RESUMEN
Stem cell (SC) differentiation and maintenance of resultant progeny underlie cell turnover in many organs, but it is difficult to pinpoint the contribution of either process. In the pituitary, a central regulator of endocrine axes, adult SCs undergo activation after target organ ablation, providing a well-characterized paradigm to study an adaptative response in a multi-organ system. Here, we used single-cell technologies to characterize SC heterogeneity and mobilization together with lineage tracing. We show that SC differentiation occurs more frequently than thought previously. In adaptative conditions, differentiation increases and is more diverse than demonstrated by the lineage tracing experiments. Detailed examination of SC progeny suggests that maintenance of selected nascent cells underlies SC output, highlighting a trophic role for the microenvironment. Analyses of cell trajectories further predict pathways and potential regulators. Our model provides a valuable system to study the influence of evolving states on the mechanisms of SC mobilization.
Asunto(s)
Células Madre , Células Madre/metabolismo , Diferenciación CelularRESUMEN
The pituitary is the master neuroendocrine gland, which regulates body homeostasis. It consists of the anterior pituitary/adenohypophysis harboring hormones producing cells and the posterior pituitary/neurohypophysis, which relays the passage of hormones from the brain to the periphery. It is accepted that the adenohypophysis originates from the oral ectoderm (Rathke's pouch), whereas the neural ectoderm contributes to the neurohypophysis. Single-cell transcriptomics of the zebrafish pituitary showed that cyp26b1-positive astroglial pituicytes of the neurohypophysis and prop1-positive adenohypophyseal progenitors expressed common markers implying lineage relatedness. Genetic tracing identifies that, in contrast to the prevailing dogma, neural plate precursors of zebrafish (her4.3+) and mouse (Sox1+) contribute to both neurohypophyseal and a subset of adenohypophyseal cells. Pituicyte-derived retinoic-acid-degrading enzyme Cyp26b1 fine-tunes differentiation of prop1+ progenitors into hormone-producing cells. These results challenge the notion that adenohypophyseal cells are exclusively derived from non-neural ectoderm and demonstrate that crosstalk between neuro- and adeno-hypophyseal cells affects differentiation of pituitary cells.
Asunto(s)
Neurohipófisis , Ratones , Animales , Pez Cebra , Placa Neural , Ácido Retinoico 4-Hidroxilasa , HormonasRESUMEN
The pituitary gland regulates growth, metabolism, reproduction, the stress response, uterine contractions, lactation, and water retention. It secretes hormones in response to hypothalamic input, end organ feedback, and diurnal cues. The mechanisms by which pituitary stem cells are recruited to proliferate, maintain quiescence, or differentiate into specific cell types, especially thyrotropes, are not well understood. We used single-cell RNA sequencing in juvenile P7 mouse pituitary cells to identify novel factors in pituitary cell populations, with a focus on thyrotropes and rare subtypes. We first observed cells coexpressing markers of both thyrotropes and gonadotropes, such as Pou1f1 and Nr5a1. This was validated in vivo by both immunohistochemistry and lineage tracing of thyrotropes derived from Nr5a1-Cre; mTmG mice and demonstrates that Nr5a1-progenitors give rise to a proportion of thyrotropes during development. Our data set also identifies novel factors expressed in pars distalis and pars tuberalis thyrotropes, including the Shox2b isoform in all thyrotropes and Sox14 specifically in Pou1f1-negative pars tuberalis thyrotropes. We have therefore used single-cell transcriptomics to determine a novel developmental trajectory for thyrotropes and potential novel regulators of thyrotrope populations.
Asunto(s)
Enfermedades de la Hipófisis , Adenohipófisis , Embarazo , Femenino , Ratones , Animales , Tirotropina/metabolismo , Hipófisis/metabolismo , Factores de Transcripción/metabolismo , Enfermedades de la Hipófisis/metabolismo , Inmunohistoquímica , Adenohipófisis/metabolismo , Factores de Transcripción SOXB2/metabolismoRESUMEN
The adult dentate gyrus (DG) of rodents hosts a neural stem cell (NSC) niche capable of generating new neurons throughout life. The embryonic origin and molecular mechanisms underlying formation of DG NSCs are still being investigated. We performed a bulk transcriptomic analysis on mouse developing archicortex conditionally deleted for Sox9, a SoxE transcription factor controlling both gliogenesis and NSC formation, and identified Hopx, a recently identified marker of both prospective adult DG NSCs and astrocytic progenitors, as being downregulated. We confirm SOX9 is required for HOPX expression in the embryonic archicortex. In particular, we found that both NSC markers are highly expressed in the cortical hem (CH), while only weakly in the adjacent dentate neuroepithelium (DNE), suggesting a potential CH embryonic origin for DG NSCs. However, we demonstrate both in vitro and in vivo that the embryonic CH, as well as its adult derivatives, lacks stem cell potential. Instead, deletion of Sox9 in the DNE affects both HOPX expression and NSC formation in the adult DG. We conclude that HOPX expression in the CH is involved in astrocytic differentiation downstream of SOX9, which we previously showed regulates DG development by inducing formation of a CH-derived astrocytic scaffold. Altogether, these results suggest that both proteins work in a dose-dependent manner to drive either astrocytic differentiation in CH or NSC formation in DNE.
Asunto(s)
Células-Madre Neurales , Ratones , Animales , Células-Madre Neurales/metabolismo , Giro Dentado , Diferenciación Celular/fisiología , Estudios Prospectivos , NeurogénesisRESUMEN
Biallelic RNPC3 variants have been reported in a few patients with growth hormone deficiency, either in isolation or in association with central hypothyroidism, congenital cataract, neuropathy, developmental delay/intellectual disability, hypogonadism, and pituitary hypoplasia. To describe a new patient with syndromic congenital hypopituitarism and diffuse brain atrophy due to RNPC3 mutations and to compare her clinical and molecular characteristics and pituitary functions with previously published patients. A 20-year-old female presented with severe growth, neuromotor, and developmental delay. Her weight, height, and head circumference were 5135 gr (-25.81 SDS), 68 cm (-16.17 SDS), and 34 cm (-17.03 SDS), respectively. She was prepubertal, and had dysmorphic facies, contractures, and spasticity in the extremities, and severe truncal hypotonia. There were no radiological signs of a skeletal dysplasia. The bone age was extremely delayed at 2 years. Investigation of pituitary function revealed growth hormone, prolactin, and thyroid-stimulating hormone deficiencies. Whole-exome sequencing revealed a novel homozygous missense (c.1328A > G; Y443C) variant in RNPC3. Cranial MRI revealed a hypoplastic anterior pituitary with diffuse cerebral and cerebellar atrophy. The Y443C variant in RNPC3 associated with syndromic congenital hypopituitarism and abnormal brain development. This report extends the RNPC3-related hypopituitarism phenotype with a severe neurodegenerative presentation.
Asunto(s)
Hormona de Crecimiento Humana , Hipopituitarismo , Hipotiroidismo , Atrofia , Femenino , Hormona del Crecimiento/genética , Homocigoto , Humanos , Hipopituitarismo/diagnóstico , Hipopituitarismo/genética , Hipotiroidismo/genética , Proteínas Nucleares/genética , Hipófisis/anomalías , Proteínas de Unión al ARN/genéticaRESUMEN
PURPOSE: We aimed to investigate the molecular basis underlying a novel phenotype including hypopituitarism associated with primary ovarian insufficiency. METHODS: We used next-generation sequencing to identify variants in all pedigrees. Expression of Rnpc3/RNPC3 was analyzed by in situ hybridization on murine/human embryonic sections. CRISPR/Cas9 was used to generate mice carrying the p.Leu483Phe pathogenic variant in the conserved murine Rnpc3 RRM2 domain. RESULTS: We described 15 patients from 9 pedigrees with biallelic pathogenic variants in RNPC3, encoding a specific protein component of the minor spliceosome, which is associated with a hypopituitary phenotype, including severe growth hormone (GH) deficiency, hypoprolactinemia, variable thyrotropin (also known as thyroid-stimulating hormone) deficiency, and anterior pituitary hypoplasia. Primary ovarian insufficiency was diagnosed in 8 of 9 affected females, whereas males had normal gonadal function. In addition, 2 affected males displayed normal growth when off GH treatment despite severe biochemical GH deficiency. In both mouse and human embryos, Rnpc3/RNPC3 was expressed in the developing forebrain, including the hypothalamus and Rathke's pouch. Female Rnpc3 mutant mice displayed a reduction in pituitary GH content but with no reproductive impairment in young mice. Male mice exhibited no obvious phenotype. CONCLUSION: Our findings suggest novel insights into the role of RNPC3 in female-specific gonadal function and emphasize a critical role for the minor spliceosome in pituitary and ovarian development and function.
Asunto(s)
Hipopituitarismo , Insuficiencia Ovárica Primaria , Animales , Femenino , Humanos , Hipopituitarismo/genética , Masculino , Ratones , Proteínas Nucleares/genética , Linaje , Fenotipo , Insuficiencia Ovárica Primaria/genética , Prolactina/genética , Proteínas de Unión al ARN/genéticaAsunto(s)
Adaptación Fisiológica/fisiología , Plasticidad de la Célula/fisiología , Hipófisis/fisiología , Vertebrados/fisiología , Adaptación Fisiológica/genética , Animales , Plasticidad de la Célula/genética , Regulación de la Expresión Génica/fisiología , Humanos , Hipófisis/citología , Transducción de Señal/genéticaRESUMEN
SOX2 is a transcription factor associated with stem cell activity in several tissues. In cancer, SOX2 expression is increased in samples from several malignancies, including glioblastoma, and high SOX2 levels are associated with the population of tumor-initiating cells and with poor patient outcome. Therefore, understanding how SOX2 is regulated in cancer cells is relevant to tackle tumorigenesis. The SOX2 regulatory region 2(SRR2) is located downstream of the SOX2 coding region and mediates SOX2 expression in embryonic and adult stem cells. In this study, we deleted SRR2 using CRISPR/Cas9 in glioblastoma cells. Importantly, SRR2-deleted glioblastoma cells presented reduced SOX2 expression and decreased proliferative activity and self-renewal capacity in vitro. In line with these results, SRR2-deleted glioblastoma cells displayed decreased tumor initiation and growth in vivo. These effects correlated with an elevation of p21CIP1 cell cycle and p27KIP1 quiescence regulators. In conclusion, our data reveal that SRR2 deletion halts malignant activity of SOX2 and confirms that the SRR2 enhancer regulates SOX2 expression in cancer.
RESUMEN
P27, a cell cycle inhibitor, is also able to drive repression of Sox2 This interaction plays a crucial role during development of p27-/- pituitary tumors because loss of one copy of Sox2 impairs tumorigenesis [H. Li et al., Cell Stem Cell 11, 845-852 (2012)]. However, SOX2 is expressed in both endocrine and stem cells (SCs), and its contribution to tumorigenesis in either cell type is unknown. We have thus explored the cellular origin and mechanisms underlying endocrine tumorigenesis in p27-/- pituitaries. We found that pituitary hyperplasia is associated with reduced cellular differentiation, in parallel with increased levels of SOX2 in stem and endocrine cells. Using conditional loss-of-function and lineage tracing approaches, we show that SOX2 is required cell autonomously in p27-/- endocrine cells for these to give rise to tumors, and in SCs for promotion of tumorigenesis. This is supported by studies deleting the Sox2 regulatory region 2 (Srr2), the target of P27 repressive action. Single cell transcriptomic analysis further reveals that activation of a SOX2-dependent MAPK pathway in SCs is important for tumorigenesis. Altogether, our data highlight different aspects of the role of SOX2 following loss of p27, according to cellular context, and uncover an unexpected SOX2-dependent tumor-promoting role for SCs. Our results imply that targeting SCs, in addition to tumor cells, may represent an efficient antitumoral strategy in certain contexts.
Asunto(s)
Carcinogénesis/metabolismo , Neoplasias Hipofisarias/metabolismo , Factores de Transcripción SOXB1/metabolismo , Animales , Carcinogénesis/genética , Linaje de la Célula , Inhibidor p27 de las Quinasas Dependientes de la Ciclina/deficiencia , Inhibidor p27 de las Quinasas Dependientes de la Ciclina/genética , Células Endocrinas/metabolismo , Mutación con Pérdida de Función , Sistema de Señalización de MAP Quinasas , Ratones , Ratones Endogámicos C57BL , Células Madre Neoplásicas/metabolismo , Neoplasias Hipofisarias/genética , Neoplasias Hipofisarias/patología , Dominios Proteicos , Factores de Transcripción SOXB1/química , Factores de Transcripción SOXB1/genéticaRESUMEN
During embryonic development, radial glial cells give rise to neurons, then to astrocytes following the gliogenic switch. Timely regulation of the switch, operated by several transcription factors, is fundamental for allowing coordinated interactions between neurons and glia. We deleted the gene for one such factor, SOX9, early during mouse brain development and observed a significantly compromised dentate gyrus (DG). We dissected the origin of the defect, targeting embryonic Sox9 deletion to either the DG neuronal progenitor domain or the adjacent cortical hem (CH). We identified in the latter previously uncharacterized ALDH1L1+ astrocytic progenitors, which form a fimbrial-specific glial scaffold necessary for neuronal progenitor migration toward the developing DG. Our results highlight an early crucial role of SOX9 for DG development through regulation of astroglial potential acquisition in the CH. Moreover, we illustrate how formation of a local network, amidst astrocytic and neuronal progenitors originating from adjacent domains, underlays brain morphogenesis.
Asunto(s)
Astrocitos/metabolismo , Giro Dentado/crecimiento & desarrollo , Animales , Femenino , Eliminación de Gen , Ratones , Neurogénesis , Neuroglía/fisiologíaRESUMEN
In the last 15 years, single-cell technologies have become robust and indispensable tools to investigate cell heterogeneity. Beyond transcriptomic, genomic and epigenome analyses, technologies are constantly evolving, in particular toward multi-omics, where analyses of different source materials from a single cell are combined, and spatial transcriptomics, where resolution of cellular heterogeneity can be detected in situ. While some of these techniques are still being optimized, single-cell RNAseq has commonly been used because the examination of transcriptomes allows characterization of cell identity and, therefore, unravel previously uncharacterized diversity within cell populations. Most endocrine organs have now been investigated using this technique, and this has given new insights into organ embryonic development, characterization of rare cell types, and disease mechanisms. Here, we highlight recent studies, particularly on the hypothalamus and pituitary, and examine recent findings on the pancreas and reproductive organs where many single-cell experiments have been performed.
Asunto(s)
Sistema Endocrino/citología , Análisis de la Célula Individual , Animales , Humanos , Morfogénesis , Reproducción , Procesos de Determinación del Sexo , Transcriptoma/genéticaRESUMEN
Hypoparathyroidism is genetically heterogeneous and characterized by low plasma calcium and parathyroid hormone (PTH) concentrations. X-linked hypoparathyroidism (XLHPT) in two American families, is associated with interstitial deletion-insertions involving deletions of chromosome Xq27.1 downstream of SOX3 and insertions of predominantly non-coding DNA from chromosome 2p25.3. These could result in loss, gain, or movement of regulatory elements, which include ultraconserved element uc482, that could alter SOX3 expression,. To investigate this, we analysed SOX3 expression in EBV-transformed lymphoblastoid cells from 3 affected males, 3 unaffected males, and 4 carrier females from one XLHPT family. SOX3 expression was similar in all individuals, indicating that the spatiotemporal effect of the interstitial deletion-insertion on SOX3 expression postulated to occur in developing parathyroids did not manifest in lymphoblastoids. Expression of SNTG2, which is duplicated and inserted into the X chromosome, and ATP11C, which is moved telomerically, were also similarly expressed in all individuals. Investigation of male hemizygous (Sox3-/Y and uc482-/Y) and female heterozygous (Sox3+/- and uc482+/-) knock-out mice, together with wild-type littermates (male Sox3+/Y and uc482+/Y, and female Sox3+/+ and uc482+/+), revealed Sox3-/Y, Sox3+/-, uc482-/Y, and uc482+/- mice to have normal plasma biochemistry, compared to their respective wild-type littermates. When challenged with a low calcium diet, all mice had hypocalcaemia, and elevated plasma PTH concentrations and alkaline phosphatase activities, and Sox3-/Y, Sox3+/-, uc482-/Y, and uc482+/- mice had similar plasma biochemistry, compared to wild-type littermates. Thus, these results indicate that absence of Sox3 or uc482 does not cause hypoparathyroidism, and that XLHPT likely reflects a more complex mechanism.
RESUMEN
The pituitary is an essential endocrine gland regulating multiple processes. Regeneration of endocrine cells is of therapeutic interest and recent studies are promising, but mechanisms of endocrine cell fate acquisition need to be better characterised. The NOTCH pathway is important during pituitary development. Here, we further characterise its role in the murine pituitary, revealing differential sensitivity within and between lineages. In progenitors, NOTCH activation blocks cell fate acquisition, with time-dependant modulation. In differentiating cells, response to activation is blunted in the POU1F1 lineage, with apparently normal cell fate specification, while POMC cells remain sensitive. Absence of apparent defects in Pou1f1-Cre; Rbpjfl/fl mice further suggests no direct role for NOTCH signalling in POU1F1 cell fate acquisition. In contrast, in the POMC lineage, NICD expression induces a regression towards a progenitor-like state, suggesting that the NOTCH pathway specifically blocks POMC cell differentiation. These results have implications for pituitary development, plasticity and regeneration. Activation of NOTCH signalling in different cell lineages of the embryonic murine pituitary uncovers an unexpected differential sensitivity, and this consequently reveals new aspects of endocrine lineages development and plasticity.
Asunto(s)
Diferenciación Celular , Linaje de la Célula , Células Endocrinas/fisiología , Hipófisis/fisiología , Receptores Notch/metabolismo , Animales , Regulación de la Expresión Génica , Ratones , Transducción de SeñalRESUMEN
Along with the sub-ventricular zone of the forebrain lateral ventricles and the sub-granular zone of the dentate gyrus in the hippocampus, the hypothalamus has recently emerged as a third gliogenic and neurogenic niche in the central nervous system. The hypothalamus is the main regulator of body homeostasis because it centralizes peripheral information to regulate crucial physiological functions through the pituitary gland and the autonomic nervous system. Its ability to sense signals originating outside the brain relies on its exposure to blood-born molecules through the median eminence, which is localized outside the blood brain barrier. Within the hypothalamus, a population of specialized radial glial cells, the tanycytes, control exposure to blood-born signals by acting both as sensors and regulators of the hypothalamic input and output. In addition, lineage-tracing experiments have recently revealed that tanycytes represent a population of hypothalamic stem cells, defining them as a pivotal cell type within the hypothalamus. Hypothalamic neurogenesis has moreover been shown to have an important role in feeding control and energy metabolism, which challenges previous knowledge and offers new therapeutic options.
Asunto(s)
Células Ependimogliales/citología , Hipotálamo/fisiología , Eminencia Media/citología , Neurogénesis , Animales , Barrera Hematoencefálica , Metabolismo Energético , Conducta Alimentaria , Homeostasis , Humanos , Hipotálamo/citología , Células Madre/citologíaRESUMEN
Sox2 mutations are associated with pituitary hormone deficiencies and the protein is required for pituitary progenitor proliferation, but its function has not been well characterized in this context. SOX2 is known to activate expression of Six6, encoding a homeodomain transcription factor, in the ventral diencephalon. Here, we find that the same relationship likely exists in the pituitary. Moreover, because Six6 deletion is associated with a similar phenotype as described here for loss of Sox2, Six6 appears to be an essential downstream target of SOX2 in the gland. We also uncover a second role for SOX2. Whereas cell differentiation is reduced in Sox2 mutants, some endocrine cells are generated, such as POMC-positive cells in the intermediate lobe. However, loss of SOX2 here results in complete downregulation of the melanotroph pioneer factor PAX7, and subsequently a switch of identity from melanotrophs to ectopic corticotrophs. Rescuing proliferation by ablating the cell cycle negative regulator p27 (also known as Cdkn1b) in Sox2 mutants does not restore melanotroph emergence. Therefore, SOX2 has two independent roles during pituitary morphogenesis; firstly, promotion of progenitor proliferation, and subsequently, acquisition of melanotroph identity.
Asunto(s)
Linaje de la Célula , Hipófisis/citología , Hipófisis/embriología , Factores de Transcripción SOXB1/metabolismo , Células Madre/citología , Animales , Recuento de Células , Proliferación Celular , Corticotrofos/citología , Corticotrofos/metabolismo , Inhibidor p27 de las Quinasas Dependientes de la Ciclina/metabolismo , Regulación hacia Abajo/genética , Eliminación de Gen , Regulación del Desarrollo de la Expresión Génica , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/metabolismo , Melanotrofos/citología , Melanotrofos/metabolismo , Ratones Endogámicos C57BL , Modelos Biológicos , Morfogénesis/genética , Factor de Transcripción PAX7/metabolismo , Proopiomelanocortina/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Proteínas Represoras/genética , Proteínas Represoras/metabolismo , Células Madre/metabolismo , Transactivadores/genética , Transactivadores/metabolismoRESUMEN
Significant progress has been made recently in unravelling the embryonic events leading to pituitary morphogenesis, both in vivo and in vitro. This includes dissection of the molecular mechanisms controlling patterning of the ventral diencephalon that regulate formation of the pituitary anlagen or Rathke's pouch. There is also a better characterisation of processes that underlie maintenance of pituitary progenitors, specification of endocrine lineages and the three-dimensional organisation of newly differentiated endocrine cells. Furthermore, a population of adult pituitary stem cells (SCs), originating from embryonic progenitors, have been described and shown to have not only regenerative potential, but also the capacity to induce tumour formation. Finally, the successful recapitulation in vitro of embryonic events leading to generation of endocrine cells from embryonic SCs, and their subsequent transplantation, represents exciting advances towards the use of regenerative medicine to treat endocrine deficits. In this review, an up-to-date description of pituitary morphogenesis will be provided and discussed with particular reference to pituitary SC studies.
Asunto(s)
Regulación del Desarrollo de la Expresión Génica , Hipófisis/embriología , Hipófisis/metabolismo , Animales , Ratones , Modelos Biológicos , Morfogénesis/genética , Hipófisis/anatomía & histología , Hipófisis/citología , Células Madre/metabolismoRESUMEN
High expression of the B1 Sox genes, Sox2 and Sox3, is associated with the development of definitive neural primordia, the neural plates, in early stage embryos. However, in the caudal lateral epiblast (CLE) where axial stem cells reside, Sox2 and Sox3 are expressed at low levels, together with Brachyury. Because axial stem cells are the bipotential precursors of the neural plate and paraxial mesoderm, we investigated the possibility that low-level B1 Sox expression in CLE may regulate the fate of axial stem cells. We combined the genetic conditions of Sox3-null and Sox2 N1 enhancer homozygous deletion (Sox2(ΔN1/ΔN1)) to decrease B1 Sox expression in CLE. At 5-7 somite stages of mouse embryogenesis, these genetic manipulations caused approximately 30% higher production of paraxial mesodermal precursors, resulting in the development of larger somites. Analysis of mitotic cell populations suggested that decrease of B1 Sox expression in CLE does not activate cell proliferation but promotes cell migration into the mesodermal compartment. Thus, the low-level B1 Sox expression in CLE regulates axial stem cells to adjust the production of paraxial mesoderm precursors to an appropriate level.
Asunto(s)
Regulación del Desarrollo de la Expresión Génica/genética , Estratos Germinativos/metabolismo , Mesodermo/metabolismo , Placa Neural/metabolismo , Factores de Transcripción SOXB1/genética , Animales , Desarrollo Embrionario/genética , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos DBA , Factores de Transcripción SOXB1/metabolismo , Somitos/metabolismo , Células Madre/metabolismoRESUMEN
Pituitary hormone deficiencies, with Growth Hormone deficiency being most frequent (1 in 3,500-10,000 births), cause significant morbidity. Regeneration of missing endocrine cells would be a significant improvement over hormone replacement therapies, which incur side effects and do not mimic physiological secretion patterns. Recent in vitro studies have identified a population of adult pituitary progenitors that express the HMG box transcription factors SOX2 and SOX9. Here, we apply cell-lineage tracing analysis to demonstrate that SOX2- and SOX9-expressing progenitors can self-renew and give rise to endocrine cells in vivo, suggesting that they are tissue stem cells. Moreover, we show that they can become mobilized and differentiate into the appropriate endocrine cell types in response to physiological stress. Our results highlight the pituitary as a model for exploring how physiological changes influence stem cell behavior and suggest that manipulation of endogenous pituitary stem cells is a potential therapeutic strategy for pituitary deficiencies.
Asunto(s)
Células Madre Adultas/citología , Células Endocrinas/citología , Hipófisis/citología , Hipófisis/fisiología , Regeneración , Células Madre Adultas/metabolismo , Animales , Células Endocrinas/metabolismo , Ratones , Ratones Transgénicos , Factor de Transcripción SOX9/metabolismo , Factores de Transcripción SOXB1/metabolismoRESUMEN
The SRY-related HMG box transcription factor Sox2 plays critical roles throughout embryogenesis. Haploinsufficiency for SOX2 results in human developmental defects including anophthalmia, microphthalmia and septo-optic dysplasia, a congenital forebrain defect. To understand how Sox2 plays a role in neurogenesis, we combined genomic and in vivo transgenic approaches to characterize genomic regions occupied by Sox2 in the developing forebrain. Six3, a homeobox gene associated with holoprosencephaly, a forebrain midline defect, was identified as a Sox2 transcriptional target. This study shows that Sox2 directly regulates a previously unidentified long-range forebrain enhancer to activate Six3 expression in the rostral diencephalon. Further biochemical and genetic evidences indicated a direct regulatory link between Sox2 and Six3 during forebrain development, providing a better understanding of a common molecular mechanism underlying these forebrain defects.
Asunto(s)
Proteínas del Ojo/metabolismo , Regulación del Desarrollo de la Expresión Génica , Proteínas de Homeodominio/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Prosencéfalo/metabolismo , Factores de Transcripción SOXB1/genética , Animales , Secuencia de Bases , Sitios de Unión , Secuencia Conservada , Embrión de Mamíferos/metabolismo , Desarrollo Embrionario , Elementos de Facilitación Genéticos , Evolución Molecular , Proteínas del Ojo/genética , Proteínas Hedgehog/genética , Proteínas Hedgehog/metabolismo , Proteínas de Homeodominio/genética , Ratones , Ratones Transgénicos , Proteínas del Tejido Nervioso/genética , Placa Neural/embriología , Placa Neural/metabolismo , Prosencéfalo/citología , Prosencéfalo/embriología , Unión Proteica , Factores de Transcripción SOXB1/metabolismo , Activación Transcripcional , Proteína Homeobox SIX3RESUMEN
The mechanisms responsible for the transcriptional silencing of pluripotency genes in differentiated cells are poorly understood. We have observed that cells lacking the tumor suppressor p27 can be reprogrammed into induced pluripotent stem cells (iPSCs) in the absence of ectopic Sox2. Interestingly, cells and tissues from p27 null mice, including brain, lung, and retina, present an elevated basal expression of Sox2, suggesting that p27 contributes to the repression of Sox2. Furthermore, p27 null iPSCs fail to fully repress Sox2 upon differentiation. Mechanistically, we have found that upon differentiation p27 associates to the SRR2 enhancer of the Sox2 gene together with a p130-E2F4-SIN3A repressive complex. Finally, Sox2 haploinsufficiency genetically rescues some of the phenotypes characteristic of p27 null mice, including gigantism, pituitary hyperplasia, pituitary tumors, and retinal defects. Collectively, these results demonstrate an unprecedented connection between p27 and Sox2 relevant for reprogramming and cancer and for understanding human pathologies associated with p27 germline mutations.