G2Cdb::Gene report

Gene id
Gene symbol
Homo sapiens
catenin (cadherin-associated protein), alpha 1, 102kDa
G00000556 (Mus musculus)

Databases (8)

ENSG00000044115 (Ensembl human gene)
1495 (Entrez Gene)
973 (G2Cdb plasticity & disease)
CTNNA1 (GeneCards)
116805 (OMIM)
Marker Symbol
HGNC:2509 (HGNC)
Protein Expression
107 (human protein atlas)
Protein Sequence
P35221 (UniProt)

Synonyms (1)

  • CAP102

Literature (86)

Pubmed - other

  • EGF-induced ERK activation promotes CK2-mediated disassociation of alpha-Catenin from beta-Catenin and transactivation of beta-Catenin.

    Ji H, Wang J, Nika H, Hawke D, Keezer S, Ge Q, Fang B, Fang X, Fang D, Litchfield DW, Aldape K and Lu Z

    Brain Tumor Center and Department of Neuro-Oncology, The University of Texas M.D. Anderson Cancer Center, Houston, 77030, USA.

    Increased transcriptional activity of beta-catenin resulting from Wnt/Wingless-dependent or -independent signaling has been detected in many types of human cancer, but the underlying mechanism of Wnt-independent regulation remains unclear. We demonstrate here that EGFR activation results in disruption of the complex of beta-catenin and alpha-catenin, thereby abrogating the inhibitory effect of alpha-catenin on beta-catenin transactivation via CK2alpha-dependent phosphorylation of alpha-catenin at S641. ERK2, which is activated by EGFR signaling, directly binds to CK2alpha via the ERK2 docking groove and phosphorylates CK2alpha primarily at T360/S362, subsequently enhancing CK2alpha activity toward alpha-catenin phosphorylation. In addition, levels of alpha-catenin S641 phosphorylation correlate with levels of ERK1/2 activity in human glioblastoma specimens and with grades of glioma malignancy. This EGFR-ERK-CK2-mediated phosphorylation of alpha-catenin promotes beta-catenin transactivation and tumor cell invasion. These findings highlight the importance of the crosstalk between EGFR and Wnt pathways in tumor development.

    Funded by: NCI NIH HHS: 5R01CA109035, R01 CA109035, R01 CA109035-05

    Molecular cell 2009;36;4;547-59

  • Alternatively activated macrophages engage in homotypic and heterotypic interactions through IL-4 and polyamine-induced E-cadherin/catenin complexes.

    Van den Bossche J, Bogaert P, van Hengel J, Guérin CJ, Berx G, Movahedi K, Van den Bergh R, Pereira-Fernandes A, Geuns JM, Pircher H, Dorny P, Grooten J, De Baetselier P and Van Ginderachter JA

    Department of Molecular and Cellular Interactions, VIB-Vrije Universiteit Brussel, Building E,Level 8, Pleinlaan 2, B-1050, Brussels, Belgium.

    Alternatively activated macrophages (AAMs), triggered by interleukin-4 (IL-4) and IL-13, play a modulating role during Th2 cytokine-driven pathologies, but their molecular armament remains poorly characterized. Here, we established E-cadherin (Cdh1) as a selective marker for IL-4/IL-13-exposed mouse and human macrophages, which is STAT6-dependently induced during polarized Th2 responses associated with Taenia crassiceps helminth infections or allergic airway inflammation. The IL-4-dependent, arginase-1/ornithine decarboxylase-mediated production of polyamines is important for maximal Cdh1 induction, unveiling a novel mechanism for IL-4-dependent gene transcription. At the macrophage surface, E-cadherin forms a functional complex with the catenins that accumulates at sites of cell contact. Macrophage-specific deletion of the Cdh1 gene illustrates the implication of E-cadherin in IL-4-driven macrophage fusion and heterotypic interactions with CD103(+) and KLRG1(+) T cells. This study identifies the E-cadherin/catenin complex as a discriminative, partly polyamine-regulated feature of IL-4/IL-13-exposed alternatively activated macrophages that contributes to homotypic and heterotypic cellular interactions.

    Blood 2009;114;21;4664-74

  • The presence of alpha-catenin in the VE-cadherin complex is required for efficient transendothelial migration of leukocytes.

    van Buul JD, van Alphen FP and Hordijk PL

    Dept. Molecular Cell Biology, Sanquin Research, and Landsteiner Laboratory, Academic Medical Center, University of Amsterdam, The Netherlands. j.vanbuul@sanquin.nl

    The majority of the leukocytes cross the endothelial lining of the vessels through cell-cell junctions. The junctional protein Vascular Endothelial (VE)-cadherin is transiently re-distributed from sites of cell-cell contacts during passage of leukocytes. VE-cadherin is part of a protein complex comprising p120-catenin and beta-catenin as intracellular partners. Beta-catenin connects VE-cadherin to alpha-catenin. This VE-cadherin-catenin complex is believed to dynamically control endothelial cell-cell junctions and to regulate the passage of leukocytes, although not much is known about the role of alpha- and beta-catenin during the process of transendothelial migration (TEM). In order to study the importance of the interaction between alpha- and beta-catenin in TEM, we used a cell-permeable version of the peptide encoding the binding site of alpha-catenin for beta-catenin (S27D). The data show that S27D interferes with the interaction between alpha- and beta-catenin and induces a reversible decrease in electrical resistance of the endothelial monolayer. In addition, S27D co-localized with beta-catenin at cell-cell junctions. Surprisingly, transmigration of neutrophils across endothelial monolayers was blocked in the presence of S27D. In conclusion, our results show for the first time that the association of alpha-catenin with the cadherin-catenin complex is required for efficient leukocyte TEM.

    International journal of biological sciences 2009;5;7;695-705

  • Loss of Coxsackie and adenovirus receptor downregulates alpha-catenin expression.

    Stecker K, Koschel A, Wiedenmann B and Anders M

    Division of Gastroenterology and Hepatology, Department of Internal Medicine, Charité Medical School, Campus Virchow, Augustenburgerplatz 1, Berlin 13353, Germany.

    Background: The Coxsackie and adenovirus receptor (CAR) has been shown to inhibit cancer cell proliferation, migration, and invasion. The underlying mechanisms, however, are poorly understood.

    Methods: The differential gene expression in the human colon cancer cell line DLD1 on RNAi-mediated functional CAR knockdown was analysed using oligo-array technology. Expression of alpha-catenin was determined by quantitative RT-PCR and western blotting. Proliferation, migration, and invasion after CAR knockdown were assessed by in vitro assays, and cell morphology in a three-dimensional context was evaluated using matrigel.

    Results: Oligo-array technology identified alpha-catenin as the strongest downregulated gene after CAR knockdown. Western blotting and quantitative RT-PCR confirmed a reduced alpha-catenin expression after CAR knockdown in DLD1 cells and in the rat intestinal cell line IEC-6. Functionally, both cell lines showed a marked increase in proliferation, migration, and invasion on CAR knockdown. In matrigel, both cell lines formed amorphous cell clusters in contrast to well-organised three-dimensional structures of CAR-expressing vector controls. Ectopic 're'-expression of alpha-catenin in DLD1 and IEC-6 CAR knockdown cells reversed these functional and morphological effects.

    Conclusion: These data suggest that an interaction of CAR and alpha-catenin mediates the impact of CAR on cell proliferation, migration, invasion, and morphology.

    British journal of cancer 2009;101;9;1574-9

  • Progressive chromatin repression and promoter methylation of CTNNA1 associated with advanced myeloid malignancies.

    Ye Y, McDevitt MA, Guo M, Zhang W, Galm O, Gore SD, Karp JE, Maciejewski JP, Kowalski J, Tsai HL, Gondek LP, Tsai HC, Wang X, Hooker C, Smith BD, Carraway HE and Herman JG

    Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA.

    Complete loss or deletion of the long arm of chromosome 5 is frequent in myelodysplastic syndrome (MDS) and acute myelogenous leukemia (AML). The putative gene(s) deleted and responsible for the pathogenesis of these poor prognosis hematologic disorders remain controversial. This study is a comprehensive analysis of previously implicated and novel genes for epigenetic inactivation in AML and MDS. In 146 AML cases, methylation of CTNNA1 was frequent, and more common in AML patients with 5q deletion (31%) than those without 5q deletion (14%), whereas no methylation of other 5q genes was observed. In 31 MDS cases, CTNNA1 methylation was only found in high-risk MDS (>or=RAEB2), but not in low-risk MDS (<RAEB2), indicating that CTNNA1 methylation might be important in the transformation of MDS to AML. CTNNA1 expression was lowest in AML/MDS patients with CTNNA1 methylation, although reduced expression was found in some patients without promoter methylation. Repressive chromatin marks (H3K27me3) at the promoter were identified in CTNNA1-repressed AML cell lines and primary leukemias, with the most repressive state correlating with DNA methylation. These results suggest progressive, acquired epigenetic inactivation at CTNNA1, including histone modifications and promoter CpG methylation, as a component of leukemia progression in patients with both 5q- and non-5q- myeloid malignancies.

    Funded by: NCI NIH HHS: P50 CA058184-090013

    Cancer research 2009;69;21;8482-90

  • Modulation of the tumor suppressor protein alpha-catenin by ischemic microenvironment.

    Plumb CL, Adamcic U, Shahrzad S, Minhas K, Adham SA and Coomber BL

    Department of Biomedical Sciences, Ontario Veterinary College, University of Guelph, Guelph, ON N1G 2W1, Canada.

    Dysregulation or mislocalization of cell adhesion molecules and their regulators, such as E-cadherin, beta-catenin, and alpha-catenin, usually correlates with loss of polarity, dedifferentiation, invasive tumor growth, and metastasis. A subpopulation of alpha-catenin-negative cells within the DLD-1 colorectal carcinoma cell line causes it to display a heterogeneous morphological makeup, thus providing an excellent model system in which to investigate the role of alpha-catenin in tumorigenesis. We re-established expression of alpha-catenin protein in an alpha-catenin-deficient subpopulation of the DLD-1 cell line and used it to demonstrate that loss of alpha-catenin resulted in increased in vitro tumorigenic characteristics (increased soft agarose colony formation, clonogenic survival after suspension, and survival in suspension). When the cells were used to form tumor xenografts, those lacking alpha-catenin showed faster growth rates because of increased cellular cycling but not increased tumor microvascular recruitment. alpha-Catenin-expressing cells were preferentially located in well perfused areas of xenografts when tumors were formed from mixed alpha-catenin-positive and -negative cells. We therefore evaluated the role of the ischemic tumor microenvironment on alpha-catenin expression and demonstrated that cells lose expression of alpha-catenin after prolonged exposure in vitro to hypoglycemic conditions. Our findings illustrate that the tumor microenvironment is a potent modulator of tumor suppressor expression, which has implications for localized nutrient deficiency and ischemia-induced cancer progression.

    The American journal of pathology 2009;175;4;1662-74

  • Tumor suppressor function of BCSC-1 in nasopharyngeal carcinoma.

    Zhou YQ, Chen SL, Ju JY, Shen L, Liu Y, Zhen S, Lv N, He ZG and Zhu LP

    Department of Immunology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences and School of Basic Medicine, Peking Union Medical College, Beijing, China.

    BCSC-1 is dramatically upregulated in CNE-2L2 human nasopharyngeal carcinoma cells with reduced malignancy (AS cells) and is proposed to be a candidate tumor suppressor gene. We therefore examined the effect of BCSC-1 expression on malignant behaviors of CNE-2L2 cells. Growth in vitro and tumorigenesis in nude mice of wild-type CNE-2L2 cells (W cells) were inhibited by ectopic BCSC-1, and those of AS cells were promoted by BCSC-1 suppression. The tumor suppressor function of BCSC-1 was further confirmed by a study showing that intratumor BCSC-1 injection caused growth suppression of the tumor from W cells inoculated in nude mice. Immunohistochemistry exhibited marked reduction of BCSC-1 expression in 11 of 39 human nasopharyngeal carcinoma specimens. Because BCSC-1 expression was as rich as that in normal cells in the rest of the carcinoma specimens and was poor in CNE-2L2 cells, HNE-1 human nasopharyngeal carcinoma cells with rich BCSC-1 expression were used as a control in the study. No effect of BCSC-1 transfection on growth of the cells was observed. The data suggest that BCSC-1 suppression might play roles in tumorigenesis of some nasopharyngeal carcinomas and that BCSC-1 might be a potential gene therapy target in nasopharyngeal carcinomas with poor BCSC-1 expression. Enhanced aggregation of cells together with increased E-cadherin and alpha-catenin expression and reduced Wnt signaling might be involved in the mechanisms of tumor suppressor function of BCSC-1.

    Cancer science 2009;100;10;1817-22

  • Loss of alpha-catenin decreases the strength of single E-cadherin bonds between human cancer cells.

    Bajpai S, Feng Y, Krishnamurthy R, Longmore GD and Wirtz D

    Department of Chemical and Biomolecular Engineering, The Johns Hopkins University, Baltimore, Maryland 21218, USA.

    The progression of several human cancers correlates with the loss of cytoplasmic protein alpha-catenin from E-cadherin-rich intercellular junctions and loss of adhesion. However, the potential role of alpha-catenin in directly modulating the adhesive function of individual E-cadherin molecules in human cancer is unknown. Here we use single-molecule force spectroscopy to probe the tensile strength, unstressed bond lifetime, and interaction energy between E-cadherins expressed on the surface of live human parental breast cancer cells lacking alpha-catenin and these cells where alpha-catenin is re-expressed. We find that the tensile strength and the lifetime of single E-cadherin/E-cadherin bonds between parental cells are significantly lower over a wide range of loading rates. Statistical analysis of the force displacement spectra reveals that single cadherin bonds between cancer cells feature an exceedingly low energy barrier against tensile forces and low molecular stiffness. Disassembly of filamentous actin using latrunculin B has no significant effect on the strength of single intercellular E-cadherin bonds. The absence of alpha-catenin causes a dominant negative effect on both global cell-cell adhesion and single E-cadherin bond strength. These results suggest that the loss of alpha-catenin alone drastically reduces the adhesive force between individual cadherin pairs on adjoining cells, explain the global loss of cell adhesion in human breast cancer cells, and show that the forced expression of alpha-catenin in cancer cells can restore both higher intercellular avidity and intercellular E-cadherin bond strength.

    Funded by: NCI NIH HHS: CA137891, R21 CA137891; NIGMS NIH HHS: GM080673, R01 GM080673

    The Journal of biological chemistry 2009;284;27;18252-9

  • Analysis of APC, alpha-, beta-catenins, and N-cadherin protein expression in aggressive fibromatosis (desmoid tumor).

    Ferenc T, Wroński JW, Kopczyński J, Kulig A, Sidor M, Stalińska L, Dziki A and Sygut J

    Department of Biology and Genetics, Medical University, Pl. Hallera 1, 90-647 Lodz, Poland. biolgen@achilles.wam.lodz.pl

    The aims of this study were to analyze the cadherin/catenin adhesion complex in cells from abdominal and extra-abdominal aggressive fibromatosis tumors, and to estimate the correlation between the expression of the tested proteins and the clinical data of the desmoid patients. Immunohistochemistry was used to examine the expression of the cadherin/catenin adhesion complex: APC protein, alpha-, beta-catenin, and N-cadherin in archival material derived from 15 cases of extra-abdominal desmoid tumor (E-AD) and 20 cases of abdominal (AD) desmoid tumor. The tested proteins demonstrated cytoplasmic (c) staining. Furthermore, nuclear (n) or cytoplasmic and nuclear (c+n) staining was observed for beta-catenin. The mean values of the percentage of positive cells for the tested proteins between E-AD vs. AD did not demonstrate any statistically significant difference except for alpha-catenin. In the E-AD group, in both cases of recurrent tumors, no alpha-catenin expression was observed but the expression of this protein was detected in primary tumors. In the groups investigated, no statistically significant correlation was found between alpha-catenin, beta-catenin (c), (n) and (c+n) expression, and tumor size (p>0.1). The results regarding beta-catenin expression obtained in our study confirm the previous findings that nuclear accumulation of this protein plays a crucial role in the pathogenesis of aggressive fibromatosis.

    Pathology, research and practice 2009;205;5;311-24

  • alpha-Catenin overrides Src-dependent activation of beta-catenin oncogenic signaling.

    Inge LJ, Rajasekaran SA, Wolle D, Barwe SP, Ryazantsev S, Ewing CM, Isaacs WB and Rajasekaran AK

    Nemours Center for Childhood Cancer Research, Alfred I. DuPont Hospital for Children, 1701 Rockland Road, Wilmington, DE 19803, USA.

    Loss of alpha-catenin is one of the characteristics of prostate cancer. The catenins (alpha and beta) associated with E-cadherin play a critical role in the regulation of cell-cell adhesion. Tyrosine phosphorylation of beta-catenin dissociates it from E-cadherin and facilitates its entry into the nucleus, where beta-catenin acts as a transcriptional activator inducing genes involved in cell proliferation. Thus, beta-catenin regulates cell-cell adhesion and cell proliferation. Mechanisms controlling the balance between these functions of beta-catenin invariably are altered in cancer. Although a wealth of information is available about beta-catenin deregulation during oncogenesis, much less is known about how or whether alpha-catenin regulates beta-catenin functions. In this study, we show that alpha-catenin acts as a switch regulating the cell-cell adhesion and proliferation functions of beta-catenin. In alpha-catenin-null prostate cancer cells, reexpression of alpha-catenin increased cell-cell adhesion and decreased beta-catenin transcriptional activity, cyclin D1 levels, and cell proliferation. Further, Src-mediated tyrosine phosphorylation of beta-catenin is a major mechanism for decreased beta-catenin interaction with E-cadherin in alpha-catenin-null cells. alpha-Catenin attenuated the effect of Src phosphorylation by increasing beta-catenin association with E-cadherin. We also show that alpha-catenin increases the sensitivity of prostate cancer cells to a Src inhibitor in suppressing cell proliferation. This study reveals for the first time that alpha-catenin is a key regulator of beta-catenin transcriptional activity and that the status of alpha-catenin expression in tumor tissues might have prognostic value for Src targeted therapy.

    Funded by: NIDDK NIH HHS: DK 56216, R01 DK056216, R01 DK056216-06, R01 DK056216-07, R01 DK056216-08; NIGMS NIH HHS: F31 GM068985, F31-GM068985

    Molecular cancer therapeutics 2008;7;6;1386-97

  • N-glycosylation affects the adhesive function of E-Cadherin through modifying the composition of adherens junctions (AJs) in human breast carcinoma cell line MDA-MB-435.

    Zhao H, Liang Y, Xu Z, Wang L, Zhou F, Li Z, Jin J, Yang Y, Fang Z, Hu Y, Zhang L, Su J and Zha X

    Department of Biochemistry and Molecular Biology, Shanghai Medical College, Fudan University, 138 Yi Xue Yuan Road, Shanghai 200032, PR China.

    E-cadherin mediates calcium-dependent cell-cell adhesion between epithelial cells. The ectodomain of human E-cadherin contains four potential N-glycosylation sites at Asn residues 554, 566, 618, and 633. In this study, the role of N-glycosylation in E-cadherin-mediated cell-cell adhesion was investigated by site-directed mutagenesis. In MDA-MB-435 cells, all four potential N-glycosylation sites of human E-cadherin were N-glycosylated. Removal of N-glycan at Asn-633 dramatically affected E-cadherin stability. In contrast, mutant E-cadherin lacking the other three N-glycans showed similar protein stability in comparison with wild-type E-cadherin. Moreover, N-glycans at Asn-554 and Asn-566 were found to affect E-cadherin-mediated calcium-dependent cell-cell adhesion, and removal of either of the two N-glycans caused a significant decrease in calcium-dependent cell-cell adhesion accompanied with elevated cell migration. Analysis of the composition of adherens junctions (AJs) revealed that removal of N-glycans on E-cadherin resulted in elevated tyrosine phosphorylation level of beta-catenin and reduced beta- and alpha-catenins at AJs. These findings demonstrate that N-glycosylation may affect the adhesive function of E-cadherin through modifying the composition of AJs.

    Journal of cellular biochemistry 2008;104;1;162-75

  • Altered expression of epithelial junctional proteins in atopic asthma: possible role in inflammation.

    de Boer WI, Sharma HS, Baelemans SM, Hoogsteden HC, Lambrecht BN and Braunstahl GJ

    Department of Pulmonary Medicine, Erasmus MC, University Medical Centre, Rotterdam, The Netherlands. deboer.pim@hetnet.nl

    Epithelial cells form a tight barrier against environmental stimuli via tight junctions (TJs) and adherence junctions (AJs). Defects in TJ and AJ proteins may cause changes in epithelial morphology and integrity and potentially lead to faster trafficking of inflammatory cells through the epithelium. Bronchial epithelial fragility has been reported in asthmatic patients, but little is known about the expression of TJ and AJ proteins in asthma. We studied epithelial expression of zonula occludens-1 (ZO-1) and AJ proteins E-cadherin, alpha-catenin, and beta-catenin in bronchial biopsies from nonatopic nonasthmatic (healthy) subjects (n = 14), and stable atopic asthmatic subjects (n = 22) at baseline conditions. Immunostaining for these proteins was semi-quantified for separate cellular compartments. E-cadherin, alpha-catenin and beta-catenin were present in the cellular membrane and less in the cytoplasm. Only beta-catenin was present in the nucleus in agreement with its potential function as transcription factor. ZO-1 was present in the apicolateral membrane of superficial cells. alpha-Catenin expression was significantly lower in subjects with asthma than without and correlated inversely with numbers of eosinophils within the epithelium. ZO-1 and E-cadherin expression were significantly lower in asthmatic than in nonasthmatic subjects. Expression of beta-catenin was not different. Our results suggest that the lower epithelial alpha-catenin, E-cadherin and (or) ZO-1 expression in patients with atopic asthma contributes to a defective airway epithelial barrier and a higher influx of eosinophils in the epithelium.

    Canadian journal of physiology and pharmacology 2008;86;3;105-12

  • Alpha(E)-Catenin induces SRF-dependent transcriptional activity through its C-terminal region and is partly RhoA/ROCK-dependent.

    Merdek KD, Jaffe AB, Dutt P, Olson MF, Hall A, Fanburg BL, Kayyali US and Toksoz D

    Physiology Department, Tufts University School of Medicine, Boston, MA 02111, USA.

    The ubiquitous alpha(E)-catenin is an essential actin cytoskeletal linker. The transcription factor, serum response factor (SRF), induces transcription via binding to the serum response element (SRE) in gene promoters, and in many cases responds to actin dynamics. Here, we report that alpha(E)-catenin expression in HEK293 cells activates the SRE.L transcriptional reporter, a reporter containing the isolated SRF-binding site, and a stably integrated SRE.L reporter in fibroblasts. alpha-Catenin-induced reporter activity appears only partly dependent on RhoA GTPase and Rho kinase function. alpha-Catenin expression has no effect on RhoA activation or localization, and alpha-catenin-induced SRE.L reporter activation is insensitive to the actin-modulating agent latrunculin B. Ectopic alpha-catenin expression was not sufficient to induce actin filament assembly as measured by stress fiber formation. SRE.L reporter is activated by the C-terminal approximately 300 residue region of alpha(E)-catenin. These results suggest induction of SRF-mediated transcription by alpha(E)-catenin either downstream of RhoA or via a parallel pathway.

    Funded by: NHLBI NIH HHS: HL32723, HL79320, R01 HL032723, R01 HL079320, R01 HL079320-04, R37 HL032723; NIDDK NIH HHS: T32 DK007542, T32DK07542

    Biochemical and biophysical research communications 2008;366;3;717-23

  • The prognostic value of E-cadherin, alpha-, beta- and gamma-catenin in bladder cancer patients who underwent radical cystectomy.

    Kashibuchi K, Tomita K, Schalken JA, Kume H, Takeuchi T and Kitamura T

    Department of Urology, Graduate School of Medicine and Faculty of Medicine, University of Tokyo, Tokyo, Japan.

    Objectives: To determine the value of the loss of expression of E-cadherin and cadherin associated molecules as useful markers for both prognosis and chemosensitivity in bladder cancer patients who have undergone radical cystectomy.

    In 55 paraffin embedded specimens of radical cystectomy at our hospital from 1982 to 2000, the expression of E-cadherin, alpha-, beta- and gamma-catenin was examined by immunohistochemical staining. To evaluate the prognostic significance of these molecules, Kaplan-Meier survival curves were constructed and a statistical analysis was calculated by a log-rank test. A multivariate test (tumor stage, tumor grade, lymph node metastasis, configuration, the expression of E-cadherin, alpha-, beta- and gamma-catenin) was performed to detect prognostic markers.

    Results: Normal expression was found in 33 cases (60.0%) for E-cadherin, 29 (52.7%) for alpha-catenin, 31 cases (56.4%) for beta-catenin, and 31 cases (56.4%) for gamma-catenin. The expression patterns for E-cadherin, alpha-, beta- and gamma-catenin were significantly correlated with each other (P < 0.01). Survival analysis showed a significant difference between normal and aberrant expression in each staining. A multivariate analysis revealed that the expression of alpha- catenin was an independent prognostic factor (P = 0.0191). In 23 patients that received adjuvant chemotherapy, there was a significant difference in survival between the normal and aberrant expression of alpha-catenin, but not other molecules.

    Conclusion: Alpha-catenin may not only be a good prognostic marker, but also one of key molecules that determine the chemosensitivities in patients with invasive bladder cancer.

    International journal of urology : official journal of the Japanese Urological Association 2007;14;9;789-94

  • Evidence that the V832M E-cadherin germ-line missense mutation does not influence the affinity of alpha -catenin for the cadherin/catenin complex.

    Curtis MW, Ly QP, Wheelock MJ and Johnson KR

    University of Nebraska Medical Center, Department of Pathology and Microbiology, Omaha, Nebraska, USA.

    Mutations in E-cadherin are associated with a number of diseases, and have been shown to contribute to disease progression. In particular, 50% of hereditary diffuse gastric cancer cases have inactivating mutations in the E-cadherin gene. An interesting mutation near the beta-catenin-binding site on the cytoplasmic domain of E-cadherin (V832M) was recently reported that produces full-length protein, but exhibits decreased binding of alpha -catenin to the cadherin/catenin complex. The study was done by transfecting mutant E-cadherin into Chinese hamster ovary fibroblast cells. Here we show that the previously reported characteristics of this mutation do not apply to human epithelial cells expressing this mutant protein and suggest that the mechanism whereby the V832M mutation in human E-cadherin promotes gastric cancer is not yet understood.

    Funded by: NIDCR NIH HHS: R01-DE12308; NIGMS NIH HHS: R01-GM51188

    Cell communication & adhesion 2007;14;2-3;45-55

  • Expression of the E-cadherin-catenins complex in sentinel node is related to tumor morphology but not to spread to nonsentinel nodes.

    Canavese G, Bernardi A, Candelaresi G, Lovadina P, Amerio S, Rossetti V, Rabagliati C and Berardengo E

    Department of Pathology, Ospedale S. Giovanni Antica Sede, Via Cavour 31,Torino, Italy. gcanavese@molinette.piemonte.it

    The sentinel node (SN) technique has gained a key role in breast cancer surgery, allowing for an accurate staging of the axillary status with a minimally invasive resection. In this study, we explored the implication of three proteins (E-cadherin, a- and b-catenins) that form the cadherin-catenin complex, a receptorial structure strictly involved in tumoral vascular invasion and embolization in this biologic event. We studied the immunohistochemical expression of the complex in patients with metastatic SN, matching the group with involved nonsentinel lymph nodes (NSNs) with that having free axillary NSNs. The simultaneous staining of the SN metastases for the three proteins has been considered an indicator of preserved function. Our data confirmed the lack of cadherin-catenin complex in tumors with lobular morphology even in SN metastasis, but statistical evaluation could not prove a significant relation between complex integrity and NSN involvement. Moreover, considering traditional histopathologic parameters, only vascular peritumoral embolization was related to an increased risk of metastatic spread to axillary NSNs.

    Pathology, research and practice 2007;203;7;517-23

  • Global, in vivo, and site-specific phosphorylation dynamics in signaling networks.

    Olsen JV, Blagoev B, Gnad F, Macek B, Kumar C, Mortensen P and Mann M

    Center for Experimental BioInformatics, Department of Biochemistry and Molecular Biology, University of Southern Denmark, DK-5230 Odense, Denmark.

    Cell signaling mechanisms often transmit information via posttranslational protein modifications, most importantly reversible protein phosphorylation. Here we develop and apply a general mass spectrometric technology for identification and quantitation of phosphorylation sites as a function of stimulus, time, and subcellular location. We have detected 6,600 phosphorylation sites on 2,244 proteins and have determined their temporal dynamics after stimulating HeLa cells with epidermal growth factor (EGF) and recorded them in the Phosida database. Fourteen percent of phosphorylation sites are modulated at least 2-fold by EGF, and these were classified by their temporal profiles. Surprisingly, a majority of proteins contain multiple phosphorylation sites showing different kinetics, suggesting that they serve as platforms for integrating signals. In addition to protein kinase cascades, the targets of reversible phosphorylation include ubiquitin ligases, guanine nucleotide exchange factors, and at least 46 different transcriptional regulators. The dynamic phosphoproteome provides a missing link in a global, integrative view of cellular regulation.

    Cell 2006;127;3;635-48

  • Direct interaction of the C-terminal domain of alpha-catenin and F-actin is necessary for stabilized cell-cell adhesion.

    Pappas DJ and Rimm DL

    The Department of Cell Biology, Yale University, New Haven, Connecticut 06520-8023, USA.

    Alpha-catenin functions to anchor adherens junctions to the filamentous actin (F-actin) cytoskeleton, through direct and indirect binding mechanisms. When truncated at amino acid 865, alpha-catenin exhibited a markedly reduced F-actin binding affinity compared to wild-type. Expression of the truncated mutant in the alpha-catenin deficient colon carcinoma cell line, Clone A, could not restore an adhesive phenotype when compared. Furthermore, the truncated alpha-catenin fusion protein failed to concentrate at sites of cell-cell contact, to promote morphological changes associated with epithelial monolayers, and to stimulate resistance to shearing forces in a hanging drop aggregation assay. Subsequent attempts to isolate single residues governing the direct F-actin interaction, using neutralizing charge or reverse charge mutations of basic residues within a homology modeled alpha-catenin C-terminal 5-helix bundle, had no effect on F-actin cosedimentation. We conclude that direct attachment of alpha-catenin to F-actin is required to promote cadherin-mediated contact formation and strong cell-cell adhesive states.

    Funded by: NIGMS NIH HHS: NIH R0-1 GM57604

    Cell communication & adhesion 2006;13;3;151-70

  • Cdc42 regulates adherens junction stability and endothelial permeability by inducing alpha-catenin interaction with the vascular endothelial cadherin complex.

    Broman MT, Kouklis P, Gao X, Ramchandran R, Neamu RF, Minshall RD and Malik AB

    Department of Pharmacology, Center for Lung and Vascular Biology, The University of Illinois College of Medicine, Chicago, IL, USA.

    The endothelial adherens junctions (AJs) consist of trans-oligomers of membrane spanning vascular endothelial (VE)-cadherin proteins, which bind beta-catenin through their cytoplasmic domain. beta-Catenin in turn binds alpha-catenin and connects the AJ complex with the actin cytoskeleton. We addressed the in vivo effects of loss of VE-cadherin interactions on lung vascular endothelial permeability and the role of specific Rho GTPase effectors in regulating the increase in permeability induced by AJ destabilization. We used cationic liposomes encapsulating the mutant of VE-cadherin lacking the extracellular domain (DeltaEXD) to interfere with AJ assembly in mouse lung endothelial cells. We observed that lung vascular permeability (quantified as microvessel filtration coefficient [K(f,c)]) was increased 5-fold in lungs expressing DeltaEXD. This did not occur to the same degree on expression of the VE-cadherin mutant, DeltaEXDDeltabeta, lacking the beta-catenin-binding site. The increased vascular permeability was the result of destabilization of VE-cadherin homotypic interaction induced by a shift in the binding of beta-catenin from wild-type VE-cadherin to the expressed DeltaEXD mutant. Because DeltaEXD expression in endothelial cells activated the Rho GTPase Cdc42, we addressed its role in the mechanism of increased endothelial permeability induced by AJ destabilization. Coexpression of dominant-negative Cdc42 (N17Cdc42) prevented the increase in K(f,c) induced by DeltaEXD. This was attributed to inhibition of the association of alpha-catenin with the DeltaEXD-beta-catenin complex. The results demonstrate that Cdc42 regulates AJ permeability by controlling the binding of alpha-catenin with beta-catenin and the consequent interaction of the VE-cadherin/catenin complex with the actin cytoskeleton.

    Circulation research 2006;98;1;73-80

  • Diversification of transcriptional modulation: large-scale identification and characterization of putative alternative promoters of human genes.

    Kimura K, Wakamatsu A, Suzuki Y, Ota T, Nishikawa T, Yamashita R, Yamamoto J, Sekine M, Tsuritani K, Wakaguri H, Ishii S, Sugiyama T, Saito K, Isono Y, Irie R, Kushida N, Yoneyama T, Otsuka R, Kanda K, Yokoi T, Kondo H, Wagatsuma M, Murakawa K, Ishida S, Ishibashi T, Takahashi-Fujii A, Tanase T, Nagai K, Kikuchi H, Nakai K, Isogai T and Sugano S

    Life Science Research Laboratory, Central Research Laboratory, Hitachi, Ltd., Kokubunji, Tokyo, 185-8601, Japan.

    By analyzing 1,780,295 5'-end sequences of human full-length cDNAs derived from 164 kinds of oligo-cap cDNA libraries, we identified 269,774 independent positions of transcriptional start sites (TSSs) for 14,628 human RefSeq genes. These TSSs were clustered into 30,964 clusters that were separated from each other by more than 500 bp and thus are very likely to constitute mutually distinct alternative promoters. To our surprise, at least 7674 (52%) human RefSeq genes were subject to regulation by putative alternative promoters (PAPs). On average, there were 3.1 PAPs per gene, with the composition of one CpG-island-containing promoter per 2.6 CpG-less promoters. In 17% of the PAP-containing loci, tissue-specific use of the PAPs was observed. The richest tissue sources of the tissue-specific PAPs were testis and brain. It was also intriguing that the PAP-containing promoters were enriched in the genes encoding signal transduction-related proteins and were rarer in the genes encoding extracellular proteins, possibly reflecting the varied functional requirement for and the restricted expression of those categories of genes, respectively. The patterns of the first exons were highly diverse as well. On average, there were 7.7 different splicing types of first exons per locus partly produced by the PAPs, suggesting that a wide variety of transcripts can be achieved by this mechanism. Our findings suggest that use of alternate promoters and consequent alternative use of first exons should play a pivotal role in generating the complexity required for the highly elaborated molecular systems in humans.

    Genome research 2006;16;1;55-65

  • Phosphoproteomic analysis of the human pituitary.

    Beranova-Giorgianni S, Zhao Y, Desiderio DM and Giorgianni F

    Charles B. Stout Neuroscience Mass Spectrometry Laboratory, The University of Tennessee Health Science Center, Memphis, TN 38163, USA.

    The pituitary is the central endocrine gland that regulates the functions of various target organs in the human body. Because of the pivotal regulatory role of the pituitary, it is essential to define on a global scale the components of the pituitary protein machinery, including a comprehensive characterization of the post-translational modifications of the pituitary proteins. Of particular interest is the examination of the phosphorylation status of the pituitary in health and disease. Towards the goal of global profiling of pituitary protein phosphorylation, we report here the application of the in-gel IEF-LC-MS/MS approach to the study of the pituitary phosphoproteome. The analytical strategy combined isoelectric focusing in immobilized pH gradient strips with immobilized metal ion affinity chromatography and mass spectrometry. With this method, a total of 50 phosphorylation sites were characterized in 26 proteins. Because the investigation involved primary tissue, the findings provide a direct glimpse into the phosphoprotein machinery operating within the human pituitary tissue microenvironment.

    Funded by: NINDS NIH HHS: NS 42843

    Pituitary 2006;9;2;109-20

  • Catenin expression in T1/2 carcinomas of the floor of the mouth.

    Fillies T, Buerger H, Gaertner C, August C, Brandt B, Joos U and Werkmeister R

    Department of Cranio-Maxillofacial Surgery, University of Muenster, Waldeyerstr. 30, D-48129 Münster, Germany. fillies@uni-muenster.de

    Reduction of the expression of catenin is a crucial step in the pathogenesis, progression and prognosis of many epithelial cancers including squamous cell carcinomas (SCCs). Catenin expression in oral carcinomas was evaluated in relation to clinico-pathological features in order to determine its value as a prognostic marker. Eighty-five patients with histologically proven T1/2 squamous cell carcinoma of the oral floor who underwent surgical treatment were eligible for the study. A tissue microarray consisting of multiple representative tissue cores of each carcinoma was composed. The expression levels of alpha, beta and gamma-catenins were determined immunohistologically. Correlation between clinical features and the expression of catenin proteins was evaluated statistically using Kaplan-Meier curves, log-rank tests and chi(2)-tests. Loss of alpha-catenin expression in carcinoma of the floor of the mouth correlated significantly with poor prognosis (P=0.05). Conversely, significantly reduced rates of lymph-node metastases were observed in alpha- and beta-catenin-positive T1 and T2 SCCs. Loss of gamma-catenin expression indicated a reduced survival rate in nodal-negative tumours (P=0.02). Catenin expression in carcinomas of the floor of the mouth seems to be a predictive parameter in the prognosis of T1 and T2 SSCs.

    International journal of oral and maxillofacial surgery 2005;34;8;907-11

  • Identification of regions of alpha-catenin required for desmosome organization in epithelial cells.

    Taniguchi T, Miyazaki M, Miyashita Y, Arima T and Ozawa M

    Department of Biochemistry and Molecular Biology, Graduate School of Medical and Dental Sciences, Kagoshima University, Kagoshima 890-8544, Japan.

    Alpha-catenin, a cadherin-associated protein, links cadherin/beta-catenin and cadherin/plakoglobin complexes to the actin cytoskeleton. This protein is required for the function of cadherins, cell adhesion molecules. We transfected an alpha-catenin-deficient colon carcinoma line, which cannot organize desmosomes, with a series of alpha-catenin mutant constructs. We examined the formation of desmosomes in these cells by immunofluorescence staining using anti-desmoglein and anti-desmoplakin antibodies. The results demonstrated that either the middle or the carboxy-terminal region of alpha-catenin was required for desmosome formation. Immunoblot analysis revealed that the amounts of desmoglein and desmoplakin did not differ significantly between cells that were capable of forming desmosomes and those that failed to form desmosomes. Cell surface biotinylation revealed that desmoglein was retained intracellularly in cells that could not organize desmosomes. The internal domain binds vinculin and alpha-actinin, actin-binding proteins, while the carboxy-terminal domain has the ability to bind and bundle actin filaments. These results indicate that the interaction of alpha-catenin and actin functions in the assembly of desmosomes in epithelial cells.

    International journal of molecular medicine 2005;16;6;1003-8

  • Towards a proteome-scale map of the human protein-protein interaction network.

    Rual JF, Venkatesan K, Hao T, Hirozane-Kishikawa T, Dricot A, Li N, Berriz GF, Gibbons FD, Dreze M, Ayivi-Guedehoussou N, Klitgord N, Simon C, Boxem M, Milstein S, Rosenberg J, Goldberg DS, Zhang LV, Wong SL, Franklin G, Li S, Albala JS, Lim J, Fraughton C, Llamosas E, Cevik S, Bex C, Lamesch P, Sikorski RS, Vandenhaute J, Zoghbi HY, Smolyar A, Bosak S, Sequerra R, Doucette-Stamm L, Cusick ME, Hill DE, Roth FP and Vidal M

    Center for Cancer Systems Biology and Department of Cancer Biology, Dana-Farber Cancer Institute, Harvard Medical School, 44 Binney Street, Boston, Massachusetts 02115, USA.

    Systematic mapping of protein-protein interactions, or 'interactome' mapping, was initiated in model organisms, starting with defined biological processes and then expanding to the scale of the proteome. Although far from complete, such maps have revealed global topological and dynamic features of interactome networks that relate to known biological properties, suggesting that a human interactome map will provide insight into development and disease mechanisms at a systems level. Here we describe an initial version of a proteome-scale map of human binary protein-protein interactions. Using a stringent, high-throughput yeast two-hybrid system, we tested pairwise interactions among the products of approximately 8,100 currently available Gateway-cloned open reading frames and detected approximately 2,800 interactions. This data set, called CCSB-HI1, has a verification rate of approximately 78% as revealed by an independent co-affinity purification assay, and correlates significantly with other biological attributes. The CCSB-HI1 data set increases by approximately 70% the set of available binary interactions within the tested space and reveals more than 300 new connections to over 100 disease-associated proteins. This work represents an important step towards a systematic and comprehensive human interactome project.

    Funded by: NCI NIH HHS: R33 CA132073; NHGRI NIH HHS: P50 HG004233, R01 HG001715, RC4 HG006066, U01 HG001715; NHLBI NIH HHS: U01 HL098166

    Nature 2005;437;7062;1173-8

  • Alpha-catenin expression is decreased in patients with gastric carcinoma.

    Zhou YN, Xu CP, Chen Y, Han B, Yang SM and Fang DC

    Department of Gastroenterology, First Hospital, Lanzhou University, Lanzhou 730000, Gansu Province, China. yongningzhou@sina.com.cn

    Aim: To assess the expression of alpha-catenin in gastric carcinoma and to determine the role of alpha-catenin expression in gastric carcinogenesis.

    Methods: alpha-catenin expression was assessed by semi-quantitative reverse transcriptase-polymerase chain reaction and immunohistochemical staining in 49 gastric carcinomas, 26 adjacent non-cancerous mucosae, and gastric biopsy specimens from 11 healthy controls.

    Results: mRNA levels of alpha-catenin were reduced or absent in 34 of 49 (69%) gastric carcinoma tissues and in 5 of 26 (19%) tumor-free gastric mucosae of carcinoma patients, respectively. Of the carcinoma samples with altered alpha-catenin mRNA levels, alpha-catenin expression was negative in 20 and decreased in 14 cases. Up to 69% of tumors were stained abnormally for alpha-catenin. Of the 34 cases whose mRNA expression of alpha-catenin was reduced, 32 (94%) showed abnormal immunostaining patterns, while only 2 showed a normal alpha-catenin expression. The frequency of reduced expression of alpha-catenin mRNA was 14% in well-differentiated carcinomas, higher than that in poorly differentiated carcinomas (86%). A significant correlation was not shown between alpha-catenin expression and both depth of invasion and lymph node metastasis. Moreover, there was no statistical difference between loss or down-regulation of alpha-catenin mRNA and Helicobacter pylori (H. pylori) infection.

    Conclusion: Downregulation of alpha-catenin expression is common in gastric carcinoma, and alpha-catenin expression may be used as a differentiation marker. Downregulation of alpha-catenin expression may be an early event in tumorigenesis. Reduced alpha-catenin expression is not correlated with H. pylori infection.

    World journal of gastroenterology 2005;11;22;3468-72

  • Regulation of beta-catenin signaling and maintenance of chondrocyte differentiation by ubiquitin-independent proteasomal degradation of alpha-catenin.

    Hwang SG, Yu SS, Ryu JH, Jeon HB, Yoo YJ, Eom SH and Chun JS

    Department of Life Science, Gwangju Institute of Science and Technology, Gwangju 500-712, Korea.

    Accumulation of beta-catenin and subsequent stimulation of beta-catenin-T cell-factor (Tcf)/lymphoid-enhancerfactor (Lef) transcriptional activity causes dedifferentiation of articular chondrocytes, which is characterized by decreased type II collagen expression and initiation of type I collagen expression. This study examined the mechanisms of alpha-catenin degradation, the role of alpha-catenin in beta-catenin signaling, and the physiological significance of alpha-catenin regulation of beta-catenin signaling in articular chondrocytes. We found that both alpha- and beta-catenin accumulated during dedifferentiation of chondrocytes by escaping from proteasomal degradation. Beta-catenin degradation was ubiquitination-dependent, whereas alpha-catenin was proteasomally degraded in a ubiquitination-independent fashion. The accumulated alpha- and beta-catenin existed as complexes in the cytosol and nucleus. The complex formation between alpha- and beta-catenin blocked proteasomal degradation of alpha-catenin and also inhibited beta-catenin-Tcf/Lef transcriptional activity and the suppression of type II collagen expression associated with ectopic expression of beta-catenin, the inhibition of proteasome, or Wnt signaling. Collectively, our results indicate that ubiquitin-independent degradation of alpha-catenin regulates beta-catenin signaling and maintenance of the differentiated phenotype of articular chondrocytes.

    The Journal of biological chemistry 2005;280;13;12758-65

  • Systematic identification and analysis of mammalian small ubiquitin-like modifier substrates.

    Gocke CB, Yu H and Kang J

    Department of Pharmacology, the University of Texas Southwestern Medical Center, Dallas, Texas 75390, USA.

    Small ubiquitin-like modifier (SUMO) regulates diverse cellular processes through its reversible, covalent attachment to target proteins. Many SUMO substrates are involved in transcription and chromatin structure. Sumoylation appears to regulate the functions of target proteins by changing their subcellular localization, increasing their stability, and/or mediating their binding to other proteins. Using an in vitro expression cloning approach, we have identified 40 human SUMO1 substrates. The spectrum of human SUMO1 substrates identified in our screen suggests general roles of sumoylation in transcription, chromosome structure, and RNA processing. We have validated the sumoylation of 24 substrates in living cells. Analysis of this panel of SUMO substrates leads to the following observations. 1) Sumoylation is more efficient in vitro than in living cells. Polysumoylation occurs on several substrates in vitro. 2) SUMO isopeptidases have little substrate specificity. 3) The SUMO ligases, PIAS1 and PIASxbeta, have broader substrate specificities than does PIASy. 4) Although SUMO1 and SUMO2 are equally efficiently conjugated to a given substrate in vitro, SUMO1 conjugation is more efficient in vivo. 5) Most SUMO substrates localize to the nucleus, and sumoylation does not generally affect their subcellular localization. Therefore, sumoylation appears to regulate the functions of its substrates through multiple, context-dependent mechanisms.

    Funded by: NIGMS NIH HHS: GM61542

    The Journal of biological chemistry 2005;280;6;5004-12

  • The HIV-1 co-receptor CCR5 binds to alpha-catenin, a component of the cellular cytoskeleton.

    Schweneker M, Bachmann AS and Moelling K

    Institute of Medical Virology, University of Zurich, 8028 Zurich, Switzerland; Institute of Biochemistry, Freie Universität Berlin, 14195 Berlin, Germany.

    The chemokine receptors CCR5 and CXCR4 belong to the family of seven transmembrane-spanning G protein-coupled receptors, which have diverse functions in host cell defense and are associated with numerous diseases. CCR5 and CXCR4 are known as co-receptors for entry of HIV-1. In this study the intracellular carboxy-terminus of CCR5, which is deleted in HIV-infected long-term non-progressors, was shown to interact with the carboxy-terminus of alpha-catenin, a component of the cytoskeleton, in a yeast two-hybrid screen. This interaction was verified in mammalian cells. Furthermore, the interaction of alpha-catenin with CCR5 and CXCR4 at endogenous protein levels was demonstrated in PM1 T-lymphocytes, a host cell line of HIV-1. Our results suggest that alpha-catenin links CCR5 and CXCR4 to the cytoskeleton and is involved in the organization of these receptors at the membrane, thereby possibly affecting HIV-1 infection.

    Biochemical and biophysical research communications 2004;325;3;751-7

  • The status, quality, and expansion of the NIH full-length cDNA project: the Mammalian Gene Collection (MGC).

    Gerhard DS, Wagner L, Feingold EA, Shenmen CM, Grouse LH, Schuler G, Klein SL, Old S, Rasooly R, Good P, Guyer M, Peck AM, Derge JG, Lipman D, Collins FS, Jang W, Sherry S, Feolo M, Misquitta L, Lee E, Rotmistrovsky K, Greenhut SF, Schaefer CF, Buetow K, Bonner TI, Haussler D, Kent J, Kiekhaus M, Furey T, Brent M, Prange C, Schreiber K, Shapiro N, Bhat NK, Hopkins RF, Hsie F, Driscoll T, Soares MB, Casavant TL, Scheetz TE, Brown-stein MJ, Usdin TB, Toshiyuki S, Carninci P, Piao Y, Dudekula DB, Ko MS, Kawakami K, Suzuki Y, Sugano S, Gruber CE, Smith MR, Simmons B, Moore T, Waterman R, Johnson SL, Ruan Y, Wei CL, Mathavan S, Gunaratne PH, Wu J, Garcia AM, Hulyk SW, Fuh E, Yuan Y, Sneed A, Kowis C, Hodgson A, Muzny DM, McPherson J, Gibbs RA, Fahey J, Helton E, Ketteman M, Madan A, Rodrigues S, Sanchez A, Whiting M, Madari A, Young AC, Wetherby KD, Granite SJ, Kwong PN, Brinkley CP, Pearson RL, Bouffard GG, Blakesly RW, Green ED, Dickson MC, Rodriguez AC, Grimwood J, Schmutz J, Myers RM, Butterfield YS, Griffith M, Griffith OL, Krzywinski MI, Liao N, Morin R, Morrin R, Palmquist D, Petrescu AS, Skalska U, Smailus DE, Stott JM, Schnerch A, Schein JE, Jones SJ, Holt RA, Baross A, Marra MA, Clifton S, Makowski KA, Bosak S, Malek J and MGC Project Team

    The National Institutes of Health's Mammalian Gene Collection (MGC) project was designed to generate and sequence a publicly accessible cDNA resource containing a complete open reading frame (ORF) for every human and mouse gene. The project initially used a random strategy to select clones from a large number of cDNA libraries from diverse tissues. Candidate clones were chosen based on 5'-EST sequences, and then fully sequenced to high accuracy and analyzed by algorithms developed for this project. Currently, more than 11,000 human and 10,000 mouse genes are represented in MGC by at least one clone with a full ORF. The random selection approach is now reaching a saturation point, and a transition to protocols targeted at the missing transcripts is now required to complete the mouse and human collections. Comparison of the sequence of the MGC clones to reference genome sequences reveals that most cDNA clones are of very high sequence quality, although it is likely that some cDNAs may carry missense variants as a consequence of experimental artifact, such as PCR, cloning, or reverse transcriptase errors. Recently, a rat cDNA component was added to the project, and ongoing frog (Xenopus) and zebrafish (Danio) cDNA projects were expanded to take advantage of the high-throughput MGC pipeline.

    Funded by: PHS HHS: N01-C0-12400

    Genome research 2004;14;10B;2121-7

  • Identification and in vivo role of the Armadillo-Legless interaction.

    Hoffmans R and Basler K

    Institut für Molekularbiologie, Universität Zürich, Winterthurerstrasse 190, 8057 Zürich, Switzerland.

    The Wnt signalling system controls many fundamental processes during animal development and its deregulation has been causally linked to colorectal cancer. Transduction of Wnt signals entails the association of beta-catenin with nuclear TCF DNA-binding factors and the subsequent activation of target genes. Using genetic assays in Drosophila, we have recently identified a presumptive adaptor protein, Legless (Lgs), that binds to beta-catenin and mediates signalling activity by recruiting the transcriptional activator Pygopus (Pygo). Here, we characterize the beta-catenin/Lgs interaction and show: (1) that it is critically dependent on two acidic amino acid residues in the first Armadillo repeat of beta-catenin; (2) that it is spatially and functionally separable from the binding sites for TCF factors, APC and E-cadherin; (3) that it is required in endogenous as well as constitutively active forms of beta-catenin for Wingless signalling output in Drosophila; and (4) that in its absence animals develop with the same phenotypic consequences as animals lacking Lgs altogether. Based on these findings, and because Lgs and Pygo have human homologues that can substitute for their Drosophila counterparts, we infer that the beta-catenin/Lgs binding site may thus serve as an attractive drug target for therapeutic intervention in beta-catenin-dependent cancer progression.

    Development (Cambridge, England) 2004;131;17;4393-400

  • Large-scale characterization of HeLa cell nuclear phosphoproteins.

    Beausoleil SA, Jedrychowski M, Schwartz D, Elias JE, Villén J, Li J, Cohn MA, Cantley LC and Gygi SP

    Department of Cell Biology, Harvard Medical School, Boston, MA 02115, USA.

    Determining the site of a regulatory phosphorylation event is often essential for elucidating specific kinase-substrate relationships, providing a handle for understanding essential signaling pathways and ultimately allowing insights into numerous disease pathologies. Despite intense research efforts to elucidate mechanisms of protein phosphorylation regulation, efficient, large-scale identification and characterization of phosphorylation sites remains an unsolved problem. In this report we describe an application of existing technology for the isolation and identification of phosphorylation sites. By using a strategy based on strong cation exchange chromatography, phosphopeptides were enriched from the nuclear fraction of HeLa cell lysate. From 967 proteins, 2,002 phosphorylation sites were determined by tandem MS. This unprecedented large collection of sites permitted a detailed accounting of known and unknown kinase motifs and substrates.

    Funded by: NHGRI NIH HHS: HG00041, K22 HG000041, T32 HG000041; NIGMS NIH HHS: GM67945, GMS6203, R01 GM056203, R01 GM067945

    Proceedings of the National Academy of Sciences of the United States of America 2004;101;33;12130-5

  • Marked allelic imbalance on chromosome 5q31 does not explain alpha-catenin expression in epithelial ovarian cancer.

    Tuhkanen H, Anttila M, Kosma VM, Puolakka J, Juhola M, Heinonen S and Mannermaa A

    Department of Pathology and Forensic Medicine, University Hospital, University of Kuopio, 70210 Kuopio, Finland.

    Objective: Human alpha-catenin gene (CTNNA1) on chromosome 5q31 is aberrantly expressed in various types of cancer including epithelial ovarian tumors. Allelic imbalance on this region has also been described in several malignant diseases. In the present work, the role of CTNNA1 as a candidate tumor suppressor gene was studied by comparing protein expression with allelic imbalance in human epithelial ovarian tumors.

    Methods: Alpha-catenin protein expression was determined from two areas of 41 tumors, and tissues from these areas were microdissected. After DNA extraction, AI analysis was carried out with eight microsatellite markers.

    Results: Altogether, 93% of the tumors (38 of 41) showed allelic imbalance at one or more loci. Two distinct common regions of allelic imbalance were identified, one comprising markers D5S2002 and D5S1995 and the other markers D5S393 and D5S476. Loss of the CTNNA1 gene did not appear to be involved in down-regulation of alpha-catenin in ovarian tumors, since allelic imbalance with a variety of markers, including CTNNA1 associated marker D5S476, was found in tumor samples independently of alpha-catenin expression. Furthermore, allelic imbalance analyses of two different samples from the same tumor revealed genetic heterogeneity.

    Conclusions: High allelic imbalance frequency indicates that chromosomal region 5q31 is functionally important in epithelial ovarian cancer. Allelic imbalance occurs at two distinct regions of which one includes the CTNNA1 gene. However, this gene is likely to be inactivated by mechanisms other than allelic imbalance. In addition, genetic heterogeneity observed in these tumors demonstrates the multiclonal nature of epithelial ovarian tumors.

    Gynecologic oncology 2004;94;2;416-21

  • Expression of alpha-catenin in alpha-catenin-deficient cells results in a reduced proliferation in three-dimensional multicellular spheroids but not in two-dimensional monolayer cultures.

    Matsubara S and Ozawa M

    Department of Biochemistry and Molecular Biology, Graduate School of Medical and Dental Science, Kagoshima University, Kagoshima 890-8544, Japan. shmlmcbd@m.kufm.kagoshima-u.ac.jp

    alpha-Catenin is an intracellular protein that associates with the carboxy-terminal region of cadherin, a cell adhesion molecule, via beta-catenin or gamma-catenin (plakoglobin). Linkage of cadherin to the cytoskeleton by catenins is required for full cadherin activity. Following transfection of an alpha-catenin-deficient colon carcinoma cell line with a series of alpha-catenin constructs, we discovered that the restoration of alpha-catenin expression results in reduced proliferation in three-dimensional multicellular spheroids, but not in two-dimensional monolayer cultures. The cellular function of alpha-catenin has not been compared between cells in three- and two-dimensional culture; this is the first evidence that growth regulation in three-dimensional cultures requires signaling mediated by alpha-catenin. Two classes of constructs, containing deletions in either the central segment or the COOH terminus of the molecule, both induced morphological changes, including cell compaction, and suppressed cell growth in three-dimensional cultures. In alpha-catenin-expressing cells, inhibition of cadherin cell adhesion by treatment with anti-E-cadherin antibodies resulted in a similar phenotype as that observed following the loss of alpha-catenin. Therefore, both the homophilic interaction of the cadherin extracellular domain and the linkage of the cadherin cytoplasmic domain to the actin cytoskeleton by alpha-catenin are necessary for growth control in three-dimensional culture.

    Oncogene 2004;23;15;2694-702

  • Complete sequencing and characterization of 21,243 full-length human cDNAs.

    Ota T, Suzuki Y, Nishikawa T, Otsuki T, Sugiyama T, Irie R, Wakamatsu A, Hayashi K, Sato H, Nagai K, Kimura K, Makita H, Sekine M, Obayashi M, Nishi T, Shibahara T, Tanaka T, Ishii S, Yamamoto J, Saito K, Kawai Y, Isono Y, Nakamura Y, Nagahari K, Murakami K, Yasuda T, Iwayanagi T, Wagatsuma M, Shiratori A, Sudo H, Hosoiri T, Kaku Y, Kodaira H, Kondo H, Sugawara M, Takahashi M, Kanda K, Yokoi T, Furuya T, Kikkawa E, Omura Y, Abe K, Kamihara K, Katsuta N, Sato K, Tanikawa M, Yamazaki M, Ninomiya K, Ishibashi T, Yamashita H, Murakawa K, Fujimori K, Tanai H, Kimata M, Watanabe M, Hiraoka S, Chiba Y, Ishida S, Ono Y, Takiguchi S, Watanabe S, Yosida M, Hotuta T, Kusano J, Kanehori K, Takahashi-Fujii A, Hara H, Tanase TO, Nomura Y, Togiya S, Komai F, Hara R, Takeuchi K, Arita M, Imose N, Musashino K, Yuuki H, Oshima A, Sasaki N, Aotsuka S, Yoshikawa Y, Matsunawa H, Ichihara T, Shiohata N, Sano S, Moriya S, Momiyama H, Satoh N, Takami S, Terashima Y, Suzuki O, Nakagawa S, Senoh A, Mizoguchi H, Goto Y, Shimizu F, Wakebe H, Hishigaki H, Watanabe T, Sugiyama A, Takemoto M, Kawakami B, Yamazaki M, Watanabe K, Kumagai A, Itakura S, Fukuzumi Y, Fujimori Y, Komiyama M, Tashiro H, Tanigami A, Fujiwara T, Ono T, Yamada K, Fujii Y, Ozaki K, Hirao M, Ohmori Y, Kawabata A, Hikiji T, Kobatake N, Inagaki H, Ikema Y, Okamoto S, Okitani R, Kawakami T, Noguchi S, Itoh T, Shigeta K, Senba T, Matsumura K, Nakajima Y, Mizuno T, Morinaga M, Sasaki M, Togashi T, Oyama M, Hata H, Watanabe M, Komatsu T, Mizushima-Sugano J, Satoh T, Shirai Y, Takahashi Y, Nakagawa K, Okumura K, Nagase T, Nomura N, Kikuchi H, Masuho Y, Yamashita R, Nakai K, Yada T, Nakamura Y, Ohara O, Isogai T and Sugano S

    Helix Research Institute, 1532-3 Yana, Kisarazu, Chiba 292-0812, Japan.

    As a base for human transcriptome and functional genomics, we created the "full-length long Japan" (FLJ) collection of sequenced human cDNAs. We determined the entire sequence of 21,243 selected clones and found that 14,490 cDNAs (10,897 clusters) were unique to the FLJ collection. About half of them (5,416) seemed to be protein-coding. Of those, 1,999 clusters had not been predicted by computational methods. The distribution of GC content of nonpredicted cDNAs had a peak at approximately 58% compared with a peak at approximately 42%for predicted cDNAs. Thus, there seems to be a slight bias against GC-rich transcripts in current gene prediction procedures. The rest of the cDNAs unique to the FLJ collection (5,481) contained no obvious open reading frames (ORFs) and thus are candidate noncoding RNAs. About one-fourth of them (1,378) showed a clear pattern of splicing. The distribution of GC content of noncoding cDNAs was narrow and had a peak at approximately 42%, relatively low compared with that of protein-coding cDNAs.

    Nature genetics 2004;36;1;40-5

  • A novel cell-cell junction system: the cortex adhaerens mosaic of lens fiber cells.

    Straub BK, Boda J, Kuhn C, Schnoelzer M, Korf U, Kempf T, Spring H, Hatzfeld M and Franke WW

    Division of Cell Biology, German Cancer Research Center, Im Neuenheimer Feld 280, 69120 Heidelberg, Germany.

    The anucleate prismoid fiber cells of the eye lens are densely packed to form a tissue in which the plasma membranes and their associated cytoplasmic coat form a single giant cell-cell adhesive complex, the cortex adhaerens. Using biochemical and immunoprecipitation methods in various species (cow, pig, rat), in combination with immunolocalization microscopy, we have identified two different major kinds of cortical complex. In one, the transmembrane glycoproteins N-cadherin and cadherin-11 [which also occur in heterotypic ('mixed') complexes] are associated with alpha- and beta-catenin, plakoglobin (proportions variable among species), p120ctn and vinculin. The other complex contains ezrin, periplakin, periaxin and desmoyokin (and so is called the EPPD complex), usually together with moesin, spectrin(s) and plectin. In sections through lens fiber tissue, the short sides of the lens fiber hexagons appear to be enriched in the cadherin-based complexes, whereas the EPPD complexes also occur on the long sides. Moreover, high resolution double-label fluorescence microscopy has revealed, on the short sides, a finer, almost regular mosaicism of blocks comprising the cadherin-based, catenin-containing complexes, alternating with patches formed by the EPPD complexes. The latter, a new type of junctional plaque ensemble of proteins hitherto known only from certain other cell types, must be added to the list of major lens cortex proteins. We here discuss its possible functional importance for the maintenance of lens structure and functions, notably clear and sharp vision.

    Journal of cell science 2003;116;Pt 24;4985-95

  • Carbonic anhydrase IX reduces E-cadherin-mediated adhesion of MDCK cells via interaction with beta-catenin.

    Svastová E, Zilka N, Zat'ovicová M, Gibadulinová A, Ciampor F, Pastorek J and Pastoreková S

    Centre of Molecular Medicine, Institute of Virology, Slovak Academy of Sciences, Dúbravská cesta 9, 845 05 Bratislava, Slovak Republic.

    Carbonic anhydrase IX (CA IX) is a cancer-associated transmembrane isoform of zinc metalloenzymes that catalyse interconversion between carbon dioxide and bicarbonate. CA IX is strongly induced by tumor hypoxia and has been proposed to participate in acidification of tumor microenvironment and in cell adhesion. To elucidate the cell adhesion-related role of CA IX, we investigated its subcellular localization and relationship to E-cadherin, a key adhesion molecule whose loss or destabilization is linked to tumor invasion. For this purpose, we generated MDCK cells with constitutive expression of human CA IX protein. During the monolayer formation, CA IX was localized to cell-cell contacts and its distribution in lateral membranes overlapped with E-cadherin. Calcium switch-triggered disruption and reconstitution of cell contacts resulted in relocalization of both CA IX and E-cadherin to cytoplasm and back to plasma membrane. A similar phenomenon was observed in hypoxia-treated and reoxygenated cells. Moreover, CA IX-expressing MDCK cells exhibited reduced cell adhesion capacity and lower levels of Triton-insoluble E-cadherin. Finally, CA IX was found to coprecipitate with beta-catenin. We conclude that CA IX has a capacity to modulate E-cadherin-mediated cell adhesion via interaction with beta-catenin, which could be of potential significance in hypoxia-induced tumor progression.

    Experimental cell research 2003;290;2;332-45

  • Relationship between expressions of E-cadherin and alpha-catenin and biological behaviors of human pancreatic cancer.

    Li YJ, Meng YX and Ji XR

    Department of Pathology, Affiliated Hospital of Medical College, Qingdao University, Qingdao 266003, China. yujun-li66@yahoo.com

    Objectives: To investigate the expressions of E-cadherin and alpha-catenin in pancreatic carcinoma and their relationship with biological behaviors, and clarify the mechanism of invasion and metastasis of pancreatic cancer.

    Methods: The expressions of E-cadherin and alpha-catenin was examined in 47 patients with infiltrative ductal adenocarcinoma of the pancreas and 12 specimens of normal pancreatic tissues by immunohistochemical technique (PicTure( trade mark ) two-step method). Proliferation cell nuclear antigen (PCNA) was tested as an index of the proliferation degree of pancreatic cancer cells.

    Results: The immunoreactivity of E-cadherin and alpha-catenin was expressed by normal ductal and acinar cells with strong membranous staining at the intercellular border in 12 specimens of normal pancreatic tissues. The abnormal rate of E-cadherin expression in pancreatic cancer was 53.2% (25/47), and it was significantly related to differentiation, high proliferation degree and lymph node and liver metastases (P<0.01, 0.05, 0.05 and 0.01, respectively). 61.7% patients with pancreatic cancer (29/47) showed abnormal expression of alpha-catenin. There was a good correlation among alpha-catenin expression, histological grade, and lymph node and liver metastases (P<0.05,0.05 and 0.01, respectively). No significant association was found among abnormal expressions of E-cadherin and alpha-catenin, tumor size, invasion, and 1-year survival rate of patients (P>0.05, all). There was a positive relationship between the expressions of E-cadherin and alpha-catenin in the 47 patients with pancreatic cancer (P<0.01, r=0.88).

    Conclusions: Pancreatic cancer likely occurs in case of the inactivation of E-cadherin and alpha-catenin genes and abnormal expression of proteins, which significantly correlate with tumorigenesis, proliferation, differentiation, and lymph node or liver metastasis of pancreatic cancer.

    Hepatobiliary & pancreatic diseases international : HBPD INT 2003;2;3;471-7

  • Characterization of cadherin-24, a novel alternatively spliced type II cadherin.

    Katafiasz BJ, Nieman MT, Wheelock MJ and Johnson KR

    University of Nebraska Medical Center, Department of Oral Biology, College of Dentistry and Eppley Cancer Center, Omaha, Nebraska 68198, USA.

    Cadherins comprise a superfamily of calcium-dependent cell-cell adhesion molecules. Within the superfamily are six subfamilies including type I and type II cadherins. Both type I and type II cadherins are composed of five extracellular repeat domains with conserved calcium-binding motifs, a single pass transmembrane domain, and a highly conserved cytoplasmic domain that interacts with beta-catenin and p120 catenin. In this study, we describe a novel cadherin, cadherin-24. It is a type II cadherin with a 781-codon open reading frame, which encodes a type II cadherin protein complete with five extracellular repeats containing calcium-binding motifs, a transmembrane domain, and a conserved cytoplasmic domain. Cadherin-24 has the unusual feature of being alternatively spliced in extracellular repeat 4. This alternative exon encodes 38 in-frame amino acids, resulting in an 819-amino-acid protein. Sequence analysis suggests the presence of beta-catenin and p120 catenin-binding sequences, and immunoprecipitation experiments confirm the ability of both forms of the novel cadherin to associate with alpha-catenin, beta-catenin, and p120 catenin. In addition, aggregation assays show that both forms of cadherin-24 mediate strong cell-cell adhesion.

    Funded by: NIGMS NIH HHS: GM51188

    The Journal of biological chemistry 2003;278;30;27513-9

  • The protein tyrosine phosphatase Pez is a major phosphatase of adherens junctions and dephosphorylates beta-catenin.

    Wadham C, Gamble JR, Vadas MA and Khew-Goodall Y

    Hanson Centre for Cancer Research, Institute of Medical and Veterinary Science, Adelaide, SA 5000, Australia.

    Cell-cell adhesion regulates processes important in embryonal development, normal physiology, and cancer progression. It is regulated by various mechanisms including tyrosine phosphorylation. We have previously shown that the protein tyrosine phosphatase Pez is concentrated at intercellular junctions in confluent, quiescent monolayers but is nuclear in cells lacking cell-cell contacts. We show here with an epithelial cell model that Pez localizes to the adherens junctions in confluent monolayers. A truncation mutant lacking the catalytic domain acts as a dominant negative mutant to upregulate tyrosine phosphorylation at adherens junctions. We identified beta-catenin, a component of adherens junctions, as a substrate of Pez by a "substrate trapping" approach and by in vitro dephosphorylation with recombinant Pez. Consistent with this, ectopic expression of the dominant negative mutant caused an increase in tyrosine phosphorylation of beta-catenin, demonstrating that Pez regulates the level of tyrosine phosphorylation of adherens junction proteins, including beta-catenin. Increased tyrosine phosphorylation of adherens junction proteins has been shown to decrease cell-cell adhesion, promoting cell migration as a result. Accordingly, the dominant negative Pez mutant enhanced cell motility in an in vitro "wound" assay. This suggests that Pez is also a regulator of cell motility, most likely through its action on cell-cell adhesion.

    Molecular biology of the cell 2003;14;6;2520-9

  • N-cadherin-catenin complexes form prior to cleavage of the proregion and transport to the plasma membrane.

    Wahl JK, Kim YJ, Cullen JM, Johnson KR and Wheelock MJ

    University of Nebraska Medical Center, College of Dentistry and Eppley Cancer Center, Omaha, Nebraska 68198-7696, USA. Jwahl@unmc.edu

    Cadherins are calcium-dependent glycoproteins that function as cell-cell adhesion molecules and are linked to the actin cytoskeleton via catenins. Newly synthesized cadherins contain a prosequence that must be proteolytically removed to generate a functional adhesion molecule. The goal of this study was to examine the proteolytic processing of N-cadherin and the assembly of the cadherin-catenin complex in cells that express endogenous N-cadherin. A monoclonal antibody specific for the proregion of human N-cadherin was generated and used to examine N-cadherin processing. Our data show that newly synthesized proN-cadherin is phosphorylated and proteolytically processed prior to transport to the plasma membrane. In addition, we show that beta-catenin and plakoglobin associate only with phosphorylated proN-cadherin, whereas p120(ctn) can associate with both phosphorylated and non-phosphorylated proN-cadherin. Immunoprecipitations using anti-proN-cadherin showed that cadherin-catenin complexes are assembled prior to localization at the plasma membrane. These data suggest that a core N-cadherin-catenin complex assembles in the endoplasmic reticulum or Golgi compartment and is transported to the plasma membrane where linkage to the actin cytoskeleton can be established.

    Funded by: NIDCR NIH HHS: DE12308; NIGMS NIH HHS: GM51188

    The Journal of biological chemistry 2003;278;19;17269-76

  • p120 Catenin-associated Fer and Fyn tyrosine kinases regulate beta-catenin Tyr-142 phosphorylation and beta-catenin-alpha-catenin Interaction.

    Piedra J, Miravet S, Castaño J, Pálmer HG, Heisterkamp N, García de Herreros A and Duñach M

    Unitat de Biofísica, Departament de Bioquímica i Biologia Molecular, Facultat de Medicina, Universitat Autònoma de Barcelona, E-08193 Bellaterra, Spain.

    beta-Catenin has a key role in the formation of adherens junction through its interactions with E-cadherin and alpha-catenin. We show here that interaction of beta-catenin with alpha-catenin is regulated by the phosphorylation of beta-catenin Tyr-142. This residue can be phosphorylated in vitro by Fer or Fyn tyrosine kinases. Transfection of these kinases to epithelial cells disrupted the association between both catenins. We have also examined whether these kinases are involved in the regulation of this interaction by K-ras. Stable transfectants of the K-ras oncogene in intestinal epithelial IEC18 cells were generated which show little alpha-catenin-beta-catenin association with respect to control clones; this effect is accompanied by increased Tyr-142 phosphorylation and activation of Fer and Fyn kinases. As reported for Fer, Fyn kinase is constitutively bound to p120 catenin; expression of K-ras induces the phosphorylation of p120 catenin on tyrosine residues increasing its affinity for E-cadherin and, consequently, promotes the association of Fyn with the adherens junction complex. Yes tyrosine kinase also binds to p120 catenin but only upon activation, and stimulates Fer and Fyn tyrosine kinases. These results indicate that p120 catenin acts as a docking protein facilitating the activation of Fer/Fyn tyrosine kinases by Yes and demonstrate the role of these p120 catenin-associated kinases in the regulation of beta-catenin-alpha-catenin interaction.

    Molecular and cellular biology 2003;23;7;2287-97

  • Comprehensive proteomic analysis of breast cancer cell membranes reveals unique proteins with potential roles in clinical cancer.

    Adam PJ, Boyd R, Tyson KL, Fletcher GC, Stamps A, Hudson L, Poyser HR, Redpath N, Griffiths M, Steers G, Harris AL, Patel S, Berry J, Loader JA, Townsend RR, Daviet L, Legrain P, Parekh R and Terrett JA

    Oxford Glycosciences, The Forum, 86 Milton Park, Abingdon, Oxford OX14 4RY, United Kingdom.

    Proteins associated with cancer cell plasma membranes are rich in known drug and antibody targets as well as other proteins known to play key roles in the abnormal signal transduction processes required for carcinogenesis. We describe here a proteomics process that comprehensively annotates the protein content of breast tumor cell membranes and defines the clinical relevance of such proteins. Tumor-derived cell lines were used to ensure an enrichment for cancer cell-specific plasma membrane proteins because it is difficult to purify cancer cells and then obtain good membrane preparations from clinical material. Multiple cell lines with different molecular pathologies were used to represent the clinical heterogeneity of breast cancer. Peptide tandem mass spectra were searched against a comprehensive data base containing known and conceptual proteins derived from many public data bases including the draft human genome sequences. This plasma membrane-enriched proteome analysis created a data base of more than 500 breast cancer cell line proteins, 27% of which were of unknown function. The value of our approach is demonstrated by further detailed analyses of three previously uncharacterized proteins whose clinical relevance has been defined by their unique cancer expression profiles and the identification of protein-binding partners that elucidate potential functionality in cancer.

    The Journal of biological chemistry 2003;278;8;6482-9

  • Relationship between expression of E-cadherin-catenin complex and clinicopathologic characteristics of pancreatic cancer.

    Li YJ and Ji XR

    Department of Pathology, the Affiliated Hospital of Medical College, Qingdao University, 16 Jiangsu Rd, Qingdao 266003, China. yunjun-Li66@yahoo.com

    Aim: To investigate the expression of E-cadherin and alpha-catenin and beta-catenin in pancreatic carcinoma and its relationship with the clinicopathologic characteristics, and clarify the mechanism of invasion and metastasis of pancreatic cancer.

    Methods: The expression of E-cadherin and alpha-, beta-catenin was examined in 47 cases of infiltrative ductal adenocarcinoma of pancreas and 12 adult normal pancreatic tissues by immunohistochemical technique.

    Results: The immunoreactivity of E-cadherin and alpha-, beta-catenin was expressed by normal ductal and acinar cells with strong membranous staining at the intercellular border in 12 cases of adult normal pancreatic tissues. Abnormal expression of E-cadherin and alpha-, beta-catenin in 47 pancreatic carcinoma tissues was demonstrated in 53.2 %, 61.7 % and 68.1 %, respectively. Both abnormal expression of E-cadherin and alpha-catenin significantly correlated with differentiation, lymph node and liver metastases (P<0.05, respectively), whereas aberrant beta-catenin expression only correlated with lymph node and liver metastases (P<0.001). Abnormal E-cadherin and alpha-, beta-catenin expression was not associated with tumor size, invasion and survival time of patients (P>0.05, all).

    Conclusion: Pancreatic cancer likely occurs in case of E-cadherin-catenin complex genes mutations or deletions and abnormal expression of proteins, which significantly correlate with the biologic character of the tumor and lymph node and liver metastases. It is suggested that the abnormal E-cadherin-catenin complex expression plays an important role in the development and progression of tumor, and thus may become a new marker in pancreatic cancer metastasis.

    World journal of gastroenterology 2003;9;2;368-72

  • The LIM protein Ajuba is recruited to cadherin-dependent cell junctions through an association with alpha-catenin.

    Marie H, Pratt SJ, Betson M, Epple H, Kittler JT, Meek L, Moss SJ, Troyanovsky S, Attwell D, Longmore GD and Braga VM

    Medical Research Council Laboratory for Molecular Cell Biology and Department of Physiology, University College London, Gower Street, United Kingdom.

    Cell-cell adhesive events affect cell growth and fate decisions and provide spatial clues for cell polarity within tissues. The complete molecular determinants required for adhesive junction formation and their function are not completely understood. LIM domain-containing proteins have been shown to be present at cell-cell contact sites and are known to shuttle into the nucleus where they can affect cell fate and growth; however, their precise localization at cell-cell contacts, how they localize to these sites, and what their functions are at these sites is unknown. Here we show that, in primary keratinocytes, the LIM domain protein Ajuba is recruited to cadherin-dependent cell-cell adhesive complexes in a regulated manner. At cadherin adhesive complexes Ajuba interacts with alpha-catenin, and alpha-catenin is required for efficient recruitment of Ajuba to cell junctions. Ajuba also interacts directly with F-actin. Keratinocytes from Ajuba null mice exhibit abnormal cell-cell junction formation and/or stability and function. These data reveal Ajuba as a new component at cadherin-mediated cell-cell junctions and suggest that Ajuba may contribute to the bridging of the cadherin adhesive complexes to the actin cytoskeleton and as such contribute to the formation or strengthening of cadherin-mediated cell-cell adhesion.

    Funded by: NCI NIH HHS: CA85839

    The Journal of biological chemistry 2003;278;2;1220-8

  • Histamine stimulates phosphorylation of adherens junction proteins and alters their link to vimentin.

    Shasby DM, Ries DR, Shasby SS and Winter MC

    Department of Internal Medicine, University of Iowa College of Medicine, Iowa City, Iowa 52242, USA. michael.shasby@uiowa.edu

    Histamine increases microvascular permeability by creating small transitory (100-400 nm) gaps between adjacent endothelial cells at sites of vascular endothelial (VE)-cadherin-based adhesion. We examined the effects of histamine on the proteins within the VE-cadherin-based adherens junction in primary human umbilical vein endothelial cells. VE-cadherin is linked not only by beta- and alpha-catenin to cortical actin but also by gamma-catenin to the intermediate filament vimentin. In mature human umbilical vein cultures, the VE-cadherin immunoprecipitate contained equivalent amounts of alpha- and beta-catenin, 130% as much beta- as gamma-catenin, and 50% as much actin as vimentin. Within 60 s, histamine decreased the fraction of VE-cadherin in the insoluble portion of the cell lysate by 35 +/- 1.5%. At the same time, histamine decreased the amount of vimentin that immunoprecipitated with VE-cadherin by 50 +/- 6%. Histamine did not affect the amount of actin or the amount of alpha-, beta-, or gamma-catenin that immunoprecipitated with VE-cadherin. Within 60 s, histamine simulated a doubling in the phosphorylation of VE-cadherin and beta- and gamma-catenin. The VE-cadherin immunoprecipitate contained kinase activity that phosphorylated VE-cadherin and gamma-catenin in vitro.

    Funded by: NHLBI NIH HHS: HL-335450, HL-63100

    American journal of physiology. Lung cellular and molecular physiology 2002;282;6;L1330-8

  • Biochemical and structural definition of the l-afadin- and actin-binding sites of alpha-catenin.

    Pokutta S, Drees F, Takai Y, Nelson WJ and Weis WI

    Department of Structural Biology, Stanford University School of Medicine, Stanford, California 94305, USA.

    alpha-Catenin is an integral component of adherens junctions, where it links cadherins to the actin cytoskeleton. alpha-Catenin is also required for the colocalization of the nectin/afadin/ponsin adhesion system to adherens junctions, and it specifically associates with the nectin-binding protein afadin. A proteolytic fragment of alpha-catenin, residues 385-651, contains the afadin-binding site. The three-dimensional structure of this fragment comprises two side-by-side four-helix bundles, both of which are required for afadin binding. The alpha-catenin fragment 385-651 binds afadin more strongly than the full-length protein, suggesting that the full-length protein harbors a cryptic binding site for afadin. Comparison of the alpha-catenin 385-651 structure with the recently solved structure of the alpha-catenin M-fragment (Yang, J., Dokurno, P., Tonks, N. K., and Barford, D. (2001) EMBO J. 20, 3645-3656) reveals a surprising flexibility in the orientation of the two four-helix bundles. alpha-Catenin and the actin-binding protein vinculin share sequence and most likely structural similarity within their actin-binding domains. Despite this homology, actin binding requires additional sequences adjacent to this region.

    Funded by: NIGMS NIH HHS: GM35227, GM56169, R01 GM035527, R01 GM056169

    The Journal of biological chemistry 2002;277;21;18868-74

  • UCS15A, a novel small molecule, SH3 domain-mediated protein-protein interaction blocking drug.

    Oneyama C, Nakano H and Sharma SV

    Tokyo Research Laboratories, Kyowa Hakko Kogyo Co., Ltd 3-6-6 Asahi-cho, Machida-shi, Tokyo 194, Japan.

    Protein-protein interactions play critical regulatory roles in mediating signal transduction. Previous studies have identified an unconventional, small-molecule, Src signal transduction inhibitor, UCS15A. UCS15A differed from conventional Src-inhibitors in that it did not alter the levels or the tyrosine kinase activity of Src. Our studies suggested that UCS15A exerted its Src-inhibitory effects by a novel mechanism that involved the disruption of protein-protein interactions mediated by Src. In the present study we have examined the ability of UCS15A to disrupt the interaction of Src-SH3 with Sam68, both in vivo and in vitro. This ability of UCS15A was not restricted to Src-SH3 mediated protein-protein interactions, since the drug was capable of disrupting the in vivo interactions of Sam68 with other SH3 domain containing proteins such as Grb2 and PLCgamma. In addition, UCS15A was capable of disrupting other typical SH3-mediated protein-protein interactions such as Grb2-Sos1, cortactin-ZO1, as well as atypical SH3-mediated protein-protein interactions such as Grb2-Gab1. However, UCS15A was unable to disrupt the non-SH3-mediated protein-protein interactions of beta-catenin, with E-cadherin and alpha-catenin. In addition, UCS15A had no effect on the SH2-mediated interaction between Grb2 and activated Epidermal Growth Factor receptor. Thus, the ability of UCS15A, to disrupt protein-protein interactions appeared to be restricted to SH3-mediated protein-protein interactions. In this regard, UCS15A represents the first example of a non-peptide, small molecule agent capable of disrupting SH3-mediated protein-protein interactions. In vitro analyses suggested that UCS15A did not bind to the SH3 domain itself but rather may interact directly with the target proline-rich domains.

    Oncogene 2002;21;13;2037-50

  • Leucine zipper domain of HIV-1 gp41 interacted specifically with alpha-catenin.

    Kim JT, Kim EM, Lee KH, Choi JE, Jhun BH and Kim JW

    Bio-Med RRC, Division of Life Sciences, Pai Chai University, Taejon 302-735, Korea.

    Interactions between viral and cellular proteins could explain the molecular mechanisms behind the viral life cycle of HIV-1. The envelope protein gp41 of HIV-1 specifically interacted with alpha-catenin, not with beta-catenin. This interaction was shown by in vitro protein assay and in vivo transfected cell systems. Microinjection of the DNA expressing HIV-1 gp160 and alpha-catenin, into the HeLa cell, resulted in the colocalization of gp41 and alpha-catenin. Interestingly the noncleavable mutant of gp160 and alpha-catenin were found to be colocalized in the cell membrane. Mapping of the interaction sites between these two proteins revealed that the leucine zipper-like structure, located between the first and second alpha-helix domains from the carboxy terminus of HIV-1 gp41, interacted strongly with the carboxy terminus of alpha-catenin.

    Biochemical and biophysical research communications 2002;291;5;1239-44

  • Polycystin: new aspects of structure, function, and regulation.

    Wilson PD

    Mount Sinai School of Medicine, 1425 Madison Avenue, New York, NY 10029, USA. pat.wilson@mssm.edu

    Polycystin-1 is a modular membrane protein with a long extracellular N-terminal portion that bears several ligand-binding domains, 11 transmembrane domains, and a > or =200 amino acid intracellular C-terminal portion with several phosphorylation signaling sites. Polycystin-1 is highly expressed in the basal membranes of ureteric bud epithelia during early development of the metanephric kidney, and disruption of the PKD1 gene in mice leads to cystic kidneys and embryonic or perinatal death. It is proposed that polycystin-1 functions as a matrix receptor to link the extracellular matrix to the actin cytoskeleton via focal adhesion proteins. Co-localization, co-sedimentation, and co-immunoprecipitation studies show that polycystin-1 forms multiprotein complexes with alpha2beta1-integrin, talin, vinculin, paxillin, p130cas, focal adhesion kinase, and c-src in normal human fetal collecting tubules and sub-confluent epithelial cultures. In normal adult kidneys and confluent epithelial cultures, polycystin-1 is downregulated and forms complexes with the cell-cell adherens junction proteins E-cadherin and beta-, gamma-, and alpha-catenin. Polycystin-1 activation at the cell membrane leads to intracellular signaling via phosphorylation through the c-Jun terminal kinase and wnt pathways leading to activation of AP-1 and TCF/LEF-dependent genes, respectively. The C-terminal of polcystin-1 has been shown to be phosphorylated by c-src at Y4237, by protein kinase A at S4252, and by focal adhesion kinase and protein kinase X at yet-to-be identified residues. Inhibition of tyrosine phosphorylation or increased cellular calcium increases polycystin-1 focal adhesion complexes versus polycystin-1 adherens junction complexes, whereas disruption of the actin cytoskeleton dissociates all polycystin-1 complexes. Genetic evidence suggests that PKD1, PKD2, NPHP1, and tensin are in the same pathway.

    Journal of the American Society of Nephrology : JASN 2001;12;4;834-45

  • Presenilin-1 binds cytoplasmic epithelial cadherin, inhibits cadherin/p120 association, and regulates stability and function of the cadherin/catenin adhesion complex.

    Baki L, Marambaud P, Efthimiopoulos S, Georgakopoulos A, Wen P, Cui W, Shioi J, Koo E, Ozawa M, Friedrich VL and Robakis NK

    Department of Psychiatry, Fishberg Research Center for Neurobiology, Mount Sinai School of Medicine, One Gustave Levy Place, New York, NY 10029, USA.

    Here we show that presenilin-1 (PS1), a protein involved in Alzheimer's disease, binds directly to epithelial cadherin (E-cadherin). This binding is mediated by the large cytoplasmic loop of PS1 and requires the membrane-proximal cytoplasmic sequence 604-615 of mature E-cadherin. This sequence is also required for E-cadherin binding of protein p120, a known regulator of cadherin-mediated cell adhesion. Using wild-type and PS1 knockout cells, we found that increasing PS1 levels suppresses p120/E-cadherin binding, and increasing p120 levels suppresses PS1/E-cadherin binding. Thus PS1 and p120 bind to and mutually compete for cellular E-cadherin. Furthermore, PS1 stimulates E-cadherin binding to beta- and gamma-catenin, promotes cytoskeletal association of the cadherin/catenin complexes, and increases Ca(2+)-dependent cell-cell aggregation. Remarkably, PS1 familial Alzheimer disease mutant DeltaE9 increased neither the levels of cadherin/catenin complexes nor cell aggregation, suggesting that this familial Alzheimer disease mutation interferes with cadherin-based cell-cell adhesion. These data identify PS1 as an E-cadherin-binding protein and a regulator of E-cadherin function in vivo.

    Funded by: NIA NIH HHS: AG-05138, AG-08200, AG-17926, P50 AG005138, R01 AG008200, R01 AG017926, R37 AG017926

    Proceedings of the National Academy of Sciences of the United States of America 2001;98;5;2381-6

  • alpha -Catenin binds directly to spectrin and facilitates spectrin-membrane assembly in vivo.

    Pradhan D, Lombardo CR, Roe S, Rimm DL and Morrow JS

    Department of Pathology, Yale University, New Haven, Connecticut 06510, USA.

    The anchorage of spectrin to biological membranes is mediated by protein and phosphoinositol phospholipid interactions. In epithelial cells, a nascent spectrin skeleton assembles in regions of cadherin-mediated cell-cell contact, and conversely, cytoskeletal assembly is required to complete the cell-adhesion process. The molecular interactions guiding these processes remain incompletely understood. We have examined the interaction of spectrin with alpha-catenin, a component of the adhesion complex. Spectrin (alphaIIbetaII) and alpha-catenin coprecipitate from extracts of confluent Madin-Darby canine kidney, HT29, and Clone A cells and from solutions of purified spectrin and alpha-catenin in vitro. By surface plasmon resonance and in vitro binding assays, we find that alpha-catenin binds alphaIIbetaII spectrin with an apparent K(d) of approximately 20-100 nm. By gel-overlay assay, alpha-catenin binds recombinant betaII-spectrin peptides that include the first 313 residues of spectrin but not to peptides that lack this region. Similarly, the binding activity of alpha-catenin is fully accounted for in recombinant peptides encompassing the NH(2)-terminal 228 amino acid region of alpha-catenin. An in vivo role for the interaction of spectrin with alpha-catenin is suggested by the impaired membrane assembly of spectrin and its enhanced detergent solubility in Clone A cells that harbor a defective alpha-catenin. Transfection of these cells with wild-type alpha-catenin reestablishes alpha-catenin at the plasma membrane and coincidentally recruits spectrin to the membrane. We propose that ankyrin-independent interactions of modest affinity between alpha-catenin and the amino-terminal domain of beta-spectrin augment the interaction between alpha-catenin and actin, and together they provide a polyvalent linkage directing the topographic assembly of a nascent spectrin-actin skeleton to membrane regions enriched in E-cadherin.

    The Journal of biological chemistry 2001;276;6;4175-81

  • Characterization of syntenin, a syndecan-binding PDZ protein, as a component of cell adhesion sites and microfilaments.

    Zimmermann P, Tomatis D, Rosas M, Grootjans J, Leenaerts I, Degeest G, Reekmans G, Coomans C and David G

    Laboratory for Glycobiology and Developmental Genetics, Center for Human Genetics, University of Leuven, Leuven, B-3000 Belgium.

    Syntenin is a PDZ protein that binds the cytoplasmic C-terminal FYA motif of the syndecans. Syntenin is widely expressed. In cell fractionation experiments, syntenin partitions between the cytosol and microsomes. Immunofluorescence microscopy localizes endogenous and epitope-tagged syntenin to cell adhesion sites, microfilaments, and the nucleus. Syntenin is composed of at least three domains. Both PDZ domains of syntenin are necessary to target reporter tags to the plasma membrane. The addition of a segment of 10 amino acids from the N-terminal domain of syntenin to these PDZ domains increases the localization of the tags to stress fibers and induces the formation of long, branching plasma membrane extensions. The addition of the complete N-terminal region, in contrast, reduces the localization of the tags to plasma membrane/adhesion sites and stress fibers, and reduces the morphotypical effects. Recombinant domains of syntenin with the highest plasma membrane localization display the lowest nuclear localization. Syndecan-1, E-cadherin, beta-catenin, and alpha-catenin colocalize with syntenin at cell-cell contacts in epithelial cells, and coimmunoprecipitate with syntenin from extracts of these cells. These results suggest a role for syntenin in the composition of adherens junctions and the regulation of plasma membrane dynamics, and imply a potential role for syntenin in nuclear processes.

    Molecular biology of the cell 2001;12;2;339-50

  • Vezatin, a novel transmembrane protein, bridges myosin VIIA to the cadherin-catenins complex.

    Küssel-Andermann P, El-Amraoui A, Safieddine S, Nouaille S, Perfettini I, Lecuit M, Cossart P, Wolfrum U and Petit C

    Unité de Génétique des Déficits Sensoriels, CNRS URA 1968 and Unité des Interactions Bactéries-Cellules, Institut Pasteur, 25-28 rue du Dr Roux, 75724 Paris cedex 15, France.

    Defects in myosin VIIA are responsible for deafness in the human and mouse. The role of this unconventional myosin in the sensory hair cells of the inner ear is not yet understood. Here we show that the C-terminal FERM domain of myosin VIIA binds to a novel transmembrane protein, vezatin, which we identified by a yeast two-hybrid screen. Vezatin is a ubiquitous protein of adherens cell-cell junctions, where it interacts with both myosin VIIA and the cadherin-catenins complex. Its recruitment to adherens junctions implicates the C-terminal region of alpha-catenin. Taken together, these data suggest that myosin VIIA, anchored by vezatin to the cadherin-catenins complex, creates a tension force between adherens junctions and the actin cytoskeleton that is expected to strengthen cell-cell adhesion. In the inner ear sensory hair cells vezatin is, in addition, concentrated at another membrane-membrane interaction site, namely at the fibrillar links interconnecting the bases of adjacent stereocilia. In myosin VIIA-defective mutants, inactivity of the vezatin-myosin VIIA complex at both sites could account for splaying out of the hair cell stereocilia.

    The EMBO journal 2000;19;22;6020-9

  • Amino-terminal domain of classic cadherins determines the specificity of the adhesive interactions.

    Klingelhöfer J, Troyanovsky RB, Laur OY and Troyanovsky S

    Division of Dermatology, Washington University Medical School, St Louis, MO 63110, USA.

    Classic cadherins are transmembrane receptors involved in cell type-specific calcium-dependent intercellular adhesion. The specificity of adhesion is mediated by homophilic interactions between cadherins extending from opposing cell surfaces. In addition, classic cadherins can self-associate forming lateral dimers. Whereas it is widely excepted that lateral dimerization of cadherins is critical for adhesion, details of this process are not known. Yet, no evidence for physical association between different classic cadherins in cells expressing complex cadherin patterns has been reported. To study lateral and adhesive intercadherin interactions, we examined interactions between two classic cadherins, E- and P-cadherins, in epithelial A-431 cells co-producing both proteins. We showed that these cells exhibited heterocomplexes consisting of laterally assembled E- and P-cadherins. These complexes were formed by a mechanism involving Trp(156) of E-cadherin. Removal of calcium ions from the culture medium triggered a novel Trp(156)-independent type of lateral E-cadherin-P-cadherin association. Notably, an antiparallel (adhesive) mode of interaction between these cadherins was negligible. The specificity of adhesive interaction was localized to the amino-terminal (EC1) domain of both cadherins. Thus, EC1 domain of classic cadherins exposes two determinants responsible for nonspecific lateral and cadherin type-specific adhesive dimerization.

    Journal of cell science 2000;113 ( Pt 16);2829-36

  • Presenilin-1 forms complexes with the cadherin/catenin cell-cell adhesion system and is recruited to intercellular and synaptic contacts.

    Georgakopoulos A, Marambaud P, Efthimiopoulos S, Shioi J, Cui W, Li HC, Schütte M, Gordon R, Holstein GR, Martinelli G, Mehta P, Friedrich VL and Robakis NK

    Department of Psychiatry, Mount Sinai School of Medicine, New York, New York 10029, USA.

    In MDCK cells, presenilin-1 (PS1) accumulates at intercellular contacts where it colocalizes with components of the cadherin-based adherens junctions. PS1 fragments form complexes with E-cadherin, beta-catenin, and alpha-catenin, all components of adherens junctions. In confluent MDCK cells, PS1 forms complexes with cell surface E-cadherin; disruption of Ca(2+)-dependent cell-cell contacts reduces surface PS1 and the levels of PS1-E-cadherin complexes. PS1 overexpression in human kidney cells enhances cell-cell adhesion. Together, these data show that PS1 incorporates into the cadherin/catenin adhesion system and regulates cell-cell adhesion. PS1 concentrates at intercellular contacts in epithelial tissue; in brain, it forms complexes with both E- and N-cadherin and concentrates at synaptic adhesions. That PS1 is a constituent of the cadherin/catenin complex makes that complex a potential target for PS1 FAD mutations.

    Funded by: NIA NIH HHS: AG-05138, AG-08200

    Molecular cell 1999;4;6;893-902

  • Antigens recognized by autologous antibody in patients with renal-cell carcinoma.

    Scanlan MJ, Gordan JD, Williamson B, Stockert E, Bander NH, Jongeneel V, Gure AO, Jäger D, Jäger E, Knuth A, Chen YT and Old LJ

    Ludwig Institute for Cancer Research, New York Branch at Memorial Sloan-Kettering Cancer Center, New York, New York 10021, USA. scanlanm@mskcc.org

    The screening of cDNA expression libraries derived from human tumors with autologous antibody (SEREX) is a powerful method for defining the structure of tumor antigens recognized by the humoral immune system. Sixty-five distinct antigens (NY-REN-1 to NY-REN-65) reactive with autologous IgG were identified by SEREX analysis of 4 renal cancer patients and were characterized in terms of cDNA sequence, mRNA expression pattern, and reactivity with allogeneic sera. REN-9, -10, -19, and -26 have a known association with human cancer. REN-9 (LUCA-15) and REN-10 (gene 21) map to the small cell lung cancer tumor suppressor gene locus on chromosome 3p21.3. REN-19 is equivalent to LKB1/STK11, a gene that is defective in Peutz-Jeghers syndrome and cancer. REN-26 is encoded by the bcr gene involved in the [t(9:22)] bcr/abl translocation. Genes encoding 3 of the antigens in the series showed differential mRNA expression; REN-3 displays a pattern of tissue-specific isoforms, and REN-21 and REN-43 are expressed at a high level in testis in comparison to 15 other normal tissues. The other 62 antigens were broadly expressed in normal tissues. With regard to immunogenicity, 20 of the 65 antigens reacted only with autologous sera. Thirty-three antigens reacted with sera from normal donors, indicating that their immunogenicity is not restricted to cancer. The remaining 12 antigens reacted with sera from 5-25% of the cancer patients but not with sera from normal donors. Seventy percent of the renal cancer patients had antibodies directed against one or more of these 12 antigens. Our results demonstrate the potential of the SEREX approach for the analysis of the humoral immune response against human cancer.

    International journal of cancer 1999;83;4;456-64

  • The cytoplasmic domain of HIV-1 gp41 interacts with the carboxyl-terminal region of alpha-catenin.

    Kim EM, Lee KH and Kim JW

    Division of Life Science, College of National Sciences, Pai Chai University, Taejon, Korea.

    To know the cellular protein interactions with the viral protein can give an insight into the molecular mechanisms of the virus life cycle. As the function of the cytoplasmic domain of human immunodeficiency virus type 1 (HIV-1) gp41 is not known clearly, we searched for a cellular protein that interacts with the cytoplasmic domain of the HIV-1 gp41 using the yeast two-hybrid assay system. Screening of HeLa cell cDNA library yielded alpha-catenin cDNA. The cytoplasmic domain of the HIV-1 gp41 and the simian immunodeficiency virus (SIV) gp41 were able to interact with the carboxyl-terminal region of alpha-catenin specifically. Mapping of the interaction sites revealed that the interaction between the domain containing the second helix structure from the carboxyl-terminus of HIV-1 gp41 and the carboxyl-terminal region of alpha-catenin was stronger than other domains of gp41.

    Molecules and cells 1999;9;3;281-5

  • Cadherin and catenin expression in normal human bronchial epithelium and non-small cell lung cancer.

    Smythe WR, Williams JP, Wheelock MJ, Johnson KR, Kaiser LR and Albelda SM

    M.D. Anderson Cancer Center, Department of Thoracic and Cardiovascular Surgery, The University of Texas, Houston 77030, USA. rsmythenotes.mdacc.tmc.edu.

    Cadherins are transmembrane cell adhesion molecules (CAMS) that mediate cell-cell interactions and are important for maintenance of epithelial cell integrity. This function is dependent on an indirect interaction between the cytoplasmic domain of the cadherin molecule with three cytoplasmic proteins known as alpha-, beta-, and gamma-catenin (-cat). Growing evidence suggests that alterations in cadherin or catenin expression or function may be important to the development of an invasive or metastatic phenotype. Immunohistochemical techniques were used to study the expression of the two major epithelial cadherins, E-cadherin (E-cad) and P-cadherin (P-cad) as well as alpha- and gamma-cat in normal bronchial epithelium and in a series of carefully TMN-staged pulmonary adenocarcinomas (n = 21) and squamous cell carcinomas (n = 7). The cadherin profile of normal pseudostratified bronchial epithelium was heterogeneous. Basilar cells strongly expressed P-cad, alpha- and gamma-cat, while columnar cells moderately expressed E-cad, alpha- and gamma-cat. In contrast to other epithelial tumors, E-cad on non-small cell lung carcinomas was actually upregulated, however, a decrease in P-cad expression was noted in 68%. At least one cadherin or catenin was downregulated, compared to normal bronchial epithelium, in 82% of tumors examined. With the exception of an association between loss of P-cad expression and poorly differentiated state, changes in cadherin and catenin expression levels were not significantly correlated to tumor stage, cell type, or nodal status. These findings illustrate that alteration of expression of cadherins and catenins are often found in non-small cell lung carcinoma when compared to the progenitor bronchial epithelium, and may play a role in the development of the malignant phenotype.

    Lung cancer (Amsterdam, Netherlands) 1999;24;3;157-68

  • The fate of E- and P-cadherin during the early stages of apoptosis.

    Schmeiser K and Grand RJ

    CRC Institute for Cancer Studies, University of Birmingham, Edgbaston, Birmingham B15 2TA, UK.

    Caspases are responsible for the proteolysis of many cytoskeletal proteins in apoptotic cells. It has been demonstrated here that during cisplatin-induced apoptosis of human embryo retinoblasts both E- and P-cadherin were degraded by caspases, giving initially major polypeptide products of apparent molecular weights 48 K and 104 K respectively. This proteolysis occurred over a similar time-scale to the observed degradation of PARP and to the onset of DNA fragmentation but appreciably later than p53 induction and cleavage of Mdm2 and p21. Addition of caspase inhibitors such as Z-VAD-FMK inhibited apoptosis and cadherin degradation. Co-immunoprecipitation studies carried out on viable cells confirmed previously observed complexes between cadherins and alpha and beta catenin and between the catenins themselves. These interactions were sustained in apoptotic cells as long as the protein components remained intact. Using confocal microscopy it has been shown that cytoskeletal changes associated with apoptosis precede degradation of catenins and cadherins by several hours. In particular, after addition of cisplatin relatively rapid (within 3 h) re-localization of adherens junction proteins from the cell periphery to the cytoplasm was observed whereas little cadherin or catenin degradation occurred until 10 h. We conclude that neither caspase-mediated degradation of cytoskeletal components nor disruption of adherens junction protein-protein interactions is required for morphological change.

    Cell death and differentiation 1999;6;4;377-86

  • Characterization of ZO-2 as a MAGUK family member associated with tight as well as adherens junctions with a binding affinity to occludin and alpha catenin.

    Itoh M, Morita K and Tsukita S

    Department of Cell Biology, Faculty of Medicine, Kyoto University, Sakyo-ku, Kyoto 606, Japan.

    ZO-2, a member of the MAGUK family, was thought to be specific for tight junctions (TJs) in contrast to ZO-1, another MAGUK family member, which is localized at TJs and adherens junctions (AJs) in epithelial and nonepithelial cells, respectively. Mouse ZO-2 cDNA was isolated, and a specific polyclonal antibody was generated using corresponding synthetic peptides as antigens. Immunofluorescence microscopy with this polyclonal antibody revealed that, similarly to ZO-1, in addition to TJs in epithelial cells, ZO-2 was also concentrated at AJs in nonepithelial cells such as fibroblasts and cardiac muscle cells lacking TJs. When NH2-terminal dlg-like and COOH-terminal non-dlg-like domains of ZO-2 (N-ZO-2 and C-ZO-2, respectively) were separately introduced into cultured cells, N-ZO-2 was colocalized with endogenous ZO-1/ZO-2, i.e. at TJs in epithelial cells and at AJs in non-epithelial cells, whereas C-ZO-2 was distributed along actin filaments. Consistently, occludin as well as alpha catenin directly bound to N-ZO-2 as well as the NH2-terminal dlg-like portion of ZO-1 (N-ZO-1) in vitro. Furthermore, immunoprecipitation experiments revealed that the second PDZ domain of ZO-2 was directly associated with N-ZO-1. These findings indicated that ZO-2 forms a complex with ZO-1/occludin or ZO-1/alpha catenin to establish TJ or AJ domains, respectively.

    The Journal of biological chemistry 1999;274;9;5981-6

  • alpha-Catenin-vinculin interaction functions to organize the apical junctional complex in epithelial cells.

    Watabe-Uchida M, Uchida N, Imamura Y, Nagafuchi A, Fujimoto K, Uemura T, Vermeulen S, van Roy F, Adamson ED and Takeichi M

    Department of Biophysics, Faculty of Science, Kyoto University, Kitashirakawa, Sakyo-ku, Kyoto 606-8502, USA.

    alphaE-catenin, a cadherin-associated protein, is required for tight junction (TJ) organization, but its role is poorly understood. We transfected an alphaE-catenin-deficient colon carcinoma line with a series of alphaE-catenin mutant constructs. The results showed that the amino acid 326-509 domain of this catenin was required to organize TJs, and its COOH-terminal domain was not essential for this process. The 326-509 internal domain was found to bind vinculin. When an NH2-terminal alphaE-catenin fragment, which is by itself unable to organize the TJ, was fused with the vinculin tail, this chimeric molecule could induce TJ assembly in the alphaE-catenin-deficient cells. In vinculin-null F9 cells, their apical junctional organization was impaired, and this phenotype was rescued by reexpression of vinculin. These results indicate that the alphaE-catenin-vinculin interaction plays a role in the assembly of the apical junctional complex in epithelia.

    The Journal of cell biology 1998;142;3;847-57

  • A mutation in alpha-catenin disrupts adhesion in clone A cells without perturbing its actin and beta-catenin binding activity.

    Roe S, Koslov ER and Rimm DL

    Department of Pathology, Yale University School of Medicine, New Haven, Connecticut 06510, USA.

    Cadherin mediated cell-cell adhesion requires cytoplasmic connections to the cytoskeleton mediated by alpha-catenin. Original descriptions of the catenins, as well as our own in vitro studies, have suggested that this connection was mediated by the interaction of alpha-catenin to actin. Loss of adhesion in the human colon carcinoma cell line "Clone A" is the result of an internal deletion mutation of 158 residues near the N-terminus of the protein resulting in an 80 kD mutated protein. Transfection of these cells with the full length protein restores the normal adhesive phenotype. We have characterized this mutant protein in efforts to understand the normal function of alpha-catenin and, in particular, the region deleted in the Clone A mutant. Co-precipitation experiments using whole cell lysates indicate that the mutant form of alpha-catenin binds beta-catenin and plakoglobin, and can form a structural complex with E-cadherin via these interactions. Actin co-sedimentation assays show that the recombinant mutant binds and bundles F-actin and binds both actin and beta-catenin simultaneously, as seen with wild type alpha-catenin. These results suggest that the stabilization of the E-cadherin-catenin complex may be mediated by factors beyond its direct interaction with actin. We conclude that a region near the N-terminus of alpha-catenin mediates additional interactions between the adhesive complex and the cytoskeleton that are critical for functional adhesion.

    Cell adhesion and communication 1998;5;4;283-96

  • E-cadherin mediated cell adhesion recruits SAP97 into the cortical cytoskeleton.

    Reuver SM and Garner CC

    Department of Neurobiology, University of Alabama at Birmingham, Birmingham, AL 35294-0021, USA.

    Members of the SAP family of synapse-associated proteins have recently emerged as central players in the molecular organization of synapses. In this study, we have examined the mechanism that localizes one member, SAP97, to sites of cell-cell contact. Utilizing epithelial CACO-2 cells and fibroblast L-cells as model systems, we demonstrate that SAP97 is associated with the submembranous cortical cytoskeleton at cell-cell adhesion sites. Furthermore, we show that its localization into this structure is triggered by E-cadherin. Although SAP97 can be found in an E-cadherin/catenin adhesion complex, this interaction seems to be mediated by the attachment of SAP97 to the cortical cytoskeleton. Our results are consistent with a model in which SAP97 is recruited to sites of cell-cell contact via an E-cadherin induced assembly of the cortical cytoskeleton.

    Funded by: NIA NIH HHS: AG 12978-02; NICHD NIH HHS: P50 HD32901

    Journal of cell science 1998;111 ( Pt 8);1071-80

  • Alteration of interendothelial adherens junctions following tumor cell-endothelial cell interaction in vitro.

    Lewalle JM, Bajou K, Desreux J, Mareel M, Dejana E, Noël A and Foidart JM

    Laboratory of Cellular Biology, University of Liége, CHU, Sart-Tilman, Belgium. JM.Lewalle@ulg.ac.be

    The integrity of the vascular endothelium is mainly dependent upon the organization of interendothelial adherens junctions (AJ). These junctions are formed by the homotypic interaction of a transmembrane protein, vascular endothelial cadherin (VE-cadherin), which is complexed to an intracellular protein network including alpha-, beta-, and gamma-catenin. Additional proteins such as vinculin and alpha-actinin have been suggested to link the VE-cadherin/catenin complex to the actin-based cytoskeleton. During the process of hematogenous metastasis, circulating tumor cells must disrupt these intercellular junctions in order to extravasate. In the present study, we have investigated the influence of tumor cell-endothelial cell interaction upon interendothelial AJ. We show that human breast adenocarcinoma cells (MCF-7), but not normal human mammary epithelial cells, induce a rapid endothelial cell (EC) dissociation which correlates with the loss of VE-cadherin expression at the site of tumor cell-EC contact and with profound changes in vinculin distribution and organization. This process could not be inhibited by metalloproteinase nor serine protease inhibitors. Immunoprecipitations and Western blot analysis demonstrate that the overall expression of VE-cadherin and vinculin as well as the composition of the VE-cadherin/catenins complex are not affected by tumor cells while the tyrosine phosphorylation status of proteins within the complex is significantly altered. Our data suggest that tumor cells modulate AJ protein distribution and phosphorylation in EC and may, thereby, facilitate EC dissociation.

    Experimental cell research 1997;237;2;347-56

  • Molecular cloning of an alternative human alphaE-catenin cDNA.

    Linkels M, Bussemakers MJ, Nollet F, Ewing CM, van Roy F and Schalken JA

    Urology Research Laboratory, University Hospital Nijmegen, The Netherlands.

    The cytoplasmic protein alpha-catenin plays a crucial role in E-cadherin mediated cell-cell adhesion by binding E-cadherin to the cytoskeleton via beta- or gamma-catenin and actin. Functional loss of one of these interacting components leads to decreased cell-cell adhesion, and therefore to loss of epithelial integrity. Northern analysis revealed two distinct alphaE-catenin transcripts in different cell lines, whereas apparently only one protein is expressed. Because of the biological importance of this protein we sought to molecularly characterize the differences between the two observed transcripts. cDNA cloning and sequence analysis revealed the earlier described 3.4 kb alphaE-catenin transcript and an alphaE-catenin transcript of approximately 3.8 kb. This larger transcript contains a 321 bp extension in the 3'UTR sequence, which probably arises as a result of alternative polyadenylation. Considering the presence of AU-rich sequences in the extension, it may be involved in mRNA stability.

    Biochemical and biophysical research communications 1997;237;1;177-81

  • Involvement of ZO-1 in cadherin-based cell adhesion through its direct binding to alpha catenin and actin filaments.

    Itoh M, Nagafuchi A, Moroi S and Tsukita S

    Department of Cell Biology, Faculty of Medicine, Kyoto University, Kyoto 606, Japan.

    ZO-1, a 220-kD peripheral membrane protein consisting of an amino-terminal half discs large (dlg)-like domain and a carboxyl-terminal half domain, is concentrated at the cadherin-based cell adhesion sites in non-epithelial cells. We introduced cDNAs encoding the full-length ZO-1, its amino-terminal half (N-ZO-1), and carboxyl-terminal half (C-ZO-1) into mouse L fibroblasts expressing exogenous E-cadherin (EL cells). The full-length ZO-1 as well as N-ZO-1 were concentrated at cadherin-based cell-cell adhesion sites. In good agreement with these observations, N-ZO-1 was specifically coimmunoprecipitated from EL transfectants expressing N-ZO-1 (NZ-EL cells) with the E-cadherin/alpha, beta catenin complex. In contrast, C-ZO-1 was localized along actin stress fibers. To examine the molecular basis of the behavior of these truncated ZO-1 molecules, N-ZO-1 and C-ZO-1 were produced in insect Sf9 cells by recombinant baculovirus infection, and their direct binding ability to the cadherin/catenin complex and the actin-based cytoskeleton, respectively, were examined in vitro. Recombinant N-ZO-1 bound directly to the glutathione-S-transferase fusion protein with alpha catenin, but not to that with beta catenin or the cytoplasmic domain of E-cadherin. The dissociation constant between N-ZO-1 and alpha catenin was approximately 0.5 nM. On the other hand, recombinant C-ZO-1 was specifically cosedimented with actin filaments in vitro with a dissociation constant of approximately 10 nM. Finally, we compared the cadherin-based cell adhesion activity of NZ-EL cells with that of parent EL cells. Cell aggregation assay revealed no significant differences among these cells, but the cadherin-dependent intercellular motility, i.e., the cell movement in a confluent monolayer, was significantly suppressed in NZ-EL cells. We conclude that in nonepithelial cells, ZO-1 works as a cross-linker between cadherin/catenin complex and the actin-based cytoskeleton through direct interaction with alpha catenin and actin filaments at its amino- and carboxyl-terminal halves, respectively, and that ZO-1 is a functional component in the cadherin-based cell adhesion system.

    The Journal of cell biology 1997;138;1;181-92

  • Induction of tyrosine phosphorylation and association of beta-catenin with EGF receptor upon tryptic digestion of quiescent cells at confluence.

    Takahashi K, Suzuki K and Tsukatani Y

    Department of Biochemistry, Kanagawa Cancer Center Research Institute, Yokohama, Japan.

    Normal human breast epithelial (HBE) cells which reached confluence ceased growth and tightly adhered to each other, forming a monolayer. In quiescent cells thus arrested by density, E-cadherin colocalized and coimmunoprecipitated with alpha- and beta-catenins in the boundary region between adjacent cells. By contrast, immunocytostaining and Western blot analyses revealed that E-cadherin colocalized and coprecipitated with beta-catenin but not with alpha-catenin in exponentially growing cells at low density. As a comparable amount of alpha-catenin was detected in the total cell lysate of cells at different densities, it is suggested that alpha-catenin is present but dissociates from the E-cadherin-beta-catenin complex in growing cells. beta-Catenin was tyrosine phosphorylated in growing cells at low density but not in quiescent cells at confluence. Tyrosine phosphorylation of beta-catenin was concomitantly induced with association of beta-catenin with EGF receptor (EGFR) when quiescent cells at confluence were dissociated into single cells by tryptic digestion, being accompanied by dissociation of alpha-catenin from E-cadherin. Both tyrosine phosphorylation and association of beta-catenin with EGFR were inhibited by tyrphostin, a specific inhibitor of the EGFR tyrosine kinase, whereas dissociation of alpha-catenin from E-cadherin was not. The results suggest that tyrosine phosphorylation of beta-catenin is achieved by EGFR upon tryptic digestion of cells and concurrent with but independent of dissociation of alpha-catenin from E-cadherin. beta-Catenin thus phosphorylated at tyrosine is suggested to play the role in preventing alpha-catenin once dissociated from reassociating with E-cadherin until cells reach confluence.

    Oncogene 1997;15;1;71-8

  • M-cadherin-mediated cell adhesion and complex formation with the catenins in myogenic mouse cells.

    Kuch C, Winnekendonk D, Butz S, Unvericht U, Kemler R and Starzinski-Powitz A

    Institut der Anthropologie und Humangenetik für Biologen, Johann Wolfgang Goethe Universität, Frankfurt am Main, Germany.

    M-cadherin is a member of the multigene family of calcium-dependent intercellular adhesion molecules, the cadherins, which are involved in morphogenetic processes. Amino acid comparisons between M-cadherin and E-, N-, and P-cadherin suggested that M-cadherin diverged phylogenetically very early from these classical cadherins. It has been shown that M-cadherin is expressed in prenatal and adult skeletal muscle. In the cerebellum, M-cadherin is present in an adherens-type junction which differs in its molecular composition from the E-cadherin-mediated adherens-type junctions. These and other findings raised the question of whether M-cadherin and the classical cadherins share basic biochemical properties, notably the calcium-dependent resistance to proteolysis, mediation of calcium-dependent intercellular adhesion, and the capability to form M-cadherin complexes with the catenins. Here we show that M-cadherin is resistant to trypsin digestion in the presence of calcium ions but at lower trypsin concentrations than E-cadherin. When ectopically expressed in LMTK- cells, M-cadherin mediated calcium-dependent cell aggregation. Finally, M-cadherin was capable of forming two distinct cytoplasmic complexes in myogenic cells, either with alpha-catenin/beta-catenin or with alpha-catenin/plakoglobin, as E-and N-cadherin, for example, have previously been shown to form. The relative amount of these complexes changed during differentiation from C2C12 myoblasts to myotubes, although the molecular composition of each complex was unaffected during differentiation. These results demonstrate that M-cadherin shares important features with the classical cadherins despite its phylogenetic divergence.

    Experimental cell research 1997;232;2;331-8

  • Identification of the domain of alpha-catenin involved in its association with beta-catenin and plakoglobin (gamma-catenin).

    Obama H and Ozawa M

    Department of Biochemistry, Faculty of Medicine, Kagoshima University, 8-35-1 Sakuragaoka, Kagoshima 890, Japan.

    alpha-Catenin is a 102-kDa protein exhibiting homology to vincuin, and it forms complexes with cadherins or the tumor-suppressor gene product adenomatous polyposis coli through binding to beta-catenin or plakoglobin (gamma-catenin). The incorporation of alpha-catenin into the cadherin-catenin complexes is a prerequisite for expression of the cell-adhesive activity of cadherins. Using an in vitro assay system involving bacterially expressed proteins, we localized a region in alpha-catenin required for molecular interaction with beta-catenin and plakoglobin. Analysis of various truncated alpha-catenin molecules revealed that amino-terminal residues 48-163 are able to bind to beta-catenin and plakoglobin. Consistent with the observation that beta-catenin and plakoglobin bind to the same region of alpha-catenin, beta-catenin competed with the binding of plakoglobin to alpha-catenin and vice versa. Under the conditions used, beta-catenin bound to alpha-catenin with higher affinity than did plakoglobin. Scatchard analysis indicated that the affinity of the interaction between alpha-catenin and beta-catenin or that between alpha-catenin and plakoglobin was moderately strong (Kd = 3. 8 x 10(-8) and 7.7 x 10(-8), respectively). When transfected into L cells expressing E-cadherin, the amino-terminal region of alpha-catenin (from residue 1 to 226) formed complexes with beta-catenin supporting the in vitro binding experiment results.

    The Journal of biological chemistry 1997;272;17;11017-20

  • The tyrosine kinase substrate p120cas binds directly to E-cadherin but not to the adenomatous polyposis coli protein or alpha-catenin.

    Daniel JM and Reynolds AB

    Department of Tumor Cell Biology, St. Jude Children's Research Hospital, Memphis, Tennessee 38105, USA.

    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.

    Funded by: NCI NIH HHS: CA55724, P30 CA21756

    Molecular and cellular biology 1995;15;9;4819-24

  • Identification of plakoglobin domains required for association with N-cadherin and alpha-catenin.

    Sacco PA, McGranahan TM, Wheelock MJ and Johnson KR

    Department of Biology, University of Toledo, Ohio 43606, USA.

    Cadherins are calcium-dependent, cell surface glycoproteins involved in cell-cell adhesion. To function in cell-cell adhesion, the transmembrane cadherin molecule must be associated with the cytoskeleton via cytoplasmic proteins known as catenins. Three catenins, alpha-catenin, beta-catenin, and gamma-catenin (also known as plakoglobin), have been identified. The domain of the cadherin molecule important for its interaction with the catenins has been mapped to the COOH-terminal 70 amino acids, but less is known about regions of the catenins that allow them to associate with one another or with the cadherin molecule. In this study we have transfected carboxyl-terminal deletions of plakoglobin into the human fibrosarcoma HT-1080 and used immunofluorescence localization and co-immunoprecipitation to map the regions of plakoglobin that allow it to associate with N-cadherin and with alpha-catenin. Plakoglobin is an armadillo family member containing 13 weakly similar internal repeats. These data show that the alpha-catenin-binding region maps within the first repeat and the N-cadherin-binding region maps within repeats 7 and 8.

    Funded by: NIGMS NIH HHS: GM51188

    The Journal of biological chemistry 1995;270;34;20201-6

  • Interaction of alpha-actinin with the cadherin/catenin cell-cell adhesion complex via alpha-catenin.

    Knudsen KA, Soler AP, Johnson KR and Wheelock MJ

    Lankenau Medical Research Center, Wynnewood, Pennsylvania 19096, USA.

    Cadherins are Ca(2+)-dependent, cell surface glycoproteins involved in cell-cell adhesion. Extracellularly, transmembrane cadherins such as E-, P-, and N-cadherin self-associate, while intracellularly they interact indirectly with the actin-based cytoskeleton. Several intracellular proteins termed catenins, including alpha-catenin, beta-catenin, and plakoglobin, are tightly associated with these cadherins and serve to link them to the cytoskeleton. Here, we present evidence that in fibroblasts alpha-actinin, but not vinculin, colocalizes extensively with the N-cadherin/catenin complex. This is in contrast to epithelial cells where both cytoskeletal proteins colocalize extensively with E-cadherin and catenins. We further show that alpha-actinin, but not vinculin, coimmunoprecipitates specifically with alpha- and beta-catenin from N- and E-cadherin-expressing cells, but only if alpha-catenin is present. Moreover, we show that alpha-actinin coimmunoprecipitates with the N-cadherin/catenin complex in an actin-independent manner. We therefore propose that cadherin/catenin complexes are linked to the actin cytoskeleton via a direct association between alpha-actinin and alpha-catenin.

    Funded by: NIGMS NIH HHS: GM 51188

    The Journal of cell biology 1995;130;1;67-77

  • Tyrosine phosphorylation regulates the adhesions of ras-transformed breast epithelia.

    Kinch MS, Clark GJ, Der CJ and Burridge K

    Department of Cell Biology and Anatomy, University of North Carolina, Chapel Hill 27599, USA.

    Transformed epithelial cells often are characterized by a fibroblastic or mesenchymal morphology. These cells exhibit altered cell-cell and cell-substrate interactions. Here we have identified changes in the adhesions and cytoskeletal interactions of transformed epithelial cells that contribute to their altered morphology. Using MCF-10A human breast epithelial cells as a model system, we have found that transformation by an activated form of ras is characterized by less developed adherens-type junctions between cells but increased focal adhesions. Contributing to the modified adherens junctions of the transformed cells are decreased interactions among beta-catenin, E-cadherin, and the actin cytoskeleton. The ras-transformed cells reveal elevated phosphotyrosine in many proteins, including beta-catenin and p120 Cas. Whereas in the normal cells beta-catenin is found in association with E-cadherin, p120 Cas is not. In the ras-transformed cells, the situation is reversed; tyrosine-phosphorylated p120 Cas, but not tyrosine-phosphorylated beta-catenin, now is detected in E-cadherin complexes. The tyrosine-phosphorylated beta-catenin also shows increased detergent solubility, suggesting a decreased association with the actin cytoskeleton. p120 Cas, whether tyrosine phosphorylated or not, partitions into the detergent soluble fraction, suggesting that it is not tightly bound to the actin cytoskeleton in either the normal or ras-transformed cells. Inhibitors of tyrosine kinases decrease the level of tyrosine phosphorylation and restore a normal epithelial morphology to the ras-transformed cells. In particular, decreased tyrosine phosphorylation of beta-catenin is accompanied by increased interaction with both E-cadherin and the detergent insoluble cytoskeletal fraction. These results suggest that elevated tyrosine phosphorylation of proteins such as beta-catenin and p120 Cas contribute to the altered adherens junctions of ras-transformed epithelia.

    Funded by: NCI NIH HHS: 5 T32 CA09156; NHLBI NIH HHS: HL-45100; NIGMS NIH HHS: GM-29860

    The Journal of cell biology 1995;130;2;461-71

  • The APC protein and E-cadherin form similar but independent complexes with alpha-catenin, beta-catenin, and plakoglobin.

    Rubinfeld B, Souza B, Albert I, Munemitsu S and Polakis P

    Onyx Pharmaceuticals, Richmond, California 94806.

    The tumor suppressor APC protein associates with the cadherin-binding proteins alpha- and beta-catenin. To examine the relationship between cadherin, catenins, and APC, we have tested combinatorial protein-protein interactions in vivo, using a yeast two-hybrid system, and in vitro, using purified proteins. beta-Catenin directly binds to APC at high and low affinity sites. alpha-Catenin cannot directly bind APC but associates with it by binding to beta-catenin. Plakoglobin, also known as gamma-catenin, directly binds to both APC and alpha-catenin and also to the APC-beta-catenin complex, but not directly to beta-catenin. beta-Catenin binds to multiple independent regions of APC, some of which include a previously identified consensus motif and others which contain the centrally located 20 amino acid repeat sequences. The APC binding site on beta-catenin may be discontinuous since neither the carboxyl- nor amino-terminal halves of beta-catenin will independently associate with APC, although the amino-terminal half independently binds alpha-catenin. The catenins bind to APC and E-cadherin in a similar fashion, but APC and E-cadherin do not associate with each other either in the presence or absence of catenins. Thus, APC forms distinct heteromeric complexes containing combinations of alpha-catenin, beta-catenin, and plakoglobin which are independent from the cadherin-catenin complexes.

    The Journal of biological chemistry 1995;270;10;5549-55

  • A truncated beta-catenin disrupts the interaction between E-cadherin and alpha-catenin: a cause of loss of intercellular adhesiveness in human cancer cell lines.

    Oyama T, Kanai Y, Ochiai A, Akimoto S, Oda T, Yanagihara K, Nagafuchi A, Tsukita S, Shibamoto S, Ito F et al.

    Pathology Division, National Cancer Center Research Institute, Tokyo, Japan.

    Cadherin cell adhesion molecules play an essential role in creating tight intercellular association and are considered to work as an invasion suppressor system of cancer cells. They form a molecular complex with catenins, a group of cytoplasmic proteins including alpha- and beta-catenins. While alpha-catenin has been demonstrated to be crucial for cadherin function, the role of beta-catenin is not yet fully understood. In this study, we analyzed the cadherin-catenin system in two human cell lines, HSC-39 and its putative subline HSC-40A, derived from a signet ring cell carcinoma of stomach. These cells grow as loose aggregates or single cells, suggesting that their cadherin system is not functional. In these cell lines, an identical 321-base pair in-frame mRNA deletion of beta-catenin was identified; this led to a 107-amino-acid deletion in the NH2-terminal region of the protein. Southern blot analysis disclosed a homozygous deletion in part of the beta-catenin gene. On the other hand, these cells expressed E-cadherin, alpha-catenin, and plakoglobin of normal size. Immunoprecipitation analyses showed that E-cadherin was coprecipitated with the mutated beta-catenin but not with alpha-catenin, and antibodies against beta-catenin did not copurify alpha-catenin. However, the recombinant fusion protein containing wild-type beta-catenin precipitated alpha-catenin from these cells. These results suggest that the dysfunction of E-cadherin in these cell lines is due primarily to its failure to interact with alpha-catenin, and that this defect results from the mutation in beta-catenin. Thus, it is most likely that the association between E-cadherin and alpha-catenin is mediated by beta-catenin, and that this process is blocked by NH2-terminal deletion in beta-catenin. These findings indicate that genetic abnormality of beta-catenin is one of the mechanisms responsible for loosening of cell-cell contact, and may be involved in enhancement of tumor invasion in human cancers.

    Cancer research 1994;54;23;6282-7

  • Assembly of the cadherin-catenin complex in vitro with recombinant proteins.

    Aberle H, Butz S, Stappert J, Weissig H, Kemler R and Hoschuetzky H

    Max-Planck-Institut für Immunbiologie, Freiburg, Germany.

    The cytoplasmic domain of classical cadherins is tightly associated with three proteins termed alpha-, beta- and gamma-catenin. These accessory proteins are of central importance for the adhesive properties of this class of cell adhesion molecules. In order to examine the molecular architecture of the cadherin-catenin complex in more detail we have expressed the catenins and the cytoplasmic domain of E-cadherin as fusion proteins in Escherichia coli, and analyzed the interaction of purified recombinant cadherin and catenins in combinatorial protein-protein interaction experiments. The cytoplasmic domain of E-cadherin cannot directly associate with alpha-catenin but interacts with high affinity with beta-catenin, whereas the binding of gamma-catenin (plakoglobin) to E-cadherin is less efficient. alpha- and beta-catenin assemble into a 1:1 heterodimeric complex. The analysis of various truncated beta-catenins revealed that an alpha-catenin binding site in beta-catenin is localized between amino acid positions 120 and 151. The central role of beta-catenin for the assembly of the heterotrimeric E-cadherin/alpha-catenin/beta-catenin complex in mixing experiments with all components was demonstrated. The reconstitution in vitro of the cadherin-catenin complex should allow the study of the interaction with signalling molecules and with the actin-based cytoskeleton.

    Journal of cell science 1994;107 ( Pt 12);3655-63

  • E-cadherin and APC compete for the interaction with beta-catenin and the cytoskeleton.

    Hülsken J, Birchmeier W and Behrens J

    Max-Delbrück-Center for Molecular Medicine, Berlin, Germany.

    beta-Catenin is involved in the formation of adherens junctions of mammalian epithelia. It interacts with the cell adhesion molecule E-cadherin and also with the tumor suppressor gene product APC, and the Drosophila homologue of beta-catenin, armadillo, mediates morphogenetic signals. We demonstrate here that E-cadherin and APC directly compete for binding to the internal, armadillo-like repeats of beta-catenin; the NH2-terminal domain of beta-catenin mediates the interaction of the alternative E-cadherin and APC complexes to the cytoskeleton by binding to alpha-catenin. Plakoglobin (gamma-catenin), which is structurally related to beta-catenin, mediates identical interactions. We thus show that the APC tumor suppressor gene product forms strikingly similar associations as found in cell junctions and suggest that beta-catenin and plakoglobin are central regulators of cell adhesion, cytoskeletal interaction, and tumor suppression.

    The Journal of cell biology 1994;127;6 Pt 2;2061-9

  • Molecular cloning reveals alternative splice forms of human alpha(E)-catenin.

    Rimm DL, Kebriaei P and Morrow JS

    Department of Pathology, Yale University School of Medicine, New Haven, CT 06510.

    At least three proteins (alpha, beta, and gamma catenin) comprise the cytoplasmic domain of the cadherin cell-cell adhesion complex. We have cloned and sequenced human epithelial alpha(E)-catenin and have identified two distinct transcripts, designated alpha 1- and alpha 2-. The human alpha 1(E)-catenin transcript predicts a 907 aa sequence 97% identical to mouse alpha-catenin. The second transcript, alpha 2(E)-catenin, displays a 24 amino acid insertion after codon 812, yielding a 931 amino acid protein (GenBank #L23805). Analysis by RT-PCR and Northern blotting detects one or both transcripts in epithelial and non-epithelial tissues. Southern blotting indicates that both arise from a single gene. The alternative transcription site in alpha-catenin is analogous to the splice site in vinculin that creates met alpha-vinculin, extending the homology between alpha-catenin and vinculin. These data with the reported structure of other catenin genes suggest that vinculin and alpha-catenin generate a superfamily of proteins mediating membrane-cytoskeletal associations.

    Biochemical and biophysical research communications 1994;203;3;1691-9

  • Assignment of the human alpha-catenin gene (CTNNA1) to chromosome 5q21-q22.

    McPherson JD, Morton RA, Ewing CM, Wasmuth JJ, Overhauser J, Nagafuchi A, Tsukita S and Isaacs WB

    Department of Biological Chemistry, College of Medicine, University of California, Irvine.

    Funded by: NCI NIH HHS: CA55231, CA58236; NHGRI NIH HHS: HG00834

    Genomics 1994;19;1;188-90

  • Structure, expression and chromosome assignment of the human catenin (cadherin-associated protein) alpha 1 gene (CTNNA1).

    Furukawa Y, Nakatsuru S, Nagafuchi A, Tsukita S, Muto T, Nakamura Y and Horii A

    Department of Biochemistry, Cancer Institute, Tokyo, Japan.

    We have isolated the human alpha-catenin gene (CTNNA1), which encodes a cadherin-associated protein, and have determined its primary structure and chromosomal localization. The transcript of CTNNA1 is 3.4 kb long and consists of 16 coding exons encoding 906 amino acids and at least one 5' noncoding exon. The 102-kDa predicted protein is the same size as the murine homolog, and the amino acid sequences of the two proteins are 99.2% homologous. Analysis by reverse transcription-PCR revealed that this gene is expressed ubiquitously in normal tissues. It was mapped to chromosome band 5q31 by fluorescent in situ hybridization.

    Cytogenetics and cell genetics 1994;65;1-2;74-8

  • Association of the APC tumor suppressor protein with catenins.

    Su LK, Vogelstein B and Kinzler KW

    Department of Oncology, Johns Hopkins University School of Medicine, Baltimore, MD 21231.

    Mutations of APC appear to initiate sporadic and inherited forms of human colorectal cancer. Although these mutations have been well characterized, little is known about the function of the APC gene product. Two cellular proteins that associate with APC were identified by nucleotide sequence analysis and peptide mapping as the E-cadherin-associated proteins alpha- and beta-catenin. A 27-residue fragment of APC containing a 15-amino acid repeat was sufficient for the interaction with the catenins. These results suggest an important link between tumor initiation and cell adhesion.

    Funded by: NCI NIH HHS: CA-57345

    Science (New York, N.Y.) 1993;262;5140;1734-7

  • Expression of E- or P-cadherin is not sufficient to modify the morphology and the tumorigenic behavior of murine spindle carcinoma cells. Possible involvement of plakoglobin.

    Navarro P, Lozano E and Cano A

    Departamento de Bioquímica, Facultad de Medicina, UAM, Madrid, Spain.

    Transfection of E- and P-cadherin cDNA has been carried out in murine spindle carcinoma cells previously shown to be deficient in both cadherins (Navarro et al., J. Cell Biol. 115, 517-533, 1991). High levels of expression of E- or P-cadherin do not significantly affect the fibroblastic morphology of the parental spindle cells. In addition, the tumorigenic behavior of these highly malignant cells is not influenced by the ectopic expression of either cadherin. Nevertheless, a fraction of the exogenous cadherins is able to associate to detergent-insoluble components of the transfectant cells, and the expression of the exogenous E-cadherin confers Ca(2+)-dependent aggregation on the spindle transfectants in an in vitro assay. Immunoprecipitation analysis of the cadherin-catenin complex of the transfectants revealed that the ectopic E-cadherin associates with the alpha- and beta-catenin proteins. However, the gamma-catenin/plakoglobin component could not be detected in the E-cadherin immunocomplexes of the spindle transfectant cells, in contrast to the epithelial cells where the three catenins appeared to be associated with E-cadherin. The lack of association of gamma-catenin is correlated with very low levels of plakoglobin in whole cell extracts of the parental spindle cells. These results indicate that the association of E-cadherin with the alpha- and beta-catenin components is not sufficient to promote a fibroblastoid-epithelial conversion of highly malignant spindle cells. The presence of plakoglobin could be required for the proper organization of E-cadherin in the transfectant cells in order to acquire an epithelioid phenotype.

    Journal of cell science 1993;105 ( Pt 4);923-34

  • Cloning of the human alpha-catenin cDNA and its aberrant mRNA in a human cancer cell line.

    Oda T, Kanai Y, Shimoyama Y, Nagafuchi A, Tsukita S and Hirohashi S

    Pathology Division, National Cancer Center Research Institute, Tokyo, Japan.

    Cadherin and catenin compose cell adhesion complex and are indispensable for tight cell-cell adhesion. Dysfunction of this adhesion complex causes dissociation of cancer cells from primary tumor nodules, thus possibly contributing to cancer invasion and metastasis. In this report, we present the human alpha-catenin sequence. Human alpha-catenin showed extensive homology with that of mouse, i.e., 91.8% and 99.3% at the nucleic acid and amino acid levels, respectively, indicating that this molecule has been evolutionarily conserved in mammals. Characterization of the mRNA sequence of alpha-catenin in PC9 was also carried out, and two distinct abnormal sequences, i.e., one of 957 bp deletion resulting in a 319-amino-acid deletion and another of 761 bp deletion resulting in a frameshift, were identified. These deletions were probably produced by an error of RNA splicing, presenting one possible mechanism for the loss of intact alpha-catenin expression.

    Biochemical and biophysical research communications 1993;193;3;897-904

  • The uvomorulin-anchorage protein alpha catenin is a vinculin homologue.

    Herrenknecht K, Ozawa M, Eckerskorn C, Lottspeich F, Lenter M and Kemler R

    Max-Planck-Institut für Immunbiologie, FG Molekulare Embryologie, Freiburg, Federal Republic of Germany.

    The cytoplasmic region of the Ca(2+)-dependent cell-adhesion molecule (CAM) uvomorulin associates with distinct cytoplasmic proteins with molecular masses of 102, 88, and 80 kDa termed alpha, beta, and gamma catenin, respectively. This complex formation links uvomorulin to the actin filament network, which seems to be of primary importance for its cell-adhesion properties. We show here that antibodies against alpha catenin also immunoprecipitate complexes that contain human N-cadherin, mouse P-cadherin, chicken A-CAM (adherens junction-specific CAM; also called N-cadherin) or Xenopus U-cadherin, demonstrating that alpha catenin is complexed with other cadherins. In immunofluorescence tests, alpha catenin is colocalized with cadherins at the plasma membrane. However, in cadherin-negative Ltk- cells, alpha catenin is found uniformly distributed in the cytoplasm, suggesting some additional biological function(s). Expression of uvomorulin in these cells results in a concentration of alpha catenin at membrane areas of cell contacts. We also have cloned and sequenced murine alpha catenin. The deduced amino acid sequence reveals a significant homology to vinculin. Our results suggest the possibility of a new vinculin-related protein family involved in the cytoplasmic anchorage of cell-cell and cell-substrate adhesion molecules.

    Proceedings of the National Academy of Sciences of the United States of America 1991;88;20;9156-60

  • The 102 kd cadherin-associated protein: similarity to vinculin and posttranscriptional regulation of expression.

    Nagafuchi A, Takeichi M and Tsukita S

    Department of Information Physiology, National Institute for Physiological Sciences, Okazaki, Japan.

    The E-cadherin cell adhesion molecule is associated with cytoplasmic polypeptides, and this association is essential for its cell-binding function. Using isolated adherens junctions of the liver, we purified a 102 kd protein that can associate with E-cadherin (CAP102) and isolated cDNAs encoding this protein. Sequence analysis of the cDNAs revealed that this protein has a similarity to vinculin. L cells not expressing endogenous cadherin express the mRNA for CAP102 but have only a trace amount of CAP102 protein. Introducing exogenous E-cadherin into these cells, however, induced a high expression of CAP102 protein without affecting the amount of its mRNA, suggesting that there is a posttranscriptional regulatory mechanism for this molecule. The same effect was observed by introducing N- or P-cadherin into L cells.

    Cell 1991;65;5;849-57

Gene lists (6)

Gene List Source Species Name Description Gene count
L00000009 G2C Homo sapiens Human PSD Human orthologues of mouse PSD adapted from Collins et al (2006) 1080
L00000016 G2C Homo sapiens Human PSP Human orthologues of mouse PSP adapted from Collins et al (2006) 1121
L00000059 G2C Homo sapiens BAYES-COLLINS-HUMAN-PSD-CONSENSUS Human cortex PSD consensus 748
L00000061 G2C Homo sapiens BAYES-COLLINS-MOUSE-PSD-CONSENSUS Mouse cortex PSD consensus (ortho) 984
L00000069 G2C Homo sapiens BAYES-COLLINS-HUMAN-PSD-FULL Human cortex biopsy PSD full list 1461
L00000071 G2C Homo sapiens BAYES-COLLINS-MOUSE-PSD-FULL Mouse cortex PSD full list (ortho) 1556
© G2C 2014. The Genes to Cognition Programme received funding from The Wellcome Trust and the EU FP7 Framework Programmes:
EUROSPIN (FP7-HEALTH-241498), SynSys (FP7-HEALTH-242167) and GENCODYS (FP7-HEALTH-241995).

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