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1.
Pemphigus vulgaris antigen, a desmoglein type of cadherin, is localized within keratinocyte desmosomes 总被引:7,自引:2,他引:7
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《The Journal of cell biology》1993,122(2):409-415
Pemphigus vulgaris antigen (PVA) is a member of the desmoglein subfamily of cadherin cell adhesion molecules. Because autoantibodies in this disease cause blisters due to loss of epidermal cell adhesion, and because desmoglein is found in the desmosome cell adhesion junction, we wanted to determine if PVA is also found in desmosomes. By immunofluorescence, PV IgG bound, in a dotted pattern, to the cell surface of cultured human keratinocytes induced to differentiate with calcium, suggesting junctional staining. However, by preembedding, immunogold electron microscopic studies only slight labeling could be detected in desmosomes, presumably because of difficulty in gold penetration of intact desmosomes. We therefore treated the keratinocytes with 0.01% trypsin in 1 mM calcium, conditions known to preserve cadherin antigenicity but that caused slight separation of desmosomes, before immunogold staining. In this case there was extensive labeling of the extracellular part of desmosomes but not of the interdesmosomal cell membrane which was stained with anti-beta 2- microglobulin antibodies. To confirm the specificity of this binding we showed that antibodies raised in rabbits against the extracellular portions of PVA also bound desmosomes in these cultures. In intact mouse epidermis we could also show slight, but specific, immunogold desmosomal labeling with PV IgG. Furthermore, neonatal mice injected with PV IgG affinity purified on PVA showed desmosomal separation with the IgG localized to desmosomal cores. These results indicate that PVA is organized and concentrated within the desmosome where it presumably functions to maintain the integrity of stratifying epithelia. 相似文献
2.
Despite the presumed importance of desmoglein, a 160-kDa glycoprotein, in desmosome formation and its possible involvement in certain blistering skin diseases, the precise location and function of this protein have not yet been firmly established. We describe here the characterization of a new monoclonal antibody, AE23, against an extracellular epitope of desmoglein. Both the AE23 epitope and another epitope, defined by the previously characterized DG3.4 antibody, reside on a 160-kDa human epidermal desmoglein as evidenced by their identical solubility profile, their coexistence in a 130-kDa desmoglein degradative product, their coadsorption by an AE23 immunoaffinity column, and the identical changes in the two antigens' electrophoretic mobility after air oxidation and deglycosylation. The AE23 epitope is resistant to various endoglycosidases, suggesting that sugar moieties are not involved. Characterization of several proteolytic fragments of this epidermal desmoglein enabled us to map the DG3.4 epitope to a 96-kDa intracellular domain and the AE23 epitope to an extracellular domain flanked by the plasma membrane and the distal N-glycosylation site(s). However, these two epitopes do not always coexist on the same desmoglein molecule. For example, tissue surveys showed that although the DG3.4 epitope is present in the desmogleins of all epithelial cell types, the AE23 epitope is limited to normal keratinocytes. Moreover, electron microscopic localization data indicate that whereas the DG3.4 epitope is detected in the submembranous plaques of desmosomes, the AE23 epitope is present in the intercellular space of both desmosomal and nondesmosomal areas. These results raise the possibility that there exist several biochemically closely related isoforms of desmoglein, one (AE23+/DG3.4+) restricted to epidermal desmosomes, one (AE23+/DG3.4-) uniformly distributed along the keratinocyte cell surface, and another (AE23-/DG3.4+) present in desmosomes of simple epithelia and basal cells of cultured keratinocytes. The uniform distribution of at least one desmoglein-related antigen in the intercellular space of keratinocytes coupled with the realization that different isoforms of desmogleins form a subfamily of cadherins suggest that desmoglein(s) may play a more general role in keratinocyte adhesion than previously appreciated. 相似文献
3.
《Cell communication & adhesion》2013,20(1):77-84
AbstractAutoantibodies from patients suffering from the autoimmune blistering skin disease pemphigus can be applied as tools to study desmosomal adhesion. These autoantibodies targeting the desmosomal cadherins desmoglein (Dsg) 1 and Dsg3 cause disruption of desmosomes and loss of intercellular cohesion. Although pemphigus autoantibodies were initially proposed to sterically hinder desmosomes, many groups have shown that they activate signaling pathways which cause disruption of desmosomes and loss of intercellular cohesion by uncoupling the desmosomal plaque from the intermediate filament cytoskeleton and/or by interfering with desmosome turnover. These studies demonstrate that desmogleins serve as receptor molecules to transmit outside-in signaling and demonstrate that desmosomal cadherins have functions in addition to their adhesive properties. Two central molecules regulating cytoskeletal anchorage and desmosome turnover are p38MAPK and PKC. As cytoskeletal uncoupling in turn enhances Dsg3 depletion from desmosomes, both mechanisms reinforce one another in a vicious cycle that compromise the integrity and number of desmosomes. 相似文献
4.
Beaudry VG Jiang D Dusek RL Park EJ Knezevich S Ridd K Vogel H Bastian BC Attardi LD 《PLoS genetics》2010,6(10):e1001168
Dysregulated cell-cell adhesion plays a critical role in epithelial cancer development. Studies of human and mouse cancers have indicated that loss of adhesion complexes known as adherens junctions contributes to tumor progression and metastasis. In contrast, little is known regarding the role of the related cell-cell adhesion junction, the desmosome, during cancer development. Studies analyzing expression of desmosome components during human cancer progression have yielded conflicting results, and therefore genetic studies using knockout mice to examine the functional consequence of desmosome inactivation for tumorigenesis are essential for elucidating the role of desmosomes in cancer development. Here, we investigate the consequences of desmosome loss for carcinogenesis by analyzing conditional knockout mice lacking Perp, a p53/p63 regulated gene that encodes an important component of desmosomes. Analysis of Perp-deficient mice in a UVB-induced squamous cell skin carcinoma model reveals that Perp ablation promotes both tumor initiation and progression. Tumor development is associated with inactivation of both of Perp's known functions, in apoptosis and cell-cell adhesion. Interestingly, Perp-deficient tumors exhibit widespread downregulation of desmosomal constituents while adherens junctions remain intact, suggesting that desmosome loss is a specific event important for tumorigenesis rather than a reflection of a general change in differentiation status. Similarly, human squamous cell carcinomas display loss of PERP expression with retention of adherens junctions components, indicating that this is a relevant stage of human cancer development. Using gene expression profiling, we show further that Perp loss induces a set of inflammation-related genes that could stimulate tumorigenesis. Together, these studies suggest that Perp-deficiency promotes cancer by enhancing cell survival, desmosome loss, and inflammation, and they highlight a fundamental role for Perp and desmosomes in tumor suppression. An understanding of the factors affecting cancer progression is important for ultimately improving the diagnosis, prognostication, and treatment of cancer. 相似文献
5.
Desmoglein 4 in hair follicle differentiation and epidermal adhesion: evidence from inherited hypotrichosis and acquired pemphigus vulgaris 总被引:14,自引:0,他引:14
Kljuic A Bazzi H Sundberg JP Martinez-Mir A O'Shaughnessy R Mahoney MG Levy M Montagutelli X Ahmad W Aita VM Gordon D Uitto J Whiting D Ott J Fischer S Gilliam TC Jahoda CA Morris RJ Panteleyev AA Nguyen VT Christiano AM 《Cell》2003,113(2):249-260
Cell adhesion and communication are interdependent aspects of cell behavior that are critical for morphogenesis and tissue architecture. In the skin, epidermal adhesion is mediated in part by specialized cell-cell junctions known as desmosomes, which are characterized by the presence of desmosomal cadherins, known as desmogleins and desmocollins. We identified a cadherin family member, desmoglein 4, which is expressed in the suprabasal epidermis and hair follicle. The essential role of desmoglein 4 in skin was established by identifying mutations in families with inherited hypotrichosis, as well as in the lanceolate hair mouse. We also show that DSG4 is an autoantigen in pemphigus vulgaris. Characterization of the phenotype of naturally occurring mutant mice revealed disruption of desmosomal adhesion and perturbations in keratinocyte behavior. We provide evidence that desmoglein 4 is a key mediator of keratinocyte cell adhesion in the hair follicle, where it coordinates the transition from proliferation to differentiation. 相似文献
6.
Desmoplakin (DP), plakoglobin (PG), and plakophilin 1 (PP1) are desmosomal components lacking a transmembrane domain, thus making them candidate linker proteins for connecting intermediate filaments and desmosomes. Using deletion and site-directed mutagenesis, we show that remarkably, removal of ~1% of DP's sequence obliterates its ability to associate with desmosomes. Conversely, when linked to a foreign protein, as few as 86 NH2-terminal DP residues are sufficient to target to desmosomes efficiently. In in vitro overlay assays, the DP head specifically associates with itself and with desmocollin 1a (Dsc1a). In similar overlay assays, PP1 binds to DP and Dsc1a, and to a lesser extent, desmoglein 1 (Dsg1), while PG binds to Dsg1 and more weakly to Dsc1a and DP. Interestingly, like DP, PG and PP1 associate with epidermal keratins, although PG is considerably weaker in its ability to do so. As judged by overlay assays, the amino terminal head domain of type II keratins appears to have a special importance in establishing these connections. Taken together, our findings provide new insights into the complexities of the links between desmosomes and intermediate filaments (IFs). Our results suggest a model whereby at desmosome sites within dividing epidermal cells, DP and PG anchor to desmosomal cadherins and to each other, forming an ordered array of nontransmembrane proteins that then bind to keratin IFs. As epidermal cells differentiate, PP1 is added as a molecular reinforcement to the plaque, enhancing anchorage to IFs and accounting at least partially for the increase in numbers and stability of desmosomes in suprabasal cells. 相似文献
7.
Identification of the plakoglobin-binding domain in desmoglein and its role in plaque assembly and intermediate filament anchorage 总被引:11,自引:8,他引:3
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《The Journal of cell biology》1994,127(1):151-160
The carboxyterminal cytoplasmic portions (tails) of desmosomal cadherins of both the desmoglein (Dsg) and desmocollin type are integral components of the desmosomal plaque and are involved in desmosome assembly and the anchorage of intermediate-sized filaments. When additional Dsg tails were introduced by cDNA transfection into cultured human epithelial cells, in the form of chimeras with the aminoterminal membrane insertion domain of rat connexin32 (Co32), the resulting stably transfected cells showed a dominant-negative defect specific for desmosomal junctions: despite the continual presence of all desmosomal proteins, the endogenous desmosomes disappeared and the formation of Co32-Dsg chimeric gap junctions was inhibited. Using cell transfection in combination with immunoprecipitation techniques, we have examined a series of deletion mutants of the Dsg1 tail in Co32-Dsg chimeras. We show that upon removal of the last 262 amino acids the truncated Dsg tail still effects the binding of plakoglobin but not of detectable amounts of any catenin and induces the dominant-negative phenotype. However, further truncation or excision of the next 41 amino acids, which correspond to the highly conserved carboxyterminus of the C-domain in other cadherins, abolishes plakoglobin binding and allows desmosomes to reform. Therefore, we conclude that this short segment provides a plakoglobin-binding site and is important for plaque assembly and the specific anchorage of either actin filaments in adherens junctions or IFs in desmosomes. 相似文献
8.
Plakophilin 1, a member of the armadillo multigene family, is a protein with dual localization in the nucleus and in desmosomes. To elucidate its role in desmosome assembly and regulation, we have analyzed its localization and binding partners in vivo. When overexpressed in HaCaT keratinocytes, plakophilin 1 localized to the nucleus and to desmosomes, and dramatically enhanced the recruitment of desmosomal proteins to the plasma membrane. This effect was mediated by plakophilin 1's head domain, which interacted with desmoglein 1, desmoplakin, and keratins in the yeast two-hybrid system. Overexpression of the armadillo repeat domain induced a striking dominant negative phenotype with the formation of filopodia and long cellular protrusions, where plakophilin 1 colocalized with actin filaments. This phenotype was strictly dependent on a conserved motif in the center of the armadillo repeat domain. Our results demonstrate that plakophilin 1 contains two functionally distinct domains: the head domain, which could play a role in organizing the desmosomal plaque in suprabasal cells, and the armadillo repeat domain, which might be involved in regulating the dynamics of the actin cytoskeleton. 相似文献
9.
In the present study, we have examined how modulation of protein kinase C (PKC) activity affected desmosome organization in HeLa cells. Immunofluorescence and electron microscopy showed that PKC activation upon short exposure to 12-O-tetradecanoylphorbol 13-acetate (TPA) resulted in a reduction of intercellular contacts, splitting of desmosomes and dislocation of desmosomal components from the cell periphery towards the cytoplasm. As determined by immunoblot analysis of Triton X-100-soluble and -insoluble pools of proteins, these morphological changes were not correlated with modifications in the extractability of both desmoglein and plakoglobin, but involved almost complete solubilization of the desmosomal plaque protein, desmoplakin. Immunoprecipitation experiments and immunoblotting with anti-phosphoserine, antiphosphothreonine and anti-phosphotyrosine antibodies revealed that desmoplakin was mainly phosphorylated on serine and tyrosine residues in both treated and untreated cells. While phosphotyrosine content was not affected by PKC activation, phosphorylation on serine residues was increased by about two-fold. This enhanced serine phosphorylation coincided with the increase in the protein solubility, suggesting that phosphorylation of desmoplakin may be a mechanism by which PKC mediates desmosome disassembly. Consistent with the loss of PKC activity, we also showed that down-modulation of the kinase (in response to prolonged TPA treatment) or its specific inhibition (by GF 109203X) had opposite effects and increased desmosome formation. Taken together, these results clearly demonstrate an important role for PKC in the regulation ofdesmosomal junctions in HeLa cells, and identify serine phosphorylation of desmoplakin as a crucial event in this pathway. 相似文献
10.
Gloushankova NA Wakatsuki T Troyanovsky RB Elson E Troyanovsky SM 《Cell and tissue research》2003,314(3):399-410
Subclones of human carcinoma-derived A-431 cell line stably producing fusion proteins consisting of the enhanced green fluorescent protein and either human desmoglein 2 (Dsg-GFP) or human plakoglobin (GFP-Pg) were used to examine the behavior of desmosomes in living cells. Immunofluorescence microscopy of the fixed cells showed that both fusion proteins, which were expressed in significantly lower levels relative to their endogenous counterparts, were efficiently recruited into desmosomes. Time-lapse confocal imaging of these cells reveals that such GFP-labeled desmosomes (GFP desmosomes) are stable structures which exhibit various dynamic and motile activities. The most notable are independent lateral mobility and fusion. Furthermore, the continual assembly of new nascent desmosomes is observed within stable contacts located at the middle of the epithelial sheet. A new GFP desmosome appears as a closely apposed group of fine patches which after a few minutes aggregate into a single structure. These three dynamic processes resulted in constant changes of desmosome distribution, numbers, and sizes. In addition, fluorescence recovery after photobleaching experiments showed that fine patches of desmosomal proteins may participate in desmosome maintenance. Such a diverse range of dynamic activities of desmosomes apparently produces flexible but tight cell-cell adhesion required for different morphogenetic events in epithelial structures.This work was supported by grant AR44016-04 from the National Institutes of Health 相似文献
11.
Desmosomes are cell junctions and cytoskeleton-anchoring structures of epithelia, the myocardium, and dendritic reticulum cells of lymphatic follicles whose major components are known. Using cultured HT-1080 SL-1 fibrosarcoma-derived cells and transfection of cDNAs encoding specific desmosomal components, we have determined a minimum ensemble of proteins sufficient to introduce de novo structures, which, by morphology and functional competence, are indistinguishable from authentic desmosomes. In a more refined analysis, the influence of the desmosomal proteins desmoplakin (Dp), plakoglobin (Pg), and plakophilin 2 (Pp2) on the lateral clustering of the desmosomal transmembrane-glycoprotein desmoglein 2 (Dsg) was examined. We found that for efficient clustering of desmoglein 2 and desmosome structure formation, all three major plaque proteins-desmoplakin, plakoglobin, and plakophilin 2- were necessary. Furthermore, in this cell model, plakophilin 2 was capable of directing desmoplakin to adhaerens junctions (AJ), whereas plakoglobin was crucial for the segregation of desmosomal and AJ components. These results are discussed with respect to the variability in cell junction composition observed in various nonepithelial tissues. 相似文献
12.
Wahl JK 《Journal of cellular biochemistry》2005,96(2):390-403
Plakophilins (pkp-1, -2, and -3) comprise a family of armadillo-repeat containing proteins that are found in the desmosomal plaque and in the nucleus. Plakophilin-1 is most highly expressed in the suprabasal layers of the epidermis and loss of plakophilin-1 expression results in skin fragility-ectodermal dysplasia syndrome, which is characterized by a reduction in the number and size of desmosomes in the epithelia of affected individuals. To investigate the role of plakophilin-1 during desmosome formation, we fused plakophilin-1 to the hormone-binding domain of the estrogen receptor to create a fusion protein (plakophilin-1/ER) that can be activated in cell culture by the addition of 4-hydroxytamoxifen. When plakophilin-1/ER was expressed in A431 cells it was incorporated into endogenous desmosomes and did not disrupt desmosome formation. A derivative of A431 cells (A431D) do not form desmosomes, even though they express all the components believed to be necessary for desmosome assembly. Expression and activation of plakophilin-1/ER in A431D cells resulted in punctate desmoplakin staining on the cell surface. Co-expression of a classical cadherin (N-cadherin) and plakophilin-1/ER in A431D cells resulted in punctate desmoplakin staining at cell-cell borders. These data suggest that plakophilin-1 can induce assembly of desmosomal components in A431D cells in the absence of a classical cadherin; however a classical cadherin (N-cadherin) is required to direct assembly of desmosomes between adjacent cells. The activatable plakophilin-1/ER system provides a unique culture system to study the assembly of the desmosomal plaque in culture. 相似文献
13.
《Cell communication & adhesion》2013,20(2):125-138
In the present study, we have examined how modulation of protein kinase C (PKC) activity affected desmosome organization in HeLa cells. Immunofluorescence and electron microscopy showed that PKC activation upon short exposure to 12-O-tetradecanoylphorbol 13-acetate (TPA) resulted in a reduction of intercellular contacts, splitting of desmosomes and dislocation of desmosomal components from the cell periphery towards the cytoplasm. As determined by immunoblot analysis of Triton X-100-soluble and -insoluble pools of proteins, these morphological changes were not correlated with modifications in the extractability of both desmoglein and plakoglobin, but involved almost complete solubilization of the desmosomal plaque protein, desmoplakin. Immunoprecipitation experiments and immunoblotting with anti-phosphoserine, anti-phosphothreonine and anti-phosphotyrosine antibodies revealed that desmoplakin was mainly phosphorylated on serine and tyrosine residues in both treated and untreated cells. While phosphotyrosine content was not affected by PKC activation, phosphorylation on serine residues was increased by about two-fold. This enhanced serine phosphorylation coincided with the increase in the protein solubility, suggesting that phosphorylation of desmoplakin may be a mechanism by which PKC mediates desmosome disassembly. Consistent with the loss of PKC activity, we also showed that down-modulation of the kinase (in response to prolonged TPA treatment) or its specific inhibition (by GF109203X) had opposite effects and increased desmosome formation. Taken together, these results clearly demonstrate an important role for PKC in the regulation of desmosomal junctions in HeLa cells, and identify serine phosphorylation of desmoplakin as a crucial event in this pathway. 相似文献
14.
A P Kowalczyk M Hatzfeld E A Bornslaeger D S Kopp J E Borgwardt C M Corcoran A Settler K J Green 《The Journal of biological chemistry》1999,274(26):18145-18148
The contribution of desmosomes to epidermal integrity is evident in the inherited blistering disorder associated with the absence of a functional gene for plakophilin-1. To define the function of plakophilin-1 in desmosome assembly, interactions among the desmosomal cadherins, desmoplakin, and the armadillo family members plakoglobin and plakophilin-1 were examined. In transient expression assays, plakophilin-1 formed complexes with a desmoplakin amino-terminal domain and enhanced its recruitment to cell-cell borders; this recruitment was not dependent on the equimolar expression of desmosomal cadherins. In contrast to desmoplakin-plakoglobin interactions, the interaction between desmoplakin and plakophilin-1 was not mediated by the armadillo repeat domain of plakophilin-1 but by the non-armadillo head domain, as assessed by yeast two-hybrid and recruitment assays. We propose a model whereby plakoglobin serves as a linker between the cadherins and desmoplakin, whereas plakophilin-1 enhances lateral interactions between desmoplakin molecules. This model suggests that epidermal lesions in patients lacking plakophilin-1 are a consequence of the loss of integrity resulting from a decrease in binding sites for desmoplakin and intermediate filaments at desmosomes. 相似文献
15.
Xinyu Chen Stefan Bonne Mechthild Hatzfeld Frans van Roy Kathleen J Green 《The Journal of biological chemistry》2002,277(12):10512-10522
Plakophilins are a subfamily of p120-related arm-repeat proteins that can be found in both desmosomes and the nucleus. Among the three known plakophilin members, plakophilin 1 has been linked to a genetic skin disorder and shown to play important roles in desmosome assembly and organization. However, little is known about the binding partners and functions of the most widely expressed member, plakophilin 2. To better understand the cellular functions of plakophilin 2, we have examined its protein interactions with other junctional molecules using co-immunoprecipitation and yeast two-hybrid assays. Here we show that plakophilin 2 can interact directly with several desmosomal components, including desmoplakin, plakoglobin, desmoglein 1 and 2, and desmocollin 1a and 2a. The head domain of plakophilin 2 is critical for most of these interactions and is sufficient to direct plakophilin 2 to cell borders. In addition, plakophilin 2 is less efficient than plakophilin 1 in localizing to the nucleus and enhancing the recruitment of excess desmoplakin to cell borders in transiently transfected COS cells. Furthermore, plakophilin 2 is able to associate with beta-catenin through its head domain, and the expression of plakophilin 2 in SW480 cells up-regulates the endogenous beta-catenin/T cell factor-signaling activity. This up-regulation by plakophilin 2 is abolished by ectopic expression of E-cadherin, suggesting that these proteins compete for the same pool of signaling active beta-catenin. Our results demonstrate that plakophilin 2 interacts with a broader repertoire of desmosomal components than plakophilin 1 and provide new insight into the possible roles of plakophilin 2 in regulating the signaling activity of beta-catenin. 相似文献
16.
17.
Desmosomes are intercellular adhesive junctions of major importance for tissue integrity. To allow cell motility and migration they are down-regulated in epidermal wound healing. Electron microscopy indicates that whole desmosomes are internalised by cells in tissues, but the mechanism of down-regulation is unclear. In this paper we provide an overview of the internalisation of half-desmosomes by cultured cells induced by calcium chelation. Our results show that: (i) half desmosome internalisation is dependent on conventional PKC isoforms; (ii) microtubules transport internalised half desmosomes to the region of the centrosome by a kinesin-dependent mechanism; (iii) desmosomal proteins remain colocalised after internalisation and are not recycled to the cell surface; (iv) internalised desmosomes are degraded by the combined action of lysosomes and proteasomes. We also confirm that half desmosome internalisation is dependent upon the actin cytoskeleton. These results suggest that half desmosomes are not disassembled and recycled during or after internalisation but instead are transported to the centrosomal region where they are degraded. These findings may have significance for the down-regulation of desmosomes in wounds. 相似文献
18.
A constitutive transmembrane glycoprotein of Mr 165,000 (desmoglein) in epidermal and non-epidermal desmosomes. II. Immunolocalization and microinjection studies 总被引:15,自引:1,他引:14
Using two monoclonal antibodies described in the preceding paper we determined by immunofluorescence microscopy the distribution of an integral membrane protein of the desmosomal domain, the major glycopolypeptide of Mr 165,000 (bovine muzzle epidermal desmosome band 3; desmoglein) in various normal tissues, tumors and cultured cell lines from several mammalian species. This protein was detected in dotted or streak-like arrays along cell boundary structures which were known to contain non-membrane-integrated desmosomal plaque proteins such as desmoplakins. This is true for epithelial, i.e. cytokeratin-expressing cell types, for the desmin-producing myocardiac and Purkinje fiber cells of the heart, and for certain vimentin-containing cells such as arachnoidal and meningiomal cells and dendritic follicular cells of lymph nodes. However, on the basis of both immunoblot and immunocytochemical reactions, the protein is absent from non-desmosomal adhering junctions, including those devoid of desmoplakin but containing another plaque protein, plakoglobin ("band 5 protein"). We have used these antibodies to localize their epitopes with respect to the cell membrane. By immunoelectron microscopy we found that both epitopes are located in the desmosomal plaques, and this was confirmed by microinjection of purified antibodies into living cultured cells which resulted in labelling of the plaques. From these findings, taken together with previous analyses and localizations of the carbohydrate moieties of this glycoprotein, we conclude that desmoglein is a transmembrane glycoprotein which projects into--and contributes to--the desmosomal plaque structure. This glycoprotein represents a general component of true desmosomes and it is coexpressed with obligatory desmosome-specific plaque proteins such as desmoplakin I. The potential value of this glycoprotein as a desmosomal and cell type marker in histology and tumor diagnosis is discussed. 相似文献
19.
The desmosome junction is an important component in the cohesion of epithelial cells, especially epidermal keratinocytes. To gain insight into the structure and function of desmosomes, their morphogenesis has been studied in a primary mouse epidermal (PME) cell culture system. When these cells are grown in approximately 0.1 mM Ca2+, they contain no desmosomes. They are induced to form desmosomes when the Ca2+ level in the culture medium is raised to approximately 1.2 mM Ca2+. PME cells in medium containing low levels of Ca2+, and then processed for indirect immunofluorescence using antibodies directed against desmoplakins (desmosomal plaque proteins), display a pattern of discrete fluorescent spots concentrated mainly in the perinuclear region. Double label immunofluorescence using keratin and desmoplakin antibodies reveals that the desmoplakin-containing spots and the cytoplasmic network of tonofibrils (bundles of intermediate filaments [IFB]) are in the same juxtanuclear region. Within 1 h after the switch to higher levels of Ca2+, the spots move toward the cell surface, primarily to areas of cell-cell contact and not to free cell surfaces. This reorganization occurs at the same time that tonofibrils also move toward cell surfaces in contact with neighboring cells. Once the desmoplakin spots have reached the cell surface, they appear to aggregate to form desmosomes. These immunofluorescence observations have been confirmed by immunogold ultrastructural localization. Preliminary biochemical and immunological studies indicate that desmoplakin appears in whole cell protein extracts and in Triton high salt insoluble residues (i.e., cytoskeletal preparations consisting primarily of IFB) prepared from PME cells maintained in medium containing both low and normal Ca2+ levels. These findings show that certain desmosome components are preformed in the cytoplasm of PME cells. These components undergo a dramatic reorganization, which parallels the changes in IFB redistribution, upon induction of desmosome formation. The reorganization depends upon both the extracellular Ca2+ level and the establishment of cell-to-cell contacts. Furthermore, the data suggests that desmosomes do not act as organizing centers for the elaboration of IFB. Indeed, we postulate that the movement of IFB and preformed desmosomal components to the cell surface is an important initiating event in desmosome morphogenesis. 相似文献
20.
To characterize the desmosome components that mediate intercellular adhesion and cytoskeletal-plasma membrane attachment, we prepared whole desmosomes and isolated desmosomal intercellular regions (desmosomal "cores") from the living cell layers of bovine muzzle epidermis. The tissue was disrupted in a nonionic detergent at low pH, sonicated, and the insoluble residue fractionated by differential centrifugation and metrizamide gradient centrifugation. Transmission electron microscopic analyses reveal that a fraction obtained after differential centrifugation is greatly enriched in whole desmosomes that possess intracellular plaques. Metrizamide gradient centrifugation removes most of the plaque material, leaving the intercellular components and the adjoining plasma membranes. Sodium dodecyl sulfate polyacrylamide gel electrophoresis coupled with methods that reveal carbohydrate-containing moieties on gels demonstrate that certain proteins present in whole desmosomes are glycosylated. These glycoproteins are specifically and greatly enriched in the desmosome cores of which they are the principal protein constituents, and thus may function as the intercellular adhesive of the desmosome. 相似文献