Desmosomes are adhesive intercellular junctions prominent in the skin and heart. Loss of desmosome function is associated with severe congenital and acquired disorders characterized by tissue fragility. Pemphigus vulgaris (PV) is an autoimmune disorder in which antibodies are directed against the desmosomal adhesion molecule Dsg3, resulting in severe mucosal erosions and epidermal blistering. To define the mechanisms by which Dsg3 autoantibodies disrupt keratinocyte adhesion, the fate of PV IgG and various desmosomal components was monitored in primary human keratinocytes exposed to PV patient IgG. PV IgG initially bound to keratinocyte cell surfaces and colocalized with desmosomal markers. Within 6 h after PV IgG binding to Dsg3, electron microscopy revealed that desmosomes were dramatically disrupted and keratinocyte adhesion was severely compromised. Immunofluorescence analysis indicated that PV IgG and Dsg3 were rapidly internalized from the cell surface in a complex with plakoglobin but not desmoplakin. Dsg3 internalization was associated with retraction of keratin filaments from cell-cell borders. Furthermore, the internalized PV IgG-Dsg3 complex colocalized with markers for both endosomes and lysosomes, suggesting that Dsg3 was targeted for degradation. Consistent with this possibility, biotinylation experiments demonstrated that soluble Dsg3 cell surface pools were rapidly depleted followed by loss of detergent-insoluble Dsg3. These findings demonstrate that Dsg3 endocytosis, keratin filament retraction, and the loss of keratinocyte cell-cell adhesion are coordinated responses to PV IgG.Desmosomes are adhesive intercellular junctions that mediate tight adhesion between epithelial cells (1, 2). Desmosomes are particularly prominent in tissues that experience mechanical stress, such as the skin and heart, and function as plasma membrane attachment sites for intermediate filaments. The importance of desmosomes in tissue function and integrity has been revealed by numerous genetic and autoimmune disorders that impact desmosomal components (3, 4). The desmosomal cadherins, the desmogleins and desmocollins, are the transmembrane components of desmosomes responsible for mediating cell-cell adhesion (5). The tails of the desmosomal cadherins interact with the cytoplasmic protein plakoglobin along with other related proteins to couple the cadherins to desmoplakin and the intermediate filament cytoskeleton (2, 6). A number of desmoglein and desmocollin isoforms have been identified, and the genes encoding these proteins are expressed in a tissue-and differentiation-specific manner (5, 7). Mutations in genes encoding desmosomal components lead to heart and skin disorders (3,8,9). Similarly, autoantibodies directed against the desmosomal cadherins lead to a class of severe epidermal blistering disorders termed pemphigus (10). These disorders underscore the importance of understanding how desmosomes assemble, disassemble, and contribute to tissue architecture and function.The mechanisms by which intercellular junctio...

Pemphigus vulgaris (PV) is a life-threatening autoimmune disease characterized by oral mucosal erosions and epidermal blistering. The autoantibodies generated target the desmosomal cadherin desmoglein-3 (Dsg3). Previous studies demonstrate that upon PV IgG binding, Dsg3 is internalized and enters an endo-lysosomal pathway where it is degraded. To define the endocytic machinery involved in PV IgG-induced Dsg3 internalization, human keratinocytes were incubated with PV IgG, and various tools were used to perturb distinct endocytic pathways. The PV IgG⅐Dsg3 complex failed to colocalize with clathrin, and inhibitors of clathrin-and dynamin-dependent pathways had little or no effect on Dsg3 internalization. In contrast, cholesterol binding agents such as filipin and nystatin and the tyrosine kinase inhibitor genistein dramatically inhibited Dsg3 internalization. Furthermore, the Dsg3 cytoplasmic tail specified sensitivity to these inhibitors. Moreover, inhibition of Dsg3 endocytosis with genistein prevented disruption of desmosomes and loss of adhesion in the presence of PV IgG. Altogether, these results suggest that PV IgG-induced Dsg3 internalization is mediated through a clathrin-and dynamin-independent pathway and that Dsg3 endocytosis is tightly coupled to the pathogenic activity of PV IgG.Desmosomes are adhesive junctions that provide robust adhesion between epithelial cells (1, 2). These organelles are prominent in tissues that experience substantial mechanical stress such as the heart, bladder, gastrointestinal mucosa, and skin. Desmosomes are comprised primarily of proteins from three major families, the desmosomal cadherins desmogleins and desmocollins, armadillo proteins such as plakoglobin and the plakophilins, and members of the plakin family of cytolinkers such as desmoplakin (1-3). Together, these proteins contribute to tissue integrity by coupling adhesive interactions mediated by the desmosomal cadherins to the keratin intermediate filament cytoskeleton, thereby integrating adhesive and cytoskeletal networks throughout the cells in a tissue. Although critical for tissue integrity, desmosomes are often remodeled and contribute to dynamic processes during development and wound healing. Furthermore, desmosomal components may also play pivotal roles in keratinocyte differentiation, morphogenesis, and tissue patterning as well as epithelial-mesenchymal transitions (4, 5).Pemphigus vulgaris (PV) 2 is a potentially fatal autoimmune skin disease in which autoantibodies are generated against the desmosomal cadherin, desmoglein-3 (Dsg3) (6 -8). Dsg3, a 130-kDa glycoprotein, is found primarily in the spinous and basal layers of the epidermis and throughout the oral mucosa (9). As a result, PV is characterized histologically by suprabasal loss of cell-cell adhesion (acantholysis) and clinically by blistering of the skin and erosion of mucous membranes (7,8). A wide range of approaches have demonstrated that Dsg3 is the key target of PV IgG (10, 11). In addition, experimentally generated mice in which the Dsg3 ge...

Porphyromonas gingivalis is one of the major pathogens associated with adult periodontitis, a major chronic inflammatory disease. Potent proteinases elaborated by these bacteria aid directly and indirectly in both the development of the pathophysiology of the disease and in host defense evasion. For these reasons they are considered key virulence factors. To investigate whether possible immune evasion mechanisms involve the dysregulation of the host cytokine network, we examined the ability of P. gingivalis cysteine proteinases, including Arg-specific gingipains HRGP and RGP2 and Lys-specific KGP, to degrade the proinflammatory cytokine tumor necrosis factor-␣ (TNF-␣). All three gingipains rapidly degraded TNF-␣ as exhibited by immunoblot analysis. Moreover, all biological activity was significantly reduced over extended incubation periods with the proteinases tested, whereas the host neutrophil proteinases were ineffective. These results indicate that the gingipain proteinases elaborated by P. gingivalis are capable of disrupting the cytokine network at the site of infection through the degradation of the proinflammatory cytokine TNF-␣, suggesting the removal of one of several mediators important to the function of polymorphonuclear leukocytes. Such a mechanism is likely to be utilized by other infective organisms not only for survival but also for growth and proliferation.

Brain angiogenesis inhibitors (BAI) are putative transmembrane proteins containing an extracellular domain with thrombospondin type-1 repeats which can exhibit anti-angiogenic activity. BAI1 mRNA is expressed mainly in the brain, while BAI2 and BAI3 mRNAs are more widely expressed. We hypothesized that the BAI family might have anti-tumoral properties and studied the expression of BAI1 protein in normal human brain and in glioblastoma multiforme. We generated an anti-BAI1 antibody and showed that BAI1 was widely expressed in normal brain but was absent in 28 glioma cell lines and in the majority of human glioblastoma investigated. BAI1 expression did not correlate with TP53 status and we did not confirm previous findings that p53 regulates BAI1 mRNA expression in glioma cells. The finding that expression of BAI proteins may be lost during tumor formation is of special interest as restoration of their function in tumors may be of therapeutic benefit.

VE-cadherin is an endothelial-specific cadherin that plays a central role in vascular barrier function and angiogenesis. The cytoplasmic domain of VE-cadherin is linked to the cytoskeleton through interactions with the armadillo family proteins β-catenin and plakoglobin. Growing evidence indicates that β-catenin and plakoglobin play important roles in epithelial growth and morphogenesis. To test the role of these proteins in vascular cells, a replication-deficient retroviral system was used to express intercellular junction proteins and mutants in the human dermal microvascular endothelial cell line (HMEC-1). A mutant VE-cadherin lacking an adhesive extracellular domain disrupted endothelial barrier function and inhibited endothelial growth. In contrast, expression of exogenous plakoglobin or metabolically stable mutants of β-catenin stimulated HMEC-1 cell growth, which suggests that the β-catenin signaling pathway was active in HMEC-1 cells. This possibility was supported by the finding that a dominant-negative mutant of the transcription factor TCF-4, designed to inhibit β-catenin signaling, also inhibited HMEC-1 cell growth. These observations suggest that intercellular junction proteins function as components of an adhesion and signaling system that regulates vascular barrier function and growth.