RESUMEN
Adrenal cortex steroids are essential for body homeostasis, and adrenal insufficiency is a life-threatening condition. Adrenal endocrine activity is maintained through recruitment of subcapsular progenitor cells that follow a unidirectional differentiation path from zona glomerulosa to zona fasciculata (zF). Here, we show that this unidirectionality is ensured by the histone methyltransferase EZH2. Indeed, we demonstrate that EZH2 maintains adrenal steroidogenic cell differentiation by preventing expression of GATA4 and WT1 that cause abnormal dedifferentiation to a progenitor-like state in Ezh2 KO adrenals. EZH2 further ensures normal cortical differentiation by programming cells for optimal response to adrenocorticotrophic hormone (ACTH)/PKA signaling. This is achieved by repression of phosphodiesterases PDE1B, 3A, and 7A and of PRKAR1B. Consequently, EZH2 ablation results in blunted zF differentiation and primary glucocorticoid insufficiency. These data demonstrate an all-encompassing role for EZH2 in programming steroidogenic cells for optimal response to differentiation signals and in maintaining their differentiated state.
Asunto(s)
Corteza Suprarrenal/enzimología , Subunidad RIbeta de la Proteína Quinasa Dependiente de AMP Cíclico/metabolismo , Proteína Potenciadora del Homólogo Zeste 2/metabolismo , Transducción de Señal , Corteza Suprarrenal/metabolismo , Animales , Diferenciación Celular , Subunidad RIbeta de la Proteína Quinasa Dependiente de AMP Cíclico/genética , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 1/genética , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 1/metabolismo , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 3/genética , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 3/metabolismo , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 7/genética , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 7/metabolismo , Proteína Potenciadora del Homólogo Zeste 2/genética , Femenino , Masculino , Ratones Endogámicos C57BL , Ratones Noqueados , Esteroides/metabolismo , Zona Fascicular/citología , Zona Fascicular/enzimología , Zona Fascicular/metabolismo , Zona Glomerular/citología , Zona Glomerular/enzimología , Zona Glomerular/metabolismoRESUMEN
SUMOylation is a highly conserved and dynamic post-translational mechanism primarily affecting nuclear programs for adapting organisms to stressful challenges. Alteration of SUMOylation cycles leads to severe developmental and homeostatic defects and malignancy, but signals coordinating SUMOylation are still unidentified. The adrenal cortex is a zonated endocrine gland that controls body homeostasis and stress response. Here, we show that in human and in mouse adrenals, SUMOylation follows a decreasing centripetal gradient that mirrors cortical differentiation flow and delimits highly and weakly SUMOylated steroidogenic compartments, overlapping glomerulosa, and fasciculata zones. Activation of PKA signaling by acute hormonal treatment, mouse genetic engineering, or in Carney complex results in repression of small ubiquitin-like modifier (SUMO) conjugation in the inner cortex by coordinating expression of SUMO pathway inducers and repressors. Conversely, genetic activation of canonical wingless-related integration site signaling maintains high SUMOylation potential in the outer neoplastic cortex. Thus, SUMOylation is tightly regulated by signaling pathways that orchestrate adrenal zonation and diseases.-Dumontet, T., Sahut-Barnola, I., Dufour, D., Lefrançois-Martinez, A.-M., Berthon, A., Montanier, N., Ragazzon, B., Djari, C., Pointud, J.-C., Roucher-Boulez, F., Batisse-Lignier, M., Tauveron, I., Bertherat, J., Val, P., Martinez, A. Hormonal and spatial control of SUMOylation in the human and mouse adrenal cortex.
Asunto(s)
Corteza Suprarrenal/metabolismo , Hormona Adrenocorticotrópica/farmacología , Procesamiento Proteico-Postraduccional/fisiología , Sumoilación/fisiología , Corteza Suprarrenal/efectos de los fármacos , Corteza Suprarrenal/ultraestructura , Neoplasias de la Corteza Suprarrenal/patología , Hormona Adrenocorticotrópica/administración & dosificación , Animales , Complejo de Carney/metabolismo , Línea Celular Tumoral , Colforsina/farmacología , Proteínas Quinasas Dependientes de AMP Cíclico/fisiología , Cicloheximida/farmacología , Dactinomicina/farmacología , Preparaciones de Acción Retardada , Dexametasona/análogos & derivados , Dexametasona/farmacología , Femenino , Humanos , Ratones , Ratones Noqueados , Ratones Transgénicos , Proteínas de Neoplasias/metabolismo , Procesamiento Proteico-Postraduccional/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Sumoilación/efectos de los fármacos , Vía de Señalización Wnt/efectos de los fármacos , Vía de Señalización Wnt/fisiología , Zona Fascicular/efectos de los fármacos , Zona Fascicular/metabolismo , Zona Glomerular/efectos de los fármacos , Zona Glomerular/metabolismo , beta Catenina/deficiencia , beta Catenina/genéticaRESUMEN
BACKGROUND: EZH2 is overexpressed and associated with poor prognosis in adrenocortical carcinoma (ACC) and its inhibition reduces growth and aggressiveness of ACC cells in culture. Although EZH2 was identified as the methyltransferase that deposits the repressive H3K27me3 histone mark, it can cooperate with transcription factors to stimulate gene transcription. METHODS: We used bioinformatics approaches on gene expression data from three cohorts of patients and a mouse model of EZH2 ablation, to identify targets and mode of action of EZH2 in ACC. This was followed by ChIP and functional assays to evaluate contribution of identified targets to ACC pathogenesis. RESULTS: We show that EZH2 mostly works as a transcriptional inducer in ACC, through cooperation with the transcription factor E2F1 and identify three positive targets involved in cell cycle regulation and mitosis i.e., RRM2, PTTG1 and ASE1/PRC1. Overexpression of these genes is associated with poor prognosis, suggesting a potential role in acquisition of aggressive ACC features. Pharmacological and siRNA-mediated inhibition of RRM2 blocks cell proliferation, induces apoptosis and inhibits cell migration, suggesting that it may be an interesting target in ACC. CONCLUSIONS: Altogether, these data show an unexpected role of EZH2 and E2F1 in stimulating expression of genes associated with ACC aggressiveness.
Asunto(s)
Neoplasias de la Corteza Suprarrenal/genética , Carcinoma Corticosuprarrenal/genética , Factor de Transcripción E2F1/genética , Proteína Potenciadora del Homólogo Zeste 2/genética , Regulación Neoplásica de la Expresión Génica , Adenosina/análogos & derivados , Adenosina/farmacología , Animales , Proteínas de Ciclo Celular/genética , Proliferación Celular/efectos de los fármacos , Proliferación Celular/genética , Inmunoprecipitación de Cromatina , Biología Computacional , Proteína Potenciadora del Homólogo Zeste 2/antagonistas & inhibidores , Humanos , Indoles/farmacología , Ratones Noqueados , Análisis Multivariante , Modelos de Riesgos Proporcionales , Ribonucleósido Difosfato Reductasa/antagonistas & inhibidores , Ribonucleósido Difosfato Reductasa/genética , Securina/genéticaRESUMEN
Adrenal Cortex Carcinoma (ACC) is an aggressive tumour with poor prognosis. Common alterations in patients include constitutive WNT/ß-catenin signalling and overexpression of the growth factor IGF2. However, the combination of both alterations in transgenic mice is not sufficient to trigger malignant tumour progression, suggesting that other alterations are required to allow development of carcinomas. Here, we have conducted a study of publicly available gene expression data from three cohorts of ACC patients to identify relevant alterations. Our data show that the histone methyltransferase EZH2 is overexpressed in ACC in the three cohorts. This overexpression is the result of deregulated P53/RB/E2F pathway activity and is associated with increased proliferation and poorer prognosis in patients. Inhibition of EZH2 by RNA interference or pharmacological treatment with DZNep inhibits cellular growth, wound healing and clonogenic growth and induces apoptosis of H295R cells in culture. Further growth inhibition is obtained when DZNep is combined with mitotane, the gold-standard treatment for ACC. Altogether, these observations suggest that overexpression of EZH2 is associated with aggressive progression and may constitute an interesting therapeutic target in the context of ACC.
Asunto(s)
Neoplasias de la Corteza Suprarrenal/genética , Proteína Potenciadora del Homólogo Zeste 2/genética , Proteína Potenciadora del Homólogo Zeste 2/metabolismo , Neoplasias de la Corteza Suprarrenal/metabolismo , Animales , Línea Celular Tumoral , Proliferación Celular , Bases de Datos de Ácidos Nucleicos , Progresión de la Enfermedad , Expresión Génica , Predisposición Genética a la Enfermedad/genética , Humanos , Factor II del Crecimiento Similar a la Insulina/genética , Factor II del Crecimiento Similar a la Insulina/metabolismo , Ratones , Ratones Transgénicos , Interferencia de ARN , Factores de Riesgo , Vía de Señalización Wnt , beta Catenina/genéticaRESUMEN
Primary pigmented nodular adrenocortical disease (PPNAD) is associated with inactivating mutations of the PRKAR1A tumor suppressor gene that encodes the regulatory subunit R1α of the cAMP-dependent protein kinase (PKA). In human and mouse adrenocortical cells, these mutations lead to increased PKA activity, which results in increased resistance to apoptosis that contributes to the tumorigenic process. We used in vitro and in vivo models to investigate the possibility of a crosstalk between PKA and mammalian target of rapamycin (mTOR) pathways in adrenocortical cells and its possible involvement in apoptosis resistance. Impact of PKA signaling on activation of the mTOR pathway and apoptosis was measured in a mouse model of PPNAD (AdKO mice), in human and mouse adrenocortical cell lines in response to pharmacological inhibitors and in PPNAD tissues by immunohistochemistry. AdKO mice showed increased mTOR complex 1 (mTORC1) pathway activity. Inhibition of mTORC1 by rapamycin restored sensitivity of adrenocortical cells to apoptosis in AdKO but not in wild-type mice. In both cell lines and mouse adrenals, rapid phosphorylation of mTORC1 targets including BAD proapoptotic protein was observed in response to PKA activation. Accordingly, BAD hyperphosphorylation, which inhibits its proapoptotic activity, was increased in both AdKO mouse adrenals and human PPNAD tissues. In conclusion, mTORC1 pathway is activated by PKA signaling in human and mouse adrenocortical cells, leading to increased cell survival, which is correlated with BAD hyperphosphorylation. These alterations could be causative of tumor formation.
Asunto(s)
Enfermedades de la Corteza Suprarrenal/metabolismo , Enfermedades de la Corteza Suprarrenal/patología , Subunidad RIalfa de la Proteína Quinasa Dependiente de AMP Cíclico/genética , Proteína Letal Asociada a bcl/metabolismo , Enfermedades de la Corteza Suprarrenal/genética , Hormona Adrenocorticotrópica/administración & dosificación , Hormona Adrenocorticotrópica/farmacología , Animales , Apoptosis/efectos de los fármacos , Línea Celular Tumoral , Subunidad RIalfa de la Proteína Quinasa Dependiente de AMP Cíclico/metabolismo , Modelos Animales de Enfermedad , Técnicas de Inactivación de Genes , Humanos , Masculino , Diana Mecanicista del Complejo 1 de la Rapamicina , Ratones , Complejos Multiproteicos/metabolismo , Fosforilación , Transducción de Señal/efectos de los fármacos , Sirolimus/farmacología , Serina-Treonina Quinasas TOR/metabolismoRESUMEN
Primary aldosteronism (PA) is the main cause of secondary hypertension, resulting from adrenal aldosterone-producing adenomas (APA) or bilateral hyperplasia. Here, we show that constitutive activation of WNT/ß-catenin signalling is the most frequent molecular alteration found in 70% of APA. We provide evidence that decreased expression of the WNT inhibitor SFRP2 may be contributing to deregulated WNT signalling and APA development in patients. This is supported by the demonstration that mice with genetic ablation of Sfrp2 have increased aldosterone production and ectopic differentiation of zona glomerulosa cells. We further show that ß-catenin plays an essential role in the control of basal and Angiotensin II-induced aldosterone secretion, by activating AT1R, CYP21 and CYP11B2 transcription. This relies on both LEF/TCF-dependent activation of AT1R and CYP21 regulatory regions and indirect activation of CYP21 and CYP11B2 promoters, through increased expression of the nuclear receptors NURR1 and NUR77. Altogether, these data show that aberrant WNT/ß-catenin activation is associated with APA development and suggest that WNT pathway may be a good therapeutic target in PA.
Asunto(s)
Neoplasias de la Corteza Suprarrenal/metabolismo , Adenoma Corticosuprarrenal/metabolismo , Aldosterona/biosíntesis , Hiperaldosteronismo/metabolismo , Vía de Señalización Wnt , Neoplasias de la Corteza Suprarrenal/complicaciones , Adenoma Corticosuprarrenal/complicaciones , Adulto , Aldosterona/sangre , Aldosterona/metabolismo , Animales , Línea Celular Tumoral , Citocromo P-450 CYP11B2/genética , Citocromo P-450 CYP11B2/metabolismo , Regulación hacia Abajo , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Hiperaldosteronismo/etiología , Masculino , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Ratones , Ratones de la Cepa 129 , Ratones Endogámicos C57BL , Ratones Noqueados , Persona de Mediana Edad , Miembro 1 del Grupo A de la Subfamilia 4 de Receptores Nucleares/genética , Miembro 1 del Grupo A de la Subfamilia 4 de Receptores Nucleares/metabolismo , Miembro 2 del Grupo A de la Subfamilia 4 de Receptores Nucleares/genética , Miembro 2 del Grupo A de la Subfamilia 4 de Receptores Nucleares/metabolismoRESUMEN
Carney complex (CNC) is an inherited neoplasia syndrome with endocrine overactivity. Its most frequent endocrine manifestation is primary pigmented nodular adrenocortical disease (PPNAD), a bilateral adrenocortical hyperplasia causing pituitary-independent Cushing's syndrome. Inactivating mutations in PRKAR1A, a gene encoding the type 1 alpha-regulatory subunit (R1alpha) of the cAMP-dependent protein kinase (PKA) have been found in 80% of CNC patients with Cushing's syndrome. To demonstrate the implication of R1alpha loss in the initiation and development of PPNAD, we generated mice lacking Prkar1a specifically in the adrenal cortex (AdKO). AdKO mice develop pituitary-independent Cushing's syndrome with increased PKA activity. This leads to autonomous steroidogenic genes expression and deregulated adreno-cortical cells differentiation, increased proliferation and resistance to apoptosis. Unexpectedly, R1alpha loss results in improper maintenance and centrifugal expansion of cortisol-producing fetal adrenocortical cells with concomitant regression of adult cortex. Our data provide the first in vivo evidence that loss of R1alpha is sufficient to induce autonomous adrenal hyper-activity and bilateral hyperplasia, both observed in human PPNAD. Furthermore, this model demonstrates that deregulated PKA activity favors the emergence of a new cell population potentially arising from the fetal adrenal, giving new insight into the mechanisms leading to PPNAD.
Asunto(s)
Corteza Suprarrenal/metabolismo , Síndrome de Cushing/metabolismo , Subunidad RIalfa de la Proteína Quinasa Dependiente de AMP Cíclico/metabolismo , Hormona Adrenocorticotrópica/metabolismo , Animales , Proliferación Celular , Síndrome de Cushing/embriología , Síndrome de Cushing/genética , Síndrome de Cushing/patología , Subunidad RIalfa de la Proteína Quinasa Dependiente de AMP Cíclico/deficiencia , Proteínas Quinasas Dependientes de AMP Cíclico/metabolismo , Femenino , Humanos , Masculino , Ratones , Ratones Noqueados , Transducción de Señal , Esteroide 17-alfa-Hidroxilasa/metabolismoRESUMEN
In the adrenal gland, adrenocorticotropin (ACTH) acting through the cAMP protein kinase (PKA) transduction pathway is the main regulator of genes involved in glucocorticoid synthesis. The prolactin (PRL) receptor is expressed in the adrenal cortex of most mammals, but experimental proof that PRL ensures direct control on glucocorticoid synthesis in rodents remains elusive. To unravel the physiological importance of PRL in adrenocortical functions, we measured steroidogenic capacity of Prlr-deficient mice (Prlr(-/-)) and explored the influence of JAK/STAT signaling, the major PRL transduction pathway, on the steroidogenic activity of adrenocortical cell cultures. We demonstrate that lack of Prlr does not affect basal (nor stress-induced) corticosterone levels in mice. PRL triggers JAK2/STAT5-dependent transcription in adrenal cells, but this does not influence corticosterone release. In contrast, pharmacological or siRNA-mediated inhibition of JAK2 reveals its essential role in both basal and ACTH/cAMP-induced steroidogenesis. We demonstrate that nuclear JAK2 regulates the amount of active transcription factor CREB (cAMP response element-binding protein) through tyrosine phosphorylation and prevention of proteasomal degradation, which in turn leads to transcriptional activation of the rate-limiting steroidogenic Star gene. Hence, we describe a novel link between PKA and JAK2 by which nuclear JAK2 signaling controls adrenal steroidogenesis by increasing the stability of CREB.
Asunto(s)
Glándulas Suprarrenales/metabolismo , Proteína de Unión a Elemento de Respuesta al AMP Cíclico/metabolismo , Proteínas Quinasas Dependientes de AMP Cíclico/metabolismo , Janus Quinasa 2/metabolismo , Esteroides/biosíntesis , Transcripción Genética , Glándulas Suprarrenales/efectos de los fármacos , Animales , Núcleo Celular/efectos de los fármacos , Núcleo Celular/enzimología , Corticosterona/biosíntesis , AMP Cíclico/farmacología , Humanos , Masculino , Ratones , Complejos Multiproteicos/metabolismo , Fosforilación/efectos de los fármacos , Prolactina/farmacología , Unión Proteica/efectos de los fármacos , Estabilidad Proteica/efectos de los fármacos , Ratas , Ovinos , Transcripción Genética/efectos de los fármacosRESUMEN
Adrenocortical carcinoma is a rare but aggressive cancer with unknown aetiology. Constitutive activation of beta-catenin is the most frequent alteration in benign and malignant adrenocortical tumours in patients. Here, we show that constitutive activation of beta-catenin in the adrenal cortex of transgenic mice resulted in progressive steroidogenic and undifferentiated spindle-shaped cells hyperplasia as well as dysplasia of the cortex and medulla. Over a 17 months time course, transgenic adrenals developed malignant characteristics such as uncontrolled neovascularization and loco-regional metastatic invasion. These oncogenic events were accompanied by ectopic differentiation of glomerulosa at the expense of fasciculata cells, which caused primary hyperaldosteronism. Altogether these observations demonstrate that constitutively active beta-catenin is an adrenal oncogene which triggers benign aldosterone-secreting tumour development and promotes malignancy.
Asunto(s)
Corteza Suprarrenal/patología , Neoplasias de las Glándulas Suprarrenales/metabolismo , Neoplasias de las Glándulas Suprarrenales/patología , beta Catenina/metabolismo , Corteza Suprarrenal/metabolismo , Neoplasias de las Glándulas Suprarrenales/fisiopatología , Aldosterona/metabolismo , Animales , Proliferación Celular , Modelos Animales de Enfermedad , Humanos , Hiperplasia , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Metástasis de la NeoplasiaRESUMEN
Excess glucocorticoid exposure affects emotional and cognitive brain functions. The extreme form, Cushing's syndrome, is adequately modelled in the AdKO2.0 mouse, consequential to adrenocortical hypertrophy and hypercorticosteronemia. We previously reported that the AdKO2.0 mouse brain undergoes volumetric changes that resemble closely those of Cushing's syndrome human patients, as well as changes in expression of glial related marker proteins. In the present work, the expression of genes related to glial and neuronal cell populations and functions was assessed in regions of the anterior brain, hippocampus, amygdala and hypothalamus. Glucocorticoid target genes were consistently regulated, including CRH mRNA suppression in the hypothalamus and induction in amygdala and hippocampus, even if glucocorticoid receptor protein was downregulated. Expression of glial genes was also affected in the AdKO2.0 mouse brain, indicating a different activation status in glial cells. Generic markers for neuronal cell populations, and cellular integrity were only slightly affected. Our findings highlight the vulnerability of glial cell populations to chronic high levels of circulating glucocorticoids.
Asunto(s)
Síndrome de Cushing , Animales , Encéfalo/metabolismo , Síndrome de Cushing/genética , Síndrome de Cushing/metabolismo , Expresión Génica , Glucocorticoides/metabolismo , Humanos , Ratones , Receptores de Glucocorticoides/genética , Receptores de Glucocorticoides/metabolismoRESUMEN
Carney complex is a rare familial multineoplastic syndrome predisposing to endocrine and nonendocrine tumors due to inactivating mutations of PRKAR1A, leading to perturbations of the cAMPâprotein kinase A signaling pathway. Skin lesions are the most common manifestation of Carney complex, including lentigines, blue nevi, and cutaneous myxomas in unusual locations such as oral and genital mucosa. Unlike endocrine disorders, the pathogenesis of skin lesions remains unexplained. In this study, we show that embryonic invalidation of the Prkar1a gene in steroidogenic factor-1âexpressing cells leads to the development of familial skin pigmentation alterations, reminiscent of those in patients with Carney complex. Immunohistological and molecular analyses, coupled with genetic monitoring of recombinant cell lineages in mouse skin, suggest that familial lentiginosis and myxomas occur in skin areas specifically enriched in dermal melanocytes. In lentigines- and blue neviâprone areas from mutant mice and patients, Prkar1a/PRKAR1A invalidation occurs in a subset of dermal fibroblasts capable of inducing, under the influence of protein kinase A signaling, the production of promelanogenic EDN3 and hepatocyte GF signals. Our model strongly suggests that the origin of the typical Carney complex cutaneous lesions is the result of noncell-autonomous promelanogenic activity of a dermal fibroblast population sharing a community of origin with steroidogenic factor-1 lineage.
Asunto(s)
Complejo de Carney , Lentigo , Mixoma , Nevo Azul , Enfermedades de la Piel , Animales , Ratones , Complejo de Carney/genética , Complejo de Carney/patología , Subunidad RIalfa de la Proteína Quinasa Dependiente de AMP Cíclico/genética , Mixoma/genética , Mixoma/patología , Síndrome , Lentigo/patologíaRESUMEN
Unlike most cancers, adrenocortical carcinomas (ACCs) are more frequent in women than in men, but the underlying mechanisms of this sexual dimorphism remain elusive. Here, we show that inactivation of Znrf3 in the mouse adrenal cortex, recapitulating the most frequent alteration in ACC patients, is associated with sexually dimorphic tumor progression. Although female knockouts develop metastatic carcinomas at 18 months, adrenal hyperplasia regresses in male knockouts. This male-specific phenotype is associated with androgen-dependent induction of senescence, recruitment, and differentiation of highly phagocytic macrophages that clear out senescent cells. In contrast, in females, macrophage recruitment is delayed and dampened, which allows for aggressive tumor progression. Consistently, analysis of TCGA-ACC data shows that phagocytic macrophages are more prominent in men and are associated with better prognosis. Together, these data show that phagocytic macrophages are key players in the sexual dimorphism of ACC that could be previously unidentified allies in the fight against this devastating cancer.
Asunto(s)
Neoplasias de la Corteza Suprarrenal , Carcinoma Corticosuprarrenal , Neoplasias de la Corteza Suprarrenal/genética , Neoplasias de la Corteza Suprarrenal/patología , Carcinoma Corticosuprarrenal/genética , Carcinoma Corticosuprarrenal/patología , Andrógenos , Animales , Femenino , Masculino , Ratones , PronósticoRESUMEN
SUMOylation is a dynamic posttranslational modification, that provides fine-tuning of protein function involved in the cellular response to stress, differentiation, and tissue development. In the adrenal cortex, an emblematic endocrine organ that mediates adaptation to physiological demands, the SUMOylation gradient is inversely correlated with the gradient of cellular differentiation raising important questions about its role in functional zonation and the response to stress. Considering that SUMO-specific protease 2 (SENP2), a deSUMOylating enzyme, is upregulated by Adrenocorticotropic Hormone (ACTH)/cAMP-dependent Protein Kinase (PKA) signalling within the zona fasciculata, we generated mice with adrenal-specific Senp2 loss to address these questions. Disruption of SENP2 activity in steroidogenic cells leads to specific hypoplasia of the zona fasciculata, a blunted reponse to ACTH and isolated glucocorticoid deficiency. Mechanistically, overSUMOylation resulting from SENP2 loss shifts the balance between ACTH/PKA and WNT/ß-catenin signalling leading to repression of PKA activity and ectopic activation of ß-catenin. At the cellular level, this blocks transdifferentiation of ß-catenin-positive zona glomerulosa cells into fasciculata cells and sensitises them to premature apoptosis. Our findings indicate that the SUMO pathway is critical for adrenal homeostasis and stress responsiveness.
Asunto(s)
Transdiferenciación Celular , Cisteína Endopeptidasas , Glucocorticoides , Animales , Ratones , Corteza Suprarrenal/metabolismo , Corticoesteroides/metabolismo , Hormona Adrenocorticotrópica/metabolismo , beta Catenina/metabolismo , Transdiferenciación Celular/genética , Cisteína Endopeptidasas/genética , Cisteína Endopeptidasas/metabolismo , Glucocorticoides/metabolismo , Vía de Señalización WntRESUMEN
Chronic exposure to high circulating levels of glucocorticoids has detrimental effects on health, including metabolic abnormalities, as exemplified in Cushing's syndrome (CS). Magnetic resonance imaging (MRI) studies have found volumetric changes in gray and white matter of the brain in CS patients during the course of active disease, but also in remission. In order to explore this further, we performed MRI-based brain volumetric analyses in the AdKO mouse model for CS, which presents its key traits. AdKO mice had reduced relative volumes in several brain regions, including the corpus callosum and cortical areas. The medial amygdala, bed nucleus of the stria terminalis, and hypothalamus were increased in relative volume. Furthermore, we found a lower immunoreactivity of myelin basic protein (MBP, an oligodendrocyte marker) in several brain regions but a paradoxically increased MBP signal in the male cingulate cortex. We also observed a decrease in the expression of glial fibrillary acidic protein (GFAP, a marker for reactive astrocytes) and ionized calcium-binding adapter molecule 1 (IBA1, a marker for activated microglia) in the cingulate regions of the anterior corpus callosum and the hippocampus. We conclude that long-term hypercorticosteronemia induced brain region-specific changes that might include aberrant myelination and a degree of white matter damage, as both repair (GFAP) and immune (IBA1) responses are decreased. These findings suggest a cause for the changes observed in the brains of human patients and serve as a background for further exploration of their subcellular and molecular mechanisms.
RESUMEN
Large-cell calcifying Sertoli cell tumors (LCCSCTs) are among the most frequent lesions occurring in male Carney complex (CNC) patients. Although they constitute a key diagnostic criterion for this rare multiple neoplasia syndrome resulting from inactivating mutations of the tumor suppressor PRKAR1A, leading to unrepressed PKA activity, LCCSCT pathogenesis and origin remain elusive. Mouse models targeting Prkar1a inactivation in all somatic populations or separately in each cell type were generated to decipher the molecular and paracrine networks involved in the induction of CNC testis lesions. We demonstrate that the Prkar1a mutation was required in both stromal and Sertoli cells for the occurrence of LCCSCTs. Integrative analyses comparing transcriptomic, immunohistological data and phenotype of mutant mouse combinations led to the understanding of human LCCSCT pathogenesis and demonstrated PKA-induced paracrine molecular circuits in which the aberrant WNT4 signal production is a limiting step in shaping intratubular lesions and tumor expansion both in a mouse model and in human CNC testes.
Asunto(s)
Complejo de Carney/metabolismo , Proteínas Quinasas Dependientes de AMP Cíclico/metabolismo , Células de Sertoli/citología , Neoplasias Testiculares/metabolismo , Proteína Wnt4/metabolismo , Animales , Apoptosis , Complejo de Carney/genética , Subunidad RIalfa de la Proteína Quinasa Dependiente de AMP Cíclico/metabolismo , Modelos Animales de Enfermedad , Perfilación de la Expresión Génica , Genes Supresores de Tumor , Humanos , Masculino , Ratones , Ratones Noqueados , Mutación , Análisis de Secuencia por Matrices de Oligonucleótidos , Comunicación Paracrina , Fenotipo , Pigmentación , Túbulos Seminíferos/metabolismo , Testículo/metabolismo , TranscriptomaRESUMEN
The Cre-loxP system combined with gene targeting strategies has proven to be very useful for gene inactivation in specific tissues and/or cell types. To achieve adrenal cortex specific recombination in vivo, we used a 0.5-kb fragment of the 5'-flanking region of the akr1b7 gene to drive Cre expression in adrenocortical cells. The resulting 0.5 akr1b7-Cre mice express Cre in all steroidogenic zones of the adrenal cortex but not in the gonads. Although recombination of the ROSA26R reporter locus was not observed in all cortical cells, we provide evidence that Cre is expressed in all the cells of the cortex in adult mice. In addition, Cre activity was found in collecting ducts and maturing glomeruli of the kidney. This line is the first to show specific Cre expression in the adrenal cortex in the absence of Cre expression in the gonads. This transgene thus provides a valuable tool for specific gene recombination in the adrenal cortex and kidney.
Asunto(s)
Corteza Suprarrenal/enzimología , Integrasas/metabolismo , Ratones Transgénicos , Corteza Suprarrenal/citología , Corteza Suprarrenal/embriología , Animales , Femenino , Gónadas/citología , Gónadas/enzimología , Integrasas/genética , Riñón/citología , Riñón/enzimología , Masculino , RatonesRESUMEN
The adrenal cortex undergoes remodeling during fetal and postnatal life. How zona reticularis emerges in the postnatal gland to support adrenarche, a process whereby higher primates increase prepubertal androgen secretion, is unknown. Using cell-fate mapping and gene deletion studies in mice, we show that activation of PKA has no effect on the fetal cortex, while it accelerates regeneration of the adult cortex, triggers zona fasciculata differentiation that is subsequently converted into a functional reticularis-like zone, and drives hypersecretion syndromes. Remarkably, PKA effects are influenced by sex. Indeed, testicular androgens increase WNT signaling that antagonizes PKA, leading to slower adrenocortical cell turnover and delayed phenotype whereas gonadectomy sensitizes males to hypercorticism and reticularis-like formation. Thus, reticularis results from ultimate centripetal conversion of adult cortex under the combined effects of PKA and cell turnover that dictate organ size. We show that PKA-induced progenitor recruitment is sexually dimorphic and may provide a paradigm for overrepresentation of women in adrenal diseases.
Asunto(s)
Subunidad RIalfa de la Proteína Quinasa Dependiente de AMP Cíclico/metabolismo , Caracteres Sexuales , Transducción de Señal/fisiología , Zona Fascicular/metabolismo , Zona Reticular/metabolismo , Adrenarquia/metabolismo , Factores de Edad , Andrógenos/metabolismo , Animales , Diferenciación Celular/fisiología , Subunidad RIalfa de la Proteína Quinasa Dependiente de AMP Cíclico/genética , Embrión de Mamíferos , Femenino , Masculino , Ratones , Ratones Noqueados , Modelos AnimalesRESUMEN
The adrenocortical gland undergoes structural and functional remodelling in the fetal and postnatal periods. After birth, the fetal zone of the gland undergoes rapid involution in favor of the definitive cortex, which reaches maturity with the emergence of the zona reticularis(zR) at the adrenarche. The mechanisms underlying the adrenarche, the process leading to pre-puberty elevation of plasma androgens in higher primates, remain unknown, largely due to lack of any experimental model. By following up fetal and definitive cortex cell lines in mice, we showed that activation of protein kinase A (PKA) signaling mainly impacts the adult cortex by stimulating centripetal regeneration, with differentiation and then conversion of the zona fasciculata into a functional zR. Animals developed Cushing syndrome associated with primary hyperaldosteronism, suggesting possible coexistence of these hypersecretions in certain patients. Remarkably, all of these traits were sex-dependent: testicular androgens promoted WNT signaling antagonism on PKA, slowing cortical renewal and delaying onset of Cushing syndrome and the establishment of the zR in male mice, this being corrected by orchidectomy. In conclusion, zR derives from centripetal conversion of the zona fasciculata under cellular renewal induced by PKA signaling, determining the size of the adult cortex. Finally, we demonstrated that this PKA-dependent mobilization of cortical progenitors is sexually dimorphic and could, if confirmed in humans, account for female preponderance in adrenocortical pathologies.
Asunto(s)
Corteza Suprarrenal/embriología , Corteza Suprarrenal/crecimiento & desarrollo , Ratones , Modelos Animales , Glándulas Suprarrenales/embriología , Glándulas Suprarrenales/crecimiento & desarrollo , Animales , Diferenciación Celular , Femenino , Humanos , Masculino , Ratones Noqueados , Maduración Sexual/fisiologíaRESUMEN
We established the first adrenocortical tumor cell lines with complete zona fasciculata (ZF) cell phenotype from tumors induced in transgenic mice by large T-antigen of simian virus 40 under the control of the aldose reductase-like akr1b7 gene promoter. Adrenocortical tumor cell lines produced high amounts of corticosterone and were responsive to ACTH. All genes that are supportive for glucocorticoid synthesis including cyp21a1 and cyp11b1 were expressed, and most of them were transiently up-regulated by ACTH at transcriptional level: stimulation culminated after 3-6 h and returned to basal levels after 24 h. Taking advantage of these cells, we have examined the effect of ACTH on DAX-1 (dosage-sensitive sex reversal-adrenal hypoplasia congenita critical region on X-chromosome, gene 1) and SF-1 (steroidogenic factor 1), two transcription factors known to respectively repress and activate adrenocortical steroidogenesis by acting on common target genes. According to their antagonistic activities, DAX-1 mRNA and protein levels were transiently down-regulated by ACTH, whereas those of SF-1 were stimulated, with kinetics paralleling those of steroidogenic genes expression, notably of two known SF-1 target genes, star and akr1b7. This suggests an essential role of SF-1/DAX-1 proteins ratio to achieve proper ACTH control of steroidogenic gene expression in cells derived from ZF. This was confirmed in mice adrenals, where repression of dax-1 gene and concomitant up-regulation of sf-1, star, and akr1b7 genes were observed in response to ACTH stimulation. In conclusion, using both unique differentiated cell lines and in vivo approaches, we provide the first evidence that hormonally induced changes in SF-1/DAX-1 ratio are part of the molecular arsenal of ZF cells to fine tune ACTH responsiveness.
Asunto(s)
Neoplasias de la Corteza Suprarrenal/metabolismo , Hormona Adrenocorticotrópica/farmacología , Corticosterona/biosíntesis , Proteínas de Unión al ADN/análisis , Proteínas de Homeodominio/análisis , Receptores Citoplasmáticos y Nucleares/análisis , Factores de Transcripción/análisis , Zona Fascicular/metabolismo , Corteza Suprarrenal , Neoplasias de la Corteza Suprarrenal/patología , Aldehído Reductasa/genética , Animales , Línea Celular Tumoral , Receptor Nuclear Huérfano DAX-1 , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/fisiología , Regulación de la Expresión Génica , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/fisiología , Humanos , Masculino , Ratones , Ratones Transgénicos , Regiones Promotoras Genéticas , ARN Mensajero/análisis , Receptores Citoplasmáticos y Nucleares/genética , Receptores Citoplasmáticos y Nucleares/fisiología , Factor Esteroidogénico 1 , Factores de Transcripción/genética , Factores de Transcripción/fisiologíaRESUMEN
Adrenal cortex tumors are divided into benign forms, such as primary hyperplasias and adrenocortical adenomas (ACAs), and malignant forms or adrenocortical carcinomas (ACCs). Primary hyperplasias are rare causes of adrenocorticotropin hormone-independent hypercortisolism. ACAs are the most common type of adrenal gland tumors and they are rarely "functional," i.e., producing steroids. When functional, adenomas result in endocrine disorders, such as Cushing's syndrome (hypercortisolism) or Conn's syndrome (hyperaldosteronism). By contrast, ACCs are extremely rare but highly aggressive tumors that may also lead to hypersecreting syndromes. Genetic analyses of patients with sporadic or familial forms of adrenocortical tumors (ACTs) led to the identification of potentially causative genes, most of them being involved in protein kinase A (PKA), Wnt/ß-catenin, and P53 signaling pathways. Development of mouse models is a crucial step to firmly establish the functional significance of candidate genes, to dissect mechanisms leading to tumors and endocrine disorders, and in fine to provide in vivo tools for therapeutic screens. In this article, we will provide an overview on the existing mouse models (xenografted and genetically engineered) of ACTs by focusing on the role of PKA and Wnt/ß-catenin pathways in this context. We will discuss the advantages and limitations of models that have been developed heretofore and we will point out necessary improvements in the development of next generation mouse models of adrenal diseases.