Plakoglobin domains that define its association with the desmosomal cadherins and the classical cadherins: identification of unique and shared domains

1996 ◽  
Vol 109 (5) ◽  
pp. 1143-1154 ◽  
Author(s):  
J.K. Wahl ◽  
P.A. Sacco ◽  
T.M. McGranahan-Sadler ◽  
L.M. Sauppe ◽  
M.J. Wheelock ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Intermediate Filament ◽  
Adherens Junction ◽  
Cell Junctions ◽  
Central Domain ◽  
Desmosomal Cadherins ◽  
Cytoplasmic Proteins ◽  
C Terminus ◽  
Classical Cadherins ◽  
N Terminus

Two cell-cell junctions, the adherens junction and the desmosome, are prominent in epithelial cells. These junctions are composed of transmembrane cadherins which interact with cytoplasmic proteins that serve to link the cadherin to the cytoskeleton. One component of both adherens junctions and desmosomes is plakoglobin. In the adherens junction plakoglobin interacts with both the classical cadherin and with alpha-catenin. Alpha-catenin in turn interacts with microfilaments. The role plakoglobin plays in the desmosome is not well understood. Plakoglobin interacts with the desmosomal cadherins, but how and if this mediates interactions with the intermediate filament cytoskeleton is not known. Here we compare the domains of plakoglobin that allow it to associate with the desmosomal cadherins with those involved in interactions with the classical cadherins. We show that three sites on plakoglobin are involved in associations with the desmosomal cadherins. A domain near the N terminus is unique to the desmosomal cadherins and overlaps with the site that interacts with alpha-catenin, suggesting that there may be competition between alpha-catenin and the desmosomal cadherins for interactions with plakoglobin. In addition, a central domain is shared with regions used by plakoglobin to associate with the classical cadherins. Finally, a domain near the C terminus is shown to strongly modulate the interactions with the desmosomal cadherins. This latter domain also contributes to the association of plakoglobin with the classical cadherins.

scholarly journals Cross-Talk between Adherens Junctions and Desmosomes Depends on Plakoglobin

1997 ◽  
Vol 136 (4) ◽  
pp. 919-934 ◽  
Author(s):  
Jani E. Lewis ◽  
James K. Wahl ◽  
Kristin M. Sass ◽  
Pamela J. Jensen ◽  
Keith R. Johnson ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Cell Line ◽  
Adherens Junctions ◽  
Adherens Junction ◽  
Epithelial Cell Line ◽  
Cell Contact ◽  
Common Component ◽  
Classical Cadherins ◽  
E Cadherin

Squamous epithelial cells have both adherens junctions and desmosomes. The ability of these cells to organize the desmosomal proteins into a functional structure depends upon their ability first to organize an adherens junction. Since the adherens junction and the desmosome are separate structures with different molecular make up, it is not immediately obvious why formation of an adherens junction is a prerequisite for the formation of a desmosome. The adherens junction is composed of a transmembrane classical cadherin (E-cadherin and/or P-cadherin in squamous epithelial cells) linked to either β-catenin or plakoglobin, which is linked to α-catenin, which is linked to the actin cytoskeleton. The desmosome is composed of transmembrane proteins of the broad cadherin family (desmogleins and desmocollins) that are linked to the intermediate filament cytoskeleton, presumably through plakoglobin and desmoplakin. To begin to study the role of adherens junctions in the assembly of desmosomes, we produced an epithelial cell line that does not express classical cadherins and hence is unable to organize desmosomes, even though it retains the requisite desmosomal components. Transfection of E-cadherin and/or P-cadherin into this cell line did not restore the ability to organize desmosomes; however, overexpression of plakoglobin, along with E-cadherin, did permit desmosome organization. These data suggest that plakoglobin, which is the only known common component to both adherens junctions and desmosomes, must be linked to E-cadherin in the adherens junction before the cell can begin to assemble desmosomal components at regions of cell–cell contact. Although adherens junctions can form in the absence of plakoglobin, making use only of β-catenin, such junctions cannot support the formation of desmosomes. Thus, we speculate that plakoglobin plays a signaling role in desmosome organization.

Molecular interactions between desmosomal cadherins

2002 ◽  
Vol 362 (2) ◽  
pp. 317-327 ◽  
Author(s):  
Shabih-e-Hassnain SYED ◽  
Brian TRINNAMAN ◽  
Stephen MARTIN ◽  
Sarah MAJOR ◽  
Jon HUTCHINSON ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Analytical Ultracentrifugation ◽  
Expression System ◽  
Quantitative Information ◽  
Cell Junctions ◽  
Protein Protein Interactions ◽  
Desmosomal Cadherins ◽  
Classical Cadherins ◽  
Escherichia Coli Expression

Desmocollins (Dscs) and desmogleins (Dsgs) are cell-adhesion molecules involved in the formation of desmosome cell—cell junctions and share structural similarities to classical cadherins such as E-cadherin. In order to identify and provide quantitative information on the types of protein—protein interactions displayed by the type 2 isoforms and investigate the role of Ca2+ in this process, we have developed an Escherichia coli expression system to generate recombinant proteins containing the first two extracellular domains, namely Dsg2(1-2) and Dsc2(1-2). Analytical ultracentrifugation, chemical cross-linking, CD, fluorescence and BIAcore have been used to provide the first direct evidence of Ca2+ binding to desmosomal cadherins. These studies suggest that Dsc2(1-2) not only exhibits homophilic interactions in solution, but can also form heterophilic interactions with Dsg2(1-2). The latter, on the other hand, shows much weaker homophilic association. Our results further demonstrate that heterophilic interactions are Ca2+-dependent, whereas the Ca2+-dependence of homophilic association is less clear. Our data indicate that the functional properties of Dsc2(1-2) are more similar to those of classical cadherins, consistent with the observation that Dsc shares a higher level of sequence homology with classical cadherins than does Dsg. In addition to corroborating the conclusions of previously reported transfection studies which suggest the formation of lateral heterodimers and homodimers, our results also provide direct quantitative information on the strength of these interactions which are essential for understanding the adhesion mechanism.

scholarly journals Desmoplakin Is Required Early in Development for Assembly of Desmosomes and Cytoskeletal Linkage

1998 ◽  
Vol 143 (7) ◽  
pp. 2009-2022 ◽  
Author(s):  
G. Ian Gallicano ◽  
Panos Kouklis ◽  
Christoph Bauer ◽  
Mei Yin ◽  
Valeri Vasioukhin ◽  
...  
Keyword(s):  
Critical Role ◽  
Early Embryo ◽  
Cell Junctions ◽  
Epithelial Polarity ◽  
Tissue Architecture ◽  
Desmosomal Cadherins ◽  
Tissue Integrity ◽  
Classical Cadherins ◽  
Mediate Cell ◽  
Cell Cell

Desmosomes first assemble in the E3.5 mouse trophectoderm, concomitant with establishment of epithelial polarity and appearance of a blastocoel cavity. Throughout development, they increase in size and number and are especially abundant in epidermis and heart muscle. Desmosomes mediate cell–cell adhesion through desmosomal cadherins, which differ from classical cadherins in their attachments to intermediate filaments (IFs), rather than actin filaments. Of the proteins implicated in making this IF connection, only desmoplakin (DP) is both exclusive to and ubiquitous among desmosomes. To explore its function and importance to tissue integrity, we ablated the desmoplakin gene. Homozygous −/− mutant embryos proceeded through implantation, but did not survive beyond E6.5. Mutant embryos proceeded through implantation, but did not survive beyond E6.5. Surprisingly, analysis of these embryos revealed a critical role for desmoplakin not only in anchoring IFs to desmosomes, but also in desmosome assembly and/or stabilization. This finding not only unveiled a new function for desmoplakin, but also provided the first opportunity to explore desmosome function during embryogenesis. While a blastocoel cavity formed and epithelial cell polarity was at least partially established in the DP (−/−) embryos, the paucity of desmosomal cell–cell junctions severely affected the modeling of tissue architecture and shaping of the early embryo.

Assembly of adherens junctions is required for sphingosine 1-phosphate-induced matriptase accumulation and activation at mammary epithelial cell-cell contacts

2004 ◽  
Vol 286 (5) ◽  
pp. C1159-C1169 ◽  
Author(s):  
Ruei-Jiun Hung ◽  
Ia-Wen J. Hsu ◽  
Jennifer L. Dreiling ◽  
Mon-Juan Lee ◽  
Cicely A. Williams ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Mammary Epithelial Cells ◽  
Adherens Junctions ◽  
Adherens Junction ◽  
Mammary Epithelial ◽  
Cell Junctions ◽  
Cell Contacts ◽  
Cytoskeletal Rearrangement ◽  
Sphingosine 1 Phosphate ◽  
Cell Cell

Sphingosine 1-phosphate (S1P), a bioactive phospholipid, simultaneously induces actin cytoskeletal rearrangements and activation of matriptase, a membrane-associated serine protease in human mammary epithelial cells. In this study, we used a monoclonal antibody selective for activated, two-chain matriptase to examine the functional relationship between these two S1P-induced events. Ten minutes after exposure of 184 A1N4 mammary epithelial cells to S1P, matriptase was observed to accumulate at cell-cell contacts. Activated matriptase first began to appear as small spots at cell-cell contacts, and then its deposits elongated along cell-cell contacts. Concomitantly, S1P induced assembly of adherens junctions and subcortical actin belts. Matriptase localization was observed to be coincident with markers of adherens junctions at cell-cell contacts but likely not to be incorporated into the tightly bound adhesion plaque. Disruption of subcortical actin belt formation and prevention of adherens junction assembly led to prevention of accumulation and activation of the protease at cell-cell contacts. These data suggest that S1P-induced accumulation and activation of matriptase depend on the S1P-induced adherens junction assembly. Although MAb M32, directed against one of the low-density lipoprotein receptor class A domains of matriptase, blocked S1P-induced activation of the enzyme, the antibody had no effect on S1P-induced actin cytoskeletal rearrangement. Together, these data indicate that actin cytoskeletal rearrangement is necessary but not sufficient for S1P-induced activation of matriptase at cell-cell contacts. The coupling of matriptase activation to adherens junction assembly and actin cytoskeletal rearrangement may serve to ensure tight control of matriptase activity, restricted to cell-cell junctions of mammary epithelial cells.

scholarly journals Breaking the connection: displacement of the desmosomal plaque protein desmoplakin from cell-cell interfaces disrupts anchorage of intermediate filament bundles and alters intercellular junction assembly.

1996 ◽  
Vol 134 (4) ◽  
pp. 985-1001 ◽  
Author(s):  
E A Bornslaeger ◽  
C M Corcoran ◽  
T S Stappenbeck ◽  
K J Green
Keyword(s):  
Electron Microscopy ◽  
Cell Lines ◽  
Intermediate Filament ◽  
Adherens Junction ◽  
Intercellular Junction ◽  
Immunogold Electron Microscopy ◽  
Desmosomal Cadherins ◽  
Cytoplasmic Tails ◽  
Filament Bundles ◽  
Cell Cell

The desmosomal plaque protein desmoplakin (DP), located at the juncture between the intermediate filament (IF) network and the cytoplasmic tails of the transmembrane desmosomal cadherins, has been proposed to link IF to the desmosomal plaque. Consistent with this hypothesis, previous studies of individual DP domains indicated that the DP COOH terminus associates with IF networks whereas NH2-terminal sequences govern the association of DP with the desmosomal plaque. Nevertheless, it had not yet been demonstrated that DP is required for attaching IF to the desmosome. To test this proposal directly, we generated A431 cell lines stably expressing DP NH2-terminal polypeptides, which were expected to compete with endogenous DP during desmosome assembly. As these polypeptides lacked the COOH-terminal IF-binding domain, this competition should result in the loss of IF anchorage if DP is required for linking IF to the desmosomal plaque. In such cells, a 70-kD DP NH2-terminal polypeptide (DP-NTP) colocalized at cell-cell interfaces with desmosomal proteins. As predicted, the distribution of endogenous DP was severely perturbed. At cell-cell borders where endogenous DP was undetectable by immunofluorescence, there was a striking absence of attached tonofibrils (IF bundles). Furthermore, DP-NTP assembled into ultrastructurally identifiable junctional structures lacking associated IF bundles. Surprisingly, immunofluorescence and immunogold electron microscopy indicated that adherens junction components were coassembled into these structures along with desmosomal components and DP-NTP. These results indicate that DP is required for anchoring IF networks to desmosomes and furthermore suggest that the DP-IF complex is important for governing the normal spatial segregation of adhesive junction components during their assembly into distinct structures.

scholarly journals Structure of APP-C991-99 and Implications for Role of Extra-Membrane Domains in Function and Oligomerization

2018 ◽  
Author(s):  
George A. Pantelopulos ◽  
John E. Straub ◽  
D. Thirumalai ◽  
Yuji Sugita
Keyword(s):  
Transmembrane Domain ◽  
Chemical Shifts ◽  
Membrane Surface ◽  
Critical Role ◽  
Amyloid Formation ◽  
Cytoplasmic Proteins ◽  
C Terminus ◽  
N Terminus ◽  
Thin Membranes

AbstractThe 99 amino acid C-terminal fragment of Amyloid Precursor Protein APP-C99 (C99) is cleaved by γ-secretase to form Aβ peptide, which plays a critical role in the etiology of Alzheimer’s Disease (AD). The structure of C99 consists of a single transmembrane domain flanked by intra and intercellular domains. While the structure of the transmembrane domain has been well characterized, little is known about the structure of the flanking domains and their role in C99 processing by γ-secretase. To gain insight into the structure of full-length C99, REMD simulations were performed for monomeric C99 in model membranes of varying thickness. We find equilibrium ensembles of C99 from simulation agree with experimentally-inferred residue insertion depths and protein backbone chemical shifts. In thin membranes, the transmembrane domain structure is correlated with extra-membrane structural states. Mean and variance of the transmembrane and G37G38 hinge angles are found to increase with thinning membrane. The N-terminus of C99 forms β-strands that may seed aggregation of Aβ on the membrane surface, promoting amyloid formation. The N-terminus, which forms α-helices that interact with the nicastrin domain of γ-secretase. The C-terminus of C99 becomes more α-helical as the membrane thickens, forming structures that may be suitable for binding by cytoplasmic proteins, while C-terminal residues essential to cytotoxic function become α-helical as the membrane thins. The heterogeneous but discrete extra-membrane domain states analyzed here open the path to new investigations of the role of C99 structure and membrane in amyloidogenesis.

scholarly journals The unique tropism ofMycobacterium lepraeto the nasal epithelial cells can be explained by the mammalian cell entry protein 1A

2018 ◽  
Author(s):  
Viesta Beby Fadlitha ◽  
Fuki Yamamoto ◽  
Irfan Idris ◽  
Haslindah Dahlan ◽  
Naoya Sato ◽  
...  
Keyword(s):  
Amino Acids ◽  
Epithelial Cell ◽  
Epithelial Cells ◽  
Recombinant Proteins ◽  
Cell Entry ◽  
Important Region ◽  
C Terminus ◽  
Protein Encoding ◽  
N Terminus ◽  
Encoding Genes

AbstractLeprosy is a chronic infection where the skin and peripheral nervous system is invaded byMycobacterium leprae. The infection mechanism remains unknown in part because culture methods have not been established yet forM.leprae.Mce1A protein (442 aa) is coded by mce1A (1326 bp) ofM.leprae. The mce1A homolog inMycobacterium tuberculosisis known to be associated withM.tuberculosisepithelial cell entry, and survival and multiplication within macrophages. Studies using recombinant proteins have indicated that mce1A ofM.lepraeis also associated with epithelial cell entry. This study is aimed at identifying particular sequences within mce1A associated withM.lepraeepithelial cell entry.Recombinant proteins having N-terminus and C-terminus truncations of the mce1A region ofM.lepraewere created inEschericia coli.Entry activity of latex beads, coated with these truncated proteins (r-lep37kDa and r-lep27kDa), into HeLa cells was observed by electron microscopy. The entry activity was preserved even when 315 bp (105 aa) and 922 bp (308 aa) was truncated from the N-terminus and C-terminus, respectively. This 316 – 921 bp region was divided into three sub-regions: 316 – 531 bp (InvX), 532 – 753 bp (InvY), and 754 – 921 bp (InvZ). Each sub-region was cloned into an AIDA vector and expressed on the surface ofE.coli.Entry of theseE.coliinto monolayer-cultured HeLa and RPMI2650 cells was observed by electron microscopy. OnlyE.coliharboring the InvX sub-region exhibited cell entry. InvX was further divided into 4 domains, InvXa - InvXd, containing sequences 1 – 24 aa, 25 – 46 aa, 47 – 57 aa, and 58 – 72 aa, respectively.RecombinantE.coli, expressing each of InvXa - InvXd on the surface, were treated with antibodies against these domains, then added to monolayer cultured RPMI cells. The effectiveness of these antibodies in preventing cell entry was studied by colony counting. Entry activity was suppressed by antibodies against InvXa, InvXb, and InvXd. This suggests that these three InvX domains of mce1A are important forM.lepraeinvasion into nasal epithelial cells.Author SummaryMce1A protein is encoded by the mce1A region of mce1 locus ofM.tuberculosisandM.leprae, and is involved in the bacteria’s invasion into epithelial cells. The present study revealed that the active sequence ofM.lepraeinvolved in the invasion into nasal mucosa epithelial cells is present in the 316-531 bp region of mce1A.The most important region of mce1A protein involved in the invasion ofM.tuberculosisinto human epithelial cells is called the InvIII region, which is located between amino acids at position 130 to 152. The InvIII region ofM.tuberculosiscorresponds to InvXb ofM.leprae. The sequences of these regions are identical between amino acids at positions 10 to position 22 as counted from the N terminus, except that amino acids at positions 1 to 3, 5, 8, 9, 13 are different betweenM.lepraeandM.tuberculosis. Suppression test results also indicated that the most important region of mce1A protein ofM.lepraeinvolved in the invasion into human epithelial cells is different from thatM.tuberculosis. WhileM.tuberculosishas 3,959 protein-encoding genes and only 6 pseudogenes,M.lepraehas only 1,604 protein-encoding genes but has 1,116 pseudogenes indicating that inM.leprae, far more proteins are inactivated as compared toM.tuberculosis. The present study also revealed that, as inM.tuberculosis,the mce1A protein is expressed on the surface of bacteria as a native protein. In light of these data, the mce1A protein is considered to be one of the most important proteins involved in the invasion ofM.lepraeinto nasal mucosa epithelial cells.

scholarly journals A polycystin multiprotein complex constitutes a cholesterol-containing signalling microdomain in human kidney epithelia

2005 ◽  
Vol 392 (1) ◽  
pp. 29-38 ◽  
Author(s):  
Tamara Roitbak ◽  
Zurab Surviladze ◽  
Ritva Tikkanen ◽  
Angela Wandinger-Ness
Keyword(s):  
Plasma Membrane ◽  
Epithelial Cells ◽  
Buoyant Density ◽  
Adherens Junction ◽  
Human Kidney ◽  
Cholesterol Depletion ◽  
Multiprotein Complex ◽  
Signalling Molecules ◽  
C Terminus ◽  
Kidney Epithelial Cells

Polycystins are plasma membrane proteins that are expressed in kidney epithelial cells and associated with the progression of ADPKD (autosomal dominant polycystic kidney disease). A polycystin multiprotein complex, including adherens junction proteins, is thought to play an important role in cell polarity and differentiation. Sucrose gradient analyses and immunoprecipitation studies of primary human kidney epithelial cells showed the polycystins and their associated proteins E-cadherin and β-catenin distributed in a complex with the raft marker flotillin-2, but not caveolin-1, in high-density gradient fractions. The integrity of the polycystin multiprotein complex was sensitive to cholesterol depletion, as shown by cyclodextrin treatment of immunoprecipitated complexes. The overexpressed C-terminus of polycystin-1 retained the ability to associate with flotillin-2. Flotillin-2 was found to contain CRAC (cholesterol recognition/interaction amino acid) cholesterol-binding domains and to promote plasma membrane cholesterol recruitment. Based on co-association of signalling molecules, such as Src kinases and phosphatases, we propose that the polycystin multiprotein complex is embedded in a cholesterol-containing signalling microdomain specified by flotillin-2, which is distinct from classical light-buoyant-density, detergent-resistant domains.

scholarly journals Vezatin, a ubiquitous protein of adherens cell–cell junctions, is exclusively expressed in germ cells in mouse testis

Reproduction ◽  
2007 ◽  
Vol 133 (3) ◽  
pp. 563-574 ◽  
Author(s):  
Vincent Hyenne ◽  
Juergen C Harf ◽  
Martin Latz ◽  
Bernard Maro ◽  
Uwe Wolfrum ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Germ Cells ◽  
Transmembrane Protein ◽  
Adherens Junctions ◽  
Adherens Junction ◽  
Reproductive Organs ◽  
Cell Junctions ◽  
Molecular Characteristics ◽  
Acrosomal Membrane ◽  
Ectoplasmic Specialization

In the male reproductive organs of mammals, the formation of spermatozoa takes place during two successive phases: differentiation (in the testis) and maturation (in the epididymis). The first phase, spermiogenesis, relies on a unique adherens junction, the apical ectoplasmic specialization linking the epithelial Sertoli cells to immature differentiating spermatids. Vezatin is a transmembrane protein associated with adherens junctions and the actin cytoskeleton in most epithelial cells. We report here the expression profile of vezatin during spermatogenesis. Vezatin is exclusively expressed in haploid germ cells. Immunocytochemical and ultrastructural analyses showed that vezatin intimately coincides, temporally and spatially, with acrosome formation. While vezatin is a transmembrane protein associated with adherens junctions in many epithelial cells, it is not seen at the ectoplasmic specializations, neither at the basal nor at the apical sites, in the seminiferous epithelium. In particular, vezatin does not colocalize with espin and myosin VIIa, two molecular markers of the ectoplasmic specialization. In differentiating spermatids, ultrastructural data indicate that vezatin localizes in the acrosome. In epididymal sperm, vezatin localizes also to the outer acrosomal membrane. Considering its developmental and molecular characteristics, vezatin may be involved in the assembly/stability of this spermatic membrane.

scholarly journals The amino- and carboxyl-terminal tails of (beta)-catenin reduce its affinity for desmoglein 2

2000 ◽  
Vol 113 (10) ◽  
pp. 1737-1745 ◽  
Author(s):  
J.K. Wahl ◽  
J.E. Nieset ◽  
P.A. Sacco-Bubulya ◽  
T.M. Sadler ◽  
K.R. Johnson ◽  
...  
Keyword(s):  
Amino Acids ◽  
Protein Interactions ◽  
Adherens Junction ◽  
Beta Catenin ◽  
Protein Protein Interactions ◽  
Desmosomal Cadherins ◽  
Classical Cadherins ◽  
Show Evidence ◽  
Carboxyl Terminal ◽  
Armadillo Repeats

beta-catenin and plakoglobin are members of the armadillo family of proteins and were first identified as components of intercellular adhering junctions. In the adherens junction beta-catenin and plakoglobin serve to link classical cadherins to the actin-based cytoskeleton. In the desmosome plakoglobin links the desmosomal cadherins, the desmogleins and the desmocollins, to the intermediate filament cytoskeleton. beta-catenin is not a component of the desmosome. Previously we have shown that the central armadillo repeat region of plakoglobin is the site for desmosomal cadherin binding. We hypothesized that the unique amino- and/or carboxyl-terminal ends of beta-catenin may regulate its exclusion from the desmosomal plaque. To test this hypothesis we used chimeras between beta-catenin and plakoglobin to identify domain(s) that modulate association with desmoglein 2. Chimeric constructs, each capable of associating with classical cadherins, were assayed for association with the desmosomal cadherin desmoglein 2. Addition of either the N- or C-terminal tail of beta-catenin to the armadillo repeats of plakoglobin did not interfere with desmoglein 2 association. However, when both beta-catenin amino terminus and carboxyl terminus were added to the plakoglobin armadillo repeats, association with desmoglein 2 was diminished. Removal of the first 26 amino acids from this construct restored association. We show evidence for direct protein-protein interactions between the amino- and carboxyl-terminal tails of beta-catenin and propose that a sequence in the first 26 amino acids of beta-catenin along with its carboxyl-terminal tail decrease its affinity for desmoglein and prevent its inclusion in the desmosome.

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