RESUMO
Photosensitivity disorders are caused by a variety of mechanisms. Three common themes are as follows: excess chromophore allowing visible light energy to cause photodynamic damage, reduced DNA repair capacity to UV-induced DNA damage, and enhanced sensitivity to light-induced allergens mediated immunologically. Although the cause of each condition may be known, the precise pathogenesis underlying the photosensitivity has taken longer to understand. By focussing on three clinical disorders under each of these themes, we have explored the following: why erythropoietic protoporphyria differs so markedly from the other cutaneous porphyrias; how a DNA repair defect was eventually revealed to be the underlying cause of the vitamin B3 deficiency disorder of pellagra; an immunological explanation for the over reactivity to photoallergens in chronic actinic dermatitis.
Assuntos
Transtornos de Fotossensibilidade/etiologia , Transtornos de Fotossensibilidade/fisiopatologia , Dano ao DNA/fisiologia , Dermatite Fotoalérgica/fisiopatologia , Humanos , Pelagra/etiologia , Pelagra/fisiopatologia , Protoporfiria Eritropoética/etiologia , Protoporfiria Eritropoética/fisiopatologiaRESUMO
BACKGROUND: Atopic dermatitis (AD) is a major inflammatory condition of the skin caused by inherited skin barrier deficiency, with mutations in the filaggrin gene predisposing to development of AD. Support for barrier deficiency initiating AD came from flaky tail mice, which have a frameshift mutation in Flg and also carry an unknown gene, matted, causing a matted hair phenotype. OBJECTIVE: We sought to identify the matted mutant gene in mice and further define whether mutations in the human gene were associated with AD. METHODS: A mouse genetics approach was used to separate the matted and Flg mutations to produce congenic single-mutant strains for genetic and immunologic analysis. Next-generation sequencing was used to identify the matted gene. Five independently recruited AD case collections were analyzed to define associations between single nucleotide polymorphisms (SNPs) in the human gene and AD. RESULTS: The matted phenotype in flaky tail mice is due to a mutation in the Tmem79/Matt gene, with no expression of the encoded protein mattrin in the skin of mutant mice. Matt(ft) mice spontaneously have dermatitis and atopy caused by a defective skin barrier, with mutant mice having systemic sensitization after cutaneous challenge with house dust mite allergens. Meta-analysis of 4,245 AD cases and 10,558 population-matched control subjects showed that a missense SNP, rs6684514, [corrected] in the human MATT gene has a small but significant association with AD. CONCLUSION: In mice mutations in Matt cause a defective skin barrier and spontaneous dermatitis and atopy. A common SNP in MATT has an association with AD in human subjects.
Assuntos
Dermatite Atópica/genética , Predisposição Genética para Doença , Proteínas de Membrana/genética , Animais , Dermatite Atópica/imunologia , Dermatite Atópica/patologia , Proteínas Filagrinas , Expressão Gênica , Humanos , Masculino , Camundongos , Mutação , Fenótipo , Mapeamento Físico do Cromossomo , Polimorfismo de Nucleotídeo Único , Pele/metabolismo , Pele/patologiaRESUMO
Palmitoylation is a key post-translational modification mediated by a family of DHHC-containing palmitoyl acyl-transferases (PATs). Unlike other lipid modifications, palmitoylation is reversible and thus often regulates dynamic protein interactions. We find that the mouse hair loss mutant, depilated, (dep) is due to a single amino acid deletion in the PAT, Zdhhc21, resulting in protein mislocalization and loss of palmitoylation activity. We examined expression of Zdhhc21 protein in skin and find it restricted to specific hair lineages. Loss of Zdhhc21 function results in delayed hair shaft differentiation, at the site of expression of the gene, but also leads to hyperplasia of the interfollicular epidermis (IFE) and sebaceous glands, distant from the expression site. The specific delay in follicle differentiation is associated with attenuated anagen propagation and is reflected by decreased levels of Lef1, nuclear beta-catenin, and Foxn1 in hair shaft progenitors. In the thickened basal compartment of mutant IFE, phospho-ERK and cell proliferation are increased, suggesting increased signaling through EGFR or integrin-related receptors, with a parallel reduction in expression of the key differentiation factor Gata3. We show that the Src-family kinase, Fyn, involved in keratinocyte differentiation, is a direct palmitoylation target of Zdhhc21 and is mislocalized in mutant follicles. This study is the first to demonstrate a key role for palmitoylation in regulating developmental signals in mammalian tissue homeostasis.
Assuntos
Aciltransferases/genética , Diferenciação Celular , Células Epidérmicas , Folículo Piloso/citologia , Homeostase , Lipoilação/fisiologia , Animais , Mutação da Fase de Leitura , Camundongos , Processamento de Proteína Pós-Traducional , Proteínas Proto-Oncogênicas c-fyn/metabolismoRESUMO
This study evaluated in vivo the potential of optical coherence tomography (OCT) to determine changes in thickness of the epidermis in response to the topically applied anti-psoriatics betamethasone dipropionate (BD), salicylic acid (SA) and also fish oil (FO). GsdmA3Dfl/+ mice have an inflammatory hair loss phenotype that includes hyperproliferation and epidermal thickening, hence a potential psoriasis model. Changes in epidermal thickness were evaluated over a period of 10 days, with the mice treated with combined BD + SA, FO + SA and BD + FO + SA. The data were validated with conventional measurement using H&E staining coupled with microscopy. Initial baseline measurement revealed an average epidermal thickness of 26.92 ± 1.17 µm. After 10 days of treatment with BD, the average epidermal thickness was reduced by 38.8% (P = 0.0001), and inversely, treatment with FO resulted in an unexpected 105% increase (P = 0.0001) in epidermal thickness. Combined BD + FO treatment did not cause any significant change (P = 0.3755) and may further indicate opposing effects on keratinocyte proliferation. The data obtained using OCT were statistically the same as those obtained by H&E/microscopy (P = 0.4325), supporting a greater role for OCT in dermatological studies, while also allowing a reduction in the number of animals used in such studies as sacrifice at individual timepoints is not necessary.
Assuntos
Betametasona/análogos & derivados , Epiderme/efeitos dos fármacos , Óleos de Peixe/farmacologia , Proteínas/genética , Psoríase/tratamento farmacológico , Ácido Salicílico/farmacologia , Tomografia de Coerência Óptica , Animais , Anti-Inflamatórios/administração & dosagem , Anti-Inflamatórios/farmacologia , Anti-Inflamatórios/uso terapêutico , Betametasona/administração & dosagem , Betametasona/farmacologia , Betametasona/uso terapêutico , Modelos Animais de Doenças , Quimioterapia Combinada , Epiderme/patologia , Óleos de Peixe/administração & dosagem , Óleos de Peixe/uso terapêutico , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Mutantes , Psoríase/genética , Ácido Salicílico/administração & dosagem , Ácido Salicílico/uso terapêutico , Coloração e Rotulagem/métodos , Resultado do TratamentoRESUMO
A large number of mutations in keratin genes underlie inherited tissue fragility disorders of epithelia. The genotype-phenotype correlations emerging from these studies provide a rich source of information about the function of keratins that would have taken decades to achieve by a purely transgenic approach. Human disease studies are being supplemented by engineered mouse mutant studies, which give access to the effects of genetic alterations unlikely to occur naturally. Evidence is emerging that the great diversity of keratins might be required to enable cells to adapt their structure in response to different signalling pathways.
Assuntos
Epiderme/patologia , Epidermólise Bolhosa/genética , Epitélio/patologia , Hiperceratose Epidermolítica/genética , Queratinas/genética , Camundongos/genética , Mutagênese , Mutação/genética , Animais , Epidermólise Bolhosa/patologia , Genótipo , Humanos , Hiperceratose Epidermolítica/patologia , Camundongos Knockout , Modelos Genéticos , FenótipoRESUMO
Plantar keratodermas can arise due to a variety of genetically inherited mutations. The need to distinguish between different plantar keratoderma disorders is becoming increasingly apparent because there is evidence that they do not respond identically to treatment. Diagnosis can be aided by observation of other clinical manifestations, such as palmar keratoderma, more widespread hyperkeratosis of the epidermis, hair and nail dystrophies, or erythroderma. However, there are frequent cases of plantar keratoderma that occur in isolation. This review focuses on the rare autosomal dominant keratin disorder pachyonychia congenita, which presents with particularly painful plantar keratoderma for which there is no specific treatment. Typically, patients regularly trim/pare/file/grind their calluses and file/grind/clip their nails. Topical agents, including keratolytics (eg, salicylic acid, urea) and moisturizers, can provide limited benefit by softening the skin. For some patients, retinoids help to thin calluses but may lead to increased pain. This finding has stimulated a drive for alternative treatment options, from gene therapy to alternative nongenetic methods that focus on novel findings regarding the pathogenesis of pachyonychia congenita and the function of the underlying genes.
Assuntos
Ceratodermia Palmar e Plantar/diagnóstico , Ceratodermia Palmar e Plantar/epidemiologia , Ceratodermia Palmar e Plantar/terapia , Paquioníquia Congênita/epidemiologia , Paquioníquia Congênita/terapia , Comorbidade , Gerenciamento Clínico , Feminino , Humanos , Ceratodermia Palmar e Plantar/psicologia , Masculino , Paquioníquia Congênita/diagnóstico , Manejo da Dor , Prognóstico , Qualidade de Vida , Medição de Risco , Índice de Gravidade de Doença , Papel do DoenteRESUMO
Psoriasis is an incurable autoimmune disease characterized by patches of abnormal red, itchy and scaly skin. This work examined the modulation of inflammation, hyperproliferation and immune cell markers following topical application of fish oil (FO) in comparison to the antipsoriatic agents, betamethasone dipropionate (BD) and salicylic acid (SA), to GsdmA3(Dfl)/+ mice, a hair loss mutant which also exhibits epidermal hyperproliferation akin to psoriasis. The mice were dosed with 100 mg of the test formulation and after 10 days, the mice were sacrificed, skin sections excised and subjected to immunohistochemical determination of COX-2, K17 and MAC-1; and immunofluorescence of Ki-67. Unchanged expression of the proinflammatory enzyme COX-2 was observed in all treatments, suggesting the noninvolvement of COX-2 in the aetiology of cutaneous aberration seen in GsdmA3(Dfl)/+ mice. Intense staining of K17 and MAC-1 in the FO-treated group mirrored the epidermal thickening seen observed in live mice by optical coherence tomography (OCT). The ratio of Ki-67-positive nuclei per 100 basal cells indicated that hyperproliferation of keratinocytes occurred in FO-treated mice and the opposite was true for BD-treated mice. There was a positive correlation (R (2) 0.995) between Ki-67 and the epidermal thickness data observed previously. In all immunochemical procedures, the combined BD, SA and FO formulation did not show any significant difference with the control group, reflecting observations seen previously. In conclusion, the epidermal changes observed following topical FO treatment on GsdmA3(Dfl)/+ mice involves an increase in cellular proliferation and macrophages, although COX-2 does not appear to play an important role.
RESUMO
Defolliculated (Dfl) is a spontaneous mouse mutant with a hair-loss phenotype that includes altered sebaceous gland differentiation, short hair shafts, aberrant catagen stage of the hair cycle, and eventual loss of the hair follicle. Recently a similar mutant, finnegan (Fgn), with an identical phenotype was discovered during a phenotypic screen for mutations induced by chemical mutagenesis. The gene underlying the phenotype of both finnegan and defolliculated has been mapped to chromosome 11 and here we show that both mice harbor mutations in gasdermin 3 (Gsdm3), a gene of unknown function. Gsdm3(Dfl) is a B2 insertion near the 3' splice site of exon 7 and Gsdm3(Fgn) is a point mutation T278P. To investigate the role of the gasdermin gene family an antiserum was raised to a peptide highly homologous to all three mouse gasdermins and human gasdermin. Immunohistochemical analysis revealed that gasdermins are expressed specifically in cells at advanced stages of differentiation in the upper epidermis, the differentiating inner root sheath and hair shaft and in the most mature sebocytes of the sebaceous gland and preputial, meibomium, ceruminous gland, and anal glands. This expression pattern suggests a role for gasdermins in differentiation of the epidermis and its appendages.
Assuntos
Alopecia/genética , Alopecia/patologia , Folículo Piloso/patologia , Proteínas/genética , Glândulas Sebáceas/patologia , Alopecia/fisiopatologia , Animais , Anticorpos , Especificidade de Anticorpos , Sequência de Bases , Carcinoma de Células Escamosas , Diferenciação Celular , Linhagem Celular Tumoral , Folículo Piloso/fisiopatologia , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Mutantes , Dados de Sequência Molecular , Fenótipo , Proteínas/imunologia , Proteínas/metabolismo , Glândulas Sebáceas/fisiopatologia , Neoplasias CutâneasRESUMO
Defolliculated is a novel spontaneous mouse mutation that maps to chromosome 11 close to the type I keratin locus. Histology shows abnormal differentiation of the sebaceous gland, with the sebocytes producing little or no sebum and undergoing abnormal cornification. The hair follicles fail to regress during catagen leading to abnormally long follicles. In contrast the hair shafts are shorter than normal, suggesting altered differentiation or proliferation of matrix cells during anagen. The shafts emerge from the follicle with cornified material still attached. The dermis contains increased numbers of immune cells, including T cells (CD4-positive), macrophages, and mast cells, at all time points examined. Complete elimination of all pelage and tail follicles occurs after two to three hair cycles, apparently by necrosis. Defolliculated may be a useful model for determining further functions of the sebaceous gland, and for understanding the regulation of catagen and hair follicle immunology.
Assuntos
Alopecia/genética , Alopecia/patologia , Folículo Piloso/anormalidades , Camundongos Mutantes/anormalidades , Glândulas Sebáceas/anormalidades , Animais , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Cromossomos , Epiderme/anormalidades , Epiderme/imunologia , Epiderme/patologia , Genes Dominantes , Folículo Piloso/imunologia , Folículo Piloso/patologia , Camundongos , Camundongos Endogâmicos BALB C , Fenótipo , Glândulas Sebáceas/imunologia , Glândulas Sebáceas/patologiaRESUMO
BACKGROUND: Recently, much interest has been generated in the use of intense pulsed light (IPL) sources in the treatment of various skin conditions. However, the underlying mechanism for its therapeutic action has not been elucidated. OBJECTIVE: To investigate the effect of IPL on the in vivo expression of transforming growth factor beta1 (TGF-ß1) and on the immunolocalization of Smad3 in biopsies obtained from perilesional skin in patients with mild-to-moderate inflammatory acne vulgaris. METHODS: Biopsies obtained from 20 patients with inflammatory acne vulgaris at baseline (B1) and post-IPL treatment (B2 = 48 h after first treatment and B3 = 1 week after final treatment) were immunohistochemically analyzed to determine the expression of TGF-ß1 and the immunolocalization of Smad3. Digital images were semiquantitatively assessed using image analysis software. RESULTS: Intense pulsed light elicited a consistent increase in epidermal TGF-ß1 expression (B2 vs. B1: P = 0.004 and B3 vs. B1: P = 0.007). Furthermore, it resulted in enhanced nuclear immunolocalization of Smad3 (B2 vs. B1: epidermis, P = 0.000055 and dermis, P = 0.014; B3 vs. B1: epidermis, P = 0.00024 and dermis, P = 0.008). CONCLUSION: Intense pulsed light upregulates TGF-ß1/Smad3 signaling in perilesional skin obtained from patients with mild-to-moderate inflammatory acne vulgaris. Further experiments on lesional skin and downstream effects are warranted to determine whether it may play a role in IPL-induced resolution of acne vulgaris.
Assuntos
Acne Vulgar/metabolismo , Acne Vulgar/patologia , Terapia de Luz Pulsada Intensa , Transdução de Sinais/efeitos da radiação , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Acne Vulgar/terapia , Núcleo Celular/química , Derme/química , Suscetibilidade a Doenças/metabolismo , Suscetibilidade a Doenças/terapia , Epiderme/química , Humanos , Proteína Smad3/análiseRESUMO
Defolliculated (Gsdma3(Dfl)/+) mice have a hair loss phenotype that involves an aberrant hair cycle, altered sebaceous gland differentiation with reduced sebum production, chronic inflammation, and ultimately the loss of the hair follicle. Hair loss in these mice is similar to that seen in primary cicatricial, or scarring alopecias in which immune targeting of hair follicle stem cells has been proposed as a key factor resulting in permanent hair follicle destruction. In this study we examine the mechanism of hair loss in GsdmA3(Dfl)/+ mice. Aberrant expression patterns of stem cell markers during the hair cycle, in addition to aberrant behavior of the melanocytes leading to ectopic pigmentation of the hair follicle and epidermis, indicated the stem cell niche was not maintained. An autoimmune mechanism was excluded by crossing the mice with rag1-/- mice. However, large numbers of macrophages and increased expression of ICAM-1 were still present and may be involved either directly or indirectly in the hair loss. Reverse transcriptase-PCR (RT-PCR) and immunohistochemistry of sebaceous gland differentiation markers revealed reduced peroxisome proliferator-activated receptor-γ (PPARγ), a potential cause of reduced sebum production, as well as the potential involvement of the innate immune system in the hair loss. As reduced PPARγ expression has recently been implicated as a cause for lichen planopilaris, these mice may be useful for testing therapies.
Assuntos
Alopecia/genética , Alopecia/fisiopatologia , Folículo Piloso/fisiopatologia , Imunidade/fisiologia , Mutação/genética , Proteínas/genética , Alopecia/metabolismo , Animais , Modelos Animais de Doenças , Folículo Piloso/metabolismo , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Molécula 1 de Adesão Intercelular/metabolismo , Macrófagos/metabolismo , Macrófagos/patologia , Melanócitos/metabolismo , Melanócitos/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Mutantes , PPAR gama/metabolismo , Proteínas/metabolismo , Pigmentação da Pele , Células-Tronco/patologiaRESUMO
When the first nomenclature of the keratin protein family was published over 20 years ago, only 19 keratins were thought to exist. Sequencing of the human genome has now revealed that there are 54 keratin genes. As a consequence, the nomenclature needed revision to apply a logical numbering system that includes the more recently identified keratins of the hair follicle.
Assuntos
Genoma Humano , Queratinas/classificação , Terminologia como Assunto , Genoma Humano/genética , Humanos , Queratinas/genética , Análise de Sequência de DNARESUMO
The outer surface of the hand, limb and body is covered by the epidermis, which is elaborated into a number of specialized appendages, evolved not only to protect and reinforce the skin but also for social signalling. The most prominent of these appendages is the hair follicle. Hair follicles are remarkable because of their prolific growth characteristics and their complexity of differentiation. After initial embryonic morphogenesis, the hair follicle undergoes repeated cycles of regression and regeneration throughout the lifetime of the organism. Studies of mouse mutants with hair loss phenotypes have suggested that the mechanisms controlling the hair cycle probably involve many of the major signalling molecules used elsewhere in development, although the complete pathway of hair follicle growth control is not yet understood. Mouse studies have also led to the discovery of genes underlying several human disorders. Future studies of mouse hair-loss mutants are likely to benefit the understanding of human hair loss as well as increasing our knowledge of mechanisms controlling morphogenesis and tumorigenesis.
Assuntos
Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Doenças do Cabelo/embriologia , Folículo Piloso/embriologia , Modelos Animais , Alopecia/genética , Animais , Fatores de Crescimento de Fibroblastos/fisiologia , Doenças do Cabelo/genética , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Nus , Morfogênese/genética , Neoplasias de Anexos e de Apêndices Cutâneos/genéticaRESUMO
The intermediate filaments (IFs) form major structural elements of the cytoskeleton. In vitro analyses of these fibrous proteins reveal very different assembly properties for the nuclear and cytoplasmic IF proteins. However, keratins in particular, the largest and most heterogenous group of cytoplasmic IF proteins, have been difficult to analyze due to their rapid assembly dynamics under the near-physiological conditions used for other IF proteins. We show here that keratins, like other cytoplasmic IF proteins, go through a stage of assembling into full-width soluble complexes, i.e., "unit-length filaments" (ULFs). In contrast to other IF proteins, however, longitudinal annealing of keratin ULFs into long filaments quasi-coincides with their formation. In vitro assembly of IF proteins into filaments can be initiated by an increase of the ionic strength and/or lowering of the pH of the assembly buffer. We now document that 23-mer peptides from the head domains of various IF proteins can induce filament formation even under conditions of low salt and high pH. This suggests that the "heads" are involved in the formation and longitudinal association of the ULFs. Using a Tris-buffering protocol that causes formation of soluble oligomers at pH 9, the epidermal keratins K5/14 form less regular filaments and less efficiently than the simple epithelial keratins K8/18. In sodium phosphate buffers (pH 7.5), however, K5/14 were able to form long partially unraveled filaments which compacted into extended, regular filaments upon addition of 20 mM KCl. Applying the same assembly regimen to mutant K14 R125H demonstrated that mutations causing a severe disease phenotype and morphological filament abnormalities can form long, regular filaments with surprising efficiency in vitro.
Assuntos
Biofísica/métodos , Queratinas/química , Proteínas Recombinantes/química , Sequência de Bases , Citoesqueleto/metabolismo , DNA Complementar/metabolismo , Humanos , Concentração de Íons de Hidrogênio , Queratina-14 , Queratina-5 , Queratina-8 , Queratinas/metabolismo , Queratinas/ultraestrutura , Microscopia Eletrônica , Dados de Sequência Molecular , Proteínas Recombinantes/metabolismo , Vimentina/químicaRESUMO
Keratins are cytoplasmic intermediate filament proteins expressed by epithelial cells. Keratin 7 (K7) is expressed in a wide range of epithelial structures in humans. We have cloned and fully sequenced the human and mouse K7 genes and mRNAs, and the K7 mRNA from the marsupial Potorous tridactylis, from which the widely used simple epithelial cell lines PtK1 and PtK2 are derived. Percentage identity plots comparing the mouse and human genomic sequences revealed a number of conserved CpG islands associated with the K7 gene. There was considerable conservation of introns between the two species, which may indicate the presence of intronic regulatory elements. Only the most proximal 500bp of the promoter was conserved, although an additional conserved sequence island was found 2-3kb upstream. Protein sequence comparisons between the three species allowed identification of conserved regions of the keratin variable domains that may be candidates for protein-protein interactions and/or regulatory modification. From the mouse sequence, we generated a polyclonal rabbit antibody specific for murine K7. This antibody was used to perform a survey of K7 expression in the mouse. The expression pattern was similar to the reported human distribution, with substantial expression observed in lung, bladder, mesothelium, hair follicle, and ductal structures. We also noted previously unreported expression of K7 in the gastrointestinal tract and filiform papillae of the tongue and specific K7 expression in a range of "hard" epithelial tissues. The distribution of K7 in mouse and availability of genomic sequence from the 129/Sv mouse strain will allow the generation and analysis of transgenic mice expressing mutant forms of K7 and to predict the phenotype of human genetic disorders caused by mutations in this keratin.
Assuntos
Queratinas/genética , Queratinas/metabolismo , Sequência de Aminoácidos , Animais , Linhagem Celular , Clonagem Molecular , Sequência Conservada , Evolução Molecular , Feminino , Biblioteca Gênica , Projeto Genoma Humano , Humanos , Queratina-7 , Queratinas/química , Macropodidae , Masculino , Camundongos , Dados de Sequência Molecular , Especificidade de Órgãos , Fragmentos de Peptídeos/química , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Especificidade da EspécieRESUMO
Desmosomal cadherins are essential cell adhesion molecules present throughout the epidermis and other organs, whose major function is to provide mechanical integrity and stability to epithelial cells in a wide variety of tissues. We recently identified a novel desmoglein family member, Desmoglein 4 (Dsg4), using a positional cloning approach in two families with localized autosomal recessive hypotrichosis (LAH) and in the lanceolate hair (lah) mouse. In this study, we report cloning and identification of the rat Dsg4 gene, in which we discovered a missense mutation in a naturally occurring lanceolate hair (lah) rat mutant. Phenotypic analysis of lah/lah mutant rats revealed a striking hair shaft defect with the appearance of a lance head within defective hair shafts. The mutation disrupts a critical calcium binding site bridging the second and third extracellular domains of Dsg4, likely disrupting extracellular interactions of the protein.