Maja M. Suter

Pemphigus vulgaris identifies plakoglobin as key suppressor of c-Myc in the skin

The autoimmune disease pemphigus vulgaris (PV) manifests as loss of keratinocyte cohesion triggered by autoantibody binding to desmoglein (Dsg)3, an intercellular adhesion molecule of mucous membranes, epidermis, and epidermal stem cells. Here we describe a so far unknown signaling cascade activated by PV antibodies. It extends from a transient enhanced turn over of cell surface‐exposed, nonkeratin‐anchored Dsg3 and associated plakoglobin (PG), through to depletion of nuclear PG, and as one of the consequences, abrogation of PG‐mediated c‐Myc suppression. In PV patients (6/6), this results in pathogenic c‐Myc overexpression in all targeted tissues, including the stem cell compartments. In summary, these results show that PV antibodies act via PG to abolish the c‐Myc suppression required for both maintenance of epidermal stem cells in their niche and controlled differentiation along the epidermal lineage. Besides a completely novel insight into PV pathogenesis, these data identify PG as a potent modulator of epithelial homeostasis via its role as a key suppressor of c‐Myc.

Proprotein cleavage of E-cadherin by furin in baculovirus over-expression system: potential role of other convertases in mammalian cells.

Sequence analysis of the adhesion molecule E‐cadherin had revealed a multibasic motif [4PArg‐Gln‐Lys‐Arg1P], reminiscent of the minimal cleavage signal for furin, the prototype of the proprotein convertase family, and/or other members sharing similar sequence specificity. Mutation of this site was sufficient to abolish processing of E‐cadherin in fibroblasts reinforcing the possibility that proprotein convertases are involved in the maturation of this adhesion molecule. Here we demonstrate that even though furin can efficiently and specifically cleave proE‐cadherin in a baculovirus‐based co‐expression system, the furin‐deficient LoVo cells were found to process endogenous E‐cadherin as efficiently as normal cell lines. This suggests, for the first time, that E‐cadherin is not only a substrate for furin but for other mammalian convertases sharing similar sequence specificity.

Missing C‐terminal filaggrin expression, NFkappaB activation and hyperproliferation identify the dog as a putative model to study epidermal dysfunction in atopic dermatitis

Please cite this paper as: Missing C‐terminal filaggrin expression, NFkappaB activation and hyperproliferation identify the dog as a putative model to study epidermal dysfunction in atopic dermatitis. Experimental Dermatology 2010; 19: e343–e346.

β-Catenin is not required for proliferation and differentiation of epidermal mouse keratinocytes

Despite the pivotal role of β-catenin in a variety of biological processes, conditional β-catenin gene ablation in the skin of transgenic mice failed to affect interfollicular epidermal morphogenesis. We elucidated the molecular mechanisms underlying this phenomenon. Long-term cultures of homozygous, heterozygous and β-catenin-null mutant keratinocytes were established to demonstrate that epidermal keratinocyte proliferation, cell cycle progression and cyclin D1 expression occur independently of β-catenin and correlate with repression of transcription from Tcf/Lef-responsive promoters. Moreover, during differentiation,β -catenin-null cells assemble normal intercellular adhesion junctions owing to the substitution of β-catenin with plakoglobin, whereas the expression of the other adhesion components remains unaffected. Taken together, our results demonstrate that epidermal proliferation and adhesion are independent of β-catenin.

The keratinocyte in epidermal renewal and defence

Traditionally, keratinocytes have been considered inert constituents of the multilayered epidermis. Todays understanding has fundamentally changed. The keratinocyte is now recognized as an active player in epidermal renewal with key functions in the skins immune defence. Under homeostatic conditions, keratinocyte progenitor cells are believed to divide symmetrically or asymmetrically, that is they continue to proliferate or go on to terminally differentiate and build up the overlaying epidermis. The fine-tuned process of epidermal renewal relies on an extraordinary network of signalling cascades which are governed by keratinocyte-receptor interactions with the environment through paracrine and autocrine circuits. Opposing this coordinated homeostatic process are signals of wounding and inflammation. They alter the fate of the keratinocyte and its response to the environment through changes in adhesion molecules and surface receptors, in addition to triggering an immediate inflammatory keratinocyte response in terms of secretion of cytokines, chemokines and antimicrobial peptides. If uncontrolled, the fundamental changes imposed by wounding and inflammation upon the homeostatic programme can lead to severe skin lesions including chronic inflammatory disorders. This review will describe the current knowledge of the regulatory signalling network which allows the keratinocyte to actively impact both epidermal homeostasis and the inflammatory response.

Keratinocyte biology and pathology

It is the saying of Socrates, ‘I know that I know nothing’, that comes to mind when reading through our review article from 1997. We realize how limited our knowledge on the biology and pathology of the epidermis was at the time and how much it has evolved since then. In our review, we had focused on the epidermis as a structural membrane protecting inner homeostasis. We also knew and had emphasized how the keratinocyte that forms this structure undergoes a highly orchestrated process of proliferation and differentiation through differential expression of structural proteins, surface molecules and enzymes. In recent years, it has become evident that the epidermis is a complex multicellular organ, in which a sophisticated crosstalk between its cells, the environment and the body’s metabolism takes place to ensure its many essential functions. While the air–liquid and the liquid–liquid barriers of the epidermis (Figure 1) form potent measures for separating the inside from the outside milieu, epidermal keratinocytes also have an immunostimulatory role to build up the defence system in co-operation with epidermal immune cells. The epidermis is, however, not just a protective measure against harmful external influences and maintenance of the inner

Plakoglobin‐dependent disruption of the desmosomal plaque in pemphigus vulgaris

Abstract:  We recently reported that the pathogenesis of pemphigus vulgaris (PV), an autoimmune blistering skin disorder, is driven by the accumulation of c‐Myc secondary to abrogation of plakoglobin (PG)‐mediated transcriptional c‐Myc suppression. PG knock‐out mouse keratinocytes express high levels of c‐Myc and resemble PVIgG‐treated wild‐type keratinocytes in most respects. However, they fail to accumulate nuclear c‐Myc and loose intercellular adhesion in response to PVIgG‐treatment like wild‐type keratinocytes. This suggested that PG is also required for propagation of the PVIgG‐induced events between augmented c‐Myc expression and acantholysis. Here, we addressed this possibility by comparing PVIgG‐induced changes in the desmosomal organization between wild‐type and PG knock‐out keratinocytes. We found that either bivalent PVIgG or monovalent PV‐Fab (known to trigger blister formation in vivo) disrupt the linear organization of all major desmosomal components along cell borders in wild‐type keratinocytes, simultaneously with a reduction in intercellular adhesive strength. In contrast, PV‐Fab failed to affect PG knock‐out keratinocytes while PVIgG cross‐linked their desmosomal cadherins without significantly affecting desmoplakin. These results identify PG as a principle effector of the PVIgG‐induced signals downstream of c‐Myc that disrupt the desmosomal plaque at the plasma membrane.

Periplakin and envoplakin are target antigens in canine and human paraneoplastic pemphigus

BACKGROUND On the basis of clinical and histopathologic similarities to human paraneoplastic pemphigus (PNP), we recently identified the first case of PNP in a nonhuman species, the dog. OBJECTIVE To determine a similar pathogenesis in both species, the present study aimed to define whether common antigens are targeted in dog and man. METHODS Canine and human PNP sera were used in parallel to immunoprecipitate 14C-labeled human keratinocyte antigens. The immunoreactive proteins were then identified by immunoprecipitation of canine keratinocyte extracts with specific antibodies to the antiplakin family members follwed by immunoblot analysis using canine and human PNP sera. RESULTS Protein bands of 210, 190, 170, and 130 kd were identified in dogs and humans. In both species, envoplakin and periplakin were demonstrated as antigens. Anti-desmoglein 3 antibodies could not be demonstrated in canine PNP, but in human PNP. CONCLUSION These results demonstrate that canine PNP closely correlates to the human counterpart and may therefore represent an excellent model for the human disease.

An adult passive transfer mouse model to study desmoglein 3 signaling in pemphigus vulgaris

Evidence has accumulated that changes in intracellular signaling downstream of desmoglein 3 (Dsg3) may play a significant role in epithelial blistering in the autoimmune disease pemphigus vulgaris (PV). Currently, most studies on PV involve passive transfer of pathogenic antibodies into neonatal mice which have not finalized epidermal morphogenesis, and do not permit analysis of mature hair follicles (HFs) and stem cell niches. To investigate Dsg3 antibody-induced signaling in the adult epidermis at defined stages of the HF cycle, we here developed a model with passive transfer of the monospecific pathogenic Dsg3 antibody AK23 into adult 8-week-old C57Bl/6J mice. Validated using histopathological and molecular methods, we found that this model faithfully recapitulates major features described in PV patients and PV models. Two hours after AK23 transfer we observed widening of intercellular spaces between desmosomes and EGFR activation, followed by increased Myc expression and epidermal hyperproliferation, desmosomal Dsg3 depletion and predominant blistering in HFs and oral mucosa. These data confirm that the adult passive transfer mouse model is ideally suited for detailed studies of Dsg3 antibody-mediated signaling in adult skin, providing the basis for investigations on novel keratinocyte-specific therapeutic strategies.

Loss of Invasiveness in Squamous Cell Carcinoma Cells Overexpressing Desmosomal Cadherins

The molecular and structural characteristics of intercellular adhesion were investigated in a squamous cell carcinoma (SCCA) cell line, originally derived from an oral tumor with an invasive growth pattern. The expression of adherens junction and desmosomal components were compared with that of cultured normal oral keratinocytes. Lack of membrane association in interdesmosomal areas, the disorganization of the actin cytoskeleton and the faster cell disassembly upon E-cadherin antibody binding in SCCA cells indicated decreased functional adherens junctions. These observations were supported by a significant reduction in E-, N-, and P-cadherin protein expression. In contrast, the level of desmosomal cadherin proteins, desmoglein 1/2 and desmocollin 2, were substantially upregulated and accompanied, ultrastructurally, by increased number and size of desmosomes. Since tumor invasion suppressor capacity has been proposed for desmosomal cadherins, we investigated the in vivo invasion potential of these SCCA cells by introducing them into SCID mice. Tumors developed, but with a benign phenotype. Based on these results, we hypothesize that the benign behavior of this SCCA cell line is a consequence of overexpressed desmosomal cadherins. This SCCA cell line, therefore, represents an excellent model system to further investigate the regulation and tumor invasion suppressor potential of desmosomal adhesion molecules.