Nathalie Jonca

Refined Characterization of Corneodesmosin Proteolysis during Terminal Differentiation of Human Epidermis and Its Relationship to Desquamation

Corneodesmosin is a putative adhesion glycoprotein located in the extracellular part of the desmosomes in the upper layers of the epidermis. Synthesized by granular keratinocytes as a 52–56-kDa protein, corneodesmosin is progressively proteolysed during corneocyte maturation. This processing is a prerequisite for desquamation. Two glycine- and serine-rich domains of the protein might take on the conformation of adhesive secondary structures similar to glycine loops. Corneodesmosin proteolysis was further characterized. Deglycosylation experiments and reactivity with lectins demonstrated that the corneodesmosin carbohydrate moiety does not prevent the proteolysis. Immunoblotting, immunohistochemistry, and immunoelectron microscopy experiments using affinity-purified anti-peptide antibodies raised to four of the five structural domains of corneodesmosin and a monoclonal antibody against its fifth central domain showed that the first step in corneodesmosin processing is the cleavage of its extremities and probably occurs before its incorporation into desmosomes. Then the glycine loop-related domains are cleaved, first the N-terminal and then part of the C-terminal domain. At the epidermis surface, the multistep proteolytic cleavage leaves intact only the central domain, which was detected on exfoliated corneocytes and probably lacks adhesive properties. Importantly, corneodesmosin was demonstrated to be a preferred substrate of two serine proteases involved in desquamation, the stratum corneum tryptic and chymotryptic enzymes.

Hypotrichosis simplex of the scalp is associated with nonsense mutations in CDSN encoding corneodesmosin

We have identified nonsense mutations in the gene CDSN (encoding corneodesmosin) in three families suffering from hypotrichosis simplex of the scalp (HSS; OMIM 146520). CDSN, a glycoprotein expressed in the epidermis and inner root sheath (IRS) of hair follicles, is a keratinocyte adhesion molecule. Truncated CDSN aggregates were detected in the superficial dermis and at the periphery of hair follicles. Our findings suggest that CDSN is important in normal scalp hair physiology.

Large-scale identification of human genes implicated in epidermal barrier function

BackgroundDuring epidermal differentiation, keratinocytes progressing through the suprabasal layers undergo complex and tightly regulated biochemical modifications leading to cornification and desquamation. The last living cells, the granular keratinocytes (GKs), produce almost all of the proteins and lipids required for the protective barrier function before their programmed cell death gives rise to corneocytes. We present here the first analysis of the transcriptome of human GKs, purified from healthy epidermis by an original approach.ResultsUsing the ORESTES method, 22,585 expressed sequence tags (ESTs) were produced that matched 3,387 genes. Despite normalization provided by this method (mean 4.6 ORESTES per gene), some highly transcribed genes, including that encoding dermokine, were overrepresented. About 330 expressed genes displayed less than 100 ESTs in UniGene clusters and are most likely to be specific for GKs and potentially involved in barrier function. This hypothesis was tested by comparing the relative expression of 73 genes in the basal and granular layers of epidermis by quantitative RT-PCR. Among these, 33 were identified as new, highly specific markers of GKs, including those encoding a protease, protease inhibitors and proteins involved in lipid metabolism and transport. We identified filaggrin 2 (also called ifapsoriasin), a poorly characterized member of the epidermal differentiation complex, as well as three new lipase genes clustered with paralogous genes on chromosome 10q23.31. A new gene of unknown function, C1orf81, is specifically disrupted in the human genome by a frameshift mutation.ConclusionThese data increase the present knowledge of genes responsible for the formation of the skin barrier and suggest new candidates for genodermatoses of unknown origin.

Corneodesmosin, a Component of Epidermal Corneocyte Desmosomes, Displays Homophilic Adhesive Properties

Corneodesmosomes, the modified desmosomes of the uppermost layers of the epidermis, play an important role in corneocyte cohesion. Corneodesmosin is a secreted glycoprotein located in the corneodesmosomal core and covalently linked to the cornified envelope of corneocytes. Its glycine- and serine-rich NH2-terminal domain may fold to give structural motifs similar to the glycine loops described in epidermal cytokeratins and loricrin and proposed to display adhesive properties. A chimeric protein comprising human corneodesmosin linked to the transmembrane and cytoplasmic domains of mouse E-cadherin was expressed in mouse fibroblasts to test the ability of corneodesmosin to promote cell-cell adhesion. Classic aggregation assays indicated that corneodesmosin mediates homophilic cell aggregation. Moreover, Ca2+depletion showed a moderate effect on aggregation. To assess the involvement of the glycine loop domain in adhesion, full-length corneodesmosin, corneodesmosin lacking this domain, or this domain alone were expressed as glutathione S-transferase fusion proteins and tested for protein-protein interactions by overlay binding assays. The results confirmed that corneodesmosin presents homophilic interactions and indicated that its NH2-terminal glycine loop domain is sufficient but not strictly necessary to promote binding. Altogether, these results provide the first experimental evidence for the adhesive properties of corneodesmosin and for the involvement of its glycine loop domain in adhesion.

Update on the epidermal differentiation complex.

On human chromosome 1q21, a 2-Mb region called the epidermal differentiation complex comprises many genes encoding structural and regulatory proteins that are of crucial importance for keratinocyte differentiation and stratum corneum properties. Apart from those for involucrin and loricrin, most of the genes are organized in four families: the genes encoding EF-hand calcium-binding proteins of the S100A family, the genes encoding the small proline rich proteins (SPRRs) and the late cornified envelope (LCE) proteins, two families of cornified cell envelope components, and the genes encoding the S100-fused type proteins (SFTPs). This review focuses on the SPRRs, LCE proteins and SFTPs. It describes their structures, their specific functions and, when known, the mechanisms involved in the regulation of their expression. It also highlights their possible involvement in skin diseases.

Corneodesmosin gene ablation induces lethal skin-barrier disruption and hair-follicle degeneration related to desmosome dysfunction

Corneodesmosin (CDSN) is specific to desmosomes of epithelia undergoing cornification, mainly the epidermis and the inner root sheath of the hair follicles. CDSN nonsense mutations are associated with hypotrichosis simplex of the scalp, a rare disease that leads to complete baldness in young adults. CDSN displays adhesive properties, mostly attributable to its N-terminal glycine-rich domain, and is sequentially proteolyzed as corneocytes migrate towards the skin surface. K14-promoter driven Cre-mediated deletion of Cdsn in mice resulted in neonatal death as a result of epidermal tearing upon minor mechanical stress. Ultrastructural analyses revealed a desmosomal break at the interface between the living and cornified layers. After grafting onto nude mice, knockout skin showed a chronic defect in the epidermal permeability barrier. The epidermis was first hyperproliferative with a thick cornified layer, then, both the epidermis and the hair follicles degenerated. In adults, Cdsn deletion resulted in similar histological abnormalities and in a lethal barrier defect. We demonstrate that Cdsn is not essential for skin-barrier formation in utero, but is vital throughout life to preserve this barrier by maintaining desmosome integrity. The strong adhesive function that the protein confers on corneodesmosomes also seems necessary for maintaining the architecture of the hair follicle.

A Novel Protease Inhibitor of the α2-Macroglobulin Family Expressed in the Human Epidermis

In the course of a large scale analysis of late-expressed genes in the human epidermis, we identified a new member of the α2-macroglobulin (α2M) protease inhibitor family, A2ML1 (for α2-macroglobulin-like 1). Like A2M and PZP, A2ML1 is located on chromosome 12p13.31. A2ML1 encodes a protein of 1454 amino acids, which fits the characteristics of α2Ms: 1) strong conservation in amino acid sequence including most of cysteine positions with α2M; 2) a putative central bait domain; 3) a typical thiol ester sequence. Northern blot and reverse transcriptase-PCR studies revealed a single 5-kb A2ML1 mRNA, mainly in the epidermis granular keratinocytes. A2ML1 is also transcribed in placenta, thymus, and testis. By Western blot analysis, α2ML1 is detected as a monomeric, ∼180-kDa protein in human epidermis. In vitro keratinocyte differentiation is associated with increased expression levels. By immunohistochemistry, α2ML1 was detected within keratinosomes in the granular layer of the epidermis, and as a secreted product in the extracellular space between the uppermost granular layer and the cornified layer. Recombinant α2ML1 displayed inhibitory activity toward chymotrypsin, papain, thermolysin, subtilisin A, and to a lesser extent, elastase but not trypsin. Incubation with chymotrypsin and the chymotrypsin-like kallikrein 7 protease indicated that α2ML1 binds covalently to these proteases, a feature shared with other members of the family. Therefore, α2ML1 is the first α2M family member detected in the epidermis. It may play an important role during desquamation by inhibiting extracellular proteases.

Corneodesmosomes and corneodesmosin: from the stratum corneum cohesion to the pathophysiology of genodermatoses

Corneodesmosin (CDSN) was identified 20 years ago by raising monoclonal antibodies against human plantar stratum corneum. The protein is specific to corneodesmosomes, cell-junction structures that, in humans, are found in the epidermis, the hard palate epithelium, and the inner root sheath of the hair follicles. Synthesized by the granular keratinocytes and secreted via the lamellar bodies, CDSN is incorporated into the desmoglea of the desmosomes, shortly before their transformation into corneodesmosomes during cornification. CDSN displays adhesive properties, mostly attributable to its N-terminal glycine-rich domain, and is sequentially proteolyzed as corneocytes migrate towards the skin surface prior to desquamation. The recent inactivation of Cdsn in mice induced a lethal epidermal barrier disruption and hair follicle degeneration, related to corneodesmosome dysfunction. That confirmed the essential role of the protein in maintaining integrity of the epidermis and the hair follicle. The CDSN gene is located in PSORS1, the major psoriasis susceptibility locus on the chromosome 6, but to date its involvement in the disease pathophysiology is not clear. By contrast, two different monogenic diseases associated with nonsense mutations in CDSN, were recently identified. First, hypotrichosis simplex of the scalp in which mutated CDSN accumulates in the dermis and forms amyloid deposits; then, peeling skin disease in which the genetic defect induces dyscohesion of the stratum corneum, responsible for abnormal desquamation and increased skin penetration of allergens.

Alterations in the desquamation-related proteolytic cleavage of corneodesmosin and other corneodesmosomal proteins in psoriatic lesional epidermis.

Background  Desquamation occurs after proteolysis of corneodesmosomal proteins, including corneodesmosin (CDSN), by proteases of the kallikrein family, particularly KLK7. Impaired desquamation is one of the features of psoriasis, and psoriasis‐associated single nucleotide polymorphisms of the CDSN gene may potentially modify the proteolysis of the encoded protein.

Identification of the first nonsense CDSN mutation with expression of a truncated protein causing peeling skin syndrome type B

Peeling skin disease (PSD), a generalized inflammatory form of peeling skin syndrome, is caused by autosomal recessive nonsense mutations in the corneodesmosin gene (CDSN).