scholarly journals Expression of beta-catenin and full-length APC protein in normal and neoplastic colonic tissues

2000 ◽  
Vol 21 (11) ◽  
pp. 1935-1940 ◽  
Author(s):  
M. Iwamoto
Keyword(s):  
Full Length ◽  
Beta Catenin ◽  
Apc Protein

Plakophilin 1 interferes with plakoglobin binding to desmoplakin, yet together with plakoglobin promotes clustering of desmosomal plaque complexes at cell-cell borders

2001 ◽  
Vol 114 (4) ◽  
pp. 727-738 ◽  
Author(s):  
E.A. Bornslaeger ◽  
L.M. Godsel ◽  
C.M. Corcoran ◽  
J.K. Park ◽  
M. Hatzfeld ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Full Length ◽  
Beta Catenin ◽  
Structural Elements ◽  
Lateral Interactions ◽  
Soluble Complex ◽  
Macromolecular Complexes ◽  
Ht1080 Cells ◽  
Associated Proteins ◽  
Filament Networks

Desmosomes are adhesive junctions that link intermediate filament networks to sites of strong intercellular adhesion. These junctions play an important role in providing strength to tissues that experience mechanical stress such as heart and epidermis. The basic structural elements of desmosomes are similar to those of the better-characterized adherens junctions, which anchor actin-containing microfilaments to cadherins at the plasma membrane. This linkage of actin to classic cadherins is thought to occur through an indirect mechanism requiring the associated proteins, alpha- and beta-catenin. In the case of desmosomes, both linear and lateral interactions have been proposed as playing an important role in formation of the plaque and linkage to the cytoskeleton. However, the precise nature of these interactions and how they cooperate in desmosome assembly are poorly understood. Here we employ a reconstitution system to examine the assembly of macromolecular complexes from components found in desmosomes of the differentiated layers of complex tissues. We demonstrate the existence of a Triton-soluble complex of proteins containing full length desmoplakin (DP), the arm protein plakoglobin, and the cytoplasmic domain of the desmosomal cadherin, desmoglein 1 (Dsg1). In addition, full length DP, but not an N-terminal plakoglobin binding domain of DP, co-immunoprecipitated with the Dsg1 tail in the absence of plakoglobin in HT1080 cells. The relative roles of the arm proteins plakoglobin and plakophilin 1 (PKP1) were also investigated. Our results suggest that, in the Triton soluble pool, PKP1 interferes with binding of plakoglobin to full length DP when these proteins are co-expressed. Nevertheless, both plakoglobin and PKP1 are required for the formation of clustered structures containing DP and the Dsg1 tail that ultrastructurally appear similar to desmosomal plaques found in the epidermis. These findings suggest that more than one armadillo family member is required for normal assembly and clustering of the desmosomal plaque in the upper layers of the epidermis.

beta-catenin signalling modulates proliferative potential of human epidermal keratinocytes independently of intercellular adhesion

Development ◽  
1999 ◽  
Vol 126 (10) ◽  
pp. 2285-2298 ◽  
Author(s):  
A.J. Zhu ◽  
F.M. Watt
Keyword(s):  
Stem Cells ◽  
Intercellular Adhesion ◽  
Dominant Negative ◽  
Full Length ◽  
Epidermal Keratinocytes ◽  
Dominant Negative Mutant ◽  
Proliferative Potential ◽  
Beta Catenin ◽  
Epidermal Stem Cells ◽  
Armadillo Repeats

We found that cultured human keratinocytes with high proliferative potential, the putative epidermal stem cells, expressed a higher level of noncadherin-associated beta-catenin than populations enriched for keratinocytes of lower proliferative potential. To investigate the physiological significance of this, a series of beta-catenin constructs was introduced into keratinocytes via retroviral infection. Full-length beta-catenin and a mutant containing only nine armadillo repeats had little effect on proliferative potential in culture, the full-length protein being rapidly degraded. However, expression of stabilised, N-terminally truncated beta-catenin increased the proportion of putative stem cells to almost 90% of the proliferative population in vitro without inducing malignant transformation, and relieved the differentiation stimulatory effect of overexpressing the E-cadherin cytoplasmic domain. Conversely, beta-catenin lacking armadillo repeats acted as a dominant negative mutant and stimulated exit from the stem cell compartment in culture. The positive and negative effects of the beta-catenin mutants on proliferative potential were independent of effects on cell-cycle kinetics, overt terminal differentiation or intercellular adhesion, and correlated with stimulation or inhibition of transactivation of a TCF/LEF reporter in basal keratinocytes. We conclude that the elevated level of cytoplasmic beta-catenin in those keratinocytes with characteristics of epidermal stem cells contributes to their high proliferative potential.

scholarly journals Expression Profiling of Proliferation and Apoptotic Markers along the Adenoma-Carcinoma Sequence in Familial Adenomatous Polyposis Patients

2013 ◽  
Vol 2013 ◽  
pp. 1-7 ◽  
Author(s):  
Jayson Wang ◽  
Nabil El-Masry ◽  
Ian Talbot ◽  
Ian Tomlinson ◽  
Malcolm R. Alison ◽  
...  
Keyword(s):  
Familial Adenomatous Polyposis ◽  
Cyclin D1 ◽  
Caspase 3 ◽  
Normal Mucosa ◽  
Beta Catenin ◽  
Adenomatous Polyposis ◽  
Ki 67 ◽  
Cd10 Expression ◽  
Apc Protein ◽  
Apoptotic Factors

Introduction. Familial adenomatous polyposis (FAP) patients have a germline mutation in the adenomatous polyposis coli (APC) gene. The APC protein interacts with beta-catenin, resulting in the activation of the Wnt signalling pathway. This results in alterations in cell proliferation and apoptosis. We investigated the expression of beta-catenin and related proliferation and apoptotic factors in FAP patients, exploring the expression along the adenoma-carcinoma sequence.Methods. The expression of beta-catenin, p53, bcl-2, cyclin-D1, caspase-3, CD10, and Ki-67 proteins was studied by immunohistochemistry in samples of colonic nonneoplastic mucosa (n=71), adenomas (n=152), and adenocarcinomas (n=19) from each of the16 FAP patients.Results. The expression of beta-catenin, caspase-3, cyclin-D1, and Ki-67 was increased in both adenomas and carcinomas in FAP patients, compared with normal mucosa. p53 and CD10 expression was only slightly increased in adenomas, but more frequently expressed in carcinomas. Bcl-2 expression was increased in adenomas, but decreased in carcinomas.Conclusion. This is the first study investigating collectively the expression of these molecules together in nonneoplastic mucosa, adenomas, and carcinomas from FAP patients. We find that beta-catenin and related proliferative and apoptotic factors (cyclin-D1, bcl-2, caspase-3, and Ki-67) are expressed early in the sequence, in adenomas. However, p53 and CD10 are often expressed later in the sequence, in carcinomas.

Restoration of full-length APC protein in SW480 colon cancer cells induces exosome-mediated secretion of DKK-4

2012 ◽  
Vol 33 (12) ◽  
pp. 1873-1880 ◽  
Author(s):  
Justin W. E. Lim ◽  
Rommel A. Mathias ◽  
Eugene A. Kapp ◽  
Meredith J. Layton ◽  
Maree C. Faux ◽  
...  
Keyword(s):  
Colon Cancer ◽  
Cancer Cells ◽  
Full Length ◽  
Colon Cancer Cells ◽  
Apc Protein

scholarly journals NH2-terminal Deletion of β-Catenin Results in Stable Colocalization of Mutant β-Catenin with Adenomatous Polyposis Coli Protein and Altered MDCK Cell Adhesion

1997 ◽  
Vol 136 (3) ◽  
pp. 693-706 ◽  
Author(s):  
Angela I.M. Barth ◽  
Anne L. Pollack ◽  
Yoram Altschuler ◽  
Keith E. Mostov ◽  
W. James Nelson
Keyword(s):  
Cell Adhesion ◽  
Adenomatous Polyposis Coli ◽  
Mdck Cells ◽  
Full Length ◽  
Terminal Deletion ◽  
Adenomatous Polyposis ◽  
Polyposis Coli ◽  
Mutant Proteins ◽  
Apc Protein ◽  
E Cadherin

β-Catenin is essential for the function of cadherins, a family of Ca2+-dependent cell–cell adhesion molecules, by linking them to α-catenin and the actin cytoskeleton. β-Catenin also binds to adenomatous polyposis coli (APC) protein, a cytosolic protein that is the product of a tumor suppressor gene mutated in colorectal adenomas. We have expressed mutant β-catenins in MDCK epithelial cells to gain insights into the regulation of β-catenin distribution between cadherin and APC protein complexes and the functions of these complexes. Full-length β-catenin, β-catenin mutant proteins with NH2-terminal deletions before (ΔN90) or after (ΔN131, ΔN151) the α-catenin binding site, or a mutant β-catenin with a COOH-terminal deletion (ΔC) were expressed in MDCK cells under the control of the tetracycline-repressible transactivator. All β-catenin mutant proteins form complexes and colocalize with E-cadherin at cell–cell contacts; ΔN90, but neither ΔN131 nor ΔN151, bind α-catenin. However, β-catenin mutant proteins containing NH2-terminal deletions also colocalize prominently with APC protein in clusters at the tips of plasma membrane protrusions; in contrast, full-length and COOH-terminal– deleted β-catenin poorly colocalize with APC protein. NH2-terminal deletions result in increased stability of β-catenin bound to APC protein and E-cadherin, compared with full-length β-catenin. At low density, MDCK cells expressing NH2-terminal–deleted β-catenin mutants are dispersed, more fibroblastic in morphology, and less efficient in forming colonies than parental MDCK cells. These results show that the NH2 terminus, but not the COOH terminus of β-catenin, regulates the dynamics of β-catenin binding to APC protein and E-cadherin. Changes in β-catenin binding to cadherin or APC protein, and the ensuing effects on cell morphology and adhesion, are independent of β-catenin binding to α-catenin. These results demonstrate that regulation of β-catenin binding to E-cadherin and APC protein is important in controlling epithelial cell adhesion.

scholarly journals Deletion of an amino-terminal sequence beta-catenin in vivo and promotes hyperphosporylation of the adenomatous polyposis coli tumor suppressor protein.

1996 ◽  
Vol 16 (8) ◽  
pp. 4088-4094 ◽  
Author(s):  
S Munemitsu ◽  
I Albert ◽  
B Rubinfeld ◽  
P Polakis
Keyword(s):  
Adenomatous Polyposis Coli ◽  
Sodium Dodecyl ◽  
Cellular Transformation ◽  
Beta Catenin ◽  
Adenomatous Polyposis ◽  
Wild Type ◽  
Polyposis Coli ◽  
High Molecular Weight Complex ◽  
Amino Terminal ◽  
Apc Protein

Regulation of cell adhesion and cell signaling by beta-catenin occurs through a mechanism likely involving the targeted degradation of the protein. Deletional analysis was used to generate a beta-catenin refractory to rapid turnover and to examine its effects on complexes containing either cadherin or the adenomatous polyposis coli (APC) protein. The results show that amino-terminal deletion of beta-catenin results in a protein with increased stability that acts in a dominant fashion with respect to wild-type beta-catenin. Constitutive expression in AtT20 cells of a beta-catenin lacking 89 N-terminal amino acids (deltaN89beta-catenin) resulted in severely reduced levels of the more labile wild-type beta-catenin. The mutant beta-catenin was expressed at endogenous levels but displaced the vast majority of wild-type beta-catenin associated with N-cadherin. The deltaN89beta-catenin accumulated on the APC protein to a level 10-fold over that of wild-type beta-catenin and recruited a kinase into the APC complex. The kinase was highly active toward APC in vitro and promoted a sodium dodecyl sulfate gel band shift that was also evident for endogenous APC from cells expressing the mutant beta-catenin. Unlike wild-type beta-catenin, which partitions solely as part of a high-molecular-weight complex, the deltaN89 mutant protein also fractionated as a stable monomer, indicating that it had escaped the requirement to associate with other proteins. That similar N-terminal mutants of beta-catenin have been implicated in cellular transformation suggests that their abnormal association with APC may, in part, be responsible for this phenotype.

In Situ Localization of PPAR Gamma and Uncoupling Protein in Mouse Embryo Sections Using Digoxigenin-Labeled Riboprobes

1996 ◽  
Vol 54 ◽  
pp. 32-33
Author(s):  
C. Jennermann ◽  
S. A. Kliewer ◽  
D. C. Morris
Keyword(s):  
Alkaline Phosphatase ◽  
Room Temperature ◽  
Uncoupling Protein ◽  
Ppar Gamma ◽  
Nuclear Hormone Receptor ◽  
Full Length ◽  
Northern Analysis ◽  
Peroxisome Proliferator

Peroxisome proliferator-activated receptor gamma (PPARg) is a member of the nuclear hormone receptor superfamily and has been shown in vitro to regulate genes involved in lipid metabolism and adipocyte differentiation. By Northern analysis, we and other researchers have shown that expression of this receptor predominates in adipose tissue in adult mice, and appears first in whole-embryo mRNA at 13.5 days postconception. In situ hybridization was used to find out in which developing tissues PPARg is specifically expressed.Digoxigenin-labeled riboprobes were generated using the Genius™ 4 RNA Labeling Kit from Boehringer Mannheim. Full length PPAR gamma, obtained by PCR from mouse liver cDNA, was inserted into pBluescript SK and used as template for the transcription reaction. Probes of average size 200 base pairs were made by partial alkaline hydrolysis of the full length transcripts. The in situ hybridization assays were performed as described previously with some modifications. Frozen sections (10 μm thick) of day 18 mouse embryos were cut, fixed with 4% paraformaldehyde and acetylated with 0.25% acetic anhydride in 1.0M triethanolamine buffer. The sections were incubated for 2 hours at room temperature in pre-hybridization buffer, and were then hybridized with a probe concentration of 200μg per ml at 70° C, overnight in a humidified chamber. Following stringent washes in SSC buffers, the immunological detection steps were performed at room temperature. The alkaline phosphatase labeled, anti-digoxigenin antibody and detection buffers were purchased from Boehringer Mannheim. The sections were treated with a blocking buffer for one hour and incubated with antibody solution at a 1:5000 dilution for 2 hours, both at room temperature. Colored precipitate was formed by exposure to the alkaline phosphatase substrate nitrobluetetrazoliumchloride/ bromo-chloroindlylphosphate.

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