RESUMEN
Epidermal development and maintenance are finely regulated events requiring a strict balance between proliferation and differentiation. Alterations in these processes give rise to human disorders such as cancer or syndromes with skin and annexes defects, known as ectodermal dysplasias (EDs). Here, we studied the functional effects of two novel receptor-interacting protein kinase 4 (RIPK4) missense mutations identified in siblings with an autosomal recessive ED with cutaneous syndactyly, palmoplantar hyperkeratosis and orofacial synechiae. Clinical overlap with distinct EDs caused by mutations in transcription factors (i.e. p63 and interferon regulatory factor 6, IRF6) or nectin adhesion molecules was noticed. Impaired activity of the RIPK4 kinase resulted both in altered epithelial differentiation and defective cell adhesion. We showed that mutant RIPK4 resulted in loss of PVRL4/nectin-4 expression in patient epidermis and primary keratinocytes, and demonstrated that PVRL4 is transcriptionally regulated by IRF6, a RIPK4 phosphorylation target. In addition, defective RIPK4 altered desmosome morphology through modulation of plakophilin-1 and desmoplakin. In conclusion, this work implicates RIPK4 kinase function in the p63-IRF6 regulatory loop that controls the proliferation/differentiation switch and cell adhesion, with implications in ectodermal development and cancer.
Asunto(s)
Displasia Ectodérmica , Factores Reguladores del Interferón , Adhesión Celular/genética , Moléculas de Adhesión Celular/metabolismo , Displasia Ectodérmica/metabolismo , Homeostasis , Humanos , Factores Reguladores del Interferón/genética , Factores Reguladores del Interferón/metabolismo , Queratinocitos/metabolismo , Nectinas , Proteínas Serina-Treonina QuinasasRESUMEN
Inborn errors of metabolism (IEMs) comprise a diverse group of monogenic disorders caused by enzyme deficiencies that result either in a toxic accumulation of metabolic intermediates or a shortage of essential end-products. Certain IEMs, like phenylketonuria (PKU), necessitate stringent dietary intervention that could lead to microbiome dysbiosis, thereby exacerbating the clinical phenotype. The objective of this systematic review was to examine the impact of PKU therapies on the intestinal microbiota. This research was conducted following the PRISMA Statement, with data from PubMed, Scopus, ScienceDirect, and Web of Science. A total of 18 articles meeting the inclusion criteria were published from 2011 to 2022. Significant reductions in several taxonomic groups in individuals with PKU when compared to the control group were detected in a quantitative analysis conducted across seven studies. The meta-analysis synthesis indicates a contrast in biodiversity between PKU subjects and the control population. Additionally, the meta-regression results, derived from the Bacillota/Bacteroidota ratio data, suggest a potential influence of diet in adult PKU populations (p = 0.004). It is worth noting that the limited number of studies calls for further research and analysis in this area. Our findings indicate the necessity of enhancing understanding of microbiota variability in reaction to treatments among PKU subjects to design tailored therapeutic and nutritional interventions to prevent complications resulting from microbiota disruption.
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Microbioma Gastrointestinal , Fenilcetonurias , Adulto , Humanos , Fenilcetonurias/complicaciones , DietaRESUMEN
Measles is characterized by fever and a maculopapular skin rash, which is accompanied by immune clearance of measles virus (MV)-infected cells. Histopathological analyses of skin biopsies from humans and non-human primates (NHPs) with measles rash have identified MV-infected keratinocytes and mononuclear cells in the epidermis, around hair follicles and near sebaceous glands. Here, we address the pathogenesis of measles skin rash by combining data from experimentally infected NHPs, ex vivo infection of human skin sheets and in vitro infection of primary human keratinocytes. Analysis of NHP skin samples collected at different time points following MV inoculation demonstrated that infection in the skin precedes onset of rash by several days. MV infection was detected in lymphoid and myeloid cells in the dermis before dissemination to the epidermal leukocytes and keratinocytes. These data were in good concordance with ex vivo MV infections of human skin sheets, in which dermal cells were more targeted than the epidermal cells. To address viral dissemination to the epidermis and to determine whether the dissemination is receptor-dependent, we performed experimental infections of primary keratinocytes collected from healthy donors. These experiments demonstrated that MV infection of keratinocytes is mainly nectin-4-dependent, and differentiated keratinocytes, which express higher levels of nectin-4, are more susceptible to MV infection than proliferating keratinocytes. Based on these data, we propose a model to explain measles skin rash: migrating MV-infected lymphocytes initiate the infection of dermal skin-resident CD150+ immune cells. The infection is subsequently disseminated from the dermal papillae to nectin-4+ keratinocytes in the basal epidermis. Lateral spread of MV infection is observed in the superficial epidermis, most likely due to the higher level of nectin-4 expression on differentiated keratinocytes. Finally, MV-infected cells are cleared by infiltrating immune cells, causing hyperemia and edema, which give the appearance of morbilliform skin rash.
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Dermis/virología , Epidermis/virología , Queratinocitos/virología , Linfocitos/virología , Sarampión/virología , Células Mieloides/virología , Piel/virología , Animales , Células Cultivadas , Dermis/patología , Epidermis/patología , Humanos , Queratinocitos/patología , Linfocitos/patología , Macaca fascicularis , Sarampión/patología , Virus del Sarampión/aislamiento & purificación , Células Mieloides/patología , Piel/patologíaRESUMEN
Leukodystrophies are a heterogeneous group of rare inherited disorders that mostly involve the white matter of the CNS. These conditions are characterized by primary glial cell and myelin sheath pathology of variable aetiology, which causes secondary axonal degeneration, generally emerging with disease progression. Whole exome sequencing performed in five large consanguineous nuclear families allowed us to identify homozygosity for two recurrent missense variants affecting highly conserved residues of RNF220 as the causative event underlying a novel form of leukodystrophy with ataxia and sensorineural deafness. We report these two homozygous missense variants (p.R363Q and p.R365Q) in the ubiquitin E3 ligase RNF220 as the underlying cause of this novel form of leukodystrophy with ataxia and sensorineural deafness that includes fibrotic cardiomyopathy and hepatopathy as associated features in seven consanguineous families. Mass spectrometry analysis identified lamin B1 as the RNF220 binding protein and co-immunoprecipitation experiments demonstrated reduced binding of both RNF220 mutants to lamin B1. We demonstrate that RNF220 silencing in Drosophila melanogaster specifically affects proper localization of lamin Dm0, the fly lamin B1 orthologue, promotes its aggregation and causes a neurodegenerative phenotype, strongly supporting the functional link between RNF220 and lamin B1. Finally, we demonstrate that RNF220 plays a crucial role in the maintenance of nuclear morphology; mutations in primary skin fibroblasts determine nuclear abnormalities such as blebs, herniations and invaginations, which are typically observed in cells of patients affected by laminopathies. Overall, our data identify RNF220 as a gene implicated in leukodystrophy with ataxia and sensorineural deafness and document a critical role of RNF220 in the regulation of nuclear lamina. Our findings provide further evidence on the direct link between nuclear lamina dysfunction and neurodegeneration.
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Alelos , Ataxia/genética , Sordera/genética , Laminopatías/genética , Mutación/genética , Ubiquitina-Proteína Ligasas/genética , Adolescente , Secuencia de Aminoácidos , Animales , Ataxia/diagnóstico , Células COS , Niño , Chlorocebus aethiops , Sordera/diagnóstico , Drosophila , Femenino , Células HEK293 , Humanos , Laminopatías/diagnóstico , Masculino , Linaje , Adulto JovenRESUMEN
Teebi hypertelorism syndrome (THS; OMIM 145420) is a rare craniofacial disorder characterized by hypertelorism, prominent forehead, short nose with broad or depressed nasal root. Some cases of THS have been attributed to SPECC1L variants. Homozygous variants in CDH11 truncating the transmembrane and intracellular domains have been implicated in Elsahy-Waters syndrome (EWS; OMIM 211380) with hypertelorism. We report THS due to CDH11 heterozygous missense variants on 19 subjects from 9 families. All affected residues in the extracellular region of Cadherin-11 (CHD11) are highly conserved across vertebrate species and classical cadherins. Six of the variants that cluster around the EC2-EC3 and EC3-EC4 linker regions are predicted to affect Ca2+ binding that is required for cadherin stability. Two of the additional variants [c.164G > C, p.(Trp55Ser) and c.418G > A, p.(Glu140Lys)] are also notable as they are predicted to directly affect trans-homodimer formation. Immunohistochemical study demonstrates that CDH11 is strongly expressed in human facial mesenchyme. Using multiple functional assays, we show that five variants from the EC1, EC2-EC3 linker, and EC3 regions significantly reduced the cell-substrate trans adhesion activity and one variant from EC3-EC4 linker results in changes in cell morphology, focal adhesion, and migration, suggesting dominant negative effect. Characteristic features in this cohort included depressed nasal root, cardiac and umbilical defects. These features distinguished this phenotype from that seen in SPECC1L-related hypertelorism syndrome and CDH11-related EWS. Our results demonstrate heterozygous variants in CDH11, which decrease cell-cell adhesion and increase cell migratory behavior, cause a form of THS, as termed CDH11-related THS.
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Anomalías Múltiples/genética , Cadherinas/genética , Adhesión Celular/genética , Anomalías Craneofaciales/genética , Deformidades Congénitas del Pie/genética , Variación Genética/genética , Deformidades Congénitas de la Mano/genética , Hipertelorismo/genética , Secuencia de Aminoácidos , Movimiento Celular/genética , Femenino , Heterocigoto , Homocigoto , Humanos , Masculino , Linaje , FenotipoRESUMEN
MicroRNAs are found throughout the genome and are processed by the microprocessor complex (MPC) from longer precursors. Some precursor miRNAs overlap intron:exon junctions. These splice site overlapping microRNAs (SO-miRNAs) are mostly located in coding genes. It has been intimated, in the rarer examples of SO-miRNAs in noncoding RNAs, that the competition between the spliceosome and the MPC modulates alternative splicing. However, the effect of this overlap on coding transcripts is unknown. Unexpectedly, we show that neither Drosha silencing nor SF3b1 silencing changed the inclusion ratio of SO-miRNA exons. Two SO-miRNAs, located in genes that code for basal membrane proteins, are known to inhibit proliferation in primary keratinocytes. These SO-miRNAs were up-regulated during differentiation and the host mRNAs were down-regulated, but again there was no change in inclusion ratio of the SO-miRNA exons. Interestingly, Drosha silencing increased nascent RNA density, on chromatin, downstream from SO-miRNA exons. Overall our data suggest a novel mechanism for regulating gene expression in which MPC-dependent cleavage of SO-miRNA exons could cause premature transcriptional termination of coding genes rather than affecting alternative splicing.
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Queratinocitos/citología , MicroARNs/genética , Fosfoproteínas/genética , Sitios de Empalme de ARN , Factores de Empalme de ARN/genética , Ribonucleasa III/genética , Empalme Alternativo , Diferenciación Celular , Proliferación Celular , Células Cultivadas , Cromatina/genética , Regulación hacia Abajo , Exones , Silenciador del Gen , Humanos , Queratinocitos/metabolismo , Empalmosomas/metabolismo , Regulación hacia ArribaRESUMEN
Variably reduced expression of the basement membrane component laminin-332 (α3aß3γ2) causes junctional epidermolysis bullosa generalized intermediate (JEB-GI), a skin fragility disorder with an increased susceptibility to squamous cell carcinoma (SCC) development in adulthood. Laminin-332 is highly expressed in several types of epithelial tumors and is central to signaling pathways that promote SCC tumorigenesis. However, laminin-332 mutations and expression in individuals affected by JEB-GI and suffering from recurrent SCCs have been poorly characterized. We studied a JEB-GI patient who developed over a hundred primary cutaneous SCCs. Molecular analysis combined with gene expression studies in patient skin and primary keratinocytes revealed that the patient is a functional hemizygous for the p.Cys1171* mutant allele which is transcribed in a stable mRNA encoding for a ß3 chain shortened of the last two C-terminal amino acids (Cys1171-Lys1172). The lack of the Cys1171 residue involved in the C-terminal disulphide bond to γ2 chain did not prevent assembly, secretion, and proteolytic processing of the heterotrimeric molecule. Immunohistochemistry of SCC specimens revealed accumulation of mutant laminin-332 at the epithelial-stromal interface of invasive front. We conclude that the C-terminal disulphide bond is a structural element crucial for laminin-332 adhesion function in-vivo. By saving laminin-332 amount, processing, and signaling role the p.Cys1171* mutation may allow intrinsic pro-tumorigenic properties of the protein to be conveyed, thus contributing to invasiveness and recurrence of SCCs in this patient.
Asunto(s)
Carcinoma de Células Escamosas , Moléculas de Adhesión Celular , Epidermólisis Ampollosa , Mutación , Proteínas de Neoplasias , Neoplasias Cutáneas , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/patología , Moléculas de Adhesión Celular/genética , Moléculas de Adhesión Celular/metabolismo , Epidermólisis Ampollosa/genética , Epidermólisis Ampollosa/metabolismo , Epidermólisis Ampollosa/patología , Humanos , Masculino , Persona de Mediana Edad , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Estabilidad del ARN/genética , ARN Mensajero/genética , ARN Mensajero/metabolismo , Neoplasias Cutáneas/genética , Neoplasias Cutáneas/metabolismo , Neoplasias Cutáneas/patología , KalininaRESUMEN
Hereditary sensory and autonomic neuropathies (HSAN) are clinically and genetically heterogeneous disorders, characterized by a progressive sensory neuropathy often complicated by ulcers and amputations, with variable motor and autonomic involvement. Several pathways have been implicated in the pathogenesis of neuronal degeneration in HSAN, while recent observations point to an emerging role of cytoskeleton organization and function. Here, we report novel biallelic mutations in the DST gene encoding dystonin, a large cytolinker protein of the plakin family, in an adult form of HSAN type VI. Affected individuals harbored the premature termination codon variant p.(Lys4330*) in trans with the p.(Ala203Glu) change affecting a highly conserved residue in an isoform-specific N-terminal region of dystonin. Functional studies showed defects in actin cytoskeleton organization and consequent delayed cell adhesion, spreading and migration, while recombinant p.Ala203Glu dystonin loses the ability to bind actin. Our data aid in the clinical and molecular delineation of HSAN-VI and suggest a central role for cell-motility and cytoskeletal defects in its pathogenesis possibly interfering with the neuronal outgrowth and guidance processes.
Asunto(s)
Citoesqueleto de Actina/patología , Distonina/genética , Genes Recesivos , Neuropatías Hereditarias Sensoriales y Autónomas/genética , Mutación/genética , Neuronas/metabolismo , Actinas/metabolismo , Adulto , Anciano , Secuencia de Aminoácidos , Animales , Células COS , Adhesión Celular , Movimiento Celular , Chlorocebus aethiops , Dermis/patología , Distonina/química , Familia , Femenino , Fibroblastos/metabolismo , Fibroblastos/patología , Células HEK293 , Humanos , Masculino , Persona de Mediana Edad , Unión Proteica , Isoformas de Proteínas/genéticaRESUMEN
Circulating anti-type VII collagen autoantibodies are frequently detected in patients with recessive dystrophic epidermolysis bullosa (RDEB). However, evidence supporting their pathogenic role in inducing epidermolysis bullosa acquisita (EBA) has been provided for only one individual with dominant dystrophic epidermolysis bullosa (DDEB). We describe here a patient who presented with dystrophic toenails since early childhood and developed trauma-induced skin blisters and oral erosions at age 26 years. Direct immunofluorescence showed IgG deposits with a u-serrated pattern along the cutaneous basement membrane zone, while no change in the expression of collagen VII could be detected by antigen mapping. High-titre anti-collagen VII antibodies were detected by enzyme-linked immunoassay (ELISA). In parallel, sequencing of epidermolysis bullosa (EB) genes identified compound heterozygous COL7A1 missense c.410G>A (p.Arg137Gln) and splicing c.3674C>T (p.Ala1225_Gln1241del) mutations, previously unrecognized in dystrophic epidermolysis bullosa (DEB). Thus, our patient had RDEB "nails-only" and developed mechanobullous EBA in adulthood. These data support a pathogenic role of circulating autoantibodies to collagen VII in inducing EBA in selected patients with DEB. Unforeseen worsening of skin symptoms in DEB should prompt laboratory investigations for EBA.
Asunto(s)
Colágeno Tipo VII/genética , Epidermólisis Ampollosa Adquirida/genética , Epidermólisis Ampollosa Distrófica/genética , Mutación Missense , Adulto , Autoanticuerpos/sangre , Biopsia , Colágeno Tipo VII/inmunología , Análisis Mutacional de ADN , Epidermólisis Ampollosa Adquirida/diagnóstico , Epidermólisis Ampollosa Adquirida/inmunología , Epidermólisis Ampollosa Distrófica/diagnóstico , Epidermólisis Ampollosa Distrófica/inmunología , Femenino , Técnica del Anticuerpo Fluorescente , Predisposición Genética a la Enfermedad , Humanos , Microscopía Electrónica de Transmisión , Uñas/inmunología , Uñas/patología , Fenotipo , Dominios Proteicos , Piel/inmunología , Piel/ultraestructuraRESUMEN
Mutations in the XPD subunit of the DNA repair/transcription factor TFIIH result in distinct clinical entities, including the cancer-prone xeroderma pigmentosum (XP) and the multisystem disorder trichothiodystrophy (TTD), which share only cutaneous photosensitivity. Gene-expression profiles of primary dermal fibroblasts revealed overexpression of matrix metalloproteinase 1 (MMP-1), the gene encoding the metalloproteinase that degrades the interstitial collagens of the extracellular matrix (ECM), in TTD patients mutated in XPD compared with their healthy parents. The defect is observed in TTD and not in XP and is specific for fibroblasts, which are the main producers of dermal ECM. MMP-1 transcriptional up-regulation in TTD is caused by an erroneous signaling mediated by retinoic acid receptors on the MMP-1 promoter and leads to hypersecretion of active MMP-1 enzyme and degradation of collagen type I in the ECM of cell/tissue systems and TTD patient skin. In agreement with the well-known role of ECM in eliciting signaling events controlling cell behavior and tissue homeostasis, ECM alterations in TTD were shown to impact on the migration and wound-healing properties of patient dermal fibroblasts. The presence of a specific inhibitor of MMP activity was sufficient to restore normal cell migration, thus providing a potential approach for therapeutic strategies. This study highlights the relevance of ECM anomalies in TTD pathogenesis and in the phenotypic differences between TTD and XP.
Asunto(s)
Matriz Extracelular/patología , Metaloproteinasa 1 de la Matriz/metabolismo , Factor de Transcripción TFIIH/fisiología , Síndromes de Tricotiodistrofia/enzimología , Humanos , Metaloproteinasa 1 de la Matriz/genética , Regiones Promotoras Genéticas , Receptores de Ácido Retinoico/metabolismo , Síndromes de Tricotiodistrofia/patología , Cicatrización de HeridasRESUMEN
Mutations in the laminin-332 (α3Aß3γ2) genes cause junctional epidermolysis bullosa (JEB), a recessively inherited disease characterized by blistering and altered wound repair. In addition, specific mutations that affect the N-terminus of the α3A chain cause a JEB-related non-blistering condition characterized by chronic production of granulation tissue, suggesting a critical role of this region in epithelial-mesenchymal communication. We report here a 9-year-old patient with JEB with a few long-standing skin ulcers with prominent granulation tissue in the absence of active blistering. He bears a homozygous missense mutation, p.Gly254Asp, within the first laminin epidermal growth factor-like (LE) repeat of the ß3 short arm. We show that p.Gly254Asp causes mis-folding of the LE motif, leading to reduced secretion of laminin-332 and structural alterations of the cutaneous basement membrane zone. These findings demonstrate, in a patient in vivo, that the ß3 short arm is also involved in the outcome of the granulation tissue response.
Asunto(s)
Moléculas de Adhesión Celular/genética , Epidermólisis Ampollosa de la Unión/genética , Niño , Tejido de Granulación , Humanos , Masculino , KalininaRESUMEN
The c.891C>T synonymous transition in SPINK5 induces exon 11 (E11) skipping and causes Netherton syndrome (NS). Using a specific RNA-protein interaction assay followed by mass spectrometry analysis along with silencing and overexpression of splicing factors, we showed that this mutation affects an exonic bifunctional splicing regulatory element composed by two partially overlapping silencer and enhancer sequences, recognized by hnRNPA1 and Tra2ß splicing factors, respectively. The C-to-T substitution concomitantly increases hnRNPA1 and weakens Tra2ß-binding sites, leading to pathological E11 skipping. In hybrid minigenes, exon-specific U1 small nuclear RNAs (ExSpe U1s) that target by complementarity intronic sequences downstream of the donor splice site rescued the E11 skipping defect caused by the c.891C>T mutation. ExSpe U1 lentiviral-mediated transduction of primary NS keratinocytes from a patient bearing the mutation recovered the correct full-length SPINK5 mRNA and the corresponding functional lympho-epithelial Kazal-type related inhibitor protein in a dose-dependent manner. This study documents the reliability of a mutation-specific, ExSpe U1-based, splicing therapy for a relatively large subset of European NS patients. Usage of ExSpe U1 may represent a general approach for correction of splicing defects affecting skin disease genes.
Asunto(s)
Empalme Alternativo , Exones , Mutación , Proteínas Inhibidoras de Proteinasas Secretoras/genética , ARN Nuclear Pequeño/genética , Secuencias Reguladoras de Ácidos Nucleicos , Línea Celular , Expresión Génica , Silenciador del Gen , Ribonucleoproteína Nuclear Heterogénea A1 , Ribonucleoproteína Heterogénea-Nuclear Grupo A-B/genética , Ribonucleoproteína Heterogénea-Nuclear Grupo A-B/metabolismo , Humanos , Queratinocitos/metabolismo , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Síndrome de Netherton/genética , Síndrome de Netherton/metabolismo , Proteínas Nucleares/metabolismo , Unión Proteica , Proteínas Inhibidoras de Proteinasas Secretoras/metabolismo , ARN Mensajero/genética , Proteínas de Unión al ARN/genética , Proteínas de Unión al ARN/metabolismo , Inhibidor de Serinpeptidasas Tipo Kazal-5 , Factores de Empalme Serina-ArgininaRESUMEN
Abnormalities in keratinocyte growth and differentiation have a pathogenic significance in many skin disorders and result in gene expression alterations detectable by quantitative real-time RT-PCR (qRT-PCR). Relative quantification based on endogenous control (EC) genes is the commonly adopted approach, and the use of multiple reference genes from independent pathways is considered a best practice guideline, unless fully validated EC genes are available. The literature on optimal reference genes during in vitro calcium-induced differentiation of normal human epidermal keratinocytes (NHEK) is inconsistent. In many studies, the expression of target genes is compared to that of housekeeping genes whose expression, however, significantly varies during keratinocyte differentiation. Here, we report the results of our investigations on the expression stability of 15 candidate EC genes, including those commonly used as reference in expression analysis by qRT-PCR, during NHEK calcium-induced differentiation. We demonstrate that YWHAZ and UBC are extremely stable genes, and therefore, they represent optimal EC genes for expression studies in proliferating and calcium-induced differentiating NHEK. Furthermore, we demonstrate that YWHAZ/14-3-3-zeta is a suitable reference for quantitative comparison of both transcript and protein levels.
Asunto(s)
Queratinocitos/citología , Queratinocitos/metabolismo , Proteínas 14-3-3/genética , Proteínas 14-3-3/metabolismo , Diferenciación Celular/genética , Proliferación Celular/genética , Células Cultivadas , Perfilación de la Expresión Génica , Humanos , Reacción en Cadena en Tiempo Real de la PolimerasaRESUMEN
Nectins are immunoglobulin-like cell adhesion molecules mainly localized in adherens junctions. The transcription factor p63 is a master regulator of gene expression in stratified epithelia and controls several molecular processes. As mutations in the Pvrl1 and Pvrl4 genes encoding for nectins cause genetic disorders with phenotypes similar to p63-related syndromes, we investigated whether these proteins might be under p63 transcriptional control. Here, we show that in p63-null skin, Pvrl1 gene expression is strongly reduced, whereas Pvrl4 expression is unaffected. In human and mouse primary keratinocytes p63 depletion leads to a specific downregulation of the Pvrl1 gene. Consistent with a direct regulation, chromatin immunoprecipitation experiments (ChIP) indicate that p63 binds to two conserved intronic Pvrl1 enhancer regions. Ankyloblepharon-ectodermal defects-cleft lip/palate (AEC) syndrome is a rare autosomal dominant disorder, caused by mutations in p63 gene, mainly characterized by skin fragility. To test whether nectins may be affected in AEC syndrome, their expression was measured in keratinocytes obtained from patients with AEC or from a conditional mouse model for AEC syndrome. Pvrl1 expression was reduced in AEC keratinocytes, consistent with impaired p63 function. Surprisingly, Pvrl4 expression was similarly affected, in parallel with decreased expression of the transcription factor Irf6. Consistent with the well-characterized role of Irf6 in keratinocyte differentiation and its strong downregulation in AEC syndrome, Irf6 depletion caused reduced expression of Pvrl4 in wild-type keratinocytes. Taken together, our results indicate that Pvrl1 is a bona fide target gene of the transcription factor p63, whereas Pvrl4 regulation is linked to epidermal differentiation and is under Irf6 control.
Asunto(s)
Moléculas de Adhesión Celular/metabolismo , Epidermis/metabolismo , Factores de Transcripción/metabolismo , Proteínas Supresoras de Tumor/metabolismo , Animales , Animales Recién Nacidos , Células Cultivadas , Epidermis/embriología , Regulación de la Expresión Génica , Regulación del Desarrollo de la Expresión Génica , Genes Reporteros , Humanos , Queratinocitos/citología , Ratones , Mutación , Nectinas , Fenotipo , Fosfoproteínas/metabolismo , Transactivadores/metabolismo , Transcripción GenéticaRESUMEN
Ichthyosis linearis circumflexa (ILC) presents as serpiginous and migratory erythematous patches with double-edged scales. ILC is rarely an isolated skin manifestation, but most commonly a part of Netherton syndrome (NS). NS is caused by SPINK5 mutations, which lead to absent or sometimes reduced expression of the serine protease inhibitor LEKTI. NS is characterised by congenital ichthyosiform erytroderma, trichorrhexis invaginata (TI) and atopy. We report 2 children who presented since the first months of life cheek erythema followed by the appearance of sparse ILC lesions on the face, trunk and proximal extremities. Erythroderma at birth, TI and atopy were absent. LEKTI immunoreactivity was reduced in patient epidermis, and serine protease activity was modestly increased, while desmoglein-1 expression remained unaffected. SPINK5 mutation and expression analysis in patient keratinocytes revealed compound heterozygous splicing variants, which allowed residual LEKTI secretion. Our results show that ILC can be the only clinical manifestation of NS.
Asunto(s)
Epidermis/química , Ictiosis/etiología , Síndrome de Netherton/complicaciones , Síndrome de Netherton/genética , Proteínas Inhibidoras de Proteinasas Secretoras/análisis , Proteínas Inhibidoras de Proteinasas Secretoras/genética , Preescolar , Desmogleína 1/análisis , Epidermis/enzimología , Femenino , Humanos , Lactante , Masculino , Mutación , Síndrome de Netherton/diagnóstico , Síndrome de Netherton/enzimología , Inhibidor de Serinpeptidasas Tipo Kazal-5 , Serina Proteasas/metabolismoRESUMEN
Lymphoepithelial Kazal-type related inhibitor (LEKTI) is a multidomain serine protease inhibitor which plays a central role in skin permeability barrier and allergy. Loss-of-function mutations in the LEKTI encoding gene SPINK5 cause Netherton syndrome, a rare and severe genetic skin disease with a profound skin barrier defect and atopic manifestations. Several studies also reported genetic association between the multifactorial disease atopic dermatitis (AD) and a frequent and non-conservative LEKTI variant, E420K, in different populations. Here, we provide evidence that the 420K variant impacts on LEKTI function by increasing the likelihood of furin-dependent LEKTI precursor cleavage within the linker region D6-D7. This results in the reversal of the cleavage priorities for LEKTI proteolytic activation and prevents the formation of the LEKTI fragment D6D9 known to display the strongest inhibitory activity against kallikrein (KLK) 5-mediated desmoglein-1 (DSG1) degradation. Using in situ and gel zymographies, we show that the modification of the subtle balance in LEKTI inhibitory fragments leads to enhanced KLK5, KLK7 and elastase-2 (ELA-2) activities in 420KK epidermis. By immunohistochemistry and western blot analyses, we found that increased epidermal protease activity correlates with reduced DSG1 protein expression and accelerated profilaggrin proteolysis. All changes determined by the presence of residue 420K within the LEKTI sequence likely contribute to defective skin barrier permeability. Remarkably, LEKTI 420KK epidermis displays an increased expression of the proallergic cytokine thymic stromal lymphopoietin (TSLP). This is the first functional evidence supporting association studies which identified the 420K LEKTI variant as a predisposing factor to AD, in combination with other genetic and environmental factors.
Asunto(s)
Dermatitis Atópica/enzimología , Dermatitis Atópica/genética , Mutación Missense , Proteínas Inhibidoras de Proteinasas Secretoras/genética , Proteínas Inhibidoras de Proteinasas Secretoras/metabolismo , Animales , Línea Celular , Dermatitis Atópica/metabolismo , Desmogleína 1/genética , Desmogleína 1/metabolismo , Epidermis/enzimología , Epidermis/metabolismo , Humanos , Calicreínas/genética , Calicreínas/metabolismo , Ratones , Proteolisis , Serina Endopeptidasas/genética , Serina Endopeptidasas/metabolismo , Inhibidor de Serinpeptidasas Tipo Kazal-5RESUMEN
Ichthyosis with confetti (IC) is a severe non-syndromic ichthyosis due to heterozygous mutations in the KRT10 gene. The disease manifests at birth with erythroderma and scaling and is characterised by the gradual development of numerous confetti-like spots of normal skin. Diagnosis of IC is frequently delayed until adolescence or even adulthood. We report 2 young children who were first diagnosed as having congenital ichthyosiform erythroderma. However, the development of thick, confluent hyperkeratotic plaques together with the histopathological finding of keratinocyte vacuolisation in the suprabasal epidermis evoked IC. Immunofluorescence analysis showed a highly reduced keratin 10 expression within the cytoplasm of suprabasal keratinocytes and its characteristic mislocalisation to the nuclei. The diagnosis was confirmed by the identification of 2 previously unreported mutations in intron 6 and exon 7 of KRT10. Careful clinical examination then showed the presence of the first spots of normal skin in both patients at the age of 2.5 and 5 years, respectively. These cases point to the usefulness of immunofluorescence analysis of keratin 10 expression for an early diagnosis of IC.
Asunto(s)
Ictiosis/genética , Queratina-10/genética , Mutación , Preescolar , Diagnóstico Diferencial , Diagnóstico Precoz , Exones , Femenino , Técnica del Anticuerpo Fluorescente , Humanos , Eritrodermia Ictiosiforme Congénita/diagnóstico , Ictiosis/diagnóstico , Inteínas , MasculinoRESUMEN
Ectodermal dysplasias form a large disease family with more than 200 members. The combination of hair and tooth abnormalities, alopecia, and cutaneous syndactyly is characteristic of ectodermal dysplasia-syndactyly syndrome (EDSS). We used a homozygosity mapping approach to map the EDSS locus to 1q23 in a consanguineous Algerian family. By candidate gene analysis, we identified a homozygous mutation in the PVRL4 gene that not only evoked an amino acid change but also led to exon skipping. In an Italian family with two siblings affected by EDSS, we further detected a missense and a frameshift mutation. PVRL4 encodes for nectin-4, a cell adhesion molecule mainly implicated in the formation of cadherin-based adherens junctions. We demonstrated high nectin-4 expression in hair follicle structures, as well as in the separating digits of murine embryos, the tissues mainly affected by the EDSS phenotype. In patient keratinocytes, mutated nectin-4 lost its capability to bind nectin-1. Additionally, in discrete structures of the hair follicle, we found alterations of the membrane localization of nectin-afadin and cadherin-catenin complexes, which are essential for adherens junction formation, and we found reorganization of actin cytoskeleton. Together with cleft lip and/or palate ectodermal dysplasia (CLPED1, or Zlotogora-Ogur syndrome) due to an impaired function of nectin-1, EDSS is the second known "nectinopathy" caused by mutations in a nectin adhesion molecule.
Asunto(s)
Moléculas de Adhesión Celular/genética , Displasia Ectodérmica/complicaciones , Displasia Ectodérmica/genética , Mutación/genética , Sindactilia/complicaciones , Sindactilia/genética , Anomalías Múltiples/genética , Adulto , Secuencia de Aminoácidos , Animales , Moléculas de Adhesión Celular/química , Moléculas de Adhesión Celular/metabolismo , Niño , Extremidades/embriología , Familia , Femenino , Regulación del Desarrollo de la Expresión Génica , Cabello/patología , Humanos , Masculino , Ratones , Datos de Secuencia Molecular , Linaje , Transporte de Proteínas , ARN Mensajero/genética , ARN Mensajero/metabolismo , Piel/patología , SíndromeRESUMEN
Anticancer agents that selectively kill tumor cells and spare normal tissues are urgently needed. Here, we engineered a cell-permeable peptidomimetic, shepherdin, modeled on the binding interface between the molecular chaperone Hsp90 and the antiapoptotic and mitotic regulator, survivin. Shepherdin makes extensive contacts with the ATP pocket of Hsp90, destabilizes its client proteins, and induces massive death of tumor cells by apoptotic and nonapoptotic mechanisms. Conversely, shepherdin does not reduce the viability of normal cells, and does not affect colony formation of purified hematopoietic progenitors. Systemic administration of shepherdin in vivo is well tolerated, and inhibits human tumor growth in mice without toxicity. Shepherdin could provide a potent and selective anticancer agent in humans.
Asunto(s)
Antimetabolitos Antineoplásicos/farmacología , Diseño de Fármacos , Fragmentos de Péptidos/farmacología , Adenosina Trifosfato/metabolismo , Animales , Proteína con Homeodominio Antennapedia , Antimetabolitos Antineoplásicos/química , Antimetabolitos Antineoplásicos/metabolismo , Apoptosis/efectos de los fármacos , Benzoquinonas , Sitios de Unión/genética , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/patología , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Citocromos c/metabolismo , Inhibidores Enzimáticos/farmacología , Femenino , Fibroblastos/efectos de los fármacos , Productos del Gen tat/genética , Proteínas HSP90 de Choque Térmico/genética , Proteínas HSP90 de Choque Térmico/metabolismo , Células HeLa , Proteínas de Homeodominio/genética , Humanos , Proteínas Inhibidoras de la Apoptosis , Lactamas Macrocíclicas , Masculino , Ratones , Ratones SCID , Proteínas Asociadas a Microtúbulos/metabolismo , Modelos Moleculares , Imitación Molecular , Proteínas de Neoplasias , Proteínas Nucleares/genética , Fragmentos de Péptidos/química , Fragmentos de Péptidos/metabolismo , Neoplasias de la Próstata/tratamiento farmacológico , Neoplasias de la Próstata/patología , Unión Proteica/efectos de los fármacos , Unión Proteica/genética , Conformación Proteica , Proteínas Proto-Oncogénicas c-bcl-2/genética , Rifabutina/análogos & derivados , Rifabutina/farmacología , Células Madre/efectos de los fármacos , Survivin , Telomerasa/metabolismo , Factores de Transcripción/genética , Proteína p53 Supresora de Tumor/genética , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Here we report a lethal case of Netherton syndrome presenting with neurologic complications, hypernatremic dehydration, failure to thrive, and episodes of sepsis. Molecular analysis of the serine protease inhibitor Kazal-type 5 gene identified a homozygous mutation (c.1111C>T, p.Arg371X). This case highlights the importance of early diagnosis to start appropriate care in a timely fashion and prevent disease complications.