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1.
Hum Mol Genet ; 28(1): 74-83, 2019 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-30239744

RESUMO

Cardio-facio-cutaneous (CFC) syndrome, a genetic disorder caused by germline mutations in BRAF, KRAS, MAP2K1 and MAP2K2, is characterized by growth retardation, heart defects, dysmorphic facial appearance and dermatologic abnormalities. We have previously reported that knock-in mice expressing the CFC syndrome-associated mutation, Braf Q241R, showed growth retardation because of gastrointestinal dysfunction. However, other factors associated with growth retardation, including chondrogenesis and endocrinological profile, have not been examined. Here, we show that 3- and 4-week-old BrafQ241R/+ mice have decreased body weight and length, as well as reduced growth plate width in the proximal tibiae. Furthermore, proliferative and hypertrophic chondrocyte zones of the growth plate were reduced in BrafQ241R/+ mice compared with Braf+/+ mice. Immunohistological analysis revealed that extracellular signal-regulated kinase (ERK) activation was enhanced in hypertrophic chondrocytes in BrafQ241R/+ mice. In accordance with growth retardation and reduced growth plate width, decreased serum levels of insulin-like growth factor 1 (IGF-1) and IGF binding protein 3 (IGFBP-3) were observed in BrafQ241R/+ mice at 3 and 4 weeks of age. Treatment with C-type natriuretic peptide (CNP), a stimulator of endochondral bone growth and a potent inhibitor of the FGFR3-RAF1-MEK/ERK signaling, increased body and tail lengths in Braf+/+ and BrafQ241R/+ mice. In conclusion, ERK activation in chondrocytes and low serum IGF-1/IGFBP-3 levels could be associated with the growth retardation observed in BrafQ241R/+ mice. Our data also suggest that CNP is a potential therapeutic target in CFC syndrome.


Assuntos
Displasia Ectodérmica/metabolismo , Insuficiência de Crescimento/metabolismo , Cardiopatias Congênitas/metabolismo , Peptídeo Natriurético Tipo C/farmacologia , Proteínas Proto-Oncogênicas B-raf/genética , Animais , Condrócitos/fisiologia , Modelos Animais de Doenças , Displasia Ectodérmica/fisiopatologia , Facies , Insuficiência de Crescimento/fisiopatologia , Mutação em Linhagem Germinativa , Transtornos do Crescimento/metabolismo , Cardiopatias Congênitas/fisiopatologia , Humanos , Fator de Crescimento Insulin-Like I/análise , MAP Quinase Quinase 1/genética , MAP Quinase Quinase 2/genética , Sistema de Sinalização das MAP Quinases/genética , Sistema de Sinalização das MAP Quinases/fisiologia , Camundongos , Camundongos Endogâmicos ICR , Mutação , Peptídeo Natriurético Tipo C/metabolismo , Proteínas Proto-Oncogênicas B-raf/fisiologia
2.
Nagoya J Med Sci ; 80(3): 299-307, 2018 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-30214079

RESUMO

The Notch signaling pathway is highly conserved and essential for animal development. It is required for cell differentiation, survival, and proliferation. Regulation of Notch signaling is a crucial process for human health. Ligands initiate a signal cascade by binding to Notch receptors expressed on a neighboring cell. Notch receptors interact with ligands through their epidermal growth factor-like repeats (EGF repeats). Most EGF repeats are modified by O-glycosylation with residues such as O-linked N-acetylglucosamine (O-GlcNAc), O-fucose, and O-glucose. These O-glycan modifications are important for Notch function. Defects in O-glycosylation affect Notch-ligand interaction, trafficking of Notch receptors, and Notch stability on the cell surface. Although the roles of each modification are not fully understood, O-fucose is essential for binding of Notch receptors to their ligands. We reported an EGF domain-specific O-GlcNAc transferase (EOGT) localized in the endoplasmic reticulum. Mutations in genes encoding EOGT or NOTCH1 cause Adams-Oliver syndrome. Dysregulation of Notch signaling because of defects or mutations in Notch receptors or Notch signal-regulating proteins, such as glycosyltransferases, induce a variety of congenital disorders. In this review, we discuss O-glycosylation of Notch receptors and congenital human diseases caused by defects in O-glycans on Notch receptors.


Assuntos
Receptores Notch/metabolismo , Animais , Displasia Ectodérmica/metabolismo , Fator de Crescimento Epidérmico/metabolismo , Glicosilação , Humanos , Deformidades Congênitas dos Membros/metabolismo , N-Acetilglucosaminiltransferases/metabolismo , Dermatoses do Couro Cabeludo/congênito , Dermatoses do Couro Cabeludo/metabolismo
3.
Cell Reprogram ; 20(4): 215-224, 2018 08.
Artigo em Inglês | MEDLINE | ID: mdl-29989433

RESUMO

Ectrodactyly-Ectodermal dysplasia-Clefting (EEC) syndrome is a rare monogenic disease with autosomal dominant inheritance caused by mutations in the TP63 gene, leading to progressive corneal keratinocyte loss, limbal stem cell deficiency (LSCD), and eventually blindness. Currently, there is no treatment available to cure or slow down the keratinocyte loss. Human oral mucosal epithelial stem cells (hOMESCs), which are a mixed population of keratinocyte precursor stem cells, are used as source of autologous tissue for treatment of bilateral LSCD. However, hOMESCs from EEC patients have a reduced life span due to TP63 mutations and cannot be used for autologous transplantation. Human induced pluripotent stem cells (hiPSCs) represent a potentially unlimited source of autologous limbal stem cell for EEC patients and can be genetically modified by genome editing technologies to correct the disease ex vivo before transplantation. In this study, we describe for the first time the generation of integration-free EEC-hiPSCs from hOMESCs of EEC patients by Sendai virus vector and episomal vector-based reprogramming. The generated hiPSC clones expressed pluripotency markers and were successfully differentiated into derivatives of the three germ layers, as well as toward corneal epithelium. These cells may be used for EEC disease modeling and open perspectives for applications in cell therapy of LSCD.


Assuntos
Biomarcadores/análise , Diferenciação Celular , Fenda Labial/patologia , Fissura Palatina/patologia , Displasia Ectodérmica/patologia , Células-Tronco Pluripotentes Induzidas/patologia , Mucosa Bucal/patologia , Células Cultivadas , Fenda Labial/genética , Fenda Labial/metabolismo , Fissura Palatina/genética , Fissura Palatina/metabolismo , Displasia Ectodérmica/genética , Displasia Ectodérmica/metabolismo , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Mucosa Bucal/metabolismo , Mutação , Fenótipo , Fatores de Transcrição/genética , Proteínas Supressoras de Tumor/genética
5.
Birth Defects Res ; 110(4): 376-381, 2018 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-29134781

RESUMO

BACKGROUND: Sensenbrenner syndrome (cranioectodermal dysplasia, CED) is a very rare autosomal recessive ciliopathy first described by Judith Sensenbrenner in 1975. CED is a complex disorder characterized by craniofacial, skeletal, and ectodermal abnormalities. The clinical symptoms are variable and the CED phenotype may present intrafamilial and interfamilial differences. Sensenbrenner syndrome belongs to a group of ciliary chondrodysplasias and is a genetically heterogeneous disease. Mutations in six genes: IFT122, WDR35, IFT43, WDR19, IFT52, and IFT140 have been associated with this disorder. All known CED genes encode proteins that are part of the intraflagellar transport complex, which plays an important role in the assembly and maintenance of cilia. CASE: We report a on 2-year-old male patient affected by Sensenbrenner syndrome. Dysmorphic features included short stature with rhizomelic shortening of limbs, short fingers, narrow chest, high forehead, epicanthal folds, telecanthus, broad nasal bridge, low-set ears, sparse hair, and widely space teeth. Craniosynostosis was surgically corrected at the age of 4 months. The patient presented chronic renal disease. Nephrologic picture showed early stages of nephronophthisis. Psychomotor development was apparently normal. Molecular analysis of the affected individual revealed compound heterozygosity for a novel nonsense p.(Arg113*) and a missense p.(Asp841Val) variant in the WDR35 gene. CONCLUSIONS: The observations of the CED patient in this study provide additional clinical data and expand the molecular spectrum of Sensenbrenner syndrome. Moreover, the two variants identified in the proband provide further evidence for the WDR35 mutations as the most common cause of this rare syndrome.


Assuntos
Osso e Ossos/anormalidades , Craniossinostoses/genética , Displasia Ectodérmica/genética , Mutação de Sentido Incorreto , Proteínas , Substituição de Aminoácidos , Osso e Ossos/metabolismo , Osso e Ossos/fisiologia , Pré-Escolar , Craniossinostoses/metabolismo , Displasia Ectodérmica/metabolismo , Proteínas Hedgehog , Humanos , Masculino
6.
Mol Psychiatry ; 23(8): 1687-1698, 2018 08.
Artigo em Inglês | MEDLINE | ID: mdl-29158583

RESUMO

Ras/MAPK pathway signaling is a major participant in neurodevelopment, and evidence suggests that BRAF, a key Ras signal mediator, influences human behavior. We studied the role of the mutation BRAFQ257R, the most common cause of cardiofaciocutaneous syndrome (CFC), in an induced pluripotent stem cell (iPSC)-derived model of human neurodevelopment. In iPSC-derived neuronal cultures from CFC subjects, we observed decreased p-AKT and p-ERK1/2 compared to controls, as well as a depleted neural progenitor pool and rapid neuronal maturation. Pharmacological PI3K/AKT pathway manipulation recapitulated cellular phenotypes in control cells and attenuated them in CFC cells. CFC cultures displayed altered cellular subtype ratios and increased intrinsic excitability. Moreover, in CFC cells, Ras/MAPK pathway activation and morphological abnormalities exhibited cell subtype-specific differences. Our results highlight the importance of exploring specific cellular subtypes and of using iPSC models to reveal relevant human-specific neurodevelopmental events.


Assuntos
Displasia Ectodérmica/metabolismo , Insuficiência de Crescimento/metabolismo , Cardiopatias Congênitas/metabolismo , Células-Tronco Pluripotentes Induzidas/metabolismo , Sistema de Sinalização das MAP Quinases , Neurogênese/fisiologia , Neurônios/metabolismo , Proteínas Proto-Oncogênicas B-raf/metabolismo , Técnicas de Cultura de Células , Displasia Ectodérmica/patologia , Facies , Insuficiência de Crescimento/patologia , Cardiopatias Congênitas/patologia , Humanos , Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Células-Tronco Pluripotentes Induzidas/patologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/fisiologia , Mutação , Neurogênese/efeitos dos fármacos , Neurogênese/genética , Neurônios/efeitos dos fármacos , Neurônios/patologia , Fenótipo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-akt/metabolismo
7.
Cell Mol Life Sci ; 75(7): 1179-1190, 2018 04.
Artigo em Inglês | MEDLINE | ID: mdl-29103147

RESUMO

The transcription factor p63 is a master regulator of epidermal development. Mutations in p63 give rise to human developmental diseases that often manifest epidermal defects. In this review, we summarize major p63 isoforms identified so far and p63 mutation-associated human diseases that show epidermal defects. We discuss key roles of p63 in epidermal keratinocyte proliferation and differentiation, emphasizing its master regulatory control of the gene expression pattern and epigenetic landscape that define epidermal fate. We subsequently review the essential function of p63 during epidermal commitment and transdifferentiation towards epithelial lineages, highlighting the notion that p63 is the guardian of the epithelial lineage. Finally, we discuss current therapeutic development strategies for p63 mutation-associated diseases. Our review proposes future directions for dissecting p63-controlled mechanisms in normal and diseased epidermal development and for developing therapeutic options.


Assuntos
Epiderme/metabolismo , Perfilação da Expressão Gênica , Queratinócitos/metabolismo , Fatores de Transcrição/genética , Proteínas Supressoras de Tumor/genética , Diferenciação Celular/genética , Proliferação de Células/genética , Displasia Ectodérmica/genética , Displasia Ectodérmica/metabolismo , Displasia Ectodérmica/terapia , Humanos , Mutação , Fatores de Transcrição/metabolismo , Proteínas Supressoras de Tumor/metabolismo
8.
Int J Mol Sci ; 18(12)2017 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-29194391

RESUMO

Cardiofaciocutaneous (CFC) syndrome is a rare genetic disorder caused by mutations in the extracellular signal-regulated kinase (ERK) signaling. However, little is known about how aberrant ERK signaling is associated with the defective bone development manifested in most CFC syndrome patients. In this study, induced pluripotent stem cells (iPSCs) were generated from dermal fibroblasts of a CFC syndrome patient having rapidly accelerated fibrosarcoma kinase B (BRAF) gain-of-function mutation. CFC-iPSCs were differentiated into mesenchymal stem cells (CFC-MSCs) and further induced to osteoblasts in vitro. The osteogenic defects of CFC-MSCs were revealed by alkaline phosphatase activity assay, mineralization assay, quantitative real-time polymerase chain reaction (qRT-PCR), and western blotting. Osteogenesis of CFC-MSCs was attenuated compared to wild-type (WT)-MSCs. In addition to activated ERK signaling, increased p-SMAD2 and decreased p-SMAD1 were observed in CFC-MSCs during osteogenesis. The defective osteogenesis of CFC-MSCs was rescued by inhibition of ERK signaling and SMAD2 signaling or activation of SMAD1 signaling. Importantly, activation of ERK signaling and SMAD2 signaling or inhibition of SMAD1 signaling recapitulated the impaired osteogenesis in WT-MSCs. Our findings indicate that SMAD2 signaling and SMAD1 signaling as well as ERK signaling are responsible for defective early bone development in CFC syndrome, providing a novel insight on the pathological mechanism and therapeutic targets.


Assuntos
Displasia Ectodérmica/patologia , Insuficiência de Crescimento/patologia , Cardiopatias Congênitas/patologia , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Mesenquimais/citologia , Osteoblastos/citologia , Proteínas Proto-Oncogênicas B-raf/genética , Animais , Diferenciação Celular , Linhagem Celular , Displasia Ectodérmica/genética , Displasia Ectodérmica/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Facies , Insuficiência de Crescimento/genética , Insuficiência de Crescimento/metabolismo , Cardiopatias Congênitas/genética , Cardiopatias Congênitas/metabolismo , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Células-Tronco Mesenquimais/metabolismo , Camundongos , Mutação , Osteoblastos/metabolismo , Osteogênese , Fosforilação , Transdução de Sinais , Proteína Smad1/metabolismo , Proteína Smad2/metabolismo
9.
Am J Med Genet A ; 173(3): 790-800, 2017 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-28160419

RESUMO

The Adams-Oliver syndrome (AOS) is defined as aplasia cutis congenita (ACC) with transverse terminal limb defects (TTLD). Frequencies of associated anomalies are not well characterized. Six causative genes have been identified: ARHGAP31, DOCK6, EOGT, RBPJ, NOTCH1, and DLL4. We review 385 previously described individuals (139 non-familial and 246 familial probands and family members) and add clinical data on 13 previously unreported individuals with AOS. In addition to ACC and TTLD, the most commonly associated anomalies included a wide variety of central nervous system (CNS) anomalies and congenital heart defects each seen in 23%. CNS anomalies included structural anomalies, microcephaly, vascular defects, and vascular sequelae. CNS migration defects were common. Cutis marmorata telangiectasia congenita (CMTC) was found in 19% of the study population and other vascular anomalies were seen in 14%. Hemorrhage was listed as the cause of death for five of 25 deaths reported. A relatively large number of non-familial probands were reported to have hepatoportal sclerosis with portal hypertension and esophageal varices. Non-familial probands were more likely to have additional anomalies than were familial probands. The data reported herein provide a basis for refining the diagnostic features of AOS and suggest management recommendations for probands newly diagnosed with AOS. © 2017 Wiley Periodicals, Inc.


Assuntos
Displasia Ectodérmica/diagnóstico , Displasia Ectodérmica/genética , Estudos de Associação Genética , Deformidades Congênitas dos Membros/diagnóstico , Deformidades Congênitas dos Membros/genética , Fenótipo , Dermatoses do Couro Cabeludo/congênito , Anormalidades Múltiplas/diagnóstico , Anormalidades Múltiplas/genética , Diagnóstico por Imagem , Displasia Ectodérmica/metabolismo , Feminino , Humanos , Deformidades Congênitas dos Membros/metabolismo , Masculino , Mutação , Receptores Notch/metabolismo , Dermatoses do Couro Cabeludo/diagnóstico , Dermatoses do Couro Cabeludo/genética , Dermatoses do Couro Cabeludo/metabolismo , Transdução de Sinais , Proteína cdc42 de Ligação ao GTP/genética , Proteína cdc42 de Ligação ao GTP/metabolismo , Proteínas rac1 de Ligação ao GTP/genética , Proteínas rac1 de Ligação ao GTP/metabolismo
10.
Cell Death Dis ; 7: e2227, 2016 05 19.
Artigo em Inglês | MEDLINE | ID: mdl-27195674

RESUMO

EEC (ectrodactily-ectodermal dysplasia and cleft lip/palate) syndrome is a rare genetic disease, autosomal dominant inherited. It is part of the ectodermal dysplasia disorders caused by heterozygous mutations in TP63 gene. EEC patients present limb malformations, orofacial clefting, skin and skin's appendages defects, ocular abnormalities. The transcription factor p63, encoded by TP63, is a master gene for the commitment of ectodermal-derived tissues, being expressed in the apical ectodermal ridge is critical for vertebrate limb formation and, at a later stage, for skin and skin's appendages development. The ΔNp63α isoform is predominantly expressed in epithelial cells and it is indispensable for preserving the self-renewal capacity of adult stem cells and to engage specific epithelial differentiation programs. Small interfering RNA (siRNA) offers a potential therapy approach for EEC patients by selectively silencing the mutant allele. Here, using a systemic screening based on a dual-luciferase reported gene assay, we have successfully identified specific siRNAs for repressing the EEC-causing p63 mutant, R304W. Upon siRNA treatment, we were able to restore ΔNp63-WT allele transcriptional function in induced pluripotent stem cells that were derived from EEC patient biopsy. This study demonstrates that siRNAs approach is promising and, may pave the way for curing/delaying major symptoms, such as cornea degeneration and skin erosions in young EEC patients.


Assuntos
Fenda Labial/genética , Fissura Palatina/genética , Displasia Ectodérmica/genética , Inativação Gênica , Células-Tronco Pluripotentes Induzidas/metabolismo , Fatores de Transcrição/genética , Transcrição Genética , Proteínas Supressoras de Tumor/genética , Alelos , Sequência de Bases , Fenda Labial/metabolismo , Fenda Labial/patologia , Fissura Palatina/metabolismo , Fissura Palatina/patologia , Displasia Ectodérmica/metabolismo , Displasia Ectodérmica/patologia , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Genes Dominantes , Genes Reporter , Humanos , Células-Tronco Pluripotentes Induzidas/patologia , Luciferases/genética , Luciferases/metabolismo , Terapia de Alvo Molecular , Mutação , Cultura Primária de Células , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Síndrome , Fatores de Transcrição/antagonistas & inibidores , Fatores de Transcrição/metabolismo , Proteínas Supressoras de Tumor/antagonistas & inibidores , Proteínas Supressoras de Tumor/metabolismo
11.
Am J Med Genet A ; 170(10): 2570-7, 2016 10.
Artigo em Inglês | MEDLINE | ID: mdl-27155212

RESUMO

Noonan, Cardio-facio-cutaneous, and Costello syndromes are disorders of the Ras/MAPK pathway that share many clinical features. This observational and anthropometric study was conducted to describe the key facial features of each syndrome in order to improve discrimination between the three conditions, particularly in young children where diagnosis is most challenging. Direct measurement of the head and face was used to enhance diagnostic accuracy, and identify the most unusual or specific dimensions. The Noonan syndrome cohort included 123 individuals, aged 6 months to 41 years. There were 20 children and adolescents with Cardio-facio-cutaneous syndrome, and 28 individuals with Costello syndrome, aged 1-32 years. The facial phenotypes of these syndromes, particularly Noonan syndrome, are well-described but objective data have not been published in peer-reviewed literature. In this study, subjective observations, in the main, were validated by anthropometry with one exception. In individuals with Costello syndrome, mouth width was normal, thus the impression of wide mouth is likely due to full lips or the mouth being viewed in relation to a narrow lower face. When the three conditions were compared objectively, syndrome-specific pattern profiles showed high concordance in early life. At older ages, Cardio-facio-cutaneous syndrome was distinguished by increased width of the mid/lower face, and reduced growth of maxillary and mandibular dimensions was noted in both Noonan and Costello syndromes. Despite substantial similarities in face shape in older individuals with these two conditions, bulbous nasal tip, full lips, and an apparently wide mouth in those with Costello Syndrome facilitate discrimination from Noonan syndrome. © 2016 Wiley Periodicals, Inc.


Assuntos
Síndrome de Costello/diagnóstico , Displasia Ectodérmica/diagnóstico , Facies , Insuficiência de Crescimento/diagnóstico , Cardiopatias Congênitas/diagnóstico , Síndrome de Noonan/diagnóstico , Adolescente , Adulto , Fatores Etários , Pesos e Medidas Corporais , Criança , Pré-Escolar , Síndrome de Costello/genética , Síndrome de Costello/metabolismo , Diagnóstico Diferencial , Displasia Ectodérmica/genética , Displasia Ectodérmica/metabolismo , Insuficiência de Crescimento/genética , Insuficiência de Crescimento/metabolismo , Feminino , Cardiopatias Congênitas/genética , Cardiopatias Congênitas/metabolismo , Humanos , Lactente , Masculino , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Síndrome de Noonan/genética , Síndrome de Noonan/metabolismo , Fenótipo , Transdução de Sinais , Adulto Jovem , Proteínas ras/metabolismo
12.
Yakugaku Zasshi ; 136(1): 29-35, 2016.
Artigo em Japonês | MEDLINE | ID: mdl-26725664

RESUMO

  Nitric oxide (NO) is historically well known as a toxic gas but now recognized as a physiological cellular mediator acting at very low concentrations. It is biosynthesized within the body, and modulates many signal transduction processes. For investigation of the functions of this gaseous mediator, it is necessary to use chemical donors that release NO specifically, and it is highly advantageous if the release can be made with precise spatiotemporal control. For this purpose, we have developed caged NO (photocontrollable NO-releasing compounds) with unique releasing mechanisms. One employs the photoinduced rearrangement of an arylnitro group and subsequent release of NO, and another uses photoinduced electron transfer to release NO. One of our caged NO was confirmed to induce a NO-dependent cellular response in vivo under photocontrol. Photocontrollable NO releasers are expected to become indispensable tools for physiological experiments, and are also potential therapeutic agents for photodynamic therapy.


Assuntos
Fenômenos Fisiológicos Celulares/fisiologia , Óxido Nítrico/fisiologia , Animais , Encéfalo/anormalidades , Encéfalo/metabolismo , Anormalidades Congênitas/metabolismo , Orelha/anormalidades , Displasia Ectodérmica/metabolismo , Doenças Genéticas Ligadas ao Cromossomo X/metabolismo , Doença de Hirschsprung/metabolismo , Humanos , Deficiência Intelectual/metabolismo , Rim/anormalidades , Rim/metabolismo , Óxido Nítrico/biossíntese , Óxido Nítrico/química , Óxido Nítrico/farmacologia , Processos Fotoquímicos , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/fisiologia , Vasodilatação/efeitos dos fármacos
13.
Hum Mol Genet ; 25(2): 340-7, 2016 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-26604144

RESUMO

The lipid phosphatase FIG4 is a subunit of the protein complex that regulates biosynthesis of the signaling lipid PI(3,5)P2. Mutations of FIG4 result in juvenile lethality and spongiform neurodegeneration in the mouse, and are responsible for the human disorders Charcot-Marie-Tooth disease, Yunis-Varon syndrome and polymicrogyria with seizures. We previously demonstrated that conditional expression of a wild-type FIG4 transgene in neurons is sufficient to rescue most of the abnormalities of Fig4 null mice, including juvenile lethality and extensive neurodegeneration. To evaluate the contribution of the phosphatase activity to the in vivo function of Fig4, we introduced the mutation p.Cys486Ser into the Sac phosphatase active-site motif CX5RT. Transfection of the Fig4(Cys486Ser) cDNA into cultured Fig4(-/-) fibroblasts was effective in preventing vacuolization. The neuronal expression of an NSE-Fig4(Cys486Ser) transgene in vivo prevented the neonatal neurodegeneration and juvenile lethality seen in Fig4 null mice. These observations demonstrate that the catalytically inactive FIG4 protein provides significant function, possibly by stabilization of the PI(3,5)P2 biosynthetic complex and/or localization of the complex to endolysosomal vesicles. Despite this partial rescue, later in life the NSE-Fig4(Cys486Ser) transgenic mice display significant abnormalities that include hydrocephalus, defective myelination and reduced lifespan. The late onset phenotype of the NSE-Fig4(Cys486Ser) transgenic mice demonstrates that the phosphatase activity of FIG4 has an essential role in vivo.


Assuntos
Flavoproteínas/genética , Hidrocefalia/genética , Mutação , Neurônios/metabolismo , Animais , Domínio Catalítico/genética , Doença de Charcot-Marie-Tooth/genética , Doença de Charcot-Marie-Tooth/metabolismo , Displasia Cleidocraniana/genética , Displasia Cleidocraniana/metabolismo , Displasia Ectodérmica/genética , Displasia Ectodérmica/metabolismo , Flavoproteínas/metabolismo , Hidrocefalia/metabolismo , Deformidades Congênitas dos Membros/genética , Deformidades Congênitas dos Membros/metabolismo , Camundongos , Camundongos Transgênicos , Micrognatismo/genética , Micrognatismo/metabolismo , Fosfatos de Fosfatidilinositol/metabolismo , Fosfatases de Fosfoinositídeos , Monoéster Fosfórico Hidrolases/genética , Monoéster Fosfórico Hidrolases/metabolismo , Polimicrogiria/genética , Polimicrogiria/metabolismo , Células de Schwann/metabolismo
14.
Aging (Albany NY) ; 7(11): 928-36, 2015 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-26554466

RESUMO

OTX proteins, homologs of the Drosophila orthodenticle (Otd), are important for the morphogenesis of the neuroectoderm, and for the central nervous system formation. OTX1 and OTX2 are important for the cochlea and macula development, indeed when OTX1 is knocked down, these organs undergo developmental failure. Moreover OTX2 transfection revert this effect in OTX1(-/-) mice. The TA isoform of TP63, involved in Notch regulation pathway, has a critical function in the cochlear neuroepithelium differentiation. TAp63 positively regulates Hes5 and Atoh1 transcription. This pathway has been also demonstrated in p63(-/-) mice, and in patients p63 mutated, affected by Ectodermal Dysplasia (ED, OMIM 129810). These patients are affected by mild sensorineural deafness, most likely related to the mutation in p63 gene impairing the Notch pathway. We demonstrated the role of OTX2 on TAp63 regulation necessary for the correct formation of macular neuroepithelium and we confirmed the impairment of vestibular function caused by p63 mutations. Although the abnormalities found in our patient were still at a subclinical extent, aging could exacerbate this impairment and cause a decrease in quality of life.


Assuntos
Cóclea/embriologia , Regulação da Expressão Gênica no Desenvolvimento , Macula Lutea/embriologia , Fatores de Transcrição Otx/fisiologia , Fatores de Transcrição/genética , Proteínas Supressoras de Tumor/genética , Animais , Diferenciação Celular , Displasia Ectodérmica/metabolismo , Humanos , Camundongos , Fatores de Transcrição Otx/química , Vestíbulo do Labirinto/fisiologia
15.
Biochem Biophys Res Commun ; 467(2): 434-40, 2015 Nov 13.
Artigo em Inglês | MEDLINE | ID: mdl-26408908

RESUMO

p63, a member of the p53 family, is a crucial transcription factor for epithelial development and skin homeostasis. Heterozygous mutations in TP63 gene have been associated with human ectodermal dysplasia disorders. Most of these TP63 mutations are missense mutations causing amino acidic substitutions at p63 DNA binding or SAM domains that reduce or abolish the transcriptional activity of mutants p63. A significant number of mutants, however, resides in part of the p63 protein that apparently do not affect DNA binding and/or transcriptional activity, such as the N-terminal domain. Here, we characterize five p63 mutations at the 5' end of TP63 gene aiming to understand the pathogenesis of the diseases and to uncover the role of ΔNp63α N-terminus residues in determining its transactivation potential.


Assuntos
Sequência de Aminoácidos , Deleção de Sequência , Fatores de Transcrição/genética , Ativação Transcricional , Proteínas Supressoras de Tumor/genética , Sítios de Ligação , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Linhagem Celular Tumoral , Inibidor de Quinase Dependente de Ciclina p21/genética , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Proteínas do Citoesqueleto/genética , Proteínas do Citoesqueleto/metabolismo , Distonina , Displasia Ectodérmica/genética , Displasia Ectodérmica/metabolismo , Displasia Ectodérmica/patologia , Genes Reporter , Células HEK293 , Humanos , Queratina-14/genética , Queratina-14/metabolismo , Luciferases/genética , Luciferases/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Dados de Sequência Molecular , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Fases de Leitura Aberta , Ligação Proteica , Precursores de Proteínas/genética , Precursores de Proteínas/metabolismo , Estrutura Terciária de Proteína , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Elementos de Resposta , Fatores de Transcrição/química , Fatores de Transcrição/metabolismo , Proteínas Supressoras de Tumor/química , Proteínas Supressoras de Tumor/metabolismo
16.
Int Rev Immunol ; 34(6): 445-59, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26269396

RESUMO

Anhidrotic Ectodermal Dysplasia with ImmunoDeficiency (EDA-ID, OMIM 300291) and Incontinentia Pigmenti (IP, OMIM 308300) are two rare diseases, caused by mutations of the IKBKG/NEMO gene. The protein NEMO/IKKγ is essential for the NF-κB activation pathway, involved in a variety of physiological and cellular processes, such as immunity, inflammation, cell proliferation, and survival. A wide spectrum of IKBKG/NEMO mutations have been identified so far, and, on the basis of their effect on NF-κB activation, they are considered hypomorphic or amorphic (loss of function) mutations. IKBKG/NEMO hypomorphic mutations, reducing but not abolishing NF-κB activation, have been identified in EDA-ID and IP patients. Instead, the amorphic mutations, abolishing NF-κB activation by complete IKBKG/NEMO gene silencing, cause only IP. Here, we present an overview of IKBKG/NEMO mutations in EDA-ID and IP patients and describe similarities and differences between the clinical/immunophenotypic and genetic aspects, highlighting any T and B lymphocyte defect, and paying particular attention to the cellular and molecular defects that underlie the pathogenesis of both diseases.


Assuntos
Displasia Ectodérmica/etiologia , Displasia Ectodérmica/metabolismo , Doenças Genéticas Ligadas ao Cromossomo X/etiologia , Doenças Genéticas Ligadas ao Cromossomo X/metabolismo , Quinase I-kappa B/genética , Síndromes de Imunodeficiência/etiologia , Síndromes de Imunodeficiência/metabolismo , Incontinência Pigmentar/etiologia , Incontinência Pigmentar/metabolismo , Mutação , NF-kappa B/metabolismo , Transdução de Sinais , Animais , Displasia Ectodérmica/diagnóstico , Estudos de Associação Genética , Doenças Genéticas Ligadas ao Cromossomo X/diagnóstico , Loci Gênicos , Genótipo , Humanos , Quinase I-kappa B/química , Quinase I-kappa B/metabolismo , Síndromes de Imunodeficiência/diagnóstico , Incontinência Pigmentar/diagnóstico , Inflamação/genética , Inflamação/imunologia , Inflamação/metabolismo , Fenótipo
17.
Stem Cells ; 33(5): 1447-55, 2015 May.
Artigo em Inglês | MEDLINE | ID: mdl-25639853

RESUMO

Cardio-facio-cutaneous (CFC) syndrome is a developmental disorder caused by constitutively active ERK signaling manifesting mainly from BRAF mutations. Little is known about the role of elevated ERK signaling in CFC syndrome during early development. Here, we show that both SMAD1 and ERK signaling pathways may contribute to the developmental defects in CFC syndrome. Induced pluripotent stem cells (iPSCs) derived from dermal fibroblasts of a CFC syndrome patient (CFC-iPSCs) revealed early developmental defects in embryoid body (EB) development, ß-catenin localization, and neuronal differentiation. Both SMAD1 and ERK signalings were significantly activated in CFC-iPSCs during EB formation. Most of the ß-catenin was dissociated from the membrane and preferentially localized into the nucleus in CFC-EBs. Furthermore, activation of SMAD1 signaling recapitulated early developmental defects in wild-type iPSCs. Intriguingly, inhibition of SMAD1 signaling in CFC-iPSCs rescued aberrant EB morphology, impaired neuronal differentiation, and altered ß-catenin localization. These results suggest that SMAD1 signaling may be a key pathway contributing the pathogenesis of CFC syndrome during early development.


Assuntos
Displasia Ectodérmica/metabolismo , Displasia Ectodérmica/patologia , Insuficiência de Crescimento/metabolismo , Insuficiência de Crescimento/patologia , Cardiopatias Congênitas/metabolismo , Cardiopatias Congênitas/patologia , Células-Tronco Pluripotentes Induzidas/metabolismo , Transdução de Sinais , Proteínas Smad/metabolismo , Diferenciação Celular , Núcleo Celular/metabolismo , Corpos Embrioides/metabolismo , Facies , Humanos , Masculino , Neurônios/patologia , Transporte Proteico , beta Catenina/metabolismo
18.
Acta Derm Venereol ; 95(4): 476-9, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25270765

RESUMO

Recently, patients with hypohidrotic/anhidrotic ectodermal dysplasia (H/AED) have been reported to have a higher prevalence of symptoms suggestive of atopic disorders than the general population. To better understand atopic diathesis in H/AED, 6 cases of clinically or genetically diagnosed H/AED were examined. The following criteria were evaluated with patient consent: sweating, blood test results, histopathology and filaggrin staining. Five of 6 H/AED cases displayed atopic dermatitis-like manifestations, and 3 of these 5 cases experienced periorbital lesions. H/AED patients tended to present with atopic dermatitis-like eruptions with characteristics potentially indicative of periorbital lesions. Atopic diathesis in H/AED appeared not to be associated with filaggrin. We could speculate that hypohidrosis or anhidrosis itself might impair skin barrier function and contribute to atopic diathesis.


Assuntos
Dermatite Atópica/complicações , Displasia Ectodérmica/complicações , Adolescente , Adulto , Criança , Pré-Escolar , Dermatite Atópica/metabolismo , Displasia Ectodérmica/metabolismo , Feminino , Humanos , Hipo-Hidrose , Imunoglobulina E/sangue , Lactente , Proteínas de Filamentos Intermediários/metabolismo , Masculino , Coloração e Rotulagem , Glândulas Sudoríparas/anormalidades , Adulto Jovem
19.
J Invest Dermatol ; 135(3): 759-767, 2015 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-25347115

RESUMO

In healthy human skin host defense molecules such as antimicrobial peptides (AMPs) contribute to skin immune homeostasis. In patients with the congenital disease ectodermal dysplasia (ED) skin integrity is disturbed and as a result patients have recurrent skin infections. The disease is characterized by developmental abnormalities of ectodermal derivatives and absent or reduced sweating. We hypothesized that ED patients have a reduced skin immune defense because of the reduced ability to sweat. Therefore, we performed a label-free quantitative proteome analysis of wash solution of human skin from ED patients or healthy individuals. A clear-cut difference between both cohorts could be observed in cellular processes related to immunity and host defense. In line with the extensive underrepresentation of proteins of the immune system, dermcidin, a sweat-derived AMP, was reduced in its abundance in the skin secretome of ED patients. In contrast, proteins involved in metabolic/catabolic and biosynthetic processes were enriched in the skin secretome of ED patients. In summary, our proteome profiling provides insights into the actual situation of healthy versus diseased skin. The systematic reduction in immune system and defense-related proteins may contribute to the high susceptibility of ED patients to skin infections and altered skin colonization.


Assuntos
Displasia Ectodérmica/imunologia , Displasia Ectodérmica/metabolismo , Peptídeos/metabolismo , Proteômica , Pele/metabolismo , Adulto , Animais , Estudos de Casos e Controles , Modelos Animais de Doenças , Feminino , Perfilação da Expressão Gênica , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Pomadas , Peptídeos/administração & dosagem , Peptídeos/uso terapêutico , Infecções Cutâneas Estafilocócicas/tratamento farmacológico , Infecções Cutâneas Estafilocócicas/microbiologia , Staphylococcus aureus , Glândulas Sudoríparas/metabolismo
20.
Hum Mol Genet ; 23(24): 6553-66, 2014 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-25035421

RESUMO

Cardio-facio-cutaneous (CFC) syndrome is one of the 'RASopathies', a group of phenotypically overlapping syndromes caused by germline mutations that encode components of the RAS-MAPK pathway. Germline mutations in BRAF cause CFC syndrome, which is characterized by heart defects, distinctive facial features and ectodermal abnormalities. To define the pathogenesis and to develop a potential therapeutic approach in CFC syndrome, we here generated new knockin mice (here Braf(Q241R/+)) expressing the Braf Q241R mutation, which corresponds to the most frequent mutation in CFC syndrome, Q257R. Braf(Q241R/+) mice manifested embryonic/neonatal lethality, showing liver necrosis, edema and craniofacial abnormalities. Histological analysis revealed multiple heart defects, including cardiomegaly, enlarged cardiac valves, ventricular noncompaction and ventricular septal defects. Braf(Q241R/+) embryos also showed massively distended jugular lymphatic sacs and subcutaneous lymphatic vessels, demonstrating lymphatic defects in RASopathy knockin mice for the first time. Prenatal treatment with a MEK inhibitor, PD0325901, rescued the embryonic lethality with amelioration of craniofacial abnormalities and edema in Braf(Q241R/+) embryos. Unexpectedly, one surviving pup was obtained after treatment with a histone 3 demethylase inhibitor, GSK-J4, or NCDM-32b. Combination treatment with PD0325901 and GSK-J4 further increased the rescue from embryonic lethality, ameliorating enlarged cardiac valves. These results suggest that our new Braf knockin mice recapitulate major features of RASopathies and that epigenetic modulation as well as the inhibition of the ERK pathway will be a potential therapeutic strategy for the treatment of CFC syndrome.


Assuntos
Benzamidas/farmacologia , Benzazepinas/farmacologia , Difenilamina/análogos & derivados , Displasia Ectodérmica/tratamento farmacológico , Displasia Ectodérmica/genética , Insuficiência de Crescimento/tratamento farmacológico , Insuficiência de Crescimento/genética , Cardiopatias Congênitas/tratamento farmacológico , Cardiopatias Congênitas/genética , Proteínas Proto-Oncogênicas B-raf/genética , Pirimidinas/farmacologia , Animais , Difenilamina/farmacologia , Modelos Animais de Doenças , Sinergismo Farmacológico , Displasia Ectodérmica/metabolismo , Displasia Ectodérmica/patologia , Embrião de Mamíferos , Facies , Insuficiência de Crescimento/metabolismo , Insuficiência de Crescimento/patologia , Feminino , Regulação da Expressão Gênica , Técnicas de Introdução de Genes , Genes Letais , Cardiopatias Congênitas/metabolismo , Cardiopatias Congênitas/patologia , Inibidores de Histona Desacetilases/farmacologia , Histona Desmetilases/antagonistas & inibidores , Histona Desmetilases/genética , Histona Desmetilases/metabolismo , Humanos , Fígado/anormalidades , Fígado/efeitos dos fármacos , MAP Quinase Quinase Quinases/antagonistas & inibidores , MAP Quinase Quinase Quinases/genética , MAP Quinase Quinase Quinases/metabolismo , Masculino , Camundongos , Camundongos Transgênicos , Mutação , Miocárdio/patologia , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas B-raf/metabolismo , Transdução de Sinais , Crânio/anormalidades , Crânio/efeitos dos fármacos
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