Characterization and Purification of Human Corneodesmosin, an Epidermal Basic Glycoprotein Associated with Corneocyte-specific Modified Desmosomes
1997; Elsevier BV; Volume: 272; Issue: 50 Linguagem: Inglês
10.1074/jbc.272.50.31770
ISSN1083-351X
AutoresMichel Simon, Martine Montézin, Marina Guerrin, Jean‐Jacques Durieux, Guy Serre,
Tópico(s)Proteins in Food Systems
ResumoUsing monoclonal antibodies, we identified a new protein in mammalian epidermis, which we called corneodesmosin. It is located in the extracellular part of the modified desmosomes in the cornified layer of the tissue, and its proteolysis (from 52–56 to 33 kDa) is thought to be a major prerequisite of desquamation. We have now further characterized human corneodesmosin. Proteolysis of purified cornified cell envelopes produced immunoreactive fragments, confirming the covalent linkage of the protein to these structures. Sequential extraction of epidermal proteins indicated that the 52–56-kDa precursor form of the protein exists in two distinct pools, one extracted with a nondenaturing hypotonic buffer, and the other with urea. Two-dimensional gel analysis and reactivity with phosphoserine-specific antibodies showed that it is a basic phosphoprotein. Deglycosylation experiments, reactivity with lectins, and chromatography on concanavalin A-Sepharose indicated that corneodesmosin is N-glycosylated. Partial sequences, 10 and 15 amino acids long, of the purified 52–56-kDa corneodesmosin showed identity with sequences predicted from a previously cloned gene, proved to be expressed in the epidermis and designated S. This indicates that corneodesmosin is probably encoded by the S gene, the function of which was unknown until now. A model of corneodesmosin maturation during cornification is proposed. Using monoclonal antibodies, we identified a new protein in mammalian epidermis, which we called corneodesmosin. It is located in the extracellular part of the modified desmosomes in the cornified layer of the tissue, and its proteolysis (from 52–56 to 33 kDa) is thought to be a major prerequisite of desquamation. We have now further characterized human corneodesmosin. Proteolysis of purified cornified cell envelopes produced immunoreactive fragments, confirming the covalent linkage of the protein to these structures. Sequential extraction of epidermal proteins indicated that the 52–56-kDa precursor form of the protein exists in two distinct pools, one extracted with a nondenaturing hypotonic buffer, and the other with urea. Two-dimensional gel analysis and reactivity with phosphoserine-specific antibodies showed that it is a basic phosphoprotein. Deglycosylation experiments, reactivity with lectins, and chromatography on concanavalin A-Sepharose indicated that corneodesmosin is N-glycosylated. Partial sequences, 10 and 15 amino acids long, of the purified 52–56-kDa corneodesmosin showed identity with sequences predicted from a previously cloned gene, proved to be expressed in the epidermis and designated S. This indicates that corneodesmosin is probably encoded by the S gene, the function of which was unknown until now. A model of corneodesmosin maturation during cornification is proposed. Corneocytes are anucleated "mummified" cells derived from keratinocytes during the late stages of terminal differentiation in cornified squamous epithelia such as the epidermis. They mainly consist of a cytokeratin-containing fibrous matrix surrounded by a highly resistant 15-nm thick protein structure called the cornified cell envelope. Stacking of the corneocytes at the outermost layer of the tissue, namely the stratum corneum (SC), 1The abbreviations used are: SC, stratum corneum; mAb, monoclonal antibody; PAGE, polyacrylamide gel electrophoresis; NEpHGE, non-equilibrium pH gel electrophoresis; ConA, concanavalin A.1The abbreviations used are: SC, stratum corneum; mAb, monoclonal antibody; PAGE, polyacrylamide gel electrophoresis; NEpHGE, non-equilibrium pH gel electrophoresis; ConA, concanavalin A. plays a critical role in the epidermal barrier function and in the mechanical protection of the body (1Holbrook K. Leigh I. Lane E. Watt F. The Keratinocyte Handbook. Cambridge University Press, Cambridge1994: 275-292Google Scholar, 2Elias P.M. J. Invest. Dermatol. 1983; 80: 44S-49SAbstract Full Text PDF PubMed Google Scholar, 3Roop D. Science. 1995; 267: 474-475Crossref PubMed Scopus (171) Google Scholar). During the normal desquamation process, the most superficial corneocytes are shed from the skin surface. The structures involved in the cohesion between individual corneocytes, and the mechanisms that lead to the detachment of these cells are poorly understood.Although intercellular structures, thought to be derived from desmosomes (4Allen T.D. Potten C.S. J. Ultrastruct. Res. 1975; 51: 94-105Crossref PubMed Scopus (71) Google Scholar, 5Menton D.N. Eisen A.Z. J. Ultrastruct. Res. 1971; 35: 247-264Crossref PubMed Scopus (80) Google Scholar), and called corneosomes or corneodesmosomes (6Chapman S.J. Walsh A. Arch. Dermatol. Res. 1990; 282: 304-310Crossref PubMed Scopus (123) Google Scholar, 7Serre G. Mils V. Haftek M. Vincent C. Croute F. Réano A. Ouhayoun J.-P. Bettinger S. Soleilhavoup J.-P. J. Invest. Dermatol. 1991; 97: 1061-1072Abstract Full Text PDF PubMed Google Scholar), have been described in the SC, they were initially considered as nonfunctional remnants. Recent studies have shown, however, their major importance in corneocyte cohesion, and there is now growing evidence that their degradation is a key event in the desquamation process (6Chapman S.J. Walsh A. Arch. Dermatol. Res. 1990; 282: 304-310Crossref PubMed Scopus (123) Google Scholar, 7Serre G. Mils V. Haftek M. Vincent C. Croute F. Réano A. Ouhayoun J.-P. Bettinger S. Soleilhavoup J.-P. J. Invest. Dermatol. 1991; 97: 1061-1072Abstract Full Text PDF PubMed Google Scholar, 8King C.S. Barton S.P. Nicholls S. Marks R. Br. J. Dermatol. 1979; 100: 165-172Crossref PubMed Scopus (70) Google Scholar, 9Skerrow C.J. Clelland D.G. Skerrow D. J. Cell Sci. 1989; 92: 667-677PubMed Google Scholar, 10Chapman S.J. Walsh A. Jackson S.M. Friedmann P.S. Arch. Dermatol. Res. 1991; 283: 167-173Crossref PubMed Scopus (82) Google Scholar, 11Rawlings A.V. Scott I.R. Harding C.R. Bowser P.A. J. Invest. Dermatol. 1994; 103: 731-740Abstract Full Text PDF PubMed Scopus (408) Google Scholar). In particular, a tight correlation seems to exist between cell dissociation and proteolysis of some corneodesmosomal components (12King I.A. Wood M.J. Fryer P.R. J. Invest. Dermatol. 1989; 92: 22-26Abstract Full Text PDF PubMed Google Scholar, 13Lundström A. Egelrud T. J. Invest. Dermatol. 1990; 94: 216-220Abstract Full Text PDF PubMed Google Scholar, 14Egelrud T. Hofer P.-A. Lundström A. Acta. Derm. Venereol. (Stockh.). 1988; 68: 93-97PubMed Google Scholar). Moreover, in plantar SC, where cell cohesion is very strong, corneodesmosomes can be detected over the whole corneocyte surface up to the top of the layer. In non-palmoplantar SC, where cohesion is weaker, they only persist at the periphery of the cells (6Chapman S.J. Walsh A. Arch. Dermatol. Res. 1990; 282: 304-310Crossref PubMed Scopus (123) Google Scholar, 7Serre G. Mils V. Haftek M. Vincent C. Croute F. Réano A. Ouhayoun J.-P. Bettinger S. Soleilhavoup J.-P. J. Invest. Dermatol. 1991; 97: 1061-1072Abstract Full Text PDF PubMed Google Scholar, 9Skerrow C.J. Clelland D.G. Skerrow D. J. Cell Sci. 1989; 92: 667-677PubMed Google Scholar). In psoriasis, various ichthyoses, and skin xerosis, characterized by an accumulation of corneocytes and by scaling, the number of corneodesmosomes is increased throughout the SC including the upper part (11Rawlings A.V. Scott I.R. Harding C.R. Bowser P.A. J. Invest. Dermatol. 1994; 103: 731-740Abstract Full Text PDF PubMed Scopus (408) Google Scholar, 15Vicanova J. Mommaas A.M. Mulder A.A. Koerten H.K. Ponec M. Cell Tissue Res. 1996; 286: 115-122Crossref PubMed Scopus (38) Google Scholar, 16Lundström A. Egelrud T. Arch. Dermatol. Res. 1990; 282: 234-237Crossref PubMed Scopus (49) Google Scholar). Several trypsin-like and chymotrypsin-like serine proteases, including the stratum corneum chymotryptic enzyme, are thought to be involved in the corneodesmosome proteolysis (17Suzuki Y. Nomura J. Hori J. Koyama J. Takahashi M. Horii I. Arch. Dermatol. Res. 1993; 285: 372-377Crossref PubMed Scopus (83) Google Scholar, 18Suzuki Y. Nomura J. Koyama J. Horii I. Arch. Dermatol. Res. 1994; 286: 249-253Crossref PubMed Scopus (92) Google Scholar, 19Hansson L. Strömqvist M. Bäckman A. Wallbrandt P. Carlstein A. Egelrud T. J. Biol. Chem. 1994; 269: 19420-19426Abstract Full Text PDF PubMed Google Scholar).Using three different monoclonal antibodies (mAbs), we recently identified a new corneodesmosome protein antigen expressed in various mammals including human, and we called it corneodesmosin (7Serre G. Mils V. Haftek M. Vincent C. Croute F. Réano A. Ouhayoun J.-P. Bettinger S. Soleilhavoup J.-P. J. Invest. Dermatol. 1991; 97: 1061-1072Abstract Full Text PDF PubMed Google Scholar, 20Montézin M. Simon M. Guerrin M. Serre G. Exp. Cell Res. 1997; 231: 132-140Crossref PubMed Scopus (20) Google Scholar). Corneodesmosin is only expressed in the cornified squamous epithelia,i.e. in man: epidermis, hard palate, epithelium, and inner root sheath of the hair follicle (7Serre G. Mils V. Haftek M. Vincent C. Croute F. Réano A. Ouhayoun J.-P. Bettinger S. Soleilhavoup J.-P. J. Invest. Dermatol. 1991; 97: 1061-1072Abstract Full Text PDF PubMed Google Scholar, 21Mils V. Vincent C. Croute F. Serre G. J. Histochem. Cytochem. 1992; 40: 1329-1337Crossref PubMed Scopus (49) Google Scholar). In human non-palmoplantar epidermis, the protein was shown to be located first in cytoplasmic vesicles, termed keratinosomes or lamellar bodies, of the upper spinous keratinocytes, then in the intercellular part of the desmosomes of the granular keratinocytes, these living cells being just beneath the SC. Lastly, it was detected in the core of corneodesmosomes in the SC (7Serre G. Mils V. Haftek M. Vincent C. Croute F. Réano A. Ouhayoun J.-P. Bettinger S. Soleilhavoup J.-P. J. Invest. Dermatol. 1991; 97: 1061-1072Abstract Full Text PDF PubMed Google Scholar). By immunofluorescence and immunoelectron microscopy, corneodesmosin was also shown to be bound to the cornified cell envelopes (22Haftek M. Serre G. Mils V. Thivolet J. J. Histochem. Cytochem. 1991; 39: 1531-1538Crossref PubMed Scopus (62) Google Scholar). When extracted from viable layers of human epidermis, corneodesmosin shows an apparent molecular mass of around 52 kDa (here designated as 52–56-kDa corneodesmosin), whereas a molecular form of 33 kDa is the major form extracted from the most superficial and less firmly attached corneocytes (7Serre G. Mils V. Haftek M. Vincent C. Croute F. Réano A. Ouhayoun J.-P. Bettinger S. Soleilhavoup J.-P. J. Invest. Dermatol. 1991; 97: 1061-1072Abstract Full Text PDF PubMed Google Scholar, 23Lundström A. Serre G. Haftek M. Egelrud T. Arch. Dermatol. Res. 1994; 286: 369-375Crossref PubMed Scopus (94) Google Scholar). Moreover, corneodesmosin is proteolysed (from 52–56 to 33 kDa) during SC maturation (23Lundström A. Serre G. Haftek M. Egelrud T. Arch. Dermatol. Res. 1994; 286: 369-375Crossref PubMed Scopus (94) Google Scholar). Some evidence was obtained in favor of a role of corneodesmosin in corneocyte cohesion, and its proteolysis was proposed as one of the major biochemical changes that lead to desquamation (23Lundström A. Serre G. Haftek M. Egelrud T. Arch. Dermatol. Res. 1994; 286: 369-375Crossref PubMed Scopus (94) Google Scholar).In this work, we have further characterized, purified, and partially sequenced the 52–56-kDa human corneodesmosin. The obtained sequences were contained in the product of the S gene, recently identified and located 160 kilobase pairs telomeric to HLA-C (24Zhou Y. Chaplin D.D. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: 9470-9474Crossref PubMed Scopus (85) Google Scholar). They confirmed our recent cloning of the human corneodesmosin cDNA.DISCUSSIONIn the studies presented above, we biochemically characterized and partially sequenced human corneodesmosin, a protein specific to cornified squamous epithelia where it is thought to play a major role in corneocyte cohesion and whose proteolysis seems to be a major prerequisite of desquamation.Our data indicate that human corneodesmosin is a glycoprotein containing mainly N-linked oligosaccharides that comprise ∼10% of the protein mass. Indeed, treatment withN-glycosidase F (a glycosidase specific forN-linked sugars) of the 52–56-kDa corneodesmosin induced a 5-kDa decrease in its apparent molecular weight, whereas extensive endo-α-N-acetylgalactosaminidase digestion (an enzyme specific for O-linked sugars) was without effect, even in the presence of neuraminidase and/or N-glycosidase F. Lectins were used to characterize the corneodesmosin carbohydrate moieties. Since the protein bound to ConA-Sepharose, and reacted with biotinylated P. sativum agglutinin, it may contain α-d-mannose and/or α-d-glucose. In addition, corneodesmosin strongly reacted with wheat germ agglutinin and, as expected for an N-glycosylated protein, may therefore contains β-N-acetylglucosamine groups. However, it seems to contain few or no galactose andN-acetylgalactosamine, since it did not react (or barely) with lectins specific for these carbohydrates. More extensive experiments will be necessary to precisely identify the oligosaccharide residues linked to corneodesmosin, and to characterize their linkage. In fact these residues may participate in the adhesion properties of the protein, as described for other corneodesmosomal adhesive proteins, desmogleins and desmocollins (26Kapprell H.-P. Cowin P. Franke W.W. Ponstingl H. Eur. J. Cell Biol. 1985; 36: 217-229PubMed Google Scholar). Alternatively, the oligosaccharide residues may transiently protect corneodesmosin against proteolysis, during maturation of the SC. Indeed, sugars have been proposed to prevent premature desquamation by protecting the desmosome and corneodesmosome core against extracellular proteases (27Walsh A. Chapman S.J. Arch. Dermatol. Res. 1991; 283: 174-179Crossref PubMed Scopus (41) Google Scholar). Differences in glycosylation between species may also explain, at least in part, differences in corneodesmosin size previously observed in various mammals (20Montézin M. Simon M. Guerrin M. Serre G. Exp. Cell Res. 1997; 231: 132-140Crossref PubMed Scopus (20) Google Scholar).Internal peptide sequences of both the purified 52–56-kDa corneodesmosin and an immunoreactive fragment derived from it, indicated 100% matching over 25 amino acids with related sequences of the predicted product of the S gene. This gene, identified by CpG island analysis of the class I region of the human major histocompatibility complex, was shown to be located 160 kilobase pairs telomeric to HLA-C (24Zhou Y. Chaplin D.D. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: 9470-9474Crossref PubMed Scopus (85) Google Scholar), but its function was unknown. Like corneodesmosin, the S gene had been shown to be highly expressed in the epidermal granular keratinocytes (24Zhou Y. Chaplin D.D. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: 9470-9474Crossref PubMed Scopus (85) Google Scholar). Therefore, our results strongly suggest that corneodesmosin is the product of the S gene. This was confirmed by the recent cloning of the human corneodesmosin cDNA we performed by immunoscreening of an expression library. Consistently with the results reported here, the corneodesmosin cDNA sequence predicts a pI of 8.5, one N-glycosylation site and several protein kinase phosphorylation sites. 2M. Guerrin, M. Simon, M. Montézin, C. Vincent, and G. Serre, submitted for publication.Furthermore, in light of the strong association of the Cw6 HLA-C allele with Psoriasis vulgaris (28Henseler T. Christophers E. J. Am. Acad. Dermatol. 1985; 13: 450-456Abstract Full Text PDF PubMed Scopus (773) Google Scholar), corneodesmosin or allelic variants of the protein may be directly involved in the pathogenesis of this common skin disorder. Indeed, psoriasis is a chronic cutaneous disease involving inflammation, hyperproliferation, and a defective program of differentiation of epidermal cells, with in particular hyperkeratosis and impaired desquamation. No direct role for the MHC peptide has been shown yet. Moreover, abnormal corneodesmosomes have been observed in psoriasis (15Vicanova J. Mommaas A.M. Mulder A.A. Koerten H.K. Ponec M. Cell Tissue Res. 1996; 286: 115-122Crossref PubMed Scopus (38) Google Scholar), reinforcing the hypothesis of corneodesmosin involvement in the pathogenesis of the disease.In view of our previously published results (7Serre G. Mils V. Haftek M. Vincent C. Croute F. Réano A. Ouhayoun J.-P. Bettinger S. Soleilhavoup J.-P. J. Invest. Dermatol. 1991; 97: 1061-1072Abstract Full Text PDF PubMed Google Scholar, 20Montézin M. Simon M. Guerrin M. Serre G. Exp. Cell Res. 1997; 231: 132-140Crossref PubMed Scopus (20) Google Scholar, 21Mils V. Vincent C. Croute F. Serre G. J. Histochem. Cytochem. 1992; 40: 1329-1337Crossref PubMed Scopus (49) Google Scholar, 22Haftek M. Serre G. Mils V. Thivolet J. J. Histochem. Cytochem. 1991; 39: 1531-1538Crossref PubMed Scopus (62) Google Scholar, 23Lundström A. Serre G. Haftek M. Egelrud T. Arch. Dermatol. Res. 1994; 286: 369-375Crossref PubMed Scopus (94) Google Scholar) and of the present data, an overall scheme of human corneodesmosin processing during SC maturation can be proposed (Fig.8). The protein is synthesized in the upper spinous and/or lower granular keratinocytes under the form of a 52–56-kDa basic precursor that is extracted with nondenaturing hypotonic buffers. As observed by immunoelectron microscopy (7Serre G. Mils V. Haftek M. Vincent C. Croute F. Réano A. Ouhayoun J.-P. Bettinger S. Soleilhavoup J.-P. J. Invest. Dermatol. 1991; 97: 1061-1072Abstract Full Text PDF PubMed Google Scholar), it is exported via keratinosomes, probably into the extracellular space (or less likely to the plasma membrane) where it is associated to the desmosome core. Then, the presence of urea, at a concentration of at least 6 m (or SDS), becomes necessary for the protein to be solubilized. During corneocyte maturation, corneodesmosin is progressively proteolysed, a molecular form of 33 kDa being the major form extracted from the most superficial and less firmly attached corneocytes (7Serre G. Mils V. Haftek M. Vincent C. Croute F. Réano A. Ouhayoun J.-P. Bettinger S. Soleilhavoup J.-P. J. Invest. Dermatol. 1991; 97: 1061-1072Abstract Full Text PDF PubMed Google Scholar, 23Lundström A. Serre G. Haftek M. Egelrud T. Arch. Dermatol. Res. 1994; 286: 369-375Crossref PubMed Scopus (94) Google Scholar). 3M. Simon, M. Montézin, M. Guerrin, and G. Serre, unpublished observations. At the same time, the corneodesmosin fragments produced are more firmly bound to corneodesmosomes by intermolecular disulfide bonds and therefore require a reducing agent to be extracted. The fragments and/or the 52–56-kDa form are also cross-linked to cornified cell envelopes, on the external face of the structures, by other covalent bonds whose nature is unknown. Preliminary experiments suggest that corneodesmosin is not a substrate of epidermal transglutaminases, the intracytoplasmic enzymes responsible for the cornified cell envelope formation. Therefore, corneodesmosin could be ester-linked to the hydroxyacyl sphingosines bound to the outside of the cornified cell envelopes. In that case, the enzymes responsible for this linkage remain to be discovered. This linkage to the envelopes may enhance corneocyte cohesion and participate in SC resistance. Since human, pig, guinea pig, mouse, and rat corneodesmosins show similar locations in epidermis, biochemical characteristics and processing (20Montézin M. Simon M. Guerrin M. Serre G. Exp. Cell Res. 1997; 231: 132-140Crossref PubMed Scopus (20) Google Scholar), the model of human corneodesmosin maturation may probably be extended to these mammals.Maturation by proteolysis is not particular to corneodesmosin. Indeed, processing of filensin and other lens-specific proteins by proteolysis during lens fiber cell differentiation has been extensively described (Ref. 29Sandilands A. Prescott A.R. Hutcheson A.M. Quilan R.A. Casselman J.T. FitzGerald P.G. Eur. J. Cell Biol. 1995; 67: 238-253PubMed Google Scholar and references therein). More closely related, desmocollin 1 undergoes limited cleavage during the later stages of epidermal differentiation, resulting in the accumulation of stable NH2-terminal fragments in the SC (12King I.A. Wood M.J. Fryer P.R. J. Invest. Dermatol. 1989; 92: 22-26Abstract Full Text PDF PubMed Google Scholar). How corneodesmosin processing is important for the function of the molecule is not clear. However, we recently proposed that desmosomal proteins may be protected and desquamation inhibited until corneodesmosin is proteolysed to 33 kDa (23Lundström A. Serre G. Haftek M. Egelrud T. Arch. Dermatol. Res. 1994; 286: 369-375Crossref PubMed Scopus (94) Google Scholar).In conclusion, our results indicate that human corneodesmosin is closely related to the product of the S gene, a gene expressed in epidermis but whose function was unknown until now. Our data also show that corneodesmosin is a basic phosphorylated and glycosylated protein. During SC maturation it is covalently linked to the corneocyte-specific structures, i.e. the corneodesmosome core and the cornified cell envelopes. This association to the superstructure (formed by the envelopes linked to the intracellular matrix, and joined together by corneodesmosomes) responsible for SC cohesion clearly indicates the involvement of corneodesmosin in this process. We think that total degradation of corneodesmosome components and in particular corneodesmosin, at the skin surface, is required to allow cell detachment, i.e. desquamation. This hypothesis is now being tested in our laboratory. Corneocytes are anucleated "mummified" cells derived from keratinocytes during the late stages of terminal differentiation in cornified squamous epithelia such as the epidermis. They mainly consist of a cytokeratin-containing fibrous matrix surrounded by a highly resistant 15-nm thick protein structure called the cornified cell envelope. Stacking of the corneocytes at the outermost layer of the tissue, namely the stratum corneum (SC), 1The abbreviations used are: SC, stratum corneum; mAb, monoclonal antibody; PAGE, polyacrylamide gel electrophoresis; NEpHGE, non-equilibrium pH gel electrophoresis; ConA, concanavalin A.1The abbreviations used are: SC, stratum corneum; mAb, monoclonal antibody; PAGE, polyacrylamide gel electrophoresis; NEpHGE, non-equilibrium pH gel electrophoresis; ConA, concanavalin A. plays a critical role in the epidermal barrier function and in the mechanical protection of the body (1Holbrook K. Leigh I. Lane E. Watt F. The Keratinocyte Handbook. Cambridge University Press, Cambridge1994: 275-292Google Scholar, 2Elias P.M. J. Invest. Dermatol. 1983; 80: 44S-49SAbstract Full Text PDF PubMed Google Scholar, 3Roop D. Science. 1995; 267: 474-475Crossref PubMed Scopus (171) Google Scholar). During the normal desquamation process, the most superficial corneocytes are shed from the skin surface. The structures involved in the cohesion between individual corneocytes, and the mechanisms that lead to the detachment of these cells are poorly understood. Although intercellular structures, thought to be derived from desmosomes (4Allen T.D. Potten C.S. J. Ultrastruct. Res. 1975; 51: 94-105Crossref PubMed Scopus (71) Google Scholar, 5Menton D.N. Eisen A.Z. J. Ultrastruct. Res. 1971; 35: 247-264Crossref PubMed Scopus (80) Google Scholar), and called corneosomes or corneodesmosomes (6Chapman S.J. Walsh A. Arch. Dermatol. Res. 1990; 282: 304-310Crossref PubMed Scopus (123) Google Scholar, 7Serre G. Mils V. Haftek M. Vincent C. Croute F. Réano A. Ouhayoun J.-P. Bettinger S. Soleilhavoup J.-P. J. Invest. Dermatol. 1991; 97: 1061-1072Abstract Full Text PDF PubMed Google Scholar), have been described in the SC, they were initially considered as nonfunctional remnants. Recent studies have shown, however, their major importance in corneocyte cohesion, and there is now growing evidence that their degradation is a key event in the desquamation process (6Chapman S.J. Walsh A. Arch. Dermatol. Res. 1990; 282: 304-310Crossref PubMed Scopus (123) Google Scholar, 7Serre G. Mils V. Haftek M. Vincent C. Croute F. Réano A. Ouhayoun J.-P. Bettinger S. Soleilhavoup J.-P. J. Invest. Dermatol. 1991; 97: 1061-1072Abstract Full Text PDF PubMed Google Scholar, 8King C.S. Barton S.P. Nicholls S. Marks R. Br. J. Dermatol. 1979; 100: 165-172Crossref PubMed Scopus (70) Google Scholar, 9Skerrow C.J. Clelland D.G. Skerrow D. J. Cell Sci. 1989; 92: 667-677PubMed Google Scholar, 10Chapman S.J. Walsh A. Jackson S.M. Friedmann P.S. Arch. Dermatol. Res. 1991; 283: 167-173Crossref PubMed Scopus (82) Google Scholar, 11Rawlings A.V. Scott I.R. Harding C.R. Bowser P.A. J. Invest. Dermatol. 1994; 103: 731-740Abstract Full Text PDF PubMed Scopus (408) Google Scholar). In particular, a tight correlation seems to exist between cell dissociation and proteolysis of some corneodesmosomal components (12King I.A. Wood M.J. Fryer P.R. J. Invest. Dermatol. 1989; 92: 22-26Abstract Full Text PDF PubMed Google Scholar, 13Lundström A. Egelrud T. J. Invest. Dermatol. 1990; 94: 216-220Abstract Full Text PDF PubMed Google Scholar, 14Egelrud T. Hofer P.-A. Lundström A. Acta. Derm. Venereol. (Stockh.). 1988; 68: 93-97PubMed Google Scholar). Moreover, in plantar SC, where cell cohesion is very strong, corneodesmosomes can be detected over the whole corneocyte surface up to the top of the layer. In non-palmoplantar SC, where cohesion is weaker, they only persist at the periphery of the cells (6Chapman S.J. Walsh A. Arch. Dermatol. Res. 1990; 282: 304-310Crossref PubMed Scopus (123) Google Scholar, 7Serre G. Mils V. Haftek M. Vincent C. Croute F. Réano A. Ouhayoun J.-P. Bettinger S. Soleilhavoup J.-P. J. Invest. Dermatol. 1991; 97: 1061-1072Abstract Full Text PDF PubMed Google Scholar, 9Skerrow C.J. Clelland D.G. Skerrow D. J. Cell Sci. 1989; 92: 667-677PubMed Google Scholar). In psoriasis, various ichthyoses, and skin xerosis, characterized by an accumulation of corneocytes and by scaling, the number of corneodesmosomes is increased throughout the SC including the upper part (11Rawlings A.V. Scott I.R. Harding C.R. Bowser P.A. J. Invest. Dermatol. 1994; 103: 731-740Abstract Full Text PDF PubMed Scopus (408) Google Scholar, 15Vicanova J. Mommaas A.M. Mulder A.A. Koerten H.K. Ponec M. Cell Tissue Res. 1996; 286: 115-122Crossref PubMed Scopus (38) Google Scholar, 16Lundström A. Egelrud T. Arch. Dermatol. Res. 1990; 282: 234-237Crossref PubMed Scopus (49) Google Scholar). Several trypsin-like and chymotrypsin-like serine proteases, including the stratum corneum chymotryptic enzyme, are thought to be involved in the corneodesmosome proteolysis (17Suzuki Y. Nomura J. Hori J. Koyama J. Takahashi M. Horii I. Arch. Dermatol. Res. 1993; 285: 372-377Crossref PubMed Scopus (83) Google Scholar, 18Suzuki Y. Nomura J. Koyama J. Horii I. Arch. Dermatol. Res. 1994; 286: 249-253Crossref PubMed Scopus (92) Google Scholar, 19Hansson L. Strömqvist M. Bäckman A. Wallbrandt P. Carlstein A. Egelrud T. J. Biol. Chem. 1994; 269: 19420-19426Abstract Full Text PDF PubMed Google Scholar). Using three different monoclonal antibodies (mAbs), we recently identified a new corneodesmosome protein antigen expressed in various mammals including human, and we called it corneodesmosin (7Serre G. Mils V. Haftek M. Vincent C. Croute F. Réano A. Ouhayoun J.-P. Bettinger S. Soleilhavoup J.-P. J. Invest. Dermatol. 1991; 97: 1061-1072Abstract Full Text PDF PubMed Google Scholar, 20Montézin M. Simon M. Guerrin M. Serre G. Exp. Cell Res. 1997; 231: 132-140Crossref PubMed Scopus (20) Google Scholar). Corneodesmosin is only expressed in the cornified squamous epithelia,i.e. in man: epidermis, hard palate, epithelium, and inner root sheath of the hair follicle (7Serre G. Mils V. Haftek M. Vincent C. Croute F. Réano A. Ouhayoun J.-P. Bettinger S. Soleilhavoup J.-P. J. Invest. Dermatol. 1991; 97: 1061-1072Abstract Full Text PDF PubMed Google Scholar, 21Mils V. Vincent C. Croute F. Serre G. J. Histochem. Cytochem. 1992; 40: 1329-1337Crossref PubMed Scopus (49) Google Scholar). In human non-palmoplantar epidermis, the protein was shown to be located first in cytoplasmic vesicles, termed keratinosomes or lamellar bodies, of the upper spinous keratinocytes, then in the intercellular part of the desmosomes of the granular keratinocytes, these living cells being just beneath the SC. Lastly, it was detected in the core of corneodesmosomes in the SC (7Serre G. Mils V. Haftek M. Vincent C. Croute F. Réano A. Ouhayoun J.-P. Bettinger S. Soleilhavoup J.-P. J. Invest. Dermatol. 1991; 97: 1061-1072Abstract Full Text PDF PubMed Google Scholar). By immunofluorescence and immunoelectron microscopy, corneodesmosin was also shown to be bound to the cornified cell envelopes (22Haftek M. Serre G. Mils V. Thivolet J. J. Histochem. Cytochem. 1991; 39: 1531-1538Crossref PubMed Scopus (62) Google Scholar). When extracted from viable layers of human epidermis, corneodesmosin shows an apparent molecular mass of around 52 kDa (here designated as 52–56-kDa corneodesmosin), whereas a molecular form of 33 kDa is the major form extracted from the most superficial and less firmly attached corneocytes (7Serre G. Mils V. Haftek M. Vincent C. Croute F. Réano A. Ouhayoun J.-P. Bettinger S. Soleilhavoup J.-P. J. Invest. Dermatol. 1991; 97: 1061-1072Abstract Full Text PDF PubMed Google Scholar, 23Lundström A. Serre G. Haftek M. Egelrud T. Arch. Dermatol. Res. 1994; 286: 369-375Crossref PubMed Scopus (94) Google Scholar). Moreover, corneodesmosin is proteolysed (from 52–56 to 33 kDa) during SC maturation (23Lundström A. Serre G. Haftek M. Egelrud T. Arch. Dermatol. Res. 1994; 286: 369-375Crossref PubMed Scopus (94) Google Scholar). Some evidence was obtained in favor of a role of corneodesmosin in corneocyte cohesion, and its proteolysis was proposed as one of the major biochemical changes that lead to desquamation (23Lundström A. Serre G. Haftek M. Egelrud T. Arch. Dermatol. Res. 1994; 286: 369-375Crossref PubMed Scopus (94) Google Scholar). In this work, we have further characterized, purified, and partially sequenced the 52–56-kDa human corneodesmosin. The obtained sequences were contained in the product of the S gene, recently identified and located 160 kilobase pairs telomeric to HLA-C (24Zhou Y. Chaplin D.D. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: 9470-9474Crossref PubMed Scopus (85) Google Scholar). They confirmed our recent cloning of the human corneodesmosin cDNA. DISCUSSIONIn the studies presented above, we biochemically characterized and partially sequenced human corneodesmosin, a protein specific to cornified squamous epithelia where it is thought to play a major role in corneocyte cohesion and whose proteolysis seems to be a major prerequisite of desquamation.Our data indicate that human corneodesmosin is a glycoprotein containing mainly N-linked oligosaccharides that comprise ∼10% of the protein mass. Indeed, treatment withN-glycosidase F (a glycosidase specific forN-linked sugars) of the 52–56-kDa corneodesmosin induced a 5-kDa decrease in its apparent molecular weight, whereas extensive endo-α-N-acetylgalactosaminidase digestion (an enzyme specific for O-linked sugars) was without effect, even in the presence of neuraminidase and/or N-glycosidase F. Lectins were used to characterize the corneodesmosin carbohydrate moieties. Since the protein bound to ConA-Sepharose, and reacted with biotinylated P. sativum agglutinin, it may contain α-d-mannose and/or α-d-glucose. In addition, corneodesmosin strongly reacted with wheat germ agglutinin and, as expected for an N-glycosylated protein, may therefore contains β-N-acetylglucosamine groups. However, it seems to contain few or no galactose andN-acetylgalactosamine, since it did not react (or barely) with lectins specific for these carbohydrates. More extensive experiments will be necessary to precisely identify the oligosaccharide residues linked to corneodesmosin, and to characterize their linkage. In fact these residues may participate in the adhesion properties of the protein, as described for other corneodesmosomal adhesive proteins, desmogleins and desmocollins (26Kapprell H.-P. Cowin P. Franke W.W. Ponstingl H. Eur. J. Cell Biol. 1985; 36: 217-229PubMed Google Scholar). Alternatively, the oligosaccharide residues may transiently protect corneodesmosin against proteolysis, during maturation of the SC. Indeed, sugars have been proposed to prevent premature desquamation by protecting the desmosome and corneodesmosome core against extracellular proteases (27Walsh A. Chapman S.J. Arch. Dermatol. Res. 1991; 283: 174-179Crossref PubMed Scopus (41) Google Scholar). Differences in glycosylation between species may also explain, at least in part, differences in corneodesmosin size previously observed in various mammals (20Montézin M. Simon M. Guerrin M. Serre G. Exp. Cell Res. 1997; 231: 132-140Crossref PubMed Scopus (20) Google Scholar).Internal peptide sequences of both the purified 52–56-kDa corneodesmosin and an immunoreactive fragment derived from it, indicated 100% matching over 25 amino acids with related sequences of the predicted product of the S gene. This gene, identified by CpG island analysis of the class I region of the human major histocompatibility complex, was shown to be located 160 kilobase pairs telomeric to HLA-C (24Zhou Y. Chaplin D.D. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: 9470-9474Crossref PubMed Scopus (85) Google Scholar), but its function was unknown. Like corneodesmosin, the S gene had been shown to be highly expressed in the epidermal granular keratinocytes (24Zhou Y. Chaplin D.D. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: 9470-9474Crossref PubMed Scopus (85) Google Scholar). Therefore, our results strongly suggest that corneodesmosin is the product of the S gene. This was confirmed by the recent cloning of the human corneodesmosin cDNA we performed by immunoscreening of an expression library. Consistently with the results reported here, the corneodesmosin cDNA sequence predicts a pI of 8.5, one N-glycosylation site and several protein kinase phosphorylation sites. 2M. Guerrin, M. Simon, M. Montézin, C. Vincent, and G. Serre, submitted for publication.Furthermore, in light of the strong association of the Cw6 HLA-C allele with Psoriasis vulgaris (28Henseler T. Christophers E. J. Am. Acad. Dermatol. 1985; 13: 450-456Abstract Full Text PDF PubMed Scopus (773) Google Scholar), corneodesmosin or allelic variants of the protein may be directly involved in the pathogenesis of this common skin disorder. Indeed, psoriasis is a chronic cutaneous disease involving inflammation, hyperproliferation, and a defective program of differentiation of epidermal cells, with in particular hyperkeratosis and impaired desquamation. No direct role for the MHC peptide has been shown yet. Moreover, abnormal corneodesmosomes have been observed in psoriasis (15Vicanova J. Mommaas A.M. Mulder A.A. Koerten H.K. Ponec M. Cell Tissue Res. 1996; 286: 115-122Crossref PubMed Scopus (38) Google Scholar), reinforcing the hypothesis of corneodesmosin involvement in the pathogenesis of the disease.In view of our previously published results (7Serre G. Mils V. Haftek M. Vincent C. Croute F. Réano A. Ouhayoun J.-P. Bettinger S. Soleilhavoup J.-P. J. Invest. Dermatol. 1991; 97: 1061-1072Abstract Full Text PDF PubMed Google Scholar, 20Montézin M. Simon M. Guerrin M. Serre G. Exp. Cell Res. 1997; 231: 132-140Crossref PubMed Scopus (20) Google Scholar, 21Mils V. Vincent C. Croute F. Serre G. J. Histochem. Cytochem. 1992; 40: 1329-1337Crossref PubMed Scopus (49) Google Scholar, 22Haftek M. Serre G. Mils V. Thivolet J. J. Histochem. Cytochem. 1991; 39: 1531-1538Crossref PubMed Scopus (62) Google Scholar, 23Lundström A. Serre G. Haftek M. Egelrud T. Arch. Dermatol. Res. 1994; 286: 369-375Crossref PubMed Scopus (94) Google Scholar) and of the present data, an overall scheme of human corneodesmosin processing during SC maturation can be proposed (Fig.8). The protein is synthesized in the upper spinous and/or lower granular keratinocytes under the form of a 52–56-kDa basic precursor that is extracted with nondenaturing hypotonic buffers. As observed by immunoelectron microscopy (7Serre G. Mils V. Haftek M. Vincent C. Croute F. Réano A. Ouhayoun J.-P. Bettinger S. Soleilhavoup J.-P. J. Invest. Dermatol. 1991; 97: 1061-1072Abstract Full Text PDF PubMed Google Scholar), it is exported via keratinosomes, probably into the extracellular space (or less likely to the plasma membrane) where it is associated to the desmosome core. Then, the presence of urea, at a concentration of at least 6 m (or SDS), becomes necessary for the protein to be solubilized. During corneocyte maturation, corneodesmosin is progressively proteolysed, a molecular form of 33 kDa being the major form extracted from the most superficial and less firmly attached corneocytes (7Serre G. Mils V. Haftek M. Vincent C. Croute F. Réano A. Ouhayoun J.-P. Bettinger S. Soleilhavoup J.-P. J. Invest. Dermatol. 1991; 97: 1061-1072Abstract Full Text PDF PubMed Google Scholar, 23Lundström A. Serre G. Haftek M. Egelrud T. Arch. Dermatol. Res. 1994; 286: 369-375Crossref PubMed Scopus (94) Google Scholar). 3M. Simon, M. Montézin, M. Guerrin, and G. Serre, unpublished observations. At the same time, the corneodesmosin fragments produced are more firmly bound to corneodesmosomes by intermolecular disulfide bonds and therefore require a reducing agent to be extracted. The fragments and/or the 52–56-kDa form are also cross-linked to cornified cell envelopes, on the external face of the structures, by other covalent bonds whose nature is unknown. Preliminary experiments suggest that corneodesmosin is not a substrate of epidermal transglutaminases, the intracytoplasmic enzymes responsible for the cornified cell envelope formation. Therefore, corneodesmosin could be ester-linked to the hydroxyacyl sphingosines bound to the outside of the cornified cell envelopes. In that case, the enzymes responsible for this linkage remain to be discovered. This linkage to the envelopes may enhance corneocyte cohesion and participate in SC resistance. Since human, pig, guinea pig, mouse, and rat corneodesmosins show similar locations in epidermis, biochemical characteristics and processing (20Montézin M. Simon M. Guerrin M. Serre G. Exp. Cell Res. 1997; 231: 132-140Crossref PubMed Scopus (20) Google Scholar), the model of human corneodesmosin maturation may probably be extended to these mammals.Maturation by proteolysis is not particular to corneodesmosin. Indeed, processing of filensin and other lens-specific proteins by proteolysis during lens fiber cell differentiation has been extensively described (Ref. 29Sandilands A. Prescott A.R. Hutcheson A.M. Quilan R.A. Casselman J.T. FitzGerald P.G. Eur. J. Cell Biol. 1995; 67: 238-253PubMed Google Scholar and references therein). More closely related, desmocollin 1 undergoes limited cleavage during the later stages of epidermal differentiation, resulting in the accumulation of stable NH2-terminal fragments in the SC (12King I.A. Wood M.J. Fryer P.R. J. Invest. Dermatol. 1989; 92: 22-26Abstract Full Text PDF PubMed Google Scholar). How corneodesmosin processing is important for the function of the molecule is not clear. However, we recently proposed that desmosomal proteins may be protected and desquamation inhibited until corneodesmosin is proteolysed to 33 kDa (23Lundström A. Serre G. Haftek M. Egelrud T. Arch. Dermatol. Res. 1994; 286: 369-375Crossref PubMed Scopus (94) Google Scholar).In conclusion, our results indicate that human corneodesmosin is closely related to the product of the S gene, a gene expressed in epidermis but whose function was unknown until now. Our data also show that corneodesmosin is a basic phosphorylated and glycosylated protein. During SC maturation it is covalently linked to the corneocyte-specific structures, i.e. the corneodesmosome core and the cornified cell envelopes. This association to the superstructure (formed by the envelopes linked to the intracellular matrix, and joined together by corneodesmosomes) responsible for SC cohesion clearly indicates the involvement of corneodesmosin in this process. We think that total degradation of corneodesmosome components and in particular corneodesmosin, at the skin surface, is required to allow cell detachment, i.e. desquamation. This hypothesis is now being tested in our laboratory. In the studies presented above, we biochemically characterized and partially sequenced human corneodesmosin, a protein specific to cornified squamous epithelia where it is thought to play a major role in corneocyte cohesion and whose proteolysis seems to be a major prerequisite of desquamation. Our data indicate that human corneodesmosin is a glycoprotein containing mainly N-linked oligosaccharides that comprise ∼10% of the protein mass. Indeed, treatment withN-glycosidase F (a glycosidase specific forN-linked sugars) of the 52–56-kDa corneodesmosin induced a 5-kDa decrease in its apparent molecular weight, whereas extensive endo-α-N-acetylgalactosaminidase digestion (an enzyme specific for O-linked sugars) was without effect, even in the presence of neuraminidase and/or N-glycosidase F. Lectins were used to characterize the corneodesmosin carbohydrate moieties. Since the protein bound to ConA-Sepharose, and reacted with biotinylated P. sativum agglutinin, it may contain α-d-mannose and/or α-d-glucose. In addition, corneodesmosin strongly reacted with wheat germ agglutinin and, as expected for an N-glycosylated protein, may therefore contains β-N-acetylglucosamine groups. However, it seems to contain few or no galactose andN-acetylgalactosamine, since it did not react (or barely) with lectins specific for these carbohydrates. More extensive experiments will be necessary to precisely identify the oligosaccharide residues linked to corneodesmosin, and to characterize their linkage. In fact these residues may participate in the adhesion properties of the protein, as described for other corneodesmosomal adhesive proteins, desmogleins and desmocollins (26Kapprell H.-P. Cowin P. Franke W.W. Ponstingl H. Eur. J. Cell Biol. 1985; 36: 217-229PubMed Google Scholar). Alternatively, the oligosaccharide residues may transiently protect corneodesmosin against proteolysis, during maturation of the SC. Indeed, sugars have been proposed to prevent premature desquamation by protecting the desmosome and corneodesmosome core against extracellular proteases (27Walsh A. Chapman S.J. Arch. Dermatol. Res. 1991; 283: 174-179Crossref PubMed Scopus (41) Google Scholar). Differences in glycosylation between species may also explain, at least in part, differences in corneodesmosin size previously observed in various mammals (20Montézin M. Simon M. Guerrin M. Serre G. Exp. Cell Res. 1997; 231: 132-140Crossref PubMed Scopus (20) Google Scholar). Internal peptide sequences of both the purified 52–56-kDa corneodesmosin and an immunoreactive fragment derived from it, indicated 100% matching over 25 amino acids with related sequences of the predicted product of the S gene. This gene, identified by CpG island analysis of the class I region of the human major histocompatibility complex, was shown to be located 160 kilobase pairs telomeric to HLA-C (24Zhou Y. Chaplin D.D. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: 9470-9474Crossref PubMed Scopus (85) Google Scholar), but its function was unknown. Like corneodesmosin, the S gene had been shown to be highly expressed in the epidermal granular keratinocytes (24Zhou Y. Chaplin D.D. Proc. Natl. Acad. Sci. U. S. A. 1993; 90: 9470-9474Crossref PubMed Scopus (85) Google Scholar). Therefore, our results strongly suggest that corneodesmosin is the product of the S gene. This was confirmed by the recent cloning of the human corneodesmosin cDNA we performed by immunoscreening of an expression library. Consistently with the results reported here, the corneodesmosin cDNA sequence predicts a pI of 8.5, one N-glycosylation site and several protein kinase phosphorylation sites. 2M. Guerrin, M. Simon, M. Montézin, C. Vincent, and G. Serre, submitted for publication. Furthermore, in light of the strong association of the Cw6 HLA-C allele with Psoriasis vulgaris (28Henseler T. Christophers E. J. Am. Acad. Dermatol. 1985; 13: 450-456Abstract Full Text PDF PubMed Scopus (773) Google Scholar), corneodesmosin or allelic variants of the protein may be directly involved in the pathogenesis of this common skin disorder. Indeed, psoriasis is a chronic cutaneous disease involving inflammation, hyperproliferation, and a defective program of differentiation of epidermal cells, with in particular hyperkeratosis and impaired desquamation. No direct role for the MHC peptide has been shown yet. Moreover, abnormal corneodesmosomes have been observed in psoriasis (15Vicanova J. Mommaas A.M. Mulder A.A. Koerten H.K. Ponec M. Cell Tissue Res. 1996; 286: 115-122Crossref PubMed Scopus (38) Google Scholar), reinforcing the hypothesis of corneodesmosin involvement in the pathogenesis of the disease. In view of our previously published results (7Serre G. Mils V. Haftek M. Vincent C. Croute F. Réano A. Ouhayoun J.-P. Bettinger S. Soleilhavoup J.-P. J. Invest. Dermatol. 1991; 97: 1061-1072Abstract Full Text PDF PubMed Google Scholar, 20Montézin M. Simon M. Guerrin M. Serre G. Exp. Cell Res. 1997; 231: 132-140Crossref PubMed Scopus (20) Google Scholar, 21Mils V. Vincent C. Croute F. Serre G. J. Histochem. Cytochem. 1992; 40: 1329-1337Crossref PubMed Scopus (49) Google Scholar, 22Haftek M. Serre G. Mils V. Thivolet J. J. Histochem. Cytochem. 1991; 39: 1531-1538Crossref PubMed Scopus (62) Google Scholar, 23Lundström A. Serre G. Haftek M. Egelrud T. Arch. Dermatol. Res. 1994; 286: 369-375Crossref PubMed Scopus (94) Google Scholar) and of the present data, an overall scheme of human corneodesmosin processing during SC maturation can be proposed (Fig.8). The protein is synthesized in the upper spinous and/or lower granular keratinocytes under the form of a 52–56-kDa basic precursor that is extracted with nondenaturing hypotonic buffers. As observed by immunoelectron microscopy (7Serre G. Mils V. Haftek M. Vincent C. Croute F. Réano A. Ouhayoun J.-P. Bettinger S. Soleilhavoup J.-P. J. Invest. Dermatol. 1991; 97: 1061-1072Abstract Full Text PDF PubMed Google Scholar), it is exported via keratinosomes, probably into the extracellular space (or less likely to the plasma membrane) where it is associated to the desmosome core. Then, the presence of urea, at a concentration of at least 6 m (or SDS), becomes necessary for the protein to be solubilized. During corneocyte maturation, corneodesmosin is progressively proteolysed, a molecular form of 33 kDa being the major form extracted from the most superficial and less firmly attached corneocytes (7Serre G. Mils V. Haftek M. Vincent C. Croute F. Réano A. Ouhayoun J.-P. Bettinger S. Soleilhavoup J.-P. J. Invest. Dermatol. 1991; 97: 1061-1072Abstract Full Text PDF PubMed Google Scholar, 23Lundström A. Serre G. Haftek M. Egelrud T. Arch. Dermatol. Res. 1994; 286: 369-375Crossref PubMed Scopus (94) Google Scholar). 3M. Simon, M. Montézin, M. Guerrin, and G. Serre, unpublished observations. At the same time, the corneodesmosin fragments produced are more firmly bound to corneodesmosomes by intermolecular disulfide bonds and therefore require a reducing agent to be extracted. The fragments and/or the 52–56-kDa form are also cross-linked to cornified cell envelopes, on the external face of the structures, by other covalent bonds whose nature is unknown. Preliminary experiments suggest that corneodesmosin is not a substrate of epidermal transglutaminases, the intracytoplasmic enzymes responsible for the cornified cell envelope formation. Therefore, corneodesmosin could be ester-linked to the hydroxyacyl sphingosines bound to the outside of the cornified cell envelopes. In that case, the enzymes responsible for this linkage remain to be discovered. This linkage to the envelopes may enhance corneocyte cohesion and participate in SC resistance. Since human, pig, guinea pig, mouse, and rat corneodesmosins show similar locations in epidermis, biochemical characteristics and processing (20Montézin M. Simon M. Guerrin M. Serre G. Exp. Cell Res. 1997; 231: 132-140Crossref PubMed Scopus (20) Google Scholar), the model of human corneodesmosin maturation may probably be extended to these mammals. Maturation by proteolysis is not particular to corneodesmosin. Indeed, processing of filensin and other lens-specific proteins by proteolysis during lens fiber cell differentiation has been extensively described (Ref. 29Sandilands A. Prescott A.R. Hutcheson A.M. Quilan R.A. Casselman J.T. FitzGerald P.G. Eur. J. Cell Biol. 1995; 67: 238-253PubMed Google Scholar and references therein). More closely related, desmocollin 1 undergoes limited cleavage during the later stages of epidermal differentiation, resulting in the accumulation of stable NH2-terminal fragments in the SC (12King I.A. Wood M.J. Fryer P.R. J. Invest. Dermatol. 1989; 92: 22-26Abstract Full Text PDF PubMed Google Scholar). How corneodesmosin processing is important for the function of the molecule is not clear. However, we recently proposed that desmosomal proteins may be protected and desquamation inhibited until corneodesmosin is proteolysed to 33 kDa (23Lundström A. Serre G. Haftek M. Egelrud T. Arch. Dermatol. Res. 1994; 286: 369-375Crossref PubMed Scopus (94) Google Scholar). In conclusion, our results indicate that human corneodesmosin is closely related to the product of the S gene, a gene expressed in epidermis but whose function was unknown until now. Our data also show that corneodesmosin is a basic phosphorylated and glycosylated protein. During SC maturation it is covalently linked to the corneocyte-specific structures, i.e. the corneodesmosome core and the cornified cell envelopes. This association to the superstructure (formed by the envelopes linked to the intracellular matrix, and joined together by corneodesmosomes) responsible for SC cohesion clearly indicates the involvement of corneodesmosin in this process. We think that total degradation of corneodesmosome components and in particular corneodesmosin, at the skin surface, is required to allow cell detachment, i.e. desquamation. This hypothesis is now being tested in our laboratory. We thank Professor M. Costagliola (Service de Chirurgie Plastique, CHU Rangueil, Toulouse, France) for providing us with human skin. We are indebted to Doctor C. Vincent for line drawings. The technical assistance of C. Pons, M. Goasampis, M.-F. Isaı̈a, and M.-P. Rué is gratefully acknowledged.
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