RESUMO
Metastatic melanoma is hallmarked by its ability of phenotype switching to more slowly proliferating, but highly invasive cells. Here, we tested the impact of signal transducer and activator of transcription 3 (STAT3) on melanoma progression in association with melanocyte inducing transcription factor (MITF) expression levels. We established a mouse melanoma model for deleting Stat3 in melanocytes with specific expression of human hyperactive NRASQ61K in an Ink4a-deficient background, two frequent driver mutations in human melanoma. Mice devoid of Stat3 showed early disease onset with higher proliferation in primary tumors, but displayed significantly diminished lung, brain, and liver metastases. Whole-genome expression profiling of tumor-derived cells also showed a reduced invasion phenotype, which was further corroborated by 3D melanoma model analysis. Notably, loss or knockdown of STAT3 in mouse or human cells resulted in the upregulation of MITF and induction of cell proliferation. Mechanistically we show that STAT3-induced CAAT Box Enhancer Binding Protein (CEBP) expression was sufficient to suppress MITF transcription. Epigenetic analysis by ATAC-seq confirmed that CEBPa/b binding to the MITF enhancer region silenced the MITF locus. Finally, by classification of patient-derived melanoma samples, we show that STAT3 and MITF act antagonistically and hence contribute differentially to melanoma progression. We conclude that STAT3 is a driver of the metastatic process in melanoma and able to antagonize MITF via direct induction of CEBP family member transcription.
Assuntos
Proteína beta Intensificadora de Ligação a CCAAT/genética , Melanoma/genética , Fator de Transcrição Associado à Microftalmia/genética , Fator de Transcrição STAT3/genética , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Modelos Animais de Doenças , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Melanócitos/efeitos dos fármacos , Melanoma/patologia , Camundongos , Metástase Neoplásica , Transdução de Sinais/efeitos dos fármacosRESUMO
Although c-Myc is essential for melanocyte development, its role in cutaneous melanoma, the most aggressive skin cancer, is only partly understood. Here we used the NrasQ61KINK4a-/- mouse melanoma model to show that c-Myc is essential for tumor initiation, maintenance, and metastasis. c-Myc-expressing melanoma cells were preferentially found at metastatic sites, correlated with increased tumor aggressiveness and high tumor initiation potential. Abrogation of c-Myc caused apoptosis in primary murine and human melanoma cells. Mechanistically, c-Myc-positive melanoma cells activated and became dependent on the metabolic energy sensor AMP-activated protein kinase (AMPK), a metabolic checkpoint kinase that plays an important role in energy and redox homeostasis under stress conditions. AMPK pathway inhibition caused apoptosis of c-Myc-expressing melanoma cells, while AMPK activation protected against cell death of c-Myc-depleted melanoma cells through suppression of oxidative stress. Furthermore, TCGA database analysis of early-stage human melanoma samples revealed an inverse correlation between C-MYC and patient survival, suggesting that C-MYC expression levels could serve as a prognostic marker for early-stage disease.
Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Transformação Celular Neoplásica/genética , Melanoma/patologia , Estresse Oxidativo/fisiologia , Proteínas Proto-Oncogênicas c-myc/metabolismo , Proteínas Quinases Ativadas por AMP/genética , Animais , Apoptose/genética , Linhagem Celular Tumoral , Sobrevivência Celular , Inibidor p16 de Quinase Dependente de Ciclina/genética , GTP Fosfo-Hidrolases/metabolismo , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Melanócitos/patologia , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Prognóstico , Proteínas Proto-Oncogênicas c-myc/genética , Interferência de RNA , RNA Interferente Pequeno/genética , Transdução de SinaisRESUMO
NRAS and its effector BRAF are frequently mutated in melanoma. Paradoxically, CRAF but not BRAF was shown to be critical for various RAS-driven cancers, raising the question of the role of RAF proteins in NRAS-induced melanoma. Here, using conditional ablation of Raf genes in NRAS-induced mouse melanoma models, we investigate their contribution in tumour progression, from the onset of benign tumours to malignant tumour maintenance. We show that BRAF expression is required for ERK activation and nevi development, demonstrating a critical role in the early stages of NRAS-driven melanoma. After melanoma formation, single Braf or Craf ablation is not sufficient to block tumour growth, showing redundant functions for RAF kinases. Finally, proliferation of resistant cells emerging in the absence of BRAF and CRAF remains dependent on ARAF-mediated ERK activation. These results reveal specific and compensatory functions for BRAF and CRAF and highlight an addiction to RAF signalling in NRAS-driven melanoma.
Assuntos
Melanoma/metabolismo , Proteínas Monoméricas de Ligação ao GTP/metabolismo , Proteínas Proto-Oncogênicas B-raf/metabolismo , Proteínas Proto-Oncogênicas c-raf/metabolismo , Proteínas ras/metabolismo , Animais , Linhagem Celular Tumoral , Progressão da Doença , Humanos , Sistema de Sinalização das MAP Quinases/genética , Melanoma/genética , Melanoma/patologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Proteínas Monoméricas de Ligação ao GTP/genética , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas c-raf/genética , Proteínas ras/genéticaRESUMO
The role of the Pax3 gene in embryonic development of pigment cells is well characterized. By contrast, the function of Pax3 in melanoma development is controversial. Indeed, data obtained from cultured cells suggest that PAX3 may contribute to melanomagenesis. PAX3 is found to be overexpressed in melanomas and also in nevi compared with normal skin samples. Pax3 homozygous loss of function is embryonic lethal. To assess the role of Pax3 in melanoma development in vivo, we analyzed Pax3 haploinsufficiency in a mouse model of melanoma predisposition. The Pax3(GFP/+) knock-in reporter system was combined with the Tyr::NRAS(Q61K); Cdkn2a(-/-) mouse melanoma model. Melanoma development was followed over 18 months. Histopathological, immunohistochemical, and molecular analyses of lesions at different stages of melanoma progression were carried out. Fluorescence-activated cell sorting on GFP of cells from primary or metastatic melanoma was followed by ex-vivo transformation tests and in-vivo passaging. We report here that Tyr::NRAS(Q61K); Cdkn2a(-/-); Pax3(GFP/+) mice developed metastasizing melanoma as their Tyr::NRAS(Q61K); Cdkn2a(-/-); littermates. Histopathology showed no differences between the two genotypes, although Pax3 mRNA and PAX3 protein levels in Pax3(GFP/+) lesions were reduced by half. The Pax3(GFP) allele proved to be a convenient marker to identify and directly sort heterogeneous populations of melanoma cells within the tumor bulk at each stage of melanoma progression. This new mouse model represents an accurate and reproducible means for identifying melanoma cells in vivo to study the mechanisms of melanoma development.
Assuntos
Transformação Celular Neoplásica/genética , Haploinsuficiência/fisiologia , Melanoma/genética , Fatores de Transcrição Box Pareados/genética , Neoplasias Cutâneas/genética , Alelos , Substituição de Aminoácidos , Animais , Separação Celular , Células Cultivadas , Feminino , Genes Reporter , Genes p16 , Genes ras , Proteínas de Fluorescência Verde/genética , Melanoma/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Monofenol Mono-Oxigenase/genética , Fator de Transcrição PAX3 , Neoplasias Cutâneas/patologiaAssuntos
Transformação Celular Neoplásica , Peptídeos e Proteínas de Sinalização Intracelular/genética , Melanoma , Proteínas Nucleares/genética , Proteínas Supressoras de Tumor/genética , Animais , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/metabolismo , Transformação Celular Neoplásica/patologia , Proteína 1 Inibidora do Crescimento , Melanoma/genética , Melanoma/metabolismo , Melanoma/patologia , Camundongos , Camundongos Knockout , Metástase NeoplásicaRESUMO
The ciliary body and iris are pigmented epithelial structures in the anterior eye segment that function to maintain correct intra-ocular pressure and regulate exposure of the internal eye structures to light, respectively. The cellular and molecular factors that mediate the development of the ciliary body and iris from the ocular pigmented epithelium remain to be fully elucidated. Here, we have investigated the role of Notch signaling during the development of the anterior pigmented epithelium by using genetic loss- and gain-of-function approaches. Loss of canonical Notch signaling results in normal iris development but absence of the ciliary body. This causes progressive hypotony and over time leads to phthisis bulbi, a condition characterized by shrinkage of the eye and loss of structure/function. Conversely, Notch gain-of-function results in aniridia and profound ciliary body hyperplasia, which causes ocular hypertension and glaucoma-like disease. Collectively, these data indicate that Notch signaling promotes ciliary body development at the expense of iris formation and reveals novel animal models of human ocular pathologies.
Assuntos
Corpo Ciliar/embriologia , Proteínas do Olho/metabolismo , Iris/embriologia , Epitélio Pigmentado Ocular/embriologia , Receptores Notch/metabolismo , Transdução de Sinais/fisiologia , Animais , Corpo Ciliar/citologia , Proteínas do Olho/genética , Humanos , Iris/citologia , Camundongos , Camundongos Transgênicos , Epitélio Pigmentado Ocular/citologia , Receptores Notch/genéticaRESUMO
Autophagy is linked to cell death, yet the associated mechanisms are largely undercharacterized. We discovered that melanoma, which is generally resistant to drug-induced apoptosis, can undergo autophagic cell death with the participation of orphan nuclear receptor TR3. A sequence of molecular events leading to cellular demise is launched by a specific chemical compound, 1-(3,4,5-trihydroxyphenyl)nonan-1-one, newly acquired from screening a library of TR3-targeting compounds. The autophagic cascade comprises TR3 translocation to mitochondria through interaction with the mitochondrial outer membrane protein Nix, crossing into the mitochondrial inner membrane through Tom40 and Tom70 channel proteins, dissipation of mitochondrial membrane potential by the permeability transition pore complex ANT1-VDAC1 and induction of autophagy. This process leads to excessive mitochondria clearance and irreversible cell death. It implicates a new approach to melanoma therapy through activation of a mitochondrial signaling pathway that integrates a nuclear receptor with autophagy for cell death.
Assuntos
Autofagia , Cetonas/química , Mitocôndrias/fisiologia , Membro 1 do Grupo A da Subfamília 4 de Receptores Nucleares/metabolismo , Pirogalol/análogos & derivados , Transdução de Sinais , Animais , Linhagem Celular Tumoral , Cristalografia por Raios X , Modelos Animais de Doenças , Humanos , Cetonas/farmacologia , Melanoma/tratamento farmacológico , Proteínas de Membrana/metabolismo , Camundongos , Conformação Proteica , Proteínas Proto-Oncogênicas/metabolismo , Pirogalol/química , Pirogalol/farmacologia , Proteínas Supressoras de Tumor/metabolismoRESUMO
The extracellular potential of excitable and nonexcitable cells with respect to ground is â¼0 mV. One of the known exceptions in mammals is the cochlear duct, where the potential is â¼80-100 mV, called the endocochlear potential (EP). The EP serves as the "battery" for transduction of sound, contributing toward the sensitivity of the auditory system. The stria vascularis (StV) of the cochlear duct is the station where the EP is generated, but the cell-specific roles in the StV are ill defined. Using the intermediate cell (IC)-specific tyrosinase promoter, under the control of diphtheria toxin (DT), we eliminated and/or halted differentiation of neural crest melanocytes after migration to the StV. The ensuing adult transgenic mice are profoundly deaf. Additionally, the EP was abolished. Expression of melanocyte early marker and Kir4.1 in ICs precedes the onset of pigment synthesis. Activation of DT leads to loss of ICs. Finally, in accord with the distinct embryology of retinal pigmented cells, transgenic mice with toxigenic ablation of neural crest-derived melanocytes have intact visual responses. We assert that the tyrosinase promoter is the distinct target for genetic manipulation of IC-specific genes.
Assuntos
Surdez/genética , Toxina Diftérica/genética , Estria Vascular/citologia , Transgenes/genética , Potenciais de Ação , Animais , Diferenciação Celular , Toxina Diftérica/metabolismo , Melanócitos/citologia , Melanócitos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Monofenol Mono-Oxigenase/genética , Crista Neural/citologia , Crista Neural/metabolismo , Canais de Potássio Corretores do Fluxo de Internalização/genética , Canais de Potássio Corretores do Fluxo de Internalização/metabolismo , Regiões Promotoras Genéticas , Epitélio Pigmentado da Retina/citologia , Epitélio Pigmentado da Retina/metabolismo , Estria Vascular/metabolismo , Estria Vascular/fisiologia , Visão Ocular/genéticaAssuntos
Inibidor p16 de Quinase Dependente de Ciclina/deficiência , Inibidor p16 de Quinase Dependente de Ciclina/genética , Melanócitos/metabolismo , Melanócitos/patologia , Mutação/genética , Proteínas ras/genética , Animais , Melanoma/patologia , Camundongos , Camundongos Transgênicos , Neoplasias Cutâneas/patologiaRESUMO
The serine protease CAP1/Prss8 is crucial for skin barrier function, lung alveolar fluid clearance and has been unveiled as diagnostic marker for specific cancer types. Here, we show that a constitutive knockout of CAP1/Prss8 leads to embryonic lethality. These embryos presented no specific defects, but it is during this period, and in particular at E13.5, that wildtype placentas show an increased expression of CAP1/Prss8, thus suggesting a placental defect in the knockout situation. The placentas of knockout embryos exhibited significantly reduced vascular development and incomplete cellular maturation. In contrary, epiblast-specific deletion of CAP1/Prss8 allowed development until birth. These CAP1/Prss8-deficient newborns presented abnormal epidermis, and died soon after birth due to impaired skin function. We thus conclude that a late placental insufficiency might be the primary cause of embryonic lethality in CAP1/Prss8 knockouts. This study highlights a novel and crucial role for CAP1/Prss8 in placental development and function.
Assuntos
Orelha Interna/crescimento & desenvolvimento , Perda do Embrião/genética , Embrião de Mamíferos/citologia , Desenvolvimento Embrionário/fisiologia , Regulação da Expressão Gênica no Desenvolvimento , Placentação/fisiologia , Serina Endopeptidases/fisiologia , Animais , Animais Recém-Nascidos , Diferenciação Celular , Embrião de Mamíferos/enzimologia , Feminino , Perfilação da Expressão Gênica , Masculino , Camundongos , Camundongos Knockout , Gravidez , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo RealRESUMO
BACKGROUND: Patent ductus arteriosus is a life-threatening condition frequent in premature newborns but also present in some term infants. Current mouse models of this malformation generally lead to perinatal death, not reproducing the full phenotypic spectrum in humans, in whom genetic inheritance appears complex. The ductus arteriosus (DA), a temporary fetal vessel that bypasses the lungs by shunting the aortic arch to the pulmonary artery, is constituted by smooth muscle cells of distinct origins (SMC1 and SMC2) and many fewer melanocytes. To understand novel mechanisms preventing DA closure at birth, we evaluated the importance of cell fate specification in SMC that form the DA during embryonic development. Upon specific Tyr::Cre-driven activation of Wnt/ß-catenin signaling at the time of cell fate specification, melanocytes replaced the SMC2 population of the DA, suggesting that SMC2 and melanocytes have a common precursor. The number of SMC1 in the DA remained similar to that in controls, but insufficient to allow full DA closure at birth. Thus, there was no cellular compensation by SMC1 for the loss of SMC2. Mice in which only melanocytes were genetically ablated after specification from their potential common precursor with SMC2, demonstrated that differentiated melanocytes themselves do not affect DA closure. Loss of the SMC2 population, independent of the presence of melanocytes, is therefore a cause of patent ductus arteriosus and premature death in the first months of life. Our results indicate that patent ductus arteriosus can result from the insufficient differentiation, proliferation, or contractility of a specific smooth muscle subpopulation that shares a common neural crest precursor with cardiovascular melanocytes.
Assuntos
Diferenciação Celular/fisiologia , Permeabilidade do Canal Arterial/fisiopatologia , Desenvolvimento Embrionário , Miócitos de Músculo Liso/patologia , Nascimento Prematuro/fisiopatologia , Animais , Linhagem da Célula , Proliferação de Células , Permeabilidade do Canal Arterial/etiologia , Feminino , Humanos , Melanócitos/citologia , Camundongos , Contração Muscular/fisiologia , Gravidez , Via de Sinalização WntRESUMO
Giant congenital naevi are pigmented childhood lesions that frequently lead to melanoma, the most aggressive skin cancer. The mechanisms underlying this malignancy are largely unknown, and there are no effective therapies. Here we describe a mouse model for giant congenital naevi and show that naevi and melanoma prominently express Sox10, a transcription factor crucial for the formation of melanocytes from the neural crest. Strikingly, Sox10 haploinsufficiency counteracts Nras(Q61K)-driven congenital naevus and melanoma formation without affecting the physiological functions of neural crest derivatives in the skin. Moreover, Sox10 is also crucial for the maintenance of neoplastic cells in vivo. In human patients, virtually all congenital naevi and melanomas are SOX10 positive. Furthermore, SOX10 silencing in human melanoma cells suppresses neural crest stem cell properties, counteracts proliferation and cell survival, and completely abolishes in vivo tumour formation. Thus, SOX10 represents a promising target for the treatment of congenital naevi and melanoma in human patients.
Assuntos
Melanoma/fisiopatologia , Nevo/patologia , Fatores de Transcrição SOXE/metabolismo , Animais , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Western Blotting , Linhagem Celular , Linhagem Celular Tumoral , Criança , Modelos Animais de Doenças , Feminino , Regulação Neoplásica da Expressão Gênica , Haploinsuficiência , Humanos , Imuno-Histoquímica , Lactente , Masculino , Melanoma/genética , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Análise em Microsséries , Nevo/fisiopatologia , Reação em Cadeia da Polimerase em Tempo Real , Fatores de Transcrição SOXE/genéticaRESUMO
The glucocorticoid-induced leucine zipper (Tsc22d3-2) is a widely expressed dexamethasone-induced transcript that has been proposed to be important in immunity, adipogenesis, and renal sodium handling based on in vitro studies. To address its function in vivo, we have used Cre/loxP technology to generate mice deficient for Tsc22d3-2. Male knockout mice were viable but surprisingly did not show any major deficiencies in immunological processes or inflammatory responses. Tsc22d3-2 knockout mice adapted to a sodium-deprived diet and to water deprivation conditions but developed a subtle deficiency in renal sodium and water handling. Moreover, the affected animals developed a mild metabolic phenotype evident by a reduction in weight from 6 months of age, mild hyperinsulinemia, and resistance to a high-fat diet. Tsc22d3-2-deficient males were infertile and exhibited severe testis dysplasia from postnatal d 10 onward with increases in apoptotic cells within seminiferous tubules, an increased number of Leydig cells, and significantly elevated FSH and testosterone levels. Thus, our analysis of the Tsc22d3-2-deficient mice demonstrated a previously uncharacterized function of glucocorticoid-induced leucine zipper protein in testis development.
Assuntos
Infertilidade Masculina/genética , Fatores de Transcrição/genética , Adipogenia , Animais , Peso Corporal , Contagem de Células , Células Cultivadas , Citocinas/metabolismo , Dexametasona/farmacologia , Feminino , Fibroblastos/fisiologia , Loci Gênicos , Hiperinsulinismo/genética , Sistema Imunitário/crescimento & desenvolvimento , Fatores Imunológicos/farmacologia , Lipopolissacarídeos/farmacologia , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Macrófagos/metabolismo , Masculino , Camundongos , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Isoformas de Proteínas/genética , Baço/patologia , Testículo/crescimento & desenvolvimento , Testículo/metabolismo , Testículo/patologia , Timo/patologia , Fatores de Transcrição/deficiênciaRESUMO
c-Myc is involved in the control of diverse cellular processes and implicated in the maintenance of different tissues including the neural crest. Here, we report that c-Myc is particularly important for pigment cell development and homeostasis. Targeting c-Myc specifically in the melanocyte lineage using the floxed allele of c-Myc and Tyr::Cre transgenic mice results in a congenital gray hair phenotype. The gray coat color is associated with a reduced number of functional melanocytes in the hair bulb and melanocyte stem cells in the hair bulge. Importantly, the gray phenotype does not progress with time, suggesting that maintenance of the melanocyte through the hair cycle does not involve c-Myc function. In embryos, at E13.5, c-Myc-deficient melanocyte precursors are affected in proliferation in concordance with a reduction in numbers, showing that c-Myc is required for the proper melanocyte development. Interestingly, melanocytes from c-Myc-deficient mice display elevated levels of the c-Myc paralog N-Myc. Double deletion of c-Myc and N-Myc results in nearly complete loss of the residual pigmentation, indicating that N-Myc is capable of compensating for c-Myc loss of function in melanocytes.
Assuntos
Genes myc , Cor de Cabelo/genética , Doenças do Cabelo/genética , Melanócitos/metabolismo , Animais , Embrião de Mamíferos , Deleção de Genes , Regulação da Expressão Gênica no Desenvolvimento , Genes myc/genética , Genes myc/fisiologia , Melanócitos/patologia , Camundongos , Camundongos Transgênicos , Especificidade de Órgãos/genética , Pigmentação/genética , Transtornos da Pigmentação/genética , Pigmentação da Pele/genéticaRESUMO
Melanoma antigen recognized by T cells 1 (MART-1) is a melanoma-specific antigen, which has been thoroughly studied in the context of immunotherapy against malignant melanoma and which is found only in the pigment cell lineage. However, its exact function and involvement in pigmentation is not clearly understood. Melanoma antigen recognized by T cells 1 has been shown to interact with the melanosomal proteins Pmel17 and OA1. To understand the function of MART-1 in pigmentation, we developed a new knockout mouse model. Mice deficient in MART-1 are viable, but loss of MART-1 leads to a coat color phenotype, with a reduction in total melanin content of the skin and hair. Lack of MART-1 did not affect localization of melanocyte-specific proteins nor maturation of Pmel17. Melanosomes of hair follicle melanocytes in MART-1 knockout mice displayed morphological abnormalities, which were exclusive to stage III and IV melanosomes. In conclusion, our results suggest that MART-1 is a pigmentation gene that is required for melanosome biogenesis and/or maintenance.
Assuntos
Cor de Cabelo/genética , Antígeno MART-1/fisiologia , Animais , Células Cultivadas/metabolismo , Vetores Genéticos , Cabelo/metabolismo , Antígeno MART-1/genética , Melaninas/metabolismo , Melanócitos/metabolismo , Melanossomas/metabolismo , Melanossomas/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fenótipo , Pele/citologia , Pele/metabolismo , Antígeno gp100 de Melanoma/metabolismoRESUMO
The incidence of melanoma has significantly increased, and a better understanding of its pathogenesis and development of new therapeutic strategies are urgently needed. Here, we describe a murine model of metastatic cutaneous melanoma using C57BL/6 mice expressing a mutated human N-Ras gene under the control of a tyrosinase promoter (TyrRas). These mice were topically exposed to 7,12- dimethylbenzanthracene (DMBA) for brief exposure periods. Cutaneous melanoma developed at the site of exposure on average by 19 weeks of age and in 80% of mice. Importantly, as in humans, melanoma development was associated with subsequent metastasis to tumor-draining lymph nodes. Critically, such metastatic behavior is transplantable, as intradermal inoculation of melanoma cells from TyrRas-DMBA mice into non-transgenic mice led to the growth of melanoma and, again, metastasis to skin-draining lymph nodes. This metastatic melanoma model closely mimics human pathology and should be a useful tool for studying melanoma pathogenesis and developing new therapies.
Assuntos
Carcinógenos/toxicidade , Transformação Celular Neoplásica/patologia , Genes ras/genética , Melanócitos/patologia , Melanoma/patologia , Mutação/genética , Neoplasias Cutâneas/patologia , 9,10-Dimetil-1,2-benzantraceno/toxicidade , Substituição de Aminoácidos/genética , Animais , Proliferação de Células , Transformação Celular Neoplásica/efeitos dos fármacos , Humanos , Melanócitos/efeitos dos fármacos , Melanócitos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Metástase Neoplásica , Transplante de Neoplasias , Neoplasias Cutâneas/metabolismoAssuntos
Melanoma Experimental/genética , Camundongos Mutantes/genética , Animais , Divisão Celular , Genes p16 , Cor de Cabelo/genética , Humanos , Sistema de Sinalização das MAP Quinases , Melanócitos/metabolismo , Camundongos , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/fisiologia , Fenótipo , Pigmentação da Pele/genética , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteínas Wnt/fisiologia , beta Catenina/fisiologiaRESUMO
Metastases are the major cause of death from melanoma, a skin cancer that has the fastest rising incidence of any malignancy in the Western world. Molecular pathways that drive melanoblast migration in development are believed to underpin the movement and ultimately the metastasis of melanoma. Here we show that mice lacking P-Rex1, a Rac-specific Rho GTPase guanine nucleotide exchange factor, have a melanoblast migration defect during development evidenced by a white belly. Moreover, these P-Rex1(-/-) mice are resistant to metastasis when crossed to a murine model of melanoma. Mechanistically, this is associated with P-Rex1 driving invasion in a Rac-dependent manner. P-Rex1 is elevated in the majority of human melanoma cell lines and tumour tissue. We conclude that P-Rex1 has an important role in melanoblast migration and cancer progression to metastasis in mice and humans.
Assuntos
Movimento Celular/fisiologia , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Melanoma/metabolismo , Melanoma/patologia , Metástase Neoplásica/fisiopatologia , Animais , Movimento Celular/genética , Células Cultivadas , Fatores de Troca do Nucleotídeo Guanina/genética , Humanos , Imuno-Histoquímica , Técnicas In Vitro , Melanoma/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Metástase Neoplásica/genética , Análise Serial de TecidosRESUMO
The number of transgenic mouse lines expressing Cre in either type of pigment cells (melanocytes and retinal pigment epithelium, RPE) is limited, and the available lines do not always offer sufficient specificity. In this study, we addressed this issue and we report on the generation of a MART-1::Cre BAC transgenic mouse line, in which the expression of Cre recombinase is controlled by regulatory elements of the pigment cell-specific gene MART-1 (mlana). When MART-1::Cre BAC transgenic mice were bred with the ROSA26-R reporter line, ß-galactosidase expression was observed in RPE from E12.5 onwards, and in melanocyte precursors from E17.5, indicating that the MART-1::Cre line provides Cre recombinase activity in pigment-producing cells rather than in a particular lineage. In addition, breeding of this mouse line to mice carrying a conditional allele of RBP-Jκ corroborated the reported phenotypes in both pigment cell lineages, inducing hair greying and microphthalmia. Our results thus suggest, that the MART-1::Cre line may serve as a novel and useful tool for functional studies in melanocytes and the RPE.genesis 49:403-409, 2011.
Assuntos
Integrases/genética , Antígeno MART-1/genética , Melanócitos/metabolismo , Epitélio Pigmentado Ocular/metabolismo , Recombinação Genética , Animais , Cromossomos Artificiais Bacterianos/genética , Embrião de Mamíferos/citologia , Embrião de Mamíferos/embriologia , Embrião de Mamíferos/metabolismo , Feminino , Imunofluorescência , Regulação da Expressão Gênica no Desenvolvimento , Integrases/metabolismo , Masculino , Melanócitos/citologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos , Camundongos Transgênicos , Epitélio Pigmentado Ocular/embriologia , Sequências Reguladoras de Ácido Nucleico/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Tempo , beta-Galactosidase/metabolismoRESUMO
B-cell-activating factor of the TNF family (BAFF)/BLyS contributes to B-cell homeostasis and function in the periphery. BAFF is expressed as a membrane-bound protein or released by proteolytic cleavage, but the functional importance of this processing event is poorly understood. Mice expressing BAFF with a mutated furin consensus cleavage site, i.e. furin-mutant BAFF (fmBAFF), were not different from BAFF-deficient mice with regard to their B-cell populations and responses to immunization. It is however noteworthy that an alternative processing event releases some soluble BAFF in fmBAFF mice. Mild overexpression (â¼ 5-fold) of fmBAFF alone generated intermediate levels of B cells without improving humoral responses to immunization. Processed BAFF was however important for B-cell homeostasis, as peripheral B-cell populations and antibody responses were readily restored by administration of soluble BAFF trimers in BAFF-deficient mice. However, the rescue of CD23 expression in B cells of BAFF-deficient mice required both soluble BAFF trimers and fmBAFF, or a polymeric form of soluble BAFF (BAFF 60-mer). These results point to a predominant role of processed BAFF for B-cell homeostasis and function, and indicate possible accessory roles for membrane-bound BAFF.