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1.
Mol Cell ; 79(3): 472-487.e10, 2020 08 06.
Artigo em Inglês | MEDLINE | ID: mdl-32531202

RESUMO

It is widely assumed that decreasing transcription factor DNA-binding affinity reduces transcription initiation by diminishing occupancy of sequence-specific regulatory elements. However, in vivo transcription factors find their binding sites while confronted with a large excess of low-affinity degenerate motifs. Here, using the melanoma lineage survival oncogene MITF as a model, we show that low-affinity binding sites act as a competitive reservoir in vivo from which transcription factors are released by mitogen-activated protein kinase (MAPK)-stimulated acetylation to promote increased occupancy of their regulatory elements. Consequently, a low-DNA-binding-affinity acetylation-mimetic MITF mutation supports melanocyte development and drives tumorigenesis, whereas a high-affinity non-acetylatable mutant does not. The results reveal a paradoxical acetylation-mediated molecular clutch that tunes transcription factor availability via genome-wide redistribution and couples BRAF to tumorigenesis. Our results further suggest that p300/CREB-binding protein-mediated transcription factor acetylation may represent a common mechanism to control transcription factor availability.


Assuntos
Regulação Neoplásica da Expressão Gênica , Genoma , Melanoma/genética , Fator de Transcrição Associado à Microftalmia/genética , Processamento de Proteína Pós-Traducional , Neoplasias Cutâneas/genética , Acetilação , Sequência de Aminoácidos , Animais , Sítios de Ligação , Linhagem Celular Tumoral , Sequência Conservada , Elementos Facilitadores Genéticos , Feminino , Xenoenxertos , Humanos , Masculino , Melanócitos/metabolismo , Melanócitos/patologia , Melanoma/metabolismo , Melanoma/patologia , Camundongos , Camundongos Nus , Fator de Transcrição Associado à Microftalmia/química , Fator de Transcrição Associado à Microftalmia/metabolismo , Motivos de Nucleotídeos , Regiões Promotoras Genéticas , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Neoplasias Cutâneas/metabolismo , Neoplasias Cutâneas/patologia , Peixe-Zebra
2.
Bull Cancer ; 107(2): 272-280, 2020 Feb.
Artigo em Francês | MEDLINE | ID: mdl-32044098

RESUMO

MiT family translocation renal cell carcinomas (tRCC) represent a rare subtype of renal cell carcinomas. These tumors have been introduced for the first time in the World Health Classification (WHO) classification of kidney cancers in 2004. tRCC are characterized by reccurent translocations involving members of the MiT family transcription factors, mainly TFE3 and TFEB. The estimated incidence of these tumors is ∼1-5 % among all renal cell carcinomas, with female prodominance. tRCC were initially described in children, and the spectrum has been expanded over time to encompass adolescents and adults. TFE3- and TFEB-rearranged RCC harbor characteristic clinicopathological and immunohistochemical features and fluorescent hybridization in situ is considered the gold standard for their diagnosis, although it has some limitations especially when the partners are located in the vicinity of TFE3. Nephron-sparing surgery is an efficient treatment of localized cases when achievable. In metastatic setting, targeted agents and immunotherapy showed modest efficacy, with response rates and median overall survival inferior to those observed in clear-cell renal cell carcinomas. Management of tRCC necessite a multidisciplinary team and accrual in clinical trials have to be encouraged when possible. Novel biological insights are urgently awaited to better understand the mechanisms associated with kidney oncogenesis in this setting, and ultimately help to identify therapeutic targets.


Assuntos
Adenocarcinoma de Células Claras , Carcinoma de Células Renais , Neoplasias Renais , Translocação Genética , Adenocarcinoma de Células Claras/diagnóstico , Adenocarcinoma de Células Claras/genética , Adenocarcinoma de Células Claras/terapia , Adolescente , Adulto , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/genética , Carcinoma de Células Renais/diagnóstico , Carcinoma de Células Renais/genética , Carcinoma de Células Renais/terapia , Criança , Feminino , Humanos , Neoplasias Renais/diagnóstico , Neoplasias Renais/genética , Neoplasias Renais/terapia , Masculino , Fator de Transcrição Associado à Microftalmia/genética
3.
BMC Med Genet ; 21(1): 1, 2020 01 02.
Artigo em Inglês | MEDLINE | ID: mdl-31898538

RESUMO

BACKGROUND: Hearing loss (HL) represents the most common congenital sensory impairment with an incidence of 1-5 per 1000 live births. Non-syndromic hearing loss (NSHL) is an isolated finding that is not part of any other disorder accounting for 70% of all genetic hearing loss cases. METHODS: In the current study, we reported a polygenic mode of inheritance in an NSHL consanguineous family using exome sequencing technology and we evaluated the possible effect of the detected single nucleotide variants (SNVs) using in silico methods. RESULTS: Two bi-allelic SNVs were detected in the affected patients; a MYO15A (. p.V485A) variant, and a novel MITF (p.P338L) variant. Along with these homozygous mutations, we detected two heterozygous variants in well described hearing loss genes (MYO7A and MYH14). The novel MITF p. Pro338Leu missense mutation was predicted to change the protein structure and function. CONCLUSION: A novel MITF mutation along with a previously described MYO15A mutation segregate with an autosomal recessive non-syndromic HL case with a post-lingual onset. The findings highlight the importance of carrying whole exome sequencing for a comprehensive assessment of HL genetic heterogeneity.


Assuntos
Heterogeneidade Genética , Perda Auditiva Neurossensorial/genética , Fator de Transcrição Associado à Microftalmia/genética , Miosinas/genética , Idade de Início , Alelos , Criança , Feminino , Predisposição Genética para Doença , Perda Auditiva Neurossensorial/fisiopatologia , Heterozigoto , Homozigoto , Humanos , Masculino , Herança Multifatorial/genética , Linhagem , Fenótipo , Polimorfismo de Nucleotídeo Único/genética , Sequenciamento Completo do Exoma
4.
Int J Mol Sci ; 21(1)2020 Jan 02.
Artigo em Inglês | MEDLINE | ID: mdl-31906440

RESUMO

Fisetin is found in many fruits and plants such as grapes and onions, and exerts anti-inflammatory, anti-proliferative, and anticancer activity. However, whether fisetin regulates melanogenesis has been rarely studied. Therefore, we evaluated the effects of fisetin on melanogenesis in B16F10 melanoma cell and zebrafish larvae. The current study revealed that fisetin slightly suppressed in vitro mushroom tyrosinase activity; however, molecular docking data showed that fisetin did not directly bind to mushroom tyrosinase. Unexpectedly, fisetin significantly increased intracellular and extracellular melanin production in B16F10 melanoma cells regardless of the presence or absence of α-melanocyte stimulating hormone (α-MSH). We also found that the expression of melanogenesis-related genes such as tyrosinase and microphthalmia-associated transcription factor (MITF), were highly increased 48 h after fisetin treatment. Pigmentation of zebrafish larvae by fisetin treatment also increased at the concentrations up to 200 µM and then slightly decreased at 400 µM, with no alteration in the heart rates. Molecular docking data also revealed that fisetin binds to glycogen synthase kinase-3ß (GSK-3ß). Therefore, we evaluated whether fisetin negatively regulated GSK-3ß, which subsequently activates ß-catenin, resulting in melanogenesis. As expected, fisetin increased the expression of ß-catenin, which was subsequently translocated into the nucleus. In the functional assay, FH535, a Wnt/ß-catenin inhibitor, significantly inhibited fisetin-mediated melanogenesis in zebrafish larvae. Our data suggested that fisetin inhibits GSK-3ß, which activates ß-catenin, resulting in melanogenesis through the revitalization of MITF and tyrosinase.


Assuntos
Flavonoides/farmacologia , Glicogênio Sintase Quinase 3 beta/antagonistas & inibidores , Melaninas/biossíntese , beta Catenina/metabolismo , Animais , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Flavonoides/química , Flavonoides/toxicidade , Glicogênio Sintase Quinase 3 beta/química , Glicogênio Sintase Quinase 3 beta/genética , Glicogênio Sintase Quinase 3 beta/metabolismo , Larva/efeitos dos fármacos , Larva/metabolismo , Melanoma Experimental , Camundongos , Fator de Transcrição Associado à Microftalmia/genética , Fator de Transcrição Associado à Microftalmia/metabolismo , Simulação de Acoplamento Molecular , Monofenol Mono-Oxigenase/antagonistas & inibidores , Monofenol Mono-Oxigenase/metabolismo , Pigmentação/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Peixe-Zebra/embriologia , Peixe-Zebra/metabolismo , alfa-MSH/farmacologia , beta Catenina/antagonistas & inibidores , beta Catenina/genética
5.
Toxicol Lett ; 319: 197-203, 2020 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-31785464

RESUMO

The chemical warfare agent sulfur mustard (SM) affects all cells in the epidermis including melanocytes which are responsible for melanin synthesis. After exposure to SM, pigment abnormalities like hypo- and hyperpigmentation can occur. The underlying molecular pathomechanisms of SM exposure on human melanogenesis have not been elucidated so far. In our study, we investigated the effect of SM on human melanocytes and melanogenesis. Normal human epidermal melanocytes (NHEM) were used as in vitro model and they were exposed to different concentrations of SM (4.5 µM-100 µM). Melanin production was analyzed by absorption measurements at 405 nm. In addition, quantitative real-time PCR (qPCR) and Western blot experiments were performed to determine the expression of essential melanogenesis-related proteins including tyrosinase (TYR), tyrosinase-related protein (TRP) 1 and 2 and microphthalmia transcription factor (MITF). Our findings demonstrated that exposure to low SM concentrations increased melanin synthesis accompanied with an increase in protein expression. In contrast, high SM concentrations led to decreased melanin content and a downregulation in expression of all investigated melanogenesis-associated proteins. We concluded that low SM concentrations may cause hyperpigmentation while high SM concentrations decreased melanin content which may explain hypopigmented skin areas in SM exposed patients.


Assuntos
Substâncias para a Guerra Química/toxicidade , Melaninas/biossíntese , Gás de Mostarda/toxicidade , Células Cultivadas , Relação Dose-Resposta a Droga , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Hiperpigmentação/induzido quimicamente , Hipopigmentação/induzido quimicamente , Oxirredutases Intramoleculares/efeitos dos fármacos , Melaninas/genética , Melanócitos/efeitos dos fármacos , Melanócitos/metabolismo , Fator de Transcrição Associado à Microftalmia/biossíntese , Fator de Transcrição Associado à Microftalmia/genética , Monofenol Mono-Oxigenase/biossíntese , Monofenol Mono-Oxigenase/genética , Tripsina/biossíntese , Tripsina/genética
6.
Nucleic Acids Res ; 48(2): 934-948, 2020 01 24.
Artigo em Inglês | MEDLINE | ID: mdl-31777941

RESUMO

Interrupted dimeric coiled coil segments are found in a broad range of proteins and generally confer selective functional properties such as binding to specific ligands. However, there is only one documented case of a basic-helix-loop-helix leucine zipper transcription factor-microphthalmia-associated transcription factor (MITF)-in which an insertion of a three-residue stammer serves as a determinant of conditional partner selectivity. To unravel the molecular principles of this selectivity, we have analyzed the high-resolution structures of stammer-containing MITF and an engineered stammer-less MITF variant, which comprises an uninterrupted symmetric coiled coil. Despite this fundamental difference, both MITF structures reveal identical flanking in-phase coiled coil arrangements, gained by helical over-winding and local asymmetry in wild-type MITF across the stammer region. These conserved structural properties allow the maintenance of a proper functional readout in terms of nuclear localization and binding to specific DNA-response motifs regardless of the presence of the stammer. By contrast, MITF heterodimer formation with other bHLH-Zip transcription factors is only permissive when both factors contain either the same type of inserted stammer or no insert. Our data illustrate a unique principle of conditional partner selectivity within the wide arsenal of transcription factors with specific partner-dependent functional readouts.


Assuntos
Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/química , Núcleo Celular/química , Fator de Transcrição Associado à Microftalmia/química , Conformação Proteica , Animais , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/genética , Ligantes , Camundongos , Fator de Transcrição Associado à Microftalmia/genética , Ligação Proteica , Domínios Proteicos/genética , Multimerização Proteica
7.
BMC Genomics ; 20(1): 962, 2019 Dec 10.
Artigo em Inglês | MEDLINE | ID: mdl-31823726

RESUMO

BACKGROUND: Melanocytes are derived from neural crest stem cells in the embryonic stage. In mature melanocytes, a series of complex enzyme-catalyzed reactions leads to the production of melanins, which determine the hair and skin colors of animals. The process of melanogenesis is complex and can be regulated by mRNA, microRNAs (miRNAs) and long noncoding RNAs (lncRNAs) genes. MiRNAs are a type of endogenous noncoding RNA approximately 22 nt in size that predominantly regulate gene expression by inhibiting translation. miR-380-3p is a candidate miRNA potentially related to melanogenesis. To better understand the mechanism of miR-380-3p melanogenesis regulation, plasmids to overexpress or knockdown miR-380-3p were transfected into alpaca melanocytes, and their effects on melanogenesis were evaluated. RESULTS: In situ hybridization identified a positive miR-380-3p signal in alpaca melanocyte cytoplasm. Luciferase activity assays confirmed that SOX6 is targeted by miR-380-3p. miR-380-3p overexpression and knockdown in alpaca melanocytes respectively downregulated and upregulated SOX6 expression at the mRNA and protein levels. Additionally, miR-380-3p overexpression and knockdown, respectively, in alpaca melanocytes decreased and increased the mRNA levels of melanin transfer-related genes, including microphthalmia-associated transcription factor (MITF), tyrosinase (TYR), tyrosine-related protein-1 (TYRP1), and dopachrome tautomerase (DCT). In contrast, miR-380-3p overexpression and knockdown respectively increased and decreased the mRNA levels of ß-catenin. Additionally, the effect of miR-380-3p on melanogenesis was assessed by Masson-Fontana melanin staining. CONCLUSIONS: The results demonstrated that miR-380-3p targeted SOX6 to regulate melanogenesis by influencing ß-catenin and MITF transcription and translation, which reduced the expression of downstream genes, including TYR, TYRP1, and DCT. These results provide insights into the mechanisms through which miR-380-3p controls melanogenesis.


Assuntos
Melaninas/metabolismo , Melanócitos/metabolismo , MicroRNAs/genética , Fatores de Transcrição SOXD/genética , Regiões 3' não Traduzidas , Animais , Camelídeos Americanos , Células Cultivadas , Expressão Gênica , Regulação da Expressão Gênica , Técnicas de Silenciamento de Genes , Oxirredutases Intramoleculares/genética , Masculino , MicroRNAs/metabolismo , Fator de Transcrição Associado à Microftalmia/genética , Fator de Transcrição Associado à Microftalmia/metabolismo , Monofenol Mono-Oxigenase/genética , Oxirredutases/genética , Fatores de Transcrição SOXD/metabolismo , Pele/citologia , Pele/metabolismo , Pele/patologia , beta Catenina/genética , beta Catenina/metabolismo
8.
PLoS Genet ; 15(12): e1008501, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31881017

RESUMO

The MITF and SOX10 transcription factors regulate the expression of genes important for melanoma proliferation, invasion and metastasis. Despite growing evidence of the contribution of long noncoding RNAs (lncRNAs) in cancer, including melanoma, their functions within MITF-SOX10 transcriptional programmes remain poorly investigated. Here we identify 245 candidate melanoma associated lncRNAs whose loci are co-occupied by MITF-SOX10 and that are enriched at active enhancer-like regions. Our work suggests that one of these, Disrupted In Renal Carcinoma 3 (DIRC3), may be a clinically important MITF-SOX10 regulated tumour suppressor. DIRC3 depletion in human melanoma cells leads to increased anchorage-independent growth, a hallmark of malignant transformation, whilst melanoma patients classified by low DIRC3 expression have decreased survival. DIRC3 is a nuclear lncRNA that activates expression of its neighbouring IGFBP5 tumour suppressor through modulating chromatin structure and suppressing SOX10 binding to putative regulatory elements within the DIRC3 locus. In turn, DIRC3 dependent regulation of IGFBP5 impacts the expression of genes involved in cancer associated processes and is needed for DIRC3 control of anchorage-independent growth. Our work indicates that lncRNA components of MITF-SOX10 networks are an important new class of melanoma regulators and candidate therapeutic targets that can act not only as downstream mediators of MITF-SOX10 function but as feedback regulators of MITF-SOX10 activity.


Assuntos
Proteína 5 de Ligação a Fator de Crescimento Semelhante à Insulina/genética , Melanoma/genética , Fator de Transcrição Associado à Microftalmia/genética , RNA Longo não Codificante/genética , Fatores de Transcrição SOXE/genética , Linhagem Celular Tumoral , Núcleo Celular/genética , Proliferação de Células , Regulação para Baixo , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Humanos , Prognóstico , Análise de Sequência de RNA , Análise de Sobrevida
9.
Elife ; 82019 12 23.
Artigo em Inglês | MEDLINE | ID: mdl-31868592

RESUMO

Preventing terminal differentiation is important in the development and progression of many cancers including melanoma. Recent identification of the BMP ligand GDF6 as a novel melanoma oncogene showed GDF6-activated BMP signaling suppresses differentiation of melanoma cells. Previous studies have identified roles for GDF6 orthologs during early embryonic and neural crest development, but have not identified direct regulation of melanocyte development by GDF6. Here, we investigate the BMP ligand gdf6a, a zebrafish ortholog of human GDF6, during the development of melanocytes from the neural crest. We establish that the loss of gdf6a or inhibition of BMP signaling during neural crest development disrupts normal pigment cell development, leading to an increase in the number of melanocytes and a corresponding decrease in iridophores, another neural crest-derived pigment cell type in zebrafish. This shift occurs as pigment cells arise from the neural crest and depends on mitfa, an ortholog of MITF, a key regulator of melanocyte development that is also targeted by oncogenic BMP signaling. Together, these results indicate that the oncogenic role ligand-dependent BMP signaling plays in suppressing differentiation in melanoma is a reiteration of its physiological roles during melanocyte development.


Assuntos
Proteínas Morfogenéticas Ósseas/genética , Fator 6 de Diferenciação de Crescimento/genética , Melanócitos/metabolismo , Fator de Transcrição Associado à Microftalmia/genética , Proteínas de Peixe-Zebra/genética , Animais , Proteínas Morfogenéticas Ósseas/antagonistas & inibidores , Diferenciação Celular/genética , Regulação da Expressão Gênica no Desenvolvimento/genética , Humanos , Ligantes , Melanócitos/patologia , Neoplasias/genética , Neoplasias/patologia , Crista Neural/crescimento & desenvolvimento , Crista Neural/metabolismo , Pigmentação/genética , Transdução de Sinais/genética , Peixe-Zebra/genética , Peixe-Zebra/crescimento & desenvolvimento
10.
Genet Sel Evol ; 51(1): 62, 2019 Nov 08.
Artigo em Inglês | MEDLINE | ID: mdl-31703548

RESUMO

BACKGROUND: White spotting of the coat is a characteristic trait of various domestic species including cattle and other mammals. It is a hallmark of Holstein-Friesian cattle, and several previous studies have detected genetic loci with major effects for white spotting in animals with Holstein-Friesian ancestry. Here, our aim was to better understand the underlying genetic and molecular mechanisms of white spotting, by conducting the largest mapping study for this trait in cattle, to date. RESULTS: Using imputed whole-genome sequence data, we conducted a genome-wide association analysis in 2973 mixed-breed cows and bulls. Highly significant quantitative trait loci (QTL) were found on chromosomes 6 and 22, highlighting the well-established coat color genes KIT and MITF as likely responsible for these effects. These results are in broad agreement with previous studies, although we also report a third significant QTL on chromosome 2 that appears to be novel. This signal maps immediately adjacent to the PAX3 gene, which encodes a known transcription factor that controls MITF expression and is the causal locus for white spotting in horses. More detailed examination of these loci revealed a candidate causal mutation in PAX3 (p.Thr424Met), and another candidate mutation (rs209784468) within a conserved element in intron 2 of MITF transcripts expressed in the skin. These analyses also revealed a mechanistic ambiguity at the chromosome 6 locus, where highly dispersed association signals suggested multiple or multiallelic QTL involving KIT and/or other genes in this region. CONCLUSIONS: Our findings extend those of previous studies that reported KIT as a likely causal gene for white spotting, and report novel associations between candidate causal mutations in both the MITF and PAX3 genes. The sizes of the effects of these QTL are substantial, and could be used to select animals with darker, or conversely whiter, coats depending on the desired characteristics.


Assuntos
Bovinos/genética , Mutação , Polimorfismo de Nucleotídeo Único , Locos de Características Quantitativas , Pigmentação da Pele/genética , Animais , Estudo de Associação Genômica Ampla , Fator de Transcrição Associado à Microftalmia/genética , Fator de Transcrição PAX3/genética , Proteínas Proto-Oncogênicas c-kit/genética
11.
Fish Shellfish Immunol ; 95: 456-463, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31669282

RESUMO

p38 mitogen-activated protein kinases (MAPKs) are involved in the response to various extracellular stimuli via regulating gene expression. In the present study, a p38 MAPK gene (MpP38) was identified from the clam Meretrix petechialis. The full-length cDNA of MpP38 measures 1,720 bp, consisting of a 134-bp 5'-UTR, a 1,095-bp ORF and a 491-bp 3'-UTR. Both the mRNA and protein expression levels of MpP38 increased after Vibrio challenge, implying that MpP38 is involved in clam immunity. Based on our previous study, a transcription factor activated by p38 MAPK, i.e. microphthalmia-associated transcription factor (MITF), participated in clam immunity by regulating the expression of phenoloxidase (PO). Coupled with other related reports, the mechanism underlying the involvement of MpP38 in clam immunity is most likely that pathogen stimuli induce the phosphorylation of p38 MAPK and thus activate MITF to regulate the expression of the immune-related gene PO. The results obtained in this study support this mechanism. First, we found that the MpP38 phosphorylation level increased in response to Vibrio challenge. Second, as revealed by a yeast two-hybrid assay, there was a direct interaction between MpP38 and MITF. Meanwhile, inhibiting the phosphorylation of MpP38 decreased the phosphorylation level of MpMITF, implying that MpP38 phosphorylation is required for MpMITF activation. Additionally, our results showed that there was a regulatory relationship between MpP38 phosphorylation level and PO expression level. With increased MpP38 phosphorylation level, the PO expression level was also increased after Vibrio challenge; when MpP38 phosphorylation was inhibited, the PO expression level was significantly decreased. This study describes the immune function of p38 MAPK in the clam for the first time and analyses its potential underlying mechanism, which will help to elucidate the immune mechanism in the clam M. petechialis.


Assuntos
Bivalves/imunologia , Bivalves/microbiologia , Interações Hospedeiro-Patógeno/imunologia , Imunidade Inata , Vibrio parahaemolyticus/patogenicidade , Proteínas Quinases p38 Ativadas por Mitógeno/imunologia , Animais , Regulação da Expressão Gênica , Fator de Transcrição Associado à Microftalmia/genética , Fator de Transcrição Associado à Microftalmia/metabolismo , Fosforilação , Técnicas do Sistema de Duplo-Híbrido , Proteínas Quinases p38 Ativadas por Mitógeno/genética
12.
Genetica ; 147(5-6): 369-379, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31625006

RESUMO

Microphthalmia-associated transcription factor (MITF) is a member of MYC superfamily, associated with melanocyte cells, as it was discovered in depigmented mice. However, over the last years it was found to be involved in many cellular signaling pathways, among which oncogenesis, osteoclast differentiation, and stress response. In mammals, Mitf gene mutations can cause diverse syndromes affecting pigmentation of eyes or skin, bone defects and melanomas. As MITF protein homologs were also found in some invertebrates, we have isolated and characterized the MITF cDNAs from the sea urchin Paracentrotus lividus, referred to as Pl-Mitf. The in silico study of the secondary and tertiary structure of Pl-Mitf protein showed high conserved regions mostly lying in the DNA binding domain. To understand the degree of evolutionary conservation of MITF, a phylogenetic analysis was performed comparing the Pl-Mitf deduced protein with proteins from different animal species. Moreover, the analysis of temporal and spatial expression pattern of Pl-Mitf mRNA showed that it was expressed from the onset of gastrulation of the sea urchin embryo to the pluteus larva, specifically in primary mesenchymes cells (PMCs), the sea urchin skeletogenic cells, and in the forming archenteron, the larval gut precursor. In silico protein-protein interactions analysis was used to understand the association of MITF with other proteins. Our results put in evidence the conservation of the MITF protein among vertebrates and invertebrates and may provide new perspectives on the pathways underlying sea urchin development, even if further functional analyses are needed.


Assuntos
Sequência Conservada , Fator de Transcrição Associado à Microftalmia/genética , Ouriços-do-Mar/genética , Animais , Fator de Transcrição Associado à Microftalmia/química , Filogenia , Domínios Proteicos , Ouriços-do-Mar/classificação
13.
Nat Commun ; 10(1): 4664, 2019 10 11.
Artigo em Inglês | MEDLINE | ID: mdl-31604935

RESUMO

Signal transduction systems enable organisms to monitor their external environments and accordingly adjust the cellular processes. In mast cells, the second messenger Ap4A binds to the histidine triad nucleotide-binding protein 1 (HINT1), disrupts its interaction with the microphthalmia-associated transcription factor (MITF), and eventually activates the transcription of genes downstream of MITF in response to immunostimulation. How the HINT1 protein recognizes and is regulated by Ap4A remain unclear. Here, using eight crystal structures, biochemical experiments, negative stain electron microscopy, and cellular experiments, we report that Ap4A specifically polymerizes HINT1 in solution and in activated rat basophilic leukemia cells. The polymerization interface overlaps with the area on HINT1 for MITF interaction, suggesting a possible competitive mechanism to release MITF for transcriptional activation. The mechanism depends precisely on the length of the phosphodiester linkage of Ap4A. These results highlight a direct polymerization signaling mechanism by the second messenger.


Assuntos
Fosfatos de Dinucleosídeos/metabolismo , Mastócitos/metabolismo , Proteínas do Tecido Nervoso/fisiologia , Linhagem Celular , Cristalografia por Raios X , Técnicas de Silenciamento de Genes , Fator de Transcrição Associado à Microftalmia/genética , Fator de Transcrição Associado à Microftalmia/metabolismo , Fator de Transcrição Associado à Microftalmia/fisiologia , Proteínas do Tecido Nervoso/química , Proteínas do Tecido Nervoso/metabolismo , Polimerização , Estrutura Terciária de Proteína , Transdução de Sinais
14.
Genes Dev ; 33(19-20): 1295-1318, 2019 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-31575676

RESUMO

An incomplete view of the mechanisms that drive metastasis, the primary cause of cancer-related death, has been a major barrier to development of effective therapeutics and prognostic diagnostics. Increasing evidence indicates that the interplay between microenvironment, genetic lesions, and cellular plasticity drives the metastatic cascade and resistance to therapies. Here, using melanoma as a model, we outline the diversity and trajectories of cell states during metastatic dissemination and therapy exposure, and highlight how understanding the magnitude and dynamics of nongenetic reprogramming in space and time at single-cell resolution can be exploited to develop therapeutic strategies that capitalize on nongenetic tumor evolution.


Assuntos
Plasticidade Celular , Melanoma/fisiopatologia , Metástase Neoplásica/fisiopatologia , Resistencia a Medicamentos Antineoplásicos , Regulação Neoplásica da Expressão Gênica , Humanos , Melanoma/terapia , Fator de Transcrição Associado à Microftalmia/genética , Fator de Transcrição Associado à Microftalmia/metabolismo , Células-Tronco Neoplásicas/citologia , Fenótipo , Microambiente Tumoral
15.
Int J Biochem Cell Biol ; 116: 105620, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31561018

RESUMO

Diazepam is a medicament of the benzodiazepine family and it typically produces a sedative effect. Researchers have revealed that diazepam can induce melanogenesis and produce dendrite-like structures in B16 melanoma cells. However, the associated mechanisms of melanogenesis and phenotypic alterations have mostly remained unknown. In this study, we determined the effects of diazepam on melanogenesis, cellular phenotypic alterations, the location of melanosomes and the expression of relevant proteins in melanocytes using Masson-Fontana ammoniacal silver staining, scanning electron microscopy, immunocytochemistry and western blot analysis. Our results collectively indicated that diazepam had a pivotal role in melanocytes by enhancing melanin synthesis, melanocyte dendricity, melanosome trafficking, and capture at the dendrite tips. These functions might be attributed to the fact that diazepam activated the peripheral benzodiazepine receptor (PBR). This increased intracellular levels of cAMP, which stimulated the phosphorylation of cAMP response element-binding (CREB). As a result, this increased the tyrosinase, microphthalmia-associated transcription factor (MITF), Rab27a, Myosin Va, Rab17 and Cdc42 expression. This caused melanogenesis and melanosome transport. Therefore, our findings may provide a potential strategy for treating anti-hypopigmentation disorders.


Assuntos
Proteínas Quinases Dependentes de AMP Cíclico/genética , AMP Cíclico/metabolismo , Diazepam/farmacologia , Hipnóticos e Sedativos/farmacologia , Melanócitos/efeitos dos fármacos , Melanossomas/efeitos dos fármacos , Receptores de GABA-A/genética , Animais , Transporte Biológico/efeitos dos fármacos , Proteína de Ligação a CREB/genética , Proteína de Ligação a CREB/metabolismo , Linhagem Celular , Linhagem Celular Tumoral , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Regulação da Expressão Gênica , Humanos , Melanócitos/citologia , Melanócitos/metabolismo , Melanossomas/metabolismo , Camundongos , Fator de Transcrição Associado à Microftalmia/genética , Fator de Transcrição Associado à Microftalmia/metabolismo , Monofenol Mono-Oxigenase/genética , Monofenol Mono-Oxigenase/metabolismo , Cadeias Pesadas de Miosina/genética , Cadeias Pesadas de Miosina/metabolismo , Miosina Tipo V/genética , Miosina Tipo V/metabolismo , Receptores de GABA-A/metabolismo , Transdução de Sinais , Proteína cdc42 de Ligação ao GTP/genética , Proteína cdc42 de Ligação ao GTP/metabolismo , Proteínas rab de Ligação ao GTP/genética , Proteínas rab de Ligação ao GTP/metabolismo , Proteínas rab27 de Ligação ao GTP/genética , Proteínas rab27 de Ligação ao GTP/metabolismo
16.
Indian J Ophthalmol ; 67(9): 1481-1483, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31436206

RESUMO

A 3-year-old girl presented with bilateral asymmetrical partial heterochromia of iris and fundus. The parents also complained of bilateral hearing loss in the child. Suspecting an auditory-pigmentary syndrome, systemic and genetic evaluation was performed. The child had profound sensory-neural hearing loss. Targeted gene sequencing revealed a novel nonsense variation in exon 9 of the MITF gene (chr3:70008440A>T) that was pathogenic for Waardenburg syndrome (WS) type 2A. This case highlights the characteristics of the iris and fundus hypochromia, which may provide a clue toward the diagnosis of WS.


Assuntos
DNA/genética , Doenças da Íris/diagnóstico , Fator de Transcrição Associado à Microftalmia/genética , Mutação , Transtornos da Pigmentação/diagnóstico , Síndrome de Waardenburg/diagnóstico , Pré-Escolar , Feminino , Humanos , Iris/patologia , Doenças da Íris/genética , Doenças da Íris/metabolismo , Fator de Transcrição Associado à Microftalmia/metabolismo , Linhagem , Fenótipo , Transtornos da Pigmentação/genética , Transtornos da Pigmentação/metabolismo , Síndrome de Waardenburg/genética , Síndrome de Waardenburg/metabolismo
17.
Int J Circumpolar Health ; 78(1): 1630219, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31213145

RESUMO

Waardenburg syndrome (WS) is an orphan genetic disease with autosomal dominant pattern of inheritance characterised by varying degrees of hearing loss accompanied by skin, hair and iris pigmentation abnormalities. Four types of WS differing in phenotypic characteristics are now described. We performed a Sanger sequencing of coding regions of genes PAX3, MITF, SOX10 and SNAI2 in the patient with WS from a Yakut family living in the Sakha Republic. No changes were found in the PAX3, SOX10 and SNAI2 coding regions while a previously reported heterozygous transition c.772C>T (p.Arg259*) in exon 8 of the MITF gene was found in this patient. This patient presents rare phenotype of WS type 2: congenital unilateral hearing loss, unilateral heterochromia of irises, and absence of skin/hair depigmentation and dystopia canthorum. Audiological variability in WS type 2, caused by the c.772C>T (p.Arg259*) variant in the MITF gene, outlines the importance of molecular analysis and careful genotype-phenotype comparisons in order to optimally inform patients about the risk of hearing loss. The results of this study confirm the association of pathogenic variants in the MITF gene with WS type 2 and expanded data on the variability of audiological features of the WS.


Assuntos
Perda Auditiva Unilateral/etiologia , Perda Auditiva Unilateral/genética , Fator de Transcrição Associado à Microftalmia/genética , Síndrome de Waardenburg/genética , Síndrome de Waardenburg/patologia , Adolescente , Humanos , Masculino , Fenótipo , Sibéria
18.
Int J Med Sci ; 16(4): 602-606, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31171912

RESUMO

Hyperpigmentation is a type of pigmentary disorder induced by overexpression of melanin content activated severe esthetic problems as melasma, freckle, ephelides, lentigo and other forms on human skin. Several whitening agents have restricted use because of their side effects or stability such as kojic acid, ascorbic acid and hydroquinone can act as cytotoxic substance which associated to dermatitis and skin cancer. To find for the safe substance, this study aimed to find for the ability of several components in Sucrier banana peel (SBP) extracts to inhibit melanogenesis process through p38 signaling pathway in B16F10 mouse melanoma cells. Tyrosinase activity and the cellular melanin content were dose dependent manner decreasing after SBP treatment. Furthermore, SBP decreased the expression of melanogenesis relate protein as microphthalmia-associated transcription factor (MITF) and tyrosinase protein after 24 hours incubation with α-melanocyte stimulating hormones (MSH) stimulating. The findings demonstrated that SBP contained an effective agent for hyperpigmentation inhibitor through p38 signaling pathways without any effect to ERK pathway, and subsequent down-regulate MITF expression and tyrosinase enzyme family production.


Assuntos
Hiperpigmentação/tratamento farmacológico , Melaninas/biossíntese , Melanoma Experimental/tratamento farmacológico , Musa/química , Animais , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Melaninas/antagonistas & inibidores , Melanoma Experimental/genética , Melanoma Experimental/patologia , Camundongos , Fator de Transcrição Associado à Microftalmia/genética , Monofenol Mono-Oxigenase/genética , Extratos Vegetais/química , Extratos Vegetais/farmacologia , alfa-MSH/farmacologia
19.
PLoS Genet ; 15(6): e1008213, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31199790

RESUMO

The neural crest (NC) is a vertebrate-specific cell type that contributes to a wide range of different tissues across all three germ layers. The gene regulatory network (GRN) responsible for the formation of neural crest is conserved across vertebrates. Central to the induction of the NC GRN are AP-2 and SoxE transcription factors. NC induction robustness is ensured through the ability of some of these transcription factors to compensate loss of function of gene family members. However the gene regulatory events underlying compensation are poorly understood. We have used gene knockout and RNA sequencing strategies to dissect NC induction and compensation in zebrafish. We genetically ablate the NC using double mutants of tfap2a;tfap2c or remove specific subsets of the NC with sox10 and mitfa knockouts and characterise genome-wide gene expression levels across multiple time points. We find that compensation through a single wild-type allele of tfap2c is capable of maintaining early NC induction and differentiation in the absence of tfap2a function, but many target genes have abnormal expression levels and therefore show sensitivity to the reduced tfap2 dosage. This separation of morphological and molecular phenotypes identifies a core set of genes required for early NC development. We also identify the 15 somites stage as the peak of the molecular phenotype which strongly diminishes at 24 hpf even as the morphological phenotype becomes more apparent. Using gene knockouts, we associate previously uncharacterised genes with pigment cell development and establish a role for maternal Hippo signalling in melanocyte differentiation. This work extends and refines the NC GRN while also uncovering the transcriptional basis of genetic compensation via paralogues.


Assuntos
Desenvolvimento Embrionário/genética , Crista Neural/crescimento & desenvolvimento , Fatores de Transcrição SOXE/genética , Fator de Transcrição AP-2/genética , Proteínas de Peixe-Zebra/genética , Animais , Diferenciação Celular/genética , Linhagem da Célula/genética , Embrião não Mamífero , Regulação da Expressão Gênica no Desenvolvimento/genética , Redes Reguladoras de Genes/genética , Melanócitos/metabolismo , Fator de Transcrição Associado à Microftalmia/genética , Crista Neural/metabolismo , Pigmentação/genética , Proteínas Serina-Treonina Quinases/genética , Peixe-Zebra/genética , Peixe-Zebra/crescimento & desenvolvimento
20.
Genes Dev ; 33(15-16): 983-1007, 2019 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-31123060

RESUMO

All transcription factors are equal, but some are more equal than others. In the 25 yr since the gene encoding the microphthalmia-associated transcription factor (MITF) was first isolated, MITF has emerged as a key coordinator of many aspects of melanocyte and melanoma biology. Like all transcription factors, MITF binds to specific DNA sequences and up-regulates or down-regulates its target genes. What marks MITF as being remarkable among its peers is the sheer range of biological processes that it appears to coordinate. These include cell survival, differentiation, proliferation, invasion, senescence, metabolism, and DNA damage repair. In this article we present our current understanding of MITF's role and regulation in development and disease, as well as those of the MITF-related factors TFEB and TFE3, and highlight key areas where our knowledge of MITF regulation and function is limited.


Assuntos
Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/metabolismo , Regulação Neoplásica da Expressão Gênica , Melanócitos/fisiologia , Melanoma/fisiopatologia , Fator de Transcrição Associado à Microftalmia/metabolismo , Animais , Genoma , Humanos , Fator de Transcrição Associado à Microftalmia/genética , Ligação Proteica , Isoformas de Proteínas
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