we describe the binding of AP2 to the endocytic dileucine motif from CD4 (ref. 2). The major recognition events are the two leucine residues binding in hydrophobic pockets on sigma2. The hydrophilic residue four residues upstream from the first leucine sits on a positively charged patch made from residues on the sigma2 and alpha subunits. Mutations in key residues inhibit the binding of AP2 to `acidic dileucine' motifs displayed in liposomes containing phosphatidylinositol-4,5-bisphosphate, but do not affect binding to YxxPhi motifs through mu2. In the `inactive' AP2 core structure both motif-binding sites are blocked by different parts of the beta2 subunit. To allow a dileucine motif to bind, the beta2 amino terminus is displaced and becomes disordered; however, in this structure the YxxPhi-binding site on mu2 remains blocked.
Localization of presynaptic components to synaptic sites is critical for hippocampal synapse formation. Cell adhesion-regulated signaling is important for synaptic development and function, but little is known about differentiation of the presynaptic compartment. In this study, we describe a pathway that promotes presynaptic development involving p120catenin (p120ctn), the cytoplasmic tyrosine kinase Fer, the protein phosphatase SHP-2, and beta-catenin. Presynaptic Fer depletion prevents localization of active zone constituents and synaptic vesicles and inhibits excitatory synapse formation and synaptic transmission. Depletion of p120ctn or SHP-2 similarly disrupts synaptic vesicle localization with active SHP-2, restoring synapse formation in the absence of Fer. Fer or SHP-2 depletion results in elevated tyrosine phosphorylation of beta-catenin. beta-Catenin overexpression restores normal synaptic vesicle localization in the absence of Fer or SHP-2. Our results indicate that a presynaptic signaling pathway through p120ctn, Fer, SHP-2, and beta-catenin promotes excitatory synapse development and function.
p120cas is a tyrosine kinase substrate implicated in ligand-induced receptor signaling through the epidermal growth factor, platelet-derived growth factor, and colony-stimulating factor receptors and in cell transformation by Src. Here we report that p120 associates with a complex containing E-cadherin, alpha-catenin, beta-catenin, and plakoglobin. Furthermore, p120 precisely colocalizes with E-cadherin and catenins in vivo in both normal and Src-transformed MDCK cells. Unlike beta-catenin and plakoglobin, p120 has at least four isoforms which are differentially expressed in a variety of cell types, suggesting novel means of modulating cadherin activities in cells. In Src-transformed MDCK cells, p120, beta-catenin, and plakoglobin were heavily phosphorylated on tyrosine, but the physical associations between these proteins were not disrupted. Association of p120 with the cadherin machinery indicates that both Src and receptor tyrosine kinases cross talk with proteins important for cadherin-mediated cell adhesion. These results also strongly suggest a role for p120 in cell adhesion.
The tyrosine kinase substrate p120cas (CAS), which is structurally similar to the cell adhesion proteins beta-catenin and plakoglobin, was recently shown to associate with the E-cadherin-catenin cell adhesion complex. beta-catenin, plakoglobin, and CAS all have an Arm domain that consists of 10 to 13 repeats of a 42-amino-acid motif originally described in the Drosophila Armadillo protein. To determine if the association of CAS with the cadherin cell adhesion machinery is similar to that of beta-catenin and plakoglobin, we examined the CAS-cadherin-catenin interactions in a number of cell lines and in the yeast two-hybrid system. In the prostate carcinoma cell line PC3, CAS associated normally with cadherin complexes despite the specific absence of alpha-catenin in these cells. However, in the colon carcinoma cell line SW480, which has negligible E-cadherin expression, CAS did not associate with beta-catenin, plakoglobin, or alpha-catenin, suggesting that E-cadherin is the protein which bridges CAS to the rest of the complex. In addition, CAS did not associate with the adenomatous polyposis coli (APC) tumor suppressor protein in any of the cell lines analyzed. Interestingly, expression of the various CAS isoforms was quite heterogeneous in these tumor cell lines, and in the colon carcinoma cell line HCT116, which expresses normal levels of E-cadherin and the catenins, the CAS1 isoforms were completely absent. By using the yeast two-hybrid system, we confirmed the direct interaction between CAS and E-cadherin and determined that CAS Arm repeats 1 to 10 are necessary and sufficient for this interaction. Hence, like beta-catenin and plakoglobin, CAS interacts directly with E-cadherin in vivo; however, unlike beta-catenin and plakoglobin, CAS does not interact with APC or alpha-catenin.
Experiments and procedures are described that greatly alleviate the sequential assignment process of uniformly 13C/15N-enriched proteins by determining the type of amino acid from experiments that correlate side chain with backbone amide resonances. A recently proposed 3D NMR experiment, CBCA(CO)NH, correlates C alpha and C beta resonances to the backbone amide 1H and 15N resonances of the next residue (Grzesiek, S. and Bax, A. (1992) J. Am. Chem. Soc., 114, 6291-6293). An extension of this experiment is described which correlates the proton H beta and H alpha resonances to the amide 1H and 15N resonances of the next amino acid, and a detailed product operator description is given. A simple 2D-edited constant-time HSQC experiment is described which rapidly identifies H beta and C beta resonances of aromatic or Asn/Asp residues. The extent to which combined knowledge of the C alpha and C beta chemical shift values determines the amino acid type is investigated, and it is demonstrated that the combined C alpha and C beta chemical shifts of three or four adjacent residues usually are sufficient for defining a unique position in the protein sequence.
Expression of the calcium-dependent adhesion molecule E-cadherin suppresses the invasion of cells in vitro, but the mechanism of this effect is unknown. To investigate this mechanism, we analyzed the effects of expressing E-cadherin in mouse L-cells and rat astrocyte-like WC5 cells. Increased cellular adhesion mediated by E-cadherin reduced invasion in WC5 cells and in some L-cells, but not in others. In all cases, suppression of invasion was correlated with decreased cell movement as assessed in an in vitro wound-filling assay and a transwell motility assay. To define the relationship between adhesion mediated by E-cadherin and suppression of motility, we analyzed the effects of deleting different regions of the E-cadherin cytoplasmic domain. E-cadherin lacking the entire cytoplasmic domain did not mediate calcium-dependent adhesion and did not reduce cell motility when expressed in WC5 cells. E-cadherin lacking a portion of the catenin-binding domain did not associate with the cytoskeleton and did not promote adhesion, yet still suppressed the motility of WC5 cells. In addition, E-cadherin that retains an intact catenin-binding domain, but lacks a juxtamembrane portion of the cytoplasmic domain, mediated effective adhesion, but did not suppress motility. These results indicate E-cadherin mediates adhesion and suppresses cell motility via distinct of E-cadherin plays a key role in suppressing motility.
In srcand ras-transformed cells, tyrosine phosphorylation of adherens junction (AJ) components is related to impairment of cell-cell adhesion. In this paper we report that in human endothelial cells (EC), tyrosine phosphorylation of AJ can be a physiological process regulated by cell density. Immunofluorescence analysis revealed that a phosphotyrosine (P-tyr) antibody could stain cell-cell junctions only in sparse or loosely confluent EC, while the staining was markedly reduced in tightly confluent cultures. This process was reversible, since on artificial wounding of EC monolayers, the cells at the migrating front reacquired P-tyr labelling at cell contacts. In EC, the major cadherin at intercellular AJ is the cell-type-specific VE-cadherin. We therefore analyzed whether this molecule was at least in part responsible for the changes in P-tyr content at cell junctions. Tyrosine phosphorylation of VE-cadherin, β-catenin and p120, occurred in looser AJ, i.e. in recently confluent cells, and was notably reduced in tightly confluent cultures. Changes in P-tyr content paralleled changes in the molecular organization of AJ. VE-cadherin was mostly associated with β-catenin and p120 in loose EC monolayers, while in long-confluent cells, these two catenins were largely replaced by plakoglobin. Inhibition of P-tyr phosphatases (PTPases) by PV markedly augmented the P-tyr content of VE-cadherin, which bound p120 and β-catenin more efficiently, but not plakoglobin.
Transfection experiments in CHO cells showed that p120 could bind to a VE-cadherin cytoplasmic region different from that responsible for β-catenin binding, and PV stabilized this association.
Overall these data indicate that endothelial AJ are dynamic structures that can be affected by the state of confluence of the cells. Tyrosine phosphorylation of VE-cadherin and its association to p120 and β-catenin characterizes early cell contacts, while the formation of mature and cytoskeleton-connected junctions is accompanied by dephosphorylation and plakoglobin association.
Cadherin cell-cell adhesion molecules form membrane-spanning molecular complexes that couple homophilic binding by the cadherin ectodomain to the actin cytoskeleton. A fundamental issue in cadherin biology is how this complex converts the weak intrinsic binding activity of the ectodomain into strong adhesion. Recently we demonstrated that cellular cadherins cluster in a ligand-dependent fashion when cells attached to substrata coated with the adhesive ectodomain of Xenopus C-cadherin (CEC1-5). Moreover, forced clustering of the ectodomain alone significantly strengthened adhesiveness (Yap, A.S., W.M. Brieher, M. Pruschy, and B.M. Gumbiner. Curr. Biol. 7:308-315). In this study we sought to identify the determinants of the cadherin cytoplasmic tail responsible for clustering activity. A deletion mutant of C-cadherin (CT669) that retained the juxtamembrane 94-amino acid region of the cytoplasmic tail, but not the beta-catenin-binding domain, clustered upon attachment to substrata coated with CEC1-5. Like wild-type C-cadherin, this clustering was ligand dependent. In contrast, mutant molecules lacking either the complete cytoplasmic tail or just the juxtamembrane region did not cluster. The juxtamembrane region was itself sufficient to induce clustering when fused to a heterologous membrane-anchored protein, albeit in a ligand-independent fashion. The CT669 cadherin mutant also displayed significant adhesive activity when tested in laminar flow detachment assays and aggregation assays. Purification of proteins binding to the juxtamembrane region revealed that the major associated protein is p120(ctn). These findings identify the juxtamembrane region of the cadherin cytoplasmic tail as a functionally active region supporting cadherin clustering and adhesive strength and raise the possibility that p120(ctn) is involved in clustering and cell adhesion.
The three-dimensional solution structure of recombinant bovine myristoylated recoverin in the Ca(2+)-free state has been refined using an array of isotope-assisted multidimensional heteronuclear NMR techniques. In some experiments, the myristoyl group covalently attached to the protein N-terminus was labeled with C and the protein was unlabeled or vice versa; in others, both were C-labeled. This differential labeling strategy was essential for structural refinement and can be applied to other acylated proteins. Stereospecific assignments of 41 pairs of beta-methylene protons and 48 methyl groups of valine and leucine were included in the structure refinement. The refined structure was constructed using a total of 3679 experimental NMR restraints, comprising 3242 approximate interproton distance restraints (including 153 between the myristoyl group and the polypeptide), 140 distance restraints for 70 backbone hydrogen bonds, and 297 torsion angle restraints. The atomic rms deviations about the average minimized coordinate positions for the secondary structure region of the N-terminal and C-terminal domains are 0.44 +/- 0.07 and 0.55 +/- 0.18 A for backbone atoms, and the 1.09 +/- 0.07 and 1.10 +/- 0.15 A for all heavy atoms, respectively. The refined structure allows for a detailed analysis of the myristoyl binding pocket. The myristoyl group is in a slightly bent conformation: the average distance between C1 and C14 atoms of the myristoyl group is 14.6 A. Hydrophobic residues Leu28, Trp31, and Tyr32 from a cluster that interacts with the front end of the myristoyl (C1-C8), whereas residues Phe49, Phe56, Tyr86, Val87, and Leu90 interact with the tail end (C9-C14). The relatively deep hydrophobic pocket that binds the myristoyl group (C14:0) could also accommodate other naturally occurring acyl groups such as C12:0, C14:1, C14:2 chains.
One of the hallmarks of polarized epithelial cells undergoing mitosis is their rounded morphology. This phenotype correlates
with a reduced cell-substratum adhesion, apparently caused by a modulation of integrin function. However, it is still unclear
whether the cadherin-mediated cell-cell adhesion is affected as well. To address this question, the cadherin complex was analyzed
in different cell cycle stages of Madin-Darby canine kidney cells. By immunofluorescence, mitotic Madin-Darby canine kidney
cells showed an increased staining of E-cadherin and the catenins (α-catenin, β-catenin, plakoglobin, p120ctn) in the cytosol, suggesting a reorganization of the cadherin-catenin complex during mitosis. Biochemical analysis revealed
that the overall amount of these components, as well as the proportion of the complex associated with the actin cytoskeleton,
remained unchanged in mitotic cells. However, we found evidence for an internalization of E-cadherin during mitosis. In addition,
the cadherin-catenin complex was analyzed for mitosis-specific changes in phosphorylation. We report a decrease in the tyrosine
phosphorylation of β-catenin, plakoglobin, and p120ctn during mitosis. Moreover, we observed a mitosis-specific Ser/Thr-phosphorylation of p120ctn, as detected by the MPM-2 antibody. Hence, the cadherin/catenin complex is a target for different posttranslational modifications
during mitosis, which may also have a profound impact on cadherin-mediated cell-cell adhesion.
p120
ctn
is an Armadillo repeat domain protein with structural similarity to the cell adhesion cofactors β-catenin and plakoglobin. All three proteins interact directly with the cytoplasmic domain of the transmembrane cell adhesion molecule E-cadherin; β-catenin and plakoglobin bind a carboxy-terminal region in a mutually exclusive manner, while p120 binds the juxtamembrane region. Unlike β-catenin and plakoglobin, p120 does not interact with α-catenin, the tumor suppressor adenomatous polyposis coli (APC), or the transcription factor Lef-1, suggesting that it has unique binding partners and plays a distinct role in the cadherin-catenin complex. Using p120 as bait, we conducted a yeast two-hybrid screen and identified a novel transcription factor which we named Kaiso. Kaiso’s deduced amino acid sequence revealed an amino-terminal BTB/POZ protein-protein interaction domain and three carboxy-terminal zinc fingers of the C
2
H
2
DNA-binding type. Kaiso thus belongs to a rapidly growing family of POZ-ZF transcription factors that include the
Drosophila
developmental regulators Tramtrak and Bric à brac, and the human oncoproteins BCL-6 and PLZF, which are causally linked to non-Hodgkins’ lymphoma and acute promyelocytic leukemia, respectively. Monoclonal antibodies to Kaiso were generated and used to immunolocalize the protein and confirm the specificity of the p120-Kaiso interaction in mammalian cells. Kaiso specifically coprecipitated with a variety of p120-specific monoclonal antibodies but not with antibodies to α- or β-catenin, E-cadherin, or APC. Like other POZ-ZF proteins, Kaiso localized to the nucleus and was associated with specific nuclear dots. Yeast two-hybrid interaction assays mapped the binding domains to Arm repeats 1 to 7 of p120 and the carboxy-terminal 200 amino acids of Kaiso. In addition, Kaiso homodimerized via its POZ domain but it did not heterodimerize with BCL-6, which heterodimerizes with PLZF. The involvement of POZ-ZF proteins in development and cancer makes Kaiso an interesting candidate for a downstream effector of cadherin and/or p120 signaling.
p120(ctn) is a catenin whose direct binding to the juxtamembrane domain of classical cadherins suggests a role in regulating cell-cell adhesion. The juxtamembrane domain has been implicated in a variety of roles including cadherin clustering, cell motility, and neuronal outgrowth, raising the possibility that p120 mediates these activities. We have generated minimal mutations in this region that uncouple the E-cadherin-p120 interaction, but do not affect interactions with other catenins. By stable transfection into E-cadherin-deficient cell lines, we show that cadherins are both necessary and sufficient for recruitment of p120 to junctions. Detergent-free subcellular fractionation studies indicated that, in contrast to previous reports, the stoichiometry of the interaction is extremely high. Unlike alpha- and beta-catenins, p120 was metabolically stable in cadherin-deficient cells, and was present at high levels in the cytoplasm. Analysis of cells expressing E-cadherin mutant constructs indicated that p120 is required for the E-cadherin-mediated transition from weak to strong adhesion. In aggregation assays, cells expressing p120-uncoupled E-cadherin formed only weak cell aggregates, which immediately dispersed into single cells upon pipetting. As an apparent consequence, the actin cytoskeleton failed to insert properly into peripheral E-cadherin plaques, resulting in the inability to form a continuous circumferential ring around cell colonies. Our data suggest that p120 directly or indirectly regulates the E-cadherin-mediated transition to tight cell-cell adhesion, possibly blocking subsequent events necessary for reorganization of the actin cytoskeleton and compaction.
p120 catenin (p120) is the prototypic member of a growing subfamily of Armadillo-domain proteins found at cell-cell junctions and in nuclei. In contrast to the functions of the classical catenins (alpha-catenin, beta-catenin, and gamma-catenin/plakoglobin), which have been studied extensively, the first clues to p120's biological function have only recently emerged, and its role remains controversial. Nonetheless, it is now clear that p120 affects cell-cell adhesion through its interaction with the highly conserved juxtamembrane domain of classical cadherins, and is likely to have additional roles in the nucleus. Here, we summarize the data on the potential involvement of p120 both in promotion of and in prevension of adhesion, and propose models that attempt to reconcile some of the disparities in the literature. We also discuss the structural relationships and functions of several known p120 family members, as well as the potential roles of p120 in signaling and cancer.
Cadherins are single pass transmembrane proteins that mediate Ca2+-dependent homophilic cell-cell adhesion by linking the cytoskeletons of adjacent cells. In adherens junctions, the cytoplasmic
domain of cadherins bind to β-catenin, which in turn binds to the actin-associated protein α-catenin. The physical properties
of the E-cadherin cytoplasmic domain and its interactions with β-catenin have been investigated. Proteolytic sensitivity,
tryptophan fluorescence, circular dichroism, and 1H NMR measurements indicate that murine E-cadherin cytoplasmic domain is unstructured. Upon binding to β-catenin, the domain
becomes resistant to proteolysis, suggesting that it structures upon binding. Cadherin-β-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 β-catenin-binding site. It is proposed that binding of cadherins to β-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.
The cytoskeletal protein alpha-catenin, which shares structural similarity with vinculin, is required for cadherin-mediated cell adhesion, and functions to modulate cell adhesive strength and to link the cadherins to the actin-based cytoskeleton. Here we describe the crystal structure of a region of alpha-catenin (residues 377-633) termed the M-fragment. The M-fragment is composed of a tandem repeat of two antiparallel four-helix bundles of virtually identical architectures that are related in structure to the dimerization domain of alpha-catenin and the tail region of vinculin. These results suggest that alpha-catenin is composed of repeating antiparallel helical domains. The region of alpha-catenin previously defined as an adhesion modulation domain corresponds to the C-terminal four-helix bundle of the M-fragment, and in the crystal lattice these domains exist as dimers. Evidence for dimerization of the M-fragment of alpha-catenin in solution was detected by chemical cross-linking experiments. The tendency of the adhesion modulation domain to form dimers may explain its biological activity of promoting cell-cell adhesiveness by inducing lateral dimerization of the associated cadherin molecule.
E-cadherin controls a wide array of cellular behaviors including cell-cell adhesion, differentiation and tissue development. Here we show that presenilin-1 (PS1), a protein involved in Alzheimer's disease, controls a gamma-secretase-like cleavage of E-cadherin. This cleavage is stimulated by apoptosis or calcium influx and occurs between human E-cadherin residues Leu731 and Arg732 at the membrane-cytoplasm interface. The PS1/gamma-secretase system cleaves both the full-length E-cadherin and a transmembrane C-terminal fragment, derived from a metalloproteinase cleavage after the E-cadherin ectodomain residue Pro700. The PS1/gamma-secretase cleavage dissociates E-cadherins, beta-catenin and alpha-catenin from the cytoskeleton, thus promoting disassembly of the E-cadherin-catenin adhesion complex. Furthermore, this cleavage releases the cytoplasmic E-cadherin to the cytosol and increases the levels of soluble beta- and alpha-catenins. Thus, the PS1/gamma-secretase system stimulates disassembly of the E-cadherin- catenin complex and increases the cytosolic pool of beta-catenin, a key regulator of the Wnt signaling pathway.
Indirect evidence suggests that p120-catenin (p120) can both positively and negatively affect cadherin adhesiveness. Here we show that the p120 gene is mutated in SW48 cells, and that the cadherin adhesion system is impaired as a direct consequence of p120 insufficiency. Restoring normal levels of p120 caused a striking reversion from poorly differentiated to cobblestone-like epithelial morphology, indicating a crucial role for p120 in reactivation of E-cadherin function. The rescue efficiency was enhanced by increased levels of p120, and reduced by the presence of the phosphorylation domain, a region previously postulated to confer negative regulation. Surprisingly, the rescue was associated with substantially increased levels of E-cadherin. E-cadherin mRNA levels were unaffected by p120 expression, but E-cadherin half-life was more than doubled. Direct p120-E-cadherin interaction was crucial, as p120 deletion analysis revealed a perfect correlation between E-cadherin binding and rescue of epithelial morphology. Interestingly, the epithelial morphology could also be rescued by forced expression of either WT E-cadherin or a p120-uncoupled mutant. Thus, the effects of uncoupling p120 from E-cadherin can be at least partially overcome by artificially maintaining high levels of cadherin expression. These data reveal a cooperative interaction between p120 and E-cadherin and a novel role for p120 that is likely indispensable in normal cells.
Cadherin-catenin complexes, localized to adherens junctions, are essential for cell-cell adhesion. One means of regulating adhesion is through the juxtamembrane domain of the cadherin cytoplasmic tail. This region is the binding site for p120, leading to the hypothesis that p120 is a key regulator of cell adhesion. p120 has also been suggested to regulate the GTPase Rho and to regulate transcription via its binding partner Kaiso. To test these hypothesized functions, we turned to Drosophila, which has only a single p120 family member. It localizes to adherens junctions and binds the juxtamembrane region of DE-cadherin (DE-cad). We generated null alleles of p120 and found that mutants are viable and fertile and have no substantial changes in junction structure or function. However, p120 mutations strongly enhance mutations in the genes encoding DE-cadherin or Armadillo, the beta-catenin homologue. Finally, we examined the localization of p120 during embryogenesis. p120 localizes to adherens junctions, but its localization there is less universal than that of core adherens junction proteins. Together, these data suggest that p120 is an important positive modulator of adhesion but that it is not an essential core component of adherens junctions.
The mechanisms by which catenins regulate cadherin function are not fully understood, and the precise function of p120 catenin (p120ctn) has remained particularly elusive. In microvascular endothelial cells, p120ctn colocalized extensively with cell surface VE-cadherin, but failed to colocalize with VE-cadherin that had entered intracellular degradative compartments. To test the possibility that p120ctn binding to VE-cadherin regulates VE-cadherin internalization, a series of approaches were undertaken to manipulate p120ctn availability to endogenous VE-cadherin. Expression of VE-cadherin mutants that competed for p120ctn binding triggered the degradation of endogenous VE-cadherin. Similarly, reducing levels of p120ctn using siRNA caused a dramatic and dose-related reduction in cellular levels of VE-cadherin. In contrast, overexpression of p120ctn increased VE-cadherin cell surface levels and inhibited entry of cell surface VE-cadherin into degradative compartments. These results demonstrate that cellular levels of p120ctn function as a set point mechanism that regulates cadherin expression levels, and that a major function of p120ctn is to control cadherin internalization and degradation.
p120cas is a tyrosine kinase substrate implicated in ligand-induced receptor signaling through the epidermal growth factor, platelet-derived growth factor, and colony-stimulating factor receptors and in cell transformation by Src. Here we report that p120 associates with a complex containing E-cadherin, alpha-catenin, beta-catenin, and plakoglobin. Furthermore, p120 precisely colocalizes with E-cadherin and catenins in vivo in both normal and Src-transformed MDCK cells. Unlike beta-catenin and plakoglobin, p120 has at least four isoforms which are differentially expressed in a variety of cell types, suggesting novel means of modulating cadherin activities in cells. In Src-transformed MDCK cells, p120, beta-catenin, and plakoglobin were heavily phosphorylated on tyrosine, but the physical associations between these proteins were not disrupted. Association of p120 with the cadherin machinery indicates that both Src and receptor tyrosine kinases cross talk with proteins important for cadherin-mediated cell adhesion. These results also strongly suggest a role for p120 in cell adhesion.
p120 catenin (p120) is the prototypic member of a growing subfamily of Armadillo-domain proteins found at cell-cell junctions and in nuclei. In contrast to the functions of the classical catenins (alpha-catenin, beta-catenin, and gamma-catenin/plakoglobin), which have been studied extensively, the first clues to p120's biological function have only recently emerged, and its role remains controversial. Nonetheless, it is now clear that p120 affects cell-cell adhesion through its interaction with the highly conserved juxtamembrane domain of classical cadherins, and is likely to have additional roles in the nucleus. Here, we summarize the data on the potential involvement of p120 both in promotion of and in prevension of adhesion, and propose models that attempt to reconcile some of the disparities in the literature. We also discuss the structural relationships and functions of several known p120 family members, as well as the potential roles of p120 in signaling and cancer.
Morphogenesis of epithelial tissues involves various forms of reshaping of cell layers, such as invagination or bending, convergent extension, and epithelial–mesenchymal transition. At the cellular level, these processes include changes in the shape, position, and assembly pattern of cells. During such morphogenetic processes, epithelial sheets in general maintain their multicellular architecture, implying that they must engage the mechanisms to change the spatial relationship with their neighbors without disrupting the junctions. A major junctional structure in epithelial tissues is the “adherens junction,” which is composed of cadherin adhesion receptors and associated proteins including F-actin. The adherens junctions are required for the firm associations between cells, as disruption of them causes disorganization of the epithelial architecture. The adherens junctions, however, appear to be a dynamic entity, allowing the rearrangement of cells within cell sheets. This dynamic nature of the adherens junctions seems to be supported by various mechanisms, such as the interactions of cadherins with actin cytoskeleton, endocytosis and recycling of cadherins, and the cooperation of cadherins with other adhesion receptors. In this chapter, we provide an overview of these mechanisms analyzed in vitro and in vivo.
In adherens junctions, α-catenin links the cadherin–β-catenin complex to the actin-based cytoskeleton. α-catenin is a homodimer in solution, but forms a 1:1 heterodimer with β-catenin. The crystal structure of the α-catenin dimerization domain, residues 82–279, shows that α-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 β-catenin. A chimera consisting of the α-catenin-binding region of β-catenin linked to the amino terminus of α-catenin 57–264 behaves as a monomer in solution, as expected, since β-catenin binding disrupts the α-catenin dimer. The crystal structure of this chimera reveals the interaction between α- and β-catenin, and provides a basis for understanding adherens junction assembly.
Evaluation of the electrostatic properties of biomolecules has become a standard practice in molecular biophysics. Foremost
among the models used to elucidate the electrostatic potential is the Poisson-Boltzmann equation; however, existing methods
for solving this equation have limited the scope of accurate electrostatic calculations to relatively small biomolecular systems.
Here we present the application of numerical methods to enable the trivially parallel solution of the Poisson-Boltzmann equation
for supramolecular structures that are orders of magnitude larger in size. As a demonstration of this methodology, electrostatic
potentials have been calculated for large microtubule and ribosome structures. The results point to the likely role of electrostatics
in a variety of activities of these structures.
In epithelial cells, tyrosine kinases induce the tyrosine phosphorylation and ubiquitination of the E-cadherin complex, which induces endocytosis of E-cadherin. With a modified yeast 2-hybrid system, we isolated Hakai, an E-cadherin binding protein, which we have identified as an E3 ubiquitin-ligase. Hakai contains SH2, RING, zinc-finger and proline-rich domains, and interacts with E-cadherin in a tyrosine phosphorylation-dependent manner, inducing ubiquitination of the E-cadherin complex. Expression of Hakai in epithelial cells disrupts cell–cell contacts and enhances endocytosis of E-cadherin and cell motility. Through dynamic recycling of E-cadherin, Hakai can thus modulate cell adhesion, and could participate in the regulation of epithelial–mesenchymal transitions in development or metastasis.
We describe a new method (T-Coffee) for multiple sequence alignment that provides a dramatic improvement in accuracy with a modest sacrifice in speed as compared to the most commonly used alternatives. The method is broadly based on the popular progressive approach to multiple alignment but avoids the most serious pitfalls caused by the greedy nature of this algorithm. With T-Coffee we pre-process a data set of all pair-wise alignments between the sequences. This provides us with a library of alignment information that can be used to guide the progressive alignment. Intermediate alignments are then based not only on the sequences to be aligned next but also on how all of the sequences align with each other. This alignment information can be derived from heterogeneous sources such as a mixture of alignment programs and/or structure superposition. Here, we illustrate the power of the approach by using a combination of local and global pair-wise alignments to generate the library. The resulting alignments are significantly more reliable, as determined by comparison with a set of 141 test cases, than any of the popular alternatives that we tried. The improvement, especially clear with the more difficult test cases, is always visible, regardless of the phylogenetic spread of the sequences in the tests.
Although p120-catenin regulates adherens junction (AJ) stability in cultured cells, genetic studies in lower eukaryotes have not revealed a role for this protein in vivo. Using conditional targeting in mice, we show that p120 null neonatal epidermis exhibits reduced intercellular AJ components but no overt disruption in barrier function or intercellular adhesion. As the mice age, however, they display epidermal hyperplasia and chronic inflammation, typified by hair degeneration and loss of body fat. Using skin engraftments and anti-inflammatory drugs, we show that these features are not attributable to reductions in junctional cadherins and catenins, but rather NFkB activation. Both in vivo and in vitro, p120 null epidermal cells activate nuclear NFkB, triggering a cascade of proinflammatory NFkB targets. Although the underlying mechanism is likely complex, we show that p120 affects NFkB activation and immune homeostasis in part through regulation of Rho GTPases. These findings provide important new insights into p120 function.
Comparative protein modeling is increasingly gaining interest since it is of great assistance during the rational design of mutagenesis experiments. The availability of this method, and the resulting models, has however been restricted by the availability of expensive computer hardware and software. To overcome these limitations, we have developed an environment for comparative protein modeling that consists of SWISS-MODEL, a server for automated comparative protein modeling and of the SWISS-PdbViewer, a sequence to structure workbench. The Swiss-PdbViewer not only acts as a client for SWISS-MODEL, but also provides a large selection of structure analysis and display tools. In addition, we provide the SWISS-MODEL Repository, a database containing more than 3500 automatically generated protein models. By making such tools freely available to the scientific community, we hope to increase the use of protein structures and models in the process of experiment design.
Classical cadherins mediate specific adhesion at intercellular adherens junctions. Interactions between cadherin ectodomains from apposed cells mediate cell-cell contact, whereas the intracellular region functionally links cadherins to the underlying cytoskeleton. Structural, biophysical, and biochemical studies have provided important insights into the mechanism and specificity of cell-cell adhesion by classical cadherins and their interplay with the cytoskeleton. Adhesive binding arises through exchange of beta strands between the first extracellular cadherin domains (EC1) of partner cadherins from adjacent cells. This "strand-swap" binding mode is common to classical and desmosomal cadherins, but sequence alignments suggest that other cadherins will bind differently. The intracellular region of classical cadherins binds to p120 and beta-catenin, and beta-catenin binds to the F-actin binding protein alpha-catenin. Rather than stably bridging beta-catenin to actin, it appears that alpha-catenin actively regulates the actin cytoskeleton at cadherin-based cell-cell contacts.
Peer Reviewed http://deepblue.lib.umich.edu/bitstream/2027.42/30326/1/0000728.pdf
Morphogenesis of epithelial tissues involves various forms of reshaping of cell layers, such as invagination or bending, convergent extension, and epithelial-mesenchymal transition. At the cellular level, these processes include changes in the shape, position, and assembly pattern of cells. During such morphogenetic processes, epithelial sheets in general maintain their multicellular architecture, implying that they must engage the mechanisms to change the spatial relationship with their neighbors without disrupting the junctions. A major junctional structure in epithelial tissues is the "adherens junction," which is composed of cadherin adhesion receptors and associated proteins including F-actin. The adherens junctions are required for the firm associations between cells, as disruption of them causes disorganization of the epithelial architecture. The adherens junctions, however, appear to be a dynamic entity, allowing the rearrangement of cells within cell sheets. This dynamic nature of the adherens junctions seems to be supported by various mechanisms, such as the interactions of cadherins with actin cytoskeleton, endocytosis and recycling of cadherins, and the cooperation of cadherins with other adhesion receptors. In this chapter, we provide an overview of these mechanisms analyzed in vitro and in vivo.
Epithelial cells contain noncentrosomal microtubules (MTs), whose minus ends are oriented apically. In contrast with the well-known interactions of the minus ends with the centrosome, little is known about the termination site of the noncentrosomal minus ends. Here we show that a population of MT minus ends is anchored at the zonula adherens (ZA), the apical-most part of the cadherin-based adherens junction, via a protein that we have termed Nezha. We initially identified PLEKHA7 as a ZA component and subsequently detected Nezha as a partner for PLEKHA7. Nezha bound MTs at their minus ends and tethered them to the ZA. Furthermore, we found that a minus end-directed motor, KIFC3, was concentrated at the ZA in a PLEKHA7/Nezha/MT-dependent manner; and depletion of any of these proteins resulted in disorganization of the ZA. We propose that the PLEKHA7/Nezha/MT complex regulates the ZA integrity by recruiting KIFC3 to this junctional site.
Once engaged by soluble or matrix-anchored ligands, cell surface proteins are commonly sorted to lysosomal degradation through several endocytic pathways. Defective vesicular trafficking of growth factor receptors, as well as unbalanced recycling of integrin- and cadherin-based adhesion complexes, has emerged in the past 5 years as a multifaceted hallmark of malignant cells. In line with the cooperative nature of endocytic machineries, multiple oncogenic alterations underlie defective endocytosis, such as altered ubiquitylation (Cbl and Nedd4 ubiquitin ligases, for example), altered cytoskeletal interactions and alterations to Rab family members. Pharmaceutical interception of the propensity of tumour cells to derail their signalling and their adhesion receptors may constitute a novel target for cancer therapy.
Synapse formation involves reciprocal interactions between cells resulting in formation of a structure optimized for efficient information transfer. Recent work has implicated constituents of the cadherin-catenin cell-adhesion complex in both synapse formation and plasticity. In this review, we describe recent interesting discoveries on mechanisms of cadherin complex function, in addition to regulating adhesion, that are relevant for understanding the role of this complex in synaptogenesis and plasticity. We describe how this complex acts via (i) recruitment/stabilization of intracellular partners; (ii) regulation of intracellular signaling pathways; (iii) regulation of cadherin surface levels, stability and turnover; (iv) stabilization of receptors; and (v) regulation of gene expression. These exciting discoveries provide insights into novel functional roles of the complex beyond regulating cell adhesion.
A novel protein tyrosine kinase (PTK) substrate, p120, has been previously implicated in ligand-induced signaling through the epidermal growth factor, platelet-derived growth factor and colony-stimulating factor 1 receptors, and in cell transformation by p60v-src. We have isolated a near full-length cDNA encoding murine p120. The encoded protein lacks significant homology with any reported protein, but it contains four copies of an imperfect 42 amino acid repeat that occurs 12.5 times in the protein encoded by Drosophila armadillo (arm), and its direct homologs, human plakoglobin (plak) and Xenopus laevis beta-catenin (beta-cat). The presence of this motif implies that p120 may share at least one aspect of its function with the arm protein and its homologs.
The cell-cell adhesion molecule N-cadherin strongly promotes neurite outgrowth in cultured retinal neurons. To test whether cadherins regulate process outgrowth in retinal neurons in vivo, we have blocked cadherin function in single cells by expression of a dominant negative N-cadherin mutant. We report that when cadherin function is inhibited, axon and dendrite outgrowth are severely impaired, particularly in retinal ganglion cells. Laminar migration and cell type specification, by contrast, appear unaffected. Further, expression of the catenin-binding domain of N-cadherin, which blocks cadherin-mediated adhesion in early embryos, does not affect axon outgrowth, suggesting that outgrowth and adhesion are mediated by distinct regions of the cytoplasmic domain. These findings indicate that cadherins play an essential role in the initiation and extension of axons from retinal ganglion cells in vivo.
Classical cadherin-based cellular adhesion is mediated by a multicomponent protein complex that links the adhesive binding activity of the cadherin ectodomain to the actin cytoskeleton. Despite the importance of cadherins in morphogenesis and development, we know very little about how cells determine and alter cadherin adhesive strength. In this study, we sought to identify specific cellular mechanisms that modulate cadherin function by studying adhesion between cells transfected with Xenopus C-cadherin mutant molecules and substrata coated with the purified ectodomain of C-cadherin.
Using the FKBP-FK1012 protein oligomerization system, we found that forced clustering, in cells, of cadherin mutants lacking the cytoplasmic tail significantly increased cellular adhesive strength. Therefore, redistribution of the adhesive binding sites of cells into clusters can influence adhesion independently of other protein interactions mediated by the cadherin cytoplasmic tail. Furthermore, cells transfected with full-length C-cadherin demonstrated dynamic changes in adhesion over time that correlated with clustering but not with changes in the surface expression of C-cadherin or in the composition of the cadherin-catenin complex. The cytoplasmic tail was, however, necessary for clustering of wild-type cadherin.
These studies directly demonstrate a fundamental role for lateral clustering in cadherin function. The distribution of cadherin binding sites presented at the cell surface, a cellular property which is regulated by the cadherin cytoplasmic tail, is an important mechanism which modulates cellular adhesion independently of cytoskeletal activity or signalling.
We wish to thank Terry Schoop of Biomed Arts Associates, San Francisco, for preparation of the figures, Cori Bargmann and Zena Werb for insightful comments on the manuscript, and Normita Santore for editorial assistance. In addition, we are indebted to Joe Harford and Richard Klausner, who allowed us to adapt and expand their depiction of the cell signaling network, and we appreciate suggestions on signaling pathways from Randy Watnick, Brian Elenbas, Bill Lundberg, Dave Morgan, and Henry Bourne. R. A. W. is a Ludwig Foundation and American Cancer Society Professor of Biology. His work has been supported by the Department of the Army and the National Institutes of Health. D. H. acknowledges the support and encouragement of the National Cancer Institute. Editorial policy has rendered the citations illustrative but not comprehensive.
Cadherins play an important role in specific cell-cell adhesion events. Their expression appears to be tightly regulated during development and each tissue or cell type shows a characteristic pattern of cadherin molecules. Inappropriate regulation of their expression levels or functionality has been observed in human malignancies, in many cases leading to aggravated cancer cell invasion and metastasis. The cadherins form a superfamily with at least six subfamilies, which can be distinguished on the basis of protein domain composition, genomic structure, and phylogenetic analysis of the protein sequences. These subfamilies comprise classical or type-I cadherins, atypical or type-II cadherins, desmocollins, desmogleins, protocadherins and Flamingo cadherins. In addition, several cadherins clearly occupy isolated positions in the cadherin superfamily (cadherin-13, -15, -16, -17, Dachsous, RET, FAT, MEGF1 and most invertebrate cadherins). We suggest a different evolutionary origin of the protocadherin and Flamingo cadherin genes versus the genes encoding desmogleins, desmocollins, classical cadherins, and atypical cadherins. The present phylogenetic analysis may accelerate the functional investigation of the whole cadherin superfamily by allowing focused research of prototype cadherins within each subfamily.
As a component of adherens junctions and the Wnt signaling pathway, beta-catenin binds cadherins, Tcf family transcription factors, and the tumor suppressor APC. We have determined the crystal structures of both unphosphorylated and phosphorylated E-cadherin cytoplasmic domain complexed with the arm repeat region of beta-catenin. The interaction spans all 12 arm repeats, and features quasi-independent binding regions that include helices which interact with both ends of the arm repeat domain and an extended stretch of 14 residues which closely resembles a portion of XTcf-3. Phosphorylation of E-cadherin results in interactions with a hydrophobic patch of beta-catenin that mimics the binding of an amphipathic XTcf-3 helix. APC contains sequences homologous to the phosphorylated region of cadherin, and is likely to bind similarly.
Cadherins are transmembrane proteins that mediate adhesion between cells in the solid tissues of animals. Here we present
the 3.1 angstrom resolution crystal structure of the whole, functional extracellular domain from C-cadherin, a representative
“classical” cadherin. The structure suggests a molecular mechanism for adhesion between cells by classical cadherins, and
it provides a new framework for understanding both cis (same cell) and trans (juxtaposed cell) cadherin interactions. The
trans adhesive interface is a twofold symmetric interaction defined by a conserved tryptophan side chain at the membrane-distal
end of a cadherin molecule from one cell, which inserts into a hydrophobic pocket at the membrane-distal end of a cadherin
molecule from the opposing cell.
Tumor progression in epithelial tissues is characterized by a series of genetic and epigenetic changes that lead ultimately to metastasis. Alterations in E-cadherin and its cytoplasmic regulators, the catenins, have been implicated as central to this process. Here, we focus on p120-catenin and its rising incidence in the pathology literature as a molecule altered in human tumors. The data show that p120 is frequently altered and/or lost in tumors of the colon, bladder, stomach, breast, prostate, lung, and pancreas. Moreover, in some cases p120 loss appears to be an early event in tumor progression, possibly preceding loss of E-cadherin. Potential roles of p120 as a tumor suppressor or metastasis promoter are discussed.