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Arrhythmogenic cardiomyopathy (AC) is a familial cardiac disease, mainly caused by mutations in desmosomal genes, which accounts for most cases of stress-related arrhythmic sudden death, in young and athletes. AC hearts display fibro-fatty lesions that generate the arrhythmic substrate and cause contractile dysfunction. A correlation between physical/emotional stresses and arrhythmias supports the involvement of sympathetic neurons (SNs) in the disease, but this has not been confirmed previously. Here, we combined molecular, in vitro and ex vivo analyses to determine the role of AC-linked DSG2 downregulation on SN biology and assess cardiac sympathetic innervation in desmoglein-2 mutant (Dsg2 mut/mut ) mice. Molecular assays showed that SNs express DSG2, implying that DSG2-mutation carriers would harbour the mutant protein in SNs. Confocal immunofluorescence of heart sections and 3-D reconstruction of SN network in clarified heart blocks revealed significant changes in the physiologialc SN topology, with massive hyperinnervation of the intact subepicardial layers and heterogeneous distribution of neurons in fibrotic areas. Cardiac SNs isolated from Dsg2 mut/mut neonatal mice, prior to the establishment of cardiac innervation, show alterations in axonal sprouting, process development and distribution of varicosities. Consistently, virus-assisted DSG2 downregulation replicated, in PC12-derived SNs, the phenotypic alterations displayed by Dsg2 mut/mut primary neurons, corroborating that AC-linked Dsg2 variants may affect SNs. Our results reveal that altered sympathetic innervation is an unrecognized feature of AC hearts, which may result from the combination of cell-autonomous and context-dependent factors implicated in myocardial remodelling. Our results favour the concept that AC is a disease of multiple cell types also hitting cardiac SNs. KEY POINTS: Arrhythmogenic cardiomyopathy is a genetically determined cardiac disease, which accounts for most cases of stress-related arrhythmic sudden death. Arrhythmogenic cardiomyopathy linked to mutations in desmoglein-2 (DSG2) is frequent and leads to a left-dominant form of the disease. Arrhythmogenic cardiomyopathy has been approached thus far as a disease of cardiomyocytes, but we here unveil that DSG2 is expressed, in addition to cardiomyocytes, by cardiac and extracardiac sympathetic neurons, although not organized into desmosomes. AC-linked DSG2 downregulation primarily affect sympathetic neurons, resulting in the significant increase in cardiac innervation density, accompanied by alterations in sympathetic neuron distribution. Our data supports the notion that AC develops with the contribution of several 'desmosomal protein-carrying' cell types and systems., (© 2024 The Author(s). The Journal of Physiology published by John Wiley & Sons Ltd on behalf of The Physiological Society.)

Proteomics studies often explore phenotypic differences between whole organs and systems. Within the heart, more subtle variation exists. To date, differences in the underlying proteome are only described between whole cardiac chambers. This study, using the bovine heart as a model, investigates inter-regional differences and assesses the feasibility of measuring detailed, cross-tissue variance in the cardiac proteome. Using a bovine heart, we created a two-dimensional section through a plane going through two chambers. This plane was further sectioned into 4 × 4 mm cubes and analysed using label-free proteomics. We identified three distinct proteomes. When mapped to the extracted sections, the proteomes corresponded largely to the outer wall of the right ventricle and secondly to the outer wall of the left ventricle, right atrial appendage, tricuspid and mitral valves, modulator band, and parts of the left atrium. The third separate proteome corresponded to the inner walls of the left and right ventricles, septum, and left atrial appendage. Differential protein abundancies indicated differences in energy metabolism between regions. Data analyses of the mitochondrial proteins revealed a variable pattern of abundances of complexes I-V between the proteomes, indicating differences in the bioenergetics of the different cardiac sub-proteomes. Mapping of disease-associated proteins interestingly showed desmoglein-2, for which defects in this protein are known to cause Arrhythmogenic Right Ventricular Dysplasia/Cardiomyopathy, which was present predominantly in the outer wall of the left ventricle. This study highlights that organs can have variable proteomes that do not necessarily correspond to anatomical features.

Arrhythmogenic cardiomyopathy (ACM) is a familial heart disease characterized by cardiac dysfunction, arrhythmias, and myocardial inflammation. Exercise and stress can influence the disease's progression. Thus, an investigation of whether a high-fat diet (HFD) contributes to ACM pathogenesis is warranted. In a robust ACM mouse model, 8-week-old Desmoglein-2 mutant ( Dsg2 mut/mut ) mice were fed either an HFD or rodent chow for 8 weeks. Chow-fed wildtype (WT) mice served as controls. Echo- and electrocardiography images pre- and post-dietary intervention were obtained, and the lipid burden, inflammatory markers, and myocardial fibrosis were assessed at the study endpoint. HFD-fed Dsg2 mut/mut mice showed numerous P-wave perturbations, reduced R-amplitude, left ventricle (LV) remodeling, and reduced ejection fraction (%LVEF). Notable elevations in plasma high-density lipoprotein (HDL) were observed, which correlated with the %LVEF. The myocardial inflammatory adipokines, adiponectin (AdipoQ) and fibroblast growth factor-1, were substantially elevated in HFD-fed Dsg2 mut/mut mice, albeit no compounding effect was observed in cardiac fibrosis. The HFD not only potentiated cardiac dysfunction but additionally promoted adverse cardiac remodeling. Further investigation is warranted, particularly given elevated AdipoQ levels and the positive correlation of HDL with the %LVEF, which may suggest a protective effect. Altogether, the HFD worsened some, but not all, disease phenotypes in Dsg2 mut/mut mice. Notwithstanding, diet may be a modifiable environmental factor in ACM disease progression.

Desmoglein-2 (DSG2) has emerged as a potential biomarker for coronavirus disease 2019 (COVID-19) complications, particularly cardiac and cardiovascular involvement. The expression of DSG2 in lung tissues has been detected at elevated levels, and circulating DSG2 levels correlate with COVID-19 severity. DSG2 may contribute to myocardial injury, cardiac dysfunction and vascular endothelial dysfunction in COVID-19. Monitoring DSG2 levels could aid in risk stratification, early detection and prognostication of COVID-19 complications. However, further research is required to validate DSG2 as a biomarker. Such research will aim to elucidate its precise role in pathogenesis, establishing standardized assays for its measurement and possibly identifying therapeutic targets.

Background: Arrhythmogenic cardiomyopathy (ACM) is a genetic heart muscle disease characterized by progressive fibro-fatty replacement of cardiac myocytes. Up to now, the existing therapeutic modalities for ACM are mostly palliative. About 50% of ACM is caused by mutations in genes encoding desmosomal proteins including Desmoglein-2 (Dsg2). In the current study, the cardiac fibrosis of ACM and its underlying mechanism were investigated by using a cardiac-specific knockout of Dsg2 mouse model., Methods: Cardiac-specific Dsg2 knockout (CS-Dsg2 -/- ) mice and wild-type (WT) mice were respectively used as the animal model of ACM and controls. The myocardial collagen volume fraction was determined by histological analysis. The expression levels of fibrotic markers such as α-SMA and Collagen I as well as signal transducers such as STAT3, SMAD3, and PPARα were measured by Western blot and quantitative real-time PCR., Results: Increased cardiac fibrosis was observed in CS-Dsg2 -/- mice according to Masson staining. PPARα deficiency and hyperactivation of STAT3 and SMAD3 were observed in the myocardium of CS-Dsg2 -/- mice. The biomarkers of fibrosis such as α-SMA and Collagen I were upregulated after gene silencing of Dsg2 in HL-1 cells. Furthermore, STAT3 gene silencing by Stat3 siRNA inhibited the expression of fibrotic markers. The activation of PPARα by fenofibrate or AAV9-Pparα improved the cardiac fibrosis and decreased the phosphorylation of STAT3, SMAD3, and AKT in CS-Dsg2 -/- mice., Conclusions: Activation of PPARα alleviates the cardiac fibrosis in ACM.

As epithelial cells in the circulation are considered to originate from the tumor, the epithelial cell adhesion molecule has been commonly used as a standard marker for circulating tumor cells (CTCs) isolation. However, it seems to disappear after the epithelial-mesenchymal transition that most cancer cells undergo for intravasation. Thus, more advanced techniques for CTC detection are needed to better understand the clinical significance of CTCs. A cancer cell-specifically-infecting or replicating virus that codes a fluorescent monitor gene can be a solution to efficiently detect CTCs. Thus, the authors designed an adenovirus to bind to desmoglein-2, which is highly expressed in most cancer cells. A cancer-specific human telomerase reverse transcriptase promoter is inserted to control a viral E1 region. The adenovirus is utilized to compare the number of CTCs from renal cell carcinoma and prostate cancer patients before and after surgery. The isolated two or three CTCs are eligible for whole genome sequencing. The genomic analysis proves the difference of variants between primary tumors and CTCs. Taken together, it is a fast and exact serial method for CTC isolation and the enriched genome sequencing may be used to determine the prognosis and as a point-of-care system for patients with cancer., (© 2022 Wiley-VCH GmbH.)

Multiple myeloma (MM) is the second most common haematological malignancy and is an incurable disease of neoplastic plasma cells (PC). Newly diagnosed MM patients currently undergo lengthy genetic testing to match chromosomal mutations with the most potent drug/s to decelerate disease progression. With only 17% of MM patients surviving 10-years postdiagnosis, faster detection and earlier intervention would unequivocally improve outcomes. Here, we show that the cell surface protein desmoglein-2 (DSG2) is overexpressed in ~ 20% of bone marrow biopsies from newly diagnosed MM patients. Importantly, DSG2 expression was strongly predictive of poor clinical outcome, with patients expressing DSG2 above the 70 th percentile exhibiting an almost 3-fold increased risk of death. As a prognostic factor, DSG2 is independent of genetic subtype as well as the routinely measured biomarkers of MM activity (e.g. paraprotein). Functional studies revealed a nonredundant role for DSG2 in adhesion of MM PC to endothelial cells. Together, our studies suggest DSG2 to be a potential cell surface biomarker that can be readily detected by flow cytometry to rapidly predict disease trajectory at the time of diagnosis., (© 2021 The Authors. Molecular Oncology published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.)

Background: High temperature requirement A (HtrA) is an active serine protease secreted by the group-I carcinogen Helicobacter pylori (H. pylori). The human cell adhesion protein and tumor suppressor E-cadherin (hCdh1) expressed on the surface of gastric epithelial cells was identified as the first HtrA substrate. HtrA-mediated hCdh1 cleavage and subsequent disruption of intercellular adhesions are considered as important steps in H. pylori pathogenesis. In this study, we performed a proteomic profiling of H. pylori HtrA (HpHtrA) to decipher the complex mechanism of H. pylori interference with the epithelial barrier integrity., Results: Using a proteomic approach we identified human desmoglein-2 (hDsg2), neuropilin-1, ephrin-B2, and semaphorin-4D as novel extracellular HpHtrA substrates and confirmed the well characterized target hCdh1. HpHtrA-mediated hDsg2 cleavage was further analyzed by in vitro cleavage assays using recombinant proteins. In infection experiments, we demonstrated hDsg2 shedding from H. pylori-colonized MKN28 and NCI-N87 cells independently of pathogen-induced matrix-metalloproteases or ADAM10 and ADAM17., Conclusions: Characterizing the substrate specificity of HpHtrA revealed efficient hDsg2 cleavage underlining the importance of HpHtrA in opening intercellular junctions. Video Abstract., (© 2021. The Author(s).)

Arrhythmogenic right ventricular cardiomyopathy (ARVC) is a progressive heart condition which causes fibro-fatty myocardial scarring, ventricular arrhythmias, and sudden cardiac death. Most cases of ARVC can be linked to pathogenic mutations in the cardiac desmosome, but the pathophysiology is not well understood, particularly in early phases when arrhythmias can develop prior to structural changes. Here, we created a novel human induced pluripotent stem cell-derived cardiomyocyte (hiPSC-CM) model of ARVC from a patient with a c.2358delA variant in desmoglein-2 ( DSG2 ). These DSG2 -mutant ( DSG2 Mut mRNA and had altered localization of desmoglein-2 protein alongside thinner, more disorganized myofibrils. No major changes in other desmosomal proteins were noted. There was increased pro-inflammatory cytokine expression that may be linked to canonical and non-canonical NFκB signaling. Action potentials in DSG2 mRNA and had altered localization of desmoglein-2 protein alongside thinner, more disorganized myofibrils. No major changes in other desmosomal proteins were noted. There was increased pro-inflammatory cytokine expression that may be linked to canonical and non-canonical NFκB signaling. Action potentials in DSG2 Mut CMs were shorter with increased upstroke heterogeneity, while time-to-peak calcium and calcium decay rate were reduced. These were accompanied by changes in ion channel and calcium handling gene expression. Lastly, suppressing DSG2 in control lines via siRNA allowed partial recapitulation of electrical anomalies noted in DSG2 Mut cells. In conclusion, the aberrant cytoskeletal organization, cytokine expression, and electrophysiology found DSG2 Mut hiPSC-CMs could underlie early mechanisms of disease manifestation in ARVC patients.

Introduction: X-linked PLAC1 is highly expressed in placenta during embryogenesis, and when ablated in mice, causes aberrant placental cell layer organization. It is also highly expressed in many types of cancer cell-lines. Although it has been shown that it promotes AKT phosphorylation in cancer cells, the exact mechanism by which it influences placental layer differentiation is unclear., Methods: To investigate the mechanism of action of PLAC1 we did cell fractionation and immunoprecipitation of the protein and Mass Spectrometry analysis to identify its interaction partners. The associated proteins were directly tested for interactions by co-transfection with PLAC1 and immunoprecipitation. Mutations in the ZP-N domain of PLAC1 were introduced to assess its involvement in the interactions., Results: We provide evidence that Desmoglein-2 (DSG2), a component of the membrane-associated desmosomal complex, directly interacts with PLAC1. Mutations of cysteines in ZP-N domain disrupt the interaction between PLAC1 and DSG-2., Discussion: Because desmosomes are responsible for establishing lateral cell-cell junctions, we suggest that direct interaction with the lateral junction protein complex may be implicated in the PLAC1 effects on cell-cell interactions, and thereby on the layer structure of the placenta., (Published by Elsevier Ltd.)

Arrhythmogenic cardiomyopathy (AC) is a familial cardiac disorder at high risk of arrhythmic sudden death in the young and athletes. AC is hallmarked by myocardial replacement with fibro-fatty tissue, favoring life-threatening cardiac arrhythmias and contractile dysfunction. The AC pathogenesis is unclear, and the disease urgently needs mechanism-driven therapies. Current AC research is mainly focused on 'desmosome-carrying' cardiomyocytes, but desmosomal proteins are also expressed by non-myocyte cells, which also harbor AC variants, including mesenchymal stromal cells (MSCs). Consistently, cardiac-MSCs contribute to adipose tissue in human AC hearts. We thus approached AC as a multicellular disorder, hypothesizing that it also affects extra-cardiac bone marrow (BM)-MSCs. Our results show changes in the desmosomal protein profile of both cardiac- and BM- MSCs, from desmoglein-2 ( Dsg2 )-mutant mice, accompanied with profound alterations in cytoskeletal organization, which are directly caused by AC-linked DSG2 downregulation. In addition, AC BM-MSCs display increased proliferation rate, both in vitro and in vivo, and, by using the principle of the competition homing assay, we demonstrated that mutant circulating BM-MSCs have increased propensity to migrate to the AC heart. Taken altogether, our results indicate that cardiac- and BM- MSCs are additional cell types affected in Dsg2 -linked AC, warranting the novel classification of AC as a multicellular and multiorgan disease.

About 50% of patients with arrhythmogenic cardiomyopathy (ACM) carry a pathogenic or likely pathogenic mutation in the desmosomal genes. However, there is a significant number of patients without positive familial anamnesis. Therefore, the molecular reasons for ACM in these patients are frequently unknown and a genetic contribution might be underestimated. Here, we used a next-generation sequencing (NGS) approach and in addition single nucleotide polymor-phism (SNP) arrays for the genetic analysis of two independent index patients without familial medical history. Of note, this genetic strategy revealed a homozygous splice site mutation (DSG2-c.378+1G>T) in the first patient and a nonsense mutation (DSG2-p.L772X) in combination with a large deletion in DSG2 in the second one. In conclusion, a recessive inheritance pattern is likely for both cases, which might contribute to the hidden medical history in both families. This is the first report about these novel loss-of-function mutations in DSG2 that have not been previously identi-fied. Therefore, we suggest performing deep genetic analyses using NGS in combination with SNP arrays also for ACM index patients without obvious familial medical history. In the future, this finding might has relevance for the genetic counseling of similar cases.

Physiological stressors, such as exercise, can precipitate sudden cardiac death or heart failure progression in patients with arrhythmogenic cardiomyopathy (ACM). Yet, whether and to what extent a highly prevalent and more elusive environmental factor, such as psychosocial stress (PSS), can also increase ACM disease progression is unexplored. Here, we first quantified perceived stress levels in patients with ACM and found these levels correlated with the extent of arrhythmias and cardiac dysfunction. To determine whether the observed correlation is due to causation, we inflicted PSS-via the resident-intruder (RI) paradigm-upon Desmoglein-2 mutant mice, a vigorously used mammalian model of ACM. We found that ACM mice succumbed to abnormally high in-trial, PSS mortality. Conversely, no sudden deaths occurred in wildtype (WT) counterparts. Desmoglein-2 mice that survived RI challenge manifested markedly worse cardiac dysfunction and remodeling, namely apoptosis and fibrosis. Furthermore, WT and ACM mice displayed similar behavior at baseline, but Desmoglein-2 mice exhibited heightened anxiety following RI-induced PSS. This outcome correlated with the worsening of cardiac phenotypes. Our mouse model demonstrates that in ACM-like subjects, PSS is incisive enough to deteriorate cardiac structure and function per se, i.e., in the absence of any pre-existing anxious behavior. Hence, PSS may represent a previously underappreciated risk factor in ACM disease penetrance.

Introduction: Coxsackievirus/adenovirus receptors (CARs) and desmoglein-2 (DSG2) are similar molecules to adenovirus-based vectors in the cell membrane. They have been found to be associated with lung epithelial cell tumorigenesis and can be useful markers in predicting survival outcome in lung adenocarcinoma (LUAD)., Methods: A gene ontology enrichment analysis disclosed that DSG2 was highly correlated with CAR. Survival analysis was then performed on 262 samples from the Cancer Genome Atlas, forming "Stage 1A" or "Stage 1B". We therefore analyzed a tissue microarray (TMA) comprised of 108 lung samples and an immunohistochemical assay. Computer counting software was used to calculate the H-score of the immune intensity. Cox regression and Kaplan-Meier analyses were used to determine the prognostic value., Results: CAR and DSG2 genes are highly co-expressed in early stage LUAD and associated with significantly poorer survival ( p = 0.0046). TMA also showed that CAR/DSG2 expressions were altered in lung cancer tissue. CAR in the TMA was correlated with proliferation, apoptosis, and epithelial-mesenchymal transition (EMT), while DSG2 was associated with proliferation only. The Kaplan-Meier survival analysis revealed that CAR, DSG2, or a co-expression of CAR/DSG2 was associated with poorer overall survival., Conclusions: The co-expression of CAR/DSG2 predicted a worse overall survival in LUAD. CAR combined with DSG2 expression can predict prognosis.

The self-renewal properties of human pluripotent stem cells (hPSCs) contribute to their efficacy in tissue regeneration applications yet increase the likelihood of teratoma formation, thereby limiting their clinical utility. To address this issue, we developed a tool to specifically target and neutralize undifferentiated hPSCs, thereby minimizing tumorigenicity risk without negatively affecting regenerated and somatic tissues. Specifically, we conjugated a monoclonal antibody (K6-1) previously generated in our laboratory against desmoglein 2 (Dsg2), which is highly differentially expressed in undifferentiated hPSCs versus somatic tissues, to the chemotherapeutic agent doxorubicin (DOX). The K6-1-DOX conjugates were selectively targeted and incorporated into Dsg2-positive hPSCs, leading to pH-dependent endosomal release and nuclear localization of DOX with subsequent cytotoxicity via an apoptotic caspase cascade. Conversely, Dsg2-negative fibroblasts showed minimal conjugate uptake or cytotoxicity, suggesting that K6-1-DOX treatment would yield few side effects owing to off-target effects. Selective removal of undifferentiated stem cells was also supported by in vivo studies using a mouse xenograft model, wherein hIgG-DOX- but not K6-1-DOX-pretreated-hPSC injection led to teratoma development. Together, these results validated the ability of the Dsg2-targeted antibody-anticancer drug conjugate to facilitate the safety of stem cell therapies., (Copyright © 2020. Published by Elsevier Ltd.)

The study of viruses causing acute respiratory distress syndromes (ARDS) is more essential than ever at a time when a virus can create a global pandemic in a matter of weeks. Among human adenoviruses, adenovirus of serotype 7 (HAdV7) is one of the most virulent serotypes. This virus regularly re-emerges in Asia and has just been the cause of several deaths in the United States. A critical step of the virus life cycle is the attachment of the knob domain of the fiber (HAd7K) to the cellular receptor desmoglein-2 (DSG2). Complexes between the fiber knob and two extracellular domains of DSG2 have been produced. Their characterization by biochemical and biophysical methods show that these two domains are sufficient for the interaction and that the trimeric HAd7K could accommodate up to three DSG2 receptor molecules. The cryo-electron microscopy (cryo-EM) structure of these complexes at 3.1 Å resolution confirmed the biochemical data, and allowed the identification of the critical amino acid residues for this interaction, which shows similarities with other DSG2 interacting adenoviruses, despite a low homology in the primary sequences.

Despite new therapies, the estimated 229 875 women living with ovarian cancer have a 5-year survival rate of 47.6%. This cavity-localized cancer lends itself to local administration of modalities, such as the oncolytic adenovirus (Ad) Ad5/3-D24-granulocyte-macrophage colony-stimulating factor virus (ONCOS-102). Its repeated administration to a patient with chemotherapy-refractory ovarian cancer induced CD8+ antitumor immune responses with the overall survival reaching 40 months. Here we probe the dominant receptor used by ONCOS-102 in four established epithelial ovarian cancer cell lines. Ad3 can use the desmoglein-2 (DSG2) and CD46 receptors on susceptible cells. DSG2 was nearly absent in A2780 cells but was expressed in more than 90% of OAW42, OVCAR3, and OV-90 cells. After 96 hours, ONCOS-102 treatment showed significant oncolytic activity (≧50%) in OAW42, OVCAR3, and OV-90 cells, but minimal activity in A2780 cells, suggesting DSG2 as the dominant receptor for ONCOS-102. Furthermore, retrospective analyses of phase I clinical trial of ONCOS-102 treatment of 12 patients with varied tumors indicated a correlation between viral genomes in blood and DSG2 RNA expression. These data support the role of DSG2 expression on cancer cells in virus infectivity and the continued development of ONCOS-102 for ovarian cancer treatment., (© 2020 The Authors. Journal of Medical Virology Published by Wiley Periodicals, Inc.)

Background: The pathogenesis and developmental mechanism of early-stage (FIGO 2009 IA2-IIA2) cervical cancer (CC) remain unclear. Seeking novel molecular biomarkers based on The Cancer Genome Atlas (TCGA) will facilitate the understanding of CC pathogenesis and help evaluate early-stage CC prognosis., Methods: To identify prognosis-related genes in early-stage CC, we analyzed TCGA mRNA-seq data and clinical data by univariate Cox and Kaplan-Meier plotter analyses. Differential expression analysis identified upregulated genes in early-stage CC. Combined with the genes correlated with unfavorable prognosis, we selected desmoglein-2 (DSG2) for further investigation. To detect DSG2 expression in early-stage CC, we used immunohistochemistry (IHC), quantitative real-time PCR (qRT-PCR) and western blotting. The relationship between the expression of DSG2 and clinical features was analyzed by the Chi square test. Cox analysis was applied to assess the relationship between CC overall survival (OS) and risk factors. The correlations between DSG2 expression and CC cell line proliferation and migration were investigated with Cell Counting Kit-8 (CCK-8) and migration assays., Results: There were 416 prognosis-related genes in early-stage CC. DSG2, matrix metallopeptidase 1 (MMP1), carbonic anhydrase IX (CA9), homeobox A1 (HOXA1), and serine protease inhibitor B3 (SERPINB3) were upregulated in early-stage CC compared with adjacent noncancerous tissue (ANT) and correlated with unfavorable prognosis. Among them, DSG2 was most significantly correlated with patient survival. Coexpression analysis indicated that DSG2 was probably involved in cell division, positive regulation of transferase activity, positive regulation of cell migration, EGFR upregulation pathway and regulation of lymphangiogenesis. IHC, qRT-PCR and western blotting showed that DSG2 expression was higher in CC than in normal tissue. Significant correlations were identified between DSG2 expression and several aggressive clinical features, including pelvic lymph node metastasis (PLNM). Multivariate Cox analysis showed that DSG2 and PLNM were independent prognostic factors for OS. DSG2 knockdown inhibited CC cell proliferation and migration., Conclusions: DSG2 is a biomarker that promotes tumor proliferation and metastasis and is correlated with poor prognosis in early-stage CC., Competing Interests: Competing interestsThe authors declare that they have no competing of interest., (© The Author(s) 2020.)

Coxsackievirus B3 (CVB3) is the primary etiologic agent of viral myocarditis, a major heart disease that occurs predominantly in children and young adolescents. In the heart, intercalated disks (ICD) are important structural formations that connect adjacent cardiomyocytes to maintain cardiac architecture and mediate signal communication. Deficiency in ICD components, such as desmosome proteins, leads to heart dysfunction. γ-catenin, a component protein of desmosomes, normally binds directly to desmocollin-2 and desmoglein-2. In this study, we found that CVB3 infection downregulated γ-catenin at the protein level but not the mRNA level in mouse HL-1 cardiomyocytes. We further found that this reduction of γ-catenin protein is a result of ubiquitin proteasome-mediated degradation, since the addition of proteasome inhibitor MG132 inhibited γ-catenin downregulation. In addition, we found that desmocollin-2 and desmoglein-2 were cleaved by both viral protease 3C and virus-activated cellular caspase, respectively. These cleavages led to the release of bound γ-catenin from the desmosome into the cytosol, resulting in rapid degradation of γ-catenin. Since γ-catenin shares high sequence homology with β-catenin in binding the TCF/LEF transcription factor, we further studied the effect of γ-catenin degradation on Wnt/β-catenin signaling. Luciferase assay showed that γ-catenin expression inhibited Wnt/β-catenin signaling. This finding was substantiated by qPCR to show that overexpression of γ-catenin downregulated transcription of Wnt signal target genes, c-myc and MMP9, while silencing γ-catenin upregulated these target genes. Finally, we demonstrated that γ-catenin expression inhibited CVB3 replication. In search for the underlying mechanism, we found that silencing γ-catenin caused down-regulation of interferon-β and its stimulated antiviral genes MDA5, MAVS, and ISG15. Taken together, our results indicate, for the first time, that CVB3 infection causes cardiomyocyte death through, at least in part, direct damage to the desmosome structure and reduction of γ-catenin protein, which in return promotes Wnt/β-catenin signaling and downregulates interferon-β stimulated immune responses., (Copyright © 2020 Zhao, Zhang, Qiu, Ye and Yang.)

Desmoglein-2 (DSG2) is an integral component of desmosomes, maintaining cell-cell adhension in multiple cancers. It has been well studied in epithelial cells, cardiomyocytes and primary prostate cancer, colon cancer, skin squamous cell carcinoma, except for cervical cancer. Hence, we performed this study to examine the function of DSG2 in cervical cancer. We used TCGA and Oncomine databases to assess the expression level of DSG2 in cervical cancer cases. Kaplan-Meier method with log-rank test was utilized to plot overall survival (OS) curve. The reverse transcription-quantitative polymerase chain reaction (qRT-PCR) and western blotting were performed to detect the expression of DSG2 in cells. Cell Counting Kit-8 (CCK-8), wound-healing analysis, and transwell assay were carried out to examine proliferation, migration, and invasion of cells. A higher level of DSG2 in cervical cancer was associated with lower OS rate. Knockdown of DSG2 inhibited cervical cancer cell proliferation, migration, and invasion, while DSG2 enhancement promoted cell proliferation, migration, and invasion. Moreover, the proteins expression of p-MEK and p-ERK that are required for mitogen-activated protein kinases (MAPK) pathway were downregulated after reducing DSG2. In conclusion, these findings illustrated the importance of DSG2 in cervical cancer development and cell behaviors by mediating MAPK signaling pathway, suggesting DSG2 maybe a novel therapeutic target in control of cervical cancer., (© 2020 The Authors. The Kaohsiung Journal of Medical Sciences published by John Wiley & Sons Australia on behalf of Kaohsiung Medical University.)

Aims: Desmoglein-2 (DSG2) mutations, which encode a heart-specific cadherin crucial for desmosomal adhesion, are frequent in arrhythmogenic right ventricular cardiomyopathy/dysplasia (ARVC/D). DSG2 mutations have been associated with higher risk of biventricular involvement. Among DSG2 mutations, mutations of the inhibitory propeptide consensus cleavage-site (Arg-X-Arg/Lys-Arg), are particularly frequent. In the present work, we explored the functional consequences of DSG2 propeptide cleavage site mutations p.Arg49His, p.Arg46Trp, and p.Arg46Gln on localization, adhesive properties, and desmosome incorporation of DSG2., Methods and Results: We studied the expression of mutant-DSG2 in human heart and in epithelial and cardiac cellular models expressing wild-type or mutant (p.Arg49His, p.Arg46Trp, and p.Arg46Gln) proDSG2-GFP fusion proteins. The consequences of the p.Arg46Trp mutation on DSG2 adhesiveness were studied by surface plasmon resonance. Incorporation of mutant p.Arg46Trp DSG2 into desmosomes was studied under low-calcium culture conditions and cyclic mechanical stress. We demonstrated in human heart and cellular models that all three mutations prevented N-terminal propeptide cleavage, but did not modify intercellular junction targeting. Surface plasmon resonance experiments showed a propeptide-dependent loss of interaction between the cadherin N-terminal extracellular 1 (EC1) domains. Additionally, proDSG2 mutant proteins were abnormally incorporated into desmosomes under low-calcium culture conditions or following mechanical stress. This was accompanied by an epidermal growth factor receptor-dependent internalization of proDSG2, suggesting increased turnover of unprocessed proDSG2., Conclusion: Our results strongly suggest weakened desmosomal adhesiveness due to abnormal incorporation of uncleaved mutant proDSG2 in cellular stress conditions. These results provide new insights into desmosomal cadherin regulation and ARVC/D pathophysiology, in particular, the potential role of mechanical stress on desmosomal dysfunction., (Published on behalf of the European Society of Cardiology. All rights reserved. © The Author(s) 2019. For permissions, please email: journals.permissions@oup.com.)

Background: Arrhythmogenic right ventricular cardiomyopathy (ARVC) is a rare cardiac disease predominantly caused by variants in desmosome genes. Variants in human Desmoglein-2 (DSG2) gene can cause ARVC with incomplete penetrance. However, it remains unknown whether ARVC would penetrate by distal myopathy., Methods: We performed targeted next-generation sequencing using a cardiomyopathy/ionchannelopathy panel in a Chinese ARVC pedigree. Plasmids of DSG2 were constructed, and pathogenicity of the DSG2 variant was investigated by real-time PCR, western blots, Immunofluorescence, and electron microscope., Results: We identified a novel nonsense variant in DSG2 (c.710T > A, p.Leu237Ter) and a reported pathogenic missense variant of distal myopathy in MYH7 (c. 1322C > T, p.Thr441Met) in the proband of an ARVC pedigree. The functional analyses suggested that the nonsense variant could affect the expression and cell location of DSG2, and the number and shape of desmosomes were affected as well, indicating that the variant was implicated in the pathogenesis of ARVC. We found only patients carrying the distal myopathy pathogenic variant would manifested early-onset severe ARVC phenotype, which suggested that MYH7-p.Thr441Met variant could induced the onset of ARVC in the DSG2-p.Leu237Ter variant carriers., Conclusions: Our study identified a novel null variant in DSG2 gene (c.710T > A, p.Leu237Ter) in an ARVC pedigree with incomplete penetrance. Only patients who carried a distal myopathy associated variant in MYH7 (c. 1322C > T, p.Thr441Met) would induce the onset of ARVC with early-onset and severe symptoms., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Desmogleins (Dsg2) are the major components of desmosomes. Dsg2 has five extracellular tandem cadherin domains (EC1-EC5) for cell-cell interaction. We had previously confirmed the Dsg2 antibody and its epitope (named KC21) derived from EC2 domain suppressing epithelial-mesenchymal transition and invasion in human cancer cell lines. Here, we screened six peptide fragments derived from EC2 domain and found that KR20, the parental peptide of KC21, was the most potent one on suppressing endothelial colony-forming cell (ECFC) tube-like structure formation. KC21 peptide also attenuated migration but did not disrupt viability and proliferation of ECFCs, consistent with the function to inhibit VEGF-mediated activation of p38 MAPK but not AKT and ERK. Animal studies showed that KC21 peptides suppressed capillary growth in Matrigel implant assay and inhibited oxygen-induced retinal neovascularization. The effects were comparable to bevacizumab (Bev). In conclusion, KC21 peptide is an angiogenic inhibitor potentially useful for treating angiogenesis-related diseases.

Background: Previous studies suggested that genetic status affects the clinical course of arrhythmogenic right ventricular cardiomyopathy/dysplasia (ARVC/D) patients. The aim of this study was to compare the outcome of desmoglein-2 (DSG2) mutation carriers to those who carry the plakophilin-2 (PKP2) mutation, the most common ARVC/D-associated gene., Methods and Results: Consecutive ARVC/D patients carrying a pathogenic mutation in PKP2 or DSG2 were selected from a national ARVC/D registry. The cumulative freedom from sustained ventricular arrhythmia and cardiac transplantation/death from heart failure (HF) during follow-up was assessed, compared between PKP2 and DSG2, and predictors for ventricular arrhythmia and HF events determined. Overall, 118 patients from 78 families were included: 27 (23%) carried a DSG2 mutation and 91 (77%) a PKP2 mutation. There were no significant differences between DSG2 and PKP2 mutation carriers concerning gender, proband status, age at diagnosis, T-wave inversion, or right ventricular dysfunction at baseline. DSG2 patients displayed more frequent epsilon wave (37% vs. 17%, P = 0.048) and left ventricular dysfunction at diagnosis (54% vs. 10%, P < 0.001). During a median follow-up of 5.6 years (2.5-16), DSG2 and PKP2 mutation carriers displayed a similar risk of sustained ventricular arrhythmia (log-rank P = 0.20), but DSG2 mutation carriers were at higher risk of transplantation/HF-related death (log-rank P < 0.001). The presence of a DSG2 mutation vs. PKP2 mutation was a predictor of transplantation/HF-related death in univariate Cox analysis (P = 0.0005)., Conclusions: In this multicentre cohort, DSG2 mutation carriers were found to be at high risk of end-stage HF compared to PKP2 mutation carriers, supporting careful haemodynamic monitoring of these patients. The benefit of early HF treatment needs to be assessed in DSG2 carriers., (© 2019 The Authors. European Journal of Heart Failure © 2019 European Society of Cardiology.)

Arrhythmogenic right ventricular cardiomyopathy is a heritable cardiac disease causing severe ventricular arrhythmias, heart failure and sudden cardiac death. It is mainly caused by mutations in genes encoding several structural proteins of the cardiac desmosomes including the DSG2 gene encoding the desmosomal cadherin desmoglein-2. Although the molecular structure of the extracellular domain of desmoglein-2 is known, it remains an open question, how mutations in DSG2 contribute to the pathogenesis of arrhythmogenic right ventricular cardiomyopathy. In the present study, we analyzed the impact of different DSG2 mutations on the glycosylation pattern using de-glycosylation assays, lectin blot analysis and genetic inhibition studies. Remarkably, wildtype and mutant desmoglein-2 displayed different glycosylation patterns, although the investigated DSG2 mutations do not directly affect the consensus sequences of the N-glycosylation sites. Our study reveals complex molecular interactions between DSG2 mutations and N-glycosylations of desmoglein-2, which may contribute to the molecular understanding of the patho-mechanisms associated with arrhythmogenic right ventricular cardiomyopathy., (Copyright © 2019 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Background: Variants in the desmoglein-2 (DSG2) gene account for a significant proportion of patients with Arrhythmogenic Right Ventricular Cardiomyopathy (ARVC). The aim of this study was to evaluate the genetic epidemiology of DSG2 and the impact of a frequent homozygous DSG2 variant in East Asia., Methods: Genetic screening of 14 ARVC related genes was performed in 118 unrelated index patients using next-generation sequencing. Following that, family screening, clinical evaluation and haplotype analysis were performed among eight probands who carry the same homozygous DSG2 variant. We also examined the histopathology and protein expression using immunofluorescence staining on the myocardial tissue of two probands undergoing heart transplant., Results: Eighteen (15.2%) patients bear rare putatively deleterious variants in DSG2, among which 8 patients shared the homozygous DSG2 p.Phe531Cys variant. Family screening demonstrated that only homozygous variant carriers exhibited definite ARVC phenotype with 100% penetrance, while heterozygous variant carriers were either unaffected or only presented mild ARVC related symptoms in 25% relatives. Left ventricular involvement and bi-ventricular failure were common among homozygous p. Phe531Cys variant patients even at early age. Haplotype analysis demonstrated p. Phe531Cys was a founder variant in East Asia population with an allele frequency of 0.12%., Conclusions: We identified, for the first time, a homozygous founder variant of DSG2 in East Asia, which was at surprisingly high frequency of 8.47% among Chinese ARVC patients with a full penetrance. This result suggested an urgent demand of genetic counseling for the probands and their relatives with heterozygous variant., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Backgrounds: This study was designed to identify the pathogenic mutations in two Chinese families of arrhythmogenic right ventricular cardiomyopathy/dysplasia (ARVC/D) using the Whole Exome Sequencing (WES)., Methods and Results: The proband 1 (Family 1, II:1) and proband 2 (Family 2, II:1) underwent the WES of DNA from peripheral blood. The genes susceptible to arrhythmias and cardiomyopathies were analyzed and both the probands carried the same exonic mutation of DSG2 p.F531C (NM&#95;001943, exon 11: c.T1592G). The proband 1 also carried the splicing mutation of DSG2 (NM&#95;001943: exon 4:c.217-1G>T), and proband 2 carried the intronic mutation of DSG2 (NM&#95;001943: exon 6: c.524-3C>G) that potentially influenced the splicing function predicted by Human Splicing Finder. The compound heterozygous mutations of the two probands inherited from their paternal and maternal side, respectively. The carriers with DSG2 p.F531C showed early abnormal electrocardiograms, characterized as the subclinical phenotype of ARVC/D., Conclusions: The DSG2 p.F531C was the main reason for ARVC/D. More severe phenotypes of ARVC/D occurred when coexisting with 217-1G>T or 524-3C>G mutation that potentially affecting the splicing function, as a compound heterozygous recessive inheritance., (Copyright © 2018. Published by Elsevier Inc.)

Pluripotent stem cells (PSCs) represent the most promising clinical source for regenerative medicine. However, given the cellular heterogeneity within cultivation and safety concerns, the development of specific and efficient tools to isolate a pure population and eliminate all residual undifferentiated PSCs from differentiated derivatives is a prerequisite for clinical applications. In this study, we raised a monoclonal antibody and identified its target antigen as desmoglein-2 (DSG2). DSG2 co-localized with human PSC (hPSC)-specific cell surface markers, and its expression was rapidly downregulated upon differentiation. The depletion of DSG2 markedly decreased hPSC proliferation and pluripotency marker expression. In addition, DSG2-negative population in hPSCs exhibited a notable suppression in embryonic body and teratoma formation. The actions of DSG2 in regulating the self-renewal and pluripotency of hPSCs were predominantly exerted through the regulation of β-catenin/Slug-mediated epithelial-to-mesenchymal transition. Our results demonstrate that DSG2 is a valuable PSC surface marker that is essential for the maintenance of PSC self-renewal., (Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.)

In mammalian pregnancy, the uterus is remodeled to become receptive to embryonic implantation. Since non-invasive placentation in marsupials is likely derived from invasive placentation, and is underpinned by intra-uterine conflict between mother and embryo, species with non-invasive placentation may employ a variety of molecular mechanisms to maintain an intact uterine epithelium and to prevent embryonic invasion. Identifying such modifications to the uterine epithelium of marsupial species with non-invasive placentation is key to understanding how conflict is mediated during pregnancy in different mammalian groups. Desmoglein-2, involved in maintaining lateral cell-cell adhesion of the uterine epithelium, is redistributed before implantation to facilitate embryo invasion in mammals with invasive placentation. We identified localization patterns of this cell adhesion molecule throughout pregnancy in two marsupial species with non-invasive placentation, the tammar wallaby (Macropus eugenii; Macropodidae), and the brushtail possum (Trichosurus vulpecula; Phalangeridae). Interestingly, Desmoglein-2 redistribution also occurs in both M. eugenii and T. vulpecula, suggesting that cell adhesion, and thus integrity of the uterine epithelium, is reduced during implantation regardless of placental type, and may be an important component of uterine remodeling. Desmoglein-2 also localizes to the mesenchymal stromal cells of M. eugenii and to epithelial cell nuclei in T. vulpecula, suggesting its involvement in cellular processes that are independent of adhesion and may compensate for reduced lateral adhesion in the uterine epithelium. We conclude that non-invasive placentation in marsupials involves diverse and complementary strategies to maintain an intact epithelial barrier., (© 2017 Wiley Periodicals, Inc.)

Krüppel-like factor 5 (KLF5) is a member of the zinc finger family of transcription factors that regulates homeostasis of the intestinal epithelium. Previous studies suggested an indispensable role of KLF5 in maintaining intestinal barrier function. In the current study, we investigated the mechanisms by which KLF5 regulates colonic barrier function in vivo and in vitro. We used an inducible and a constitutive intestine-specific Klf5 knockout mouse models ( Villin-CreER T2 ;Klf5 fl/fl designated as Klf5 ΔIND and Villin-Cre;Klf5 fl/fl as Klf5 ΔIS ) and studied an inducible KLF5 knockdown in Caco-2 BBe cells using a lentiviral Tet-on system (Caco-2 BBe KLF5ΔIND ). Specific knockout of Klf5 in colonic tissues, either inducible or constitutive, resulted in increased intestinal permeability. The phenotype was accompanied by a significant reduction in Dsg2 , which encodes desmoglein-2, a desmosomal cadherin, at both mRNA and protein levels. Transmission electron microscopy showed alterations of desmosomal morphology in both KLF5 knockdown Caco-2 BBe cells and Klf5 knockout mouse colonic tissues. Inducible knockdown of KLF5 in Caco-2BBe cells grown on Transwell plates led to impaired barrier function as evidenced by decreased transepithelial electrical resistance and increased paracellular permeability to fluorescein isothiocyanate-4 kDa dextran. Furthermore, DSG2 was significantly decreased in KLF5 knockdown cells, and DSG2 overexpression partially rescued the impaired barrier function caused by KLF5 knockdown. Electron microscopy studies demonstrated altered desmosomal morphology after KLF5 knockdown. In combination with chromatin immunoprecipitation analysis and promoter study, our data show that KLF5 regulates intestinal barrier function by mediating the transcription of DSG2 , a gene encoding a major component of desmosome structures. NEW & NOTEWORTHY The study is original research on the direct function of a Krüppel-like factor on intestinal barrier function, which is commonly exerted by cell junctions, including tight junctions, adherens junctions, and desmosomes. Numerous previous studies were focused on tight junctions and adherens junctions. However, this study provided a new perspective on how the intestinal barrier function is regulated by KLF5 through DSG2, a component of desmosome complexes., (Copyright © 2017 the American Physiological Society.)

Dilated cardiomyopathy (DCM) leads to disturbed contraction and force transduction, and is associated with substantial mortality in all age groups. Involvement of a disrupted composition of the intercalated disc (ID) has been reported. However, in children, little is established about such subcellular changes during disease, because of the pathological mix-up with the ongoing cardiac maturation. This leaves maladaptive remodeling often undetected. We aimed at illustrating subcellular alterations in children diagnosed with DCM compared to age-matched controls, focusing on ID proteins known to be crucially stable under healthy conditions and destabilized during cardiac injury in adults. Left ventricular or septal pediatric specimens were collected from 7 individuals diagnosed with DCM (age: 23 weeks in utero to 8 weeks postnatal) and age-matched controls that died of non-cardiovascular cause. We determined the amount of fibrosis and localization of ID proteins by immunohistochemistry. In pediatric DCM, most ID proteins follow similar spatiotemporal changes in localization as in controls. However, although no mutations were found, the signal of the desmosomal protein Desmoglein-2 was reduced in all pediatric DCM specimens, but not in controls or adult DCM patients. Endocardial and transmural fibrosis was increased in all pediatric DCM patients compared to age-matched controls. Composition of the ID in pediatric DCM patients is similar to controls, except for the localization of Desmoglein-2 and presence of severe fibrosis. This suggests that the architecture of desmosomes is already disturbed in the early stages of DCM. These findings contribute to the understanding of pediatric DCM., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Purpose: Desmoglein-2 (Dsg2) is a cell adhesion protein of the cadherin superfamily. Altered Dsg2 expression is associated with tumorigenesis. This study determined Dsg2 expression in non-small cell lung cancer (NSCLC) tissue specimens for association with clinicopathological and survival data and then assessed the effect of Dsg2 knockdown on regulation of NSCLC cell malignant behaviors in vitro and in nude mouse xenografts., Methods: qRT-PCR and Western blot were used to detect Dsg2 expression in 28 paired NSCLC and normal tissue samples. Immunohistochemistry was used to detect Dsg2 expression in 70 cases of paraffin-embedded NSCLC tissues. NSCLC A549, H1703, and H1299 cells were cultured with Dsg2 knockdown performed using Dsg2 siRNA. Cell viability, cell cycle, apoptosis, and colony formation were assessed. siRNA-transfected A549 cells were also used to generate tumor xenografts in nude mice., Results: Both Dsg2 mRNA and protein were highly expressed in NSCLC tissues and associated with NSCLC size, but not with overall survival of patients. Moreover, knockdown of Dsg2 expression reduced NSCLC cell proliferation and arrested them at the G1 phase of the cell cycle, but did not significantly affect NSCLC cell apoptosis. Dsg2 knockdown downregulated cyclin-dependent kinase 2 expression and upregulated p27 expression. Nude mouse xenograft assays showed that Dsg2 knockdown inhibited NSCLC xenograft growth in vivo., Conclusion: This study revealed the importance of Dsg2 in suppression of NSCLC development and progression. Further studies will explore whether restoration of Dsg2 expression is a novel strategy in control of NSCLC.

Desmogleins (DSG) are a family of cadherin adhesion proteins that were first identified in desmosomes and provide cardiomyocytes and epithelial cells with the junctional stability to tolerate mechanical stress. However, one member of this family, DSG2, is emerging as a protein with additional biological functions on a broader range of cells. Here we reveal that DSG2 is expressed by non-desmosome-forming human endothelial progenitor cells as well as their mature counterparts [endothelial cells (ECs)] in human tissue from healthy individuals and cancer patients. Analysis of normal blood and bone marrow showed that DSG2 is also expressed by CD34(+)CD45(dim) hematopoietic progenitor cells. An inability to detect other desmosomal components, i.e., DSG1, DSG3 and desmocollin (DSC)2/3, on these cells supports a solitary role for DSG2 outside of desmosomes. Functionally, we show that CD34(+)CD45(dim)DSG2(+) progenitor cells are multi-potent and pro-angiogenic in vitro. Using a 'knockout-first' approach, we generated a Dsg2 loss-of-function strain of mice (Dsg2 (lo/lo)) and observed that, in response to reduced levels of Dsg2: (i) CD31(+) ECs in the pancreas are hypertrophic and exhibit altered morphology, (ii) bone marrow-derived endothelial colony formation is impaired, (iii) ex vivo vascular sprouting from aortic rings is reduced, and (iv) vessel formation in vitro and in vivo is attenuated. Finally, knockdown of DSG2 in a human bone marrow EC line reveals a reduction in an in vitro angiogenesis assay as well as relocalisation of actin and VE-cadherin away from the cell junctions, reduced cell-cell adhesion and increased invasive properties by these cells. In summary, we have identified DSG2 expression in distinct progenitor cell subpopulations and show that, independent from its classical function as a component of desmosomes, this cadherin also plays a critical role in the vasculature., Competing Interests: The authors indicate no potential conflicts of interest.

The desmosomal cadherin, desmoglein 2 (Dsg2), is deregulated in a variety of human cancers including those of the skin. When ectopically expressed in the epidermis of transgenic mice, Dsg2 activates multiple mitogenic signaling pathways and increases susceptibility to tumorigenesis. However, the molecular mechanism responsible for Dsg2-mediated cellular signaling is poorly understood. Here we show overexpression as well as co-localization of Dsg2 and EGFR in cutaneous SCCs in vivo. Using HaCaT keratinocytes, knockdown of Dsg2 decreases EGFR expression and abrogates the activation of EGFR, c-Src and Stat3, but not Erk1/2 or Akt, in response to EGF ligand stimulation. To determine whether Dsg2 mediates signaling through lipid microdomains, sucrose density fractionation illustrated that Dsg2 is recruited to and displaces Cav1, EGFR and c-Src from light density lipid raft fractions. STED imaging confirmed that the presence of Dsg2 disperses Cav1 from the cell-cell borders. Perturbation of lipid rafts with the cholesterol-chelating agent MβCD also shifts Cav1, c-Src and EGFR out of the rafts and activates signaling pathways. Functionally, overexpression of Dsg2 in human SCC A431 cells enhances EGFR activation and increases cell proliferation and migration through a c-Src and EGFR dependent manner. In summary, our data suggest that Dsg2 stimulates cell growth and migration by positively regulating EGFR level and signaling through a c-Src and Cav1-dependent mechanism using lipid rafts as signal modulatory platforms., Competing Interests: The authors declare that there are no competing financial interests in relation to the work described.

Desmoglein-2 (DSG2), a member of the desmosomal cadherin superfamily, has been linked to arrhythmogenic right ventricular cardiomyopathy (ARVC)which may cause life-threatening ventricular arrhythmias and sudden death. Fatal arrhythmias resulting in sudden death also occur in the absence of morphologic cardiac abnormalities at autopsy. We sequenced all 15 exons of DSG2 in DNA extracted from post-mortem heart tissues of 25 patients dying with ARVC and 25 from sudden unexplained death (SUD). The primers were designed using the Primer Express 3.0 software. Direct sequencing for both sense and antisense strands was performed with a BigDye Terminator DNA sequencing kit on a 3130 xl Genetic Analyzer. Mutation damage prediction was made using Mutation Taster, Polyphen and SIFT software. 2 DSG2 mutations (p. S1026Q fsX12, p. G678R)in two ARVC samples and 2 DSG2 mutations(p. E 896K, p. A858 V) in two SUD samples were identified, all the mutations were novel. We concluded that DSG2 mutations may not specific for ARVC and may be related to the fatal arrhythmic events even in patients with a morphological normal heart., (Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.)

Attachment of the blastocyst and formation of the placenta during pregnancy is dependent on structural and cellular changes occurring in the uterine epithelium and in particular to the plasma membrane of these uterine cells. Desmosome expression decreases during pregnancy in eutherians and some squamates, presumably allowing for remodeling of the uterine epithelium and invasion of the trophoblast during implantation. Marsupials are a distinct mammalian amniote lineage of viviparity, with a short implantation or attachment period and varying levels of invasive placentation. To test the generality of changes to the uterine epithelium during pregnancy across mammals, we characterized the distribution of desmosomes in the uterine epithelial cells of a marsupial, Sminthopsis crassicaudata, using electron microscopy and immunohistochemistry. The absolute number of desmosomes along the lateral plasma membrane decreases during pregnancy and desmosomes are redistributed towards the apical region of the lateral plasma membrane as pregnancy proceeds, similar to what occurs during pregnancy in eutherian mammals. Despite the lower level of maternal investment in pregnancy and the noninvasive structure of fetal membranes in marsupials there are similarities in number and redistribution of desmosomes along the plasma membrane and changes to the morphology of the uterine epithelial cells suggesting that similar plasma membrane changes occur across all lineages of amniote vertebrates., (© 2014 Wiley Periodicals, Inc.)

Aims: We determined the contribution of the desmosomal cadherin desmoglein-2 to cell-cell cohesion in cardiomyocytes. In the intercalated disc, providing mechanical strength and electrical communication between adjacent cardiomyocytes, desmoglein-2 is closely associated with N-cadherin and gap junctions., Methods and Results: We studied intercalated discs of HL-1 cardiomyocytes by immunostaining of desmoglein-2 and N-cadherin. Cohesion was measured using a liberase-based dissociation-assay and compared with cell-free single-molecule atomic force microscopy measurements. L-tryptophan caused irregular desmoglein-2 condensation, weakened cell-cell cohesion and impaired both homophilic desmoglein-2 and N-cadherin trans-interaction, whereas l-phenylalanine had no effect. L-tryptophan did not affect N-cadherin localization and its inhibitory effect on cell-cohesion and desmoglein-2 binding, but not on N-cadherin interaction, was blocked by a desmoglein-specific tandem peptide. Moreover, Ca(2+)-depletion, desmoglein-2 knockdown, a desmoglein-specific single peptide and certain desmoglein-2 mutations associated with arrhythmogenic cardiomyopathy reduced cell-cell cohesion, whereas cell adhesion was strengthened by desmoglein-2 overexpression. Since single peptide did not interfere with N-cadherin trans-interaction, these data indicate that (i) desmoglein-2 binding is crucial for cardiomyocyte cohesion and (ii) L-tryptophan reduced both desmoglein-2 and N-cadherin binding, whereas single and tandem peptide can be used to specifically target desmoglein-2-mediated adhesion. L-tryptophan and single peptide also induced ultrastructural alterations of areae compositae. Functional analyses at the organ level revealed reduced cardiomyocyte function and inefficient response to adrenergic stimulation in both L-tryptophan- and single peptide-challenged murine Langendorff hearts paralleled by redistribution of connexin 43 in L-tryptophan-treated heart slices., Conclusion: Our data demonstrate that desmoglein-2 plays a critical role in cardiomyocyte cohesion and function., (Published on behalf of the European Society of Cardiology. All rights reserved. © The Author 2014. For permissions please email: journals.permissions@oup.com.)

The desmosomal cadherins, desmogleins, and desmocollins mediate strong intercellular adhesion. Human intestinal epithelial cells express the desmoglein-2 isoform. A proteomic screen for Dsg2-associated proteins in intestinal epithelial cells identified a lectin referred to as galectin-3 (Gal3). Gal3 bound to N-linked β-galactosides in Dsg2 extracellular domain and co-sedimented with caveolin-1 in lipid rafts. Down-regulation of Gal3 protein or incubation with lactose, a galactose-containing disaccharide that competitively inhibits galectin binding to Dsg2, decreased intercellular adhesion in intestinal epithelial cells. In the absence of functional Gal3, Dsg2 protein was internalized from the plasma membrane and degraded in the proteasome. These results report a novel role of Gal3 in stabilizing a desmosomal cadherin and intercellular adhesion in intestinal epithelial cells.

The secretopeptidome comprises endogenous peptides derived from proteins secreted into the tumour microenvironment through classical and non-classical secretion. This study characterised the low-Mr (<3kDa) component of the human colon tumour (LIM1215, LIM1863) secretopeptidome, as a first step towards gaining insights into extracellular proteolytic cleavage events in the tumour microenvironment. Based on two biological replicates, this secretopeptidome isolation strategy utilised differential centrifugal ultrafiltration in combination with analytical RP-HPLC and nanoLC-MS/MS. Secreted peptides were identified using a combination of Mascot and post-processing analyses including MSPro re-scoring, extended feature sets and Percolator, resulting in 474 protein identifications from 1228 peptides (≤1% q-value, ≤5% PEP) - a 36% increase in peptide identifications when compared with conventional Mascot (homology ionscore thresholding). In both colon tumour models, 122 identified peptides were derived from 41 cell surface protein ectodomains, 23 peptides (12 proteins) from regulated intramembrane proteolysis (RIP), and 12 peptides (9 proteins) generated from intracellular domain proteolysis. Further analyses using the protease/substrate database MEROPS, (http://merops.sanger.ac.uk/), revealed 335 (71%) proteins classified as originating from classical/non-classical secretion, or the cell membrane. Of these, peptides were identified from 42 substrates in MEROPS with defined protease cleavage sites, while peptides generated from a further 205 substrates were fragmented by hitherto unknown proteases. A salient finding was the identification of peptides from 88 classical/non-classical secreted substrates in MEROPS, implicated in tumour progression and angiogenesis (FGFBP1, PLXDC2), cell-cell recognition and signalling (DDR1, GPA33), and tumour invasiveness and metastasis (MACC1, SMAGP); the nature of the proteases responsible for these proteolytic events is unknown. To confirm reproducibility of peptide fragment abundance in this study, we report the identification of a specific cleaved peptide fragment in the secretopeptidome from the colon-specific GPA33 antigen in 4/14 human CRC models. This improved secretopeptidome isolation and characterisation strategy has extended our understanding of endogenous peptides generated through proteolysis of classical/non-classical secreted proteins, extracellular proteolytic processing of cell surface membrane proteins, and peptides generated through RIP. The novel peptide cleavage site information in this study provides a useful first step in detailing proteolytic cleavage associated with tumourigenesis and the extracellular environment. This article is part of a Special Issue entitled: An Updated Secretome., (Copyright © 2013 Elsevier B.V. All rights reserved.)