RESUMEN
While the major drivers of melanoma initiation, including activation of NRAS/BRAF and loss of PTEN or CDKN2A, have been identified, the role of key transcription factors that impose altered transcriptional states in response to deregulated signaling is not well understood. The POU domain transcription factor BRN2 is a key regulator of melanoma invasion, yet its role in melanoma initiation remains unknown. Here, in a BrafV600E PtenF/+ context, we show that BRN2 haplo-insufficiency promotes melanoma initiation and metastasis. However, metastatic colonization is less efficient in the absence of Brn2. Mechanistically, BRN2 directly induces PTEN expression and in consequence represses PI3K signaling. Moreover, MITF, a BRN2 target, represses PTEN transcription. Collectively, our results suggest that on a PTEN heterozygous background somatic deletion of one BRN2 allele and temporal regulation of the other allele elicits melanoma initiation and progression.
Asunto(s)
Carcinogénesis/metabolismo , Proliferación Celular/genética , Regulación Neoplásica de la Expresión Génica/genética , Genes Supresores de Tumor , Proteínas de Homeodominio/metabolismo , Melanoma/metabolismo , Factores del Dominio POU/metabolismo , Neoplasias Cutáneas/metabolismo , Animales , Carcinogénesis/genética , Línea Celular Tumoral , Inmunoprecipitación de Cromatina , Estudios de Cohortes , Variaciones en el Número de Copia de ADN , Progresión de la Enfermedad , Técnicas de Silenciamiento del Gen , Haploinsuficiencia , Proteínas de Homeodominio/genética , Humanos , Inmunohistoquímica , Melanoma/genética , Melanoma/mortalidad , Melanoma/secundario , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Análisis por Micromatrices , Factor de Transcripción Asociado a Microftalmía/metabolismo , Mutación , Factores del Dominio POU/genética , Fosfohidrolasa PTEN/genética , Fosfohidrolasa PTEN/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Proteínas Proto-Oncogénicas B-raf/genética , ARN Interferente Pequeño , Neoplasias Cutáneas/genética , Neoplasias Cutáneas/mortalidad , Neoplasias Cutáneas/secundario , Melanoma Cutáneo MalignoRESUMEN
MIcrophthalmia-associated Transcription Factor (MITF) regulates melanocyte and melanoma physiology. We show that MITF associates the NURF chromatin-remodelling factor in melanoma cells. ShRNA-mediated silencing of the NURF subunit BPTF revealed its essential role in several melanoma cell lines and in untransformed melanocytes in vitro. Comparative RNA-seq shows that MITF and BPTF co-regulate overlapping gene expression programs in cell lines in vitro. Somatic and specific inactivation of Bptf in developing murine melanoblasts in vivo shows that Bptf regulates their proliferation, migration and morphology. Once born, Bptf-mutant mice display premature greying where the second post-natal coat is white. This second coat is normally pigmented by differentiated melanocytes derived from the adult melanocyte stem cell (MSC) population that is stimulated to proliferate and differentiate at anagen. An MSC population is established and maintained throughout the life of the Bptf-mutant mice, but these MSCs are abnormal and at anagen, give rise to reduced numbers of transient amplifying cells (TACs) that do not express melanocyte markers and fail to differentiate into mature melanin producing melanocytes. MSCs display a transcriptionally repressed chromatin state and Bptf is essential for reactivation of the melanocyte gene expression program at anagen, the subsequent normal proliferation of TACs and their differentiation into mature melanocytes.
Asunto(s)
Antígenos Nucleares/genética , Ensamble y Desensamble de Cromatina/genética , Melanoma/genética , Células Madre Mesenquimatosas , Factor de Transcripción Asociado a Microftalmía/genética , Proteínas del Tejido Nervioso/genética , Factores de Transcripción/genética , Animales , Ciclo Celular/genética , Diferenciación Celular/genética , División Celular/genética , Regulación del Desarrollo de la Expresión Génica , Folículo Piloso , Melanocitos/metabolismo , Melanoma/patología , RatonesRESUMEN
We show, for the first time, that melanocytes can form a primary cilium in vitro, corresponding to an immotile or sensory cilium. Such cilia are observed when melanocytes reach confluence or when medium nutrient levels are insufficient. This observation should greatly improve our understanding of the signal transduction processes potentially occurring in these cells during embryonic development, homeostasis in adulthood and melanomagenesis.
Asunto(s)
Senescencia Celular/fisiología , Cilios/patología , Melanocitos/patología , Animales , Línea Celular , Proliferación Celular/fisiología , Células Cultivadas , Cilios/fisiología , Humanos , Técnicas In Vitro , Melanocitos/fisiología , Ratones , Transducción de Señal/fisiologíaRESUMEN
DNA methylation is a major epigenetic mechanism for gene silencing. Whereas methyltransferases mediate cytosine methylation, it is less clear how unmethylated regions in mammalian genomes are protected from de novo methylation and whether an active demethylating activity is involved. Here, we show that either knockout or catalytic inactivation of the DNA repair enzyme thymine DNA glycosylase (TDG) leads to embryonic lethality in mice. TDG is necessary for recruiting p300 to retinoic acid (RA)-regulated promoters, protection of CpG islands from hypermethylation, and active demethylation of tissue-specific developmentally and hormonally regulated promoters and enhancers. TDG interacts with the deaminase AID and the damage response protein GADD45a. These findings highlight a dual role for TDG in promoting proper epigenetic states during development and suggest a two-step mechanism for DNA demethylation in mammals, whereby 5-methylcytosine and 5-hydroxymethylcytosine are first deaminated by AID to thymine and 5-hydroxymethyluracil, respectively, followed by TDG-mediated thymine and 5-hydroxymethyluracil excision repair.
