Regulation of intestinal epithelial intercellular adhesion and barrier function by desmosomal cadherin desmocollin-2.

The role of desmosomal cadherin desmocollin-2 (Dsc2) in regulating barrier function in intestinal epithelial cells (IECs) is not well understood. Here, we report the consequences of silencing Dsc2 on IEC barrier function in vivo using mice with inducible intestinal-epithelial-specific Dsc2 knockdown (KD) (Dsc2ERΔIEC). While the small intestinal gross architecture was maintained, loss of epithelial Dsc2 influenced desmosomal plaque structure, which was smaller in size and had increased intermembrane space between adjacent epithelial cells. Functional analysis revealed that loss of Dsc2 increased intestinal permeability in vivo, supporting a role for Dsc2 in the regulation of intestinal epithelial barrier function. These results were corroborated in model human IECs in which Dsc2 KD resulted in decreased cell-cell adhesion and impaired barrier function. It is noteworthy that Dsc2 KD cells exhibited delayed recruitment of desmoglein-2 (Dsg2) to the plasma membrane after calcium switch-induced intercellular junction reassembly, while E-cadherin accumulation was unaffected. Mechanistically, loss of Dsc2 increased desmoplakin (DP I/II) protein expression and promoted intermediate filament interaction with DP I/II and was associated with enhanced tension on desmosomes as measured by a Dsg2-tension sensor. In conclusion, we provide new insights on Dsc2 regulation of mechanical tension, adhesion, and barrier function in IECs.

Pemphigus Vulgaris and Foliaceus IgG Autoantibodies Directly Block Heterophilic Transinteraction between Desmoglein and Desmocollin.

Anti-desmoglein (Dsg) 1 and Dsg3 IgG autoantibodies in pemphigus foliaceus and pemphigus vulgaris cause blisters through loss of desmosomal adhesion. It is controversial whether blister formation is due to direct inhibition of Dsg, intracellular signaling events causing desmosome destabilization, or both. Recent studies show that heterophilic binding between Dsg and desmocollin (Dsc) is the fundamental adhesive unit of desmosomes. To eliminate cellular contributions to potential pathogenicity of pemphigus antibodies, bead assays coated with recombinant Dsg1, Dsc1, Dsg3, or Dsc3 ectodomains were developed. A mixture of Dsg beads and Dsc beads formed large aggregates, confirming that the heterophilic binding is dominant. The pathogenic anti-Dsg1 and anti-Dsg3 mAbs, which bind the transadhesive interface, blocked the aggregation of Dsg1/Dsc1 and Dsg3/Dsc3 beads, respectively, whereas nonpathogenic mAbs did not. All sera tested from eight patients with pemphigus foliaceus and eight patients with mucosal pemphigus vulgaris with active disease inhibited the adhesion of Dsg1/Dsc1 and Dsg3/Dsc3 beads, respectively. When paired sera obtained from seven patients with pemphigus foliaceus and six patients with pemphigus vulgaris in active disease and remission were compared, the former inhibited aggregation better than the latter. These findings strongly suggest that steric hindrance of heterophilic transinteraction between Dsg and Dsc is important for disease pathology in both pemphigus foliaceus and pemphigus vulgaris.

Desmocollin­2 affects the adhesive strength and cytoskeletal arrangement in esophageal squamous cell carcinoma cells.

Desmocollin­2 (DSC2), a transmembrane glycoprotein belonging to the desmosomal cadherin family, has been found to be differentially expressed in several types of cancer and to be involved in tumor progression. The tumor metastasis suppressing property of DSC2 in esophageal squamous cell carcinoma (ESCC) has been described, however, its contribution to cell cohesion in ESCC remains to be elucidated. In the present study, using RNA interference (RNAi), the expression of DSC2 was silenced in SHEEC and KYSE510 cells. Hanging drop and fragmentation assays were performed to investigate the role of DSC2 in cell­cell adhesion. Western blot analysis and confocal microscopy were used to analyze the expression and localization of cell adhesion molecules and cytoskeletal arrangement. The results demonstrated that DSC2 knock down by RNAi caused defects in cell­cell adhesion and a concomitant reduction in desmosomal protein expression and adherens junction molecule distribution. A decrease in the expression of DSC2 caused an increase in free γ­catenin levels, thus promoting its recruitment to the adherens junction complex. In addition, the RNAi­mediated inhibition of DSC2 led to keratin intermediate filament retraction and filamentous­actin cytoskeleton rearrangement. Taken together, these data support our previous findings and the proposal that DSC2 may be involved in the regulation of the invasive behavior of cells by a mechanism that controls cell­cell attachment and cytoskeleton rearrangement.

The p53 target gene desmocollin 3 acts as a novel tumor suppressor through inhibiting EGFR/ERK pathway in human lung cancer.

Desmosomes are intercellular junctions that confer strong cell-cell adhesion. Altered expression of desmocollin 3 (DSC3), a member of the desmosomal cadherin family, was found in various cancers; however, its functional involvement in carcinogenesis has not yet been elucidated. Expression/localization of DSC3 was analyzed by real-time reverse transcription-PCR, western blotting, immunofluorescence and immunohistochemistry. Methylation status of DSC3 was examined by demethylation tests, methylation-specific PCR and bisulfite sequencing. It turned out that downregulation of DSC3 in lung cancer cells was associated with DNA hypermethylation. In primary lung tumors, DSC3 was a potential diagnostic marker for lung squamous cell carcinoma, and DSC3 DNA hypermethylation was correlated with poor clinical outcome. To investigate the effect of the tumor suppressor gene p53 on DSC3, transient transfection with a wild-type p53-expression vector was performed. Overexpression of p53 resulted in an increased expression of DSC3 in a DSC3-unmethylated lung cancer cell line H2170, but not in H1299, a DSC3-methylated cell line. However, combination of p53 transfection with demethylation agent 5-aza-2'-deoxycytidine treatment led to increased expression of DSC3 in H1299 cells. Furthermore, functional studies after stable transfection of a DSC3 expression vector showed that ectopic expression of DSC3 inhibited cell proliferation, anchorage-independent growth, migration, as well as invasion, and most interestingly led to reduced phosphorylation levels of extracellular signal-regulated kinase1/2. Taken together, our data suggested that DSC3 acts as a novel tumor suppressor gene through inhibition of epidermal growth factor receptor/extracellular signal-regulated kinase signaling in lung cancer cells.

Desmocollin 3-mediated binding is crucial for keratinocyte cohesion and is impaired in pemphigus.

Desmocollin (Dsc) 1-3 and desmoglein (Dsg) 1-4, transmembrane proteins of the cadherin family, form the adhesive core of desmosomes. Here we provide evidence that Dsc3 homo- and heterophilic trans-interaction is crucial for epidermal integrity. Single molecule atomic force microscopy (AFM) revealed homophilic trans-interaction of Dsc3. Dsc3 displayed heterophilic interaction with Dsg1 but not with Dsg3. A monoclonal antibody targeted against the extracellular domain reduced homophilic and heterophilic binding as measured by AFM, caused intraepidermal blistering in a model of human skin, and a loss of intercellular adhesion in cultured keratinocytes. Because autoantibodies against Dsg1 are associated with skin blistering in pemphigus, we characterized the role of Dsc3 binding for pemphigus pathogenesis. In contrast to AFM experiments, laser tweezer trapping revealed that pemphigus autoantibodies reduced binding of Dsc3-coated beads to the keratinocyte cell surface. These data indicate that loss of heterophilic Dsc3/Dsg1 binding may contribute to pemphigus skin blistering.