RESUMEN
The glucocorticoid (GC) receptor (GR) suppresses inflammation by activating anti-inflammatory and repressing pro-inflammatory genes. GR-interacting protein-1 (GRIP1) is a GR corepressor in macrophages, however, whether GRIP1 mediates GR-activated transcription, and what dictates its coactivator versus corepressor properties is unknown. Here we report that GRIP1 loss in macrophages attenuates glucocorticoid induction of several anti-inflammatory targets, and that GC treatment of quiescent macrophages globally directs GRIP1 toward GR binding sites dominated by palindromic GC response elements (GRE), suggesting a non-redundant GRIP1 function as a GR coactivator. Interestingly, GRIP1 is phosphorylated at an N-terminal serine cluster by cyclin-dependent kinase-9 (CDK9), which is recruited into GC-induced GR:GRIP1:CDK9 hetero-complexes, producing distinct GRE-specific GRIP1 phospho-isoforms. Phosphorylation potentiates GRIP1 coactivator but, remarkably, not its corepressor properties. Consistently, phospho-GRIP1 and CDK9 are not detected at GR transrepression sites near pro-inflammatory genes. Thus, GR restricts actions of its own coregulator via CDK9-mediated phosphorylation to a subset of anti-inflammatory genes.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/metabolismo , Proteínas Portadoras/metabolismo , Quinasa 9 Dependiente de la Ciclina/metabolismo , Glucocorticoides/metabolismo , Macrófagos/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Proteínas Adaptadoras Transductoras de Señales/deficiencia , Proteínas Adaptadoras Transductoras de Señales/genética , Animales , Sitios de Unión/genética , Proteínas Portadoras/antagonistas & inhibidores , Proteínas Portadoras/genética , Línea Celular , Células Cultivadas , Dexametasona/farmacología , Técnicas de Silenciamiento del Gen , Glucocorticoides/farmacología , Humanos , Inflamación/genética , Inflamación/metabolismo , Macrófagos/efectos de los fármacos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Modelos Biológicos , Proteínas del Tejido Nervioso/antagonistas & inhibidores , Proteínas del Tejido Nervioso/deficiencia , Proteínas del Tejido Nervioso/genética , Fosforilación , Receptores de Glucocorticoides/metabolismo , Elementos de Respuesta , Activación TranscripcionalRESUMEN
The risk posed by complex chemical mixtures in the environment to wildlife and humans is increasingly debated, but has been rarely tested under environmentally relevant scenarios. To address this issue, two mixtures of 14 or 19 substances of concern (pesticides, pharmaceuticals, heavy metals, polyaromatic hydrocarbons, a surfactant, and a plasticizer), each present at its safety limit concentration imposed by the European legislation, were prepared and tested for their toxic effects. The effects of the mixtures were assessed in 35 bioassays, based on 11 organisms representing different trophic levels. A consortium of 16 laboratories was involved in performing the bioassays. The mixtures elicited quantifiable toxic effects on some of the test systems employed, including i) changes in marine microbial composition, ii) microalgae toxicity, iii) immobilization in the crustacean Daphnia magna, iv) fish embryo toxicity, v) impaired frog embryo development, and vi) increased expression on oxidative stress-linked reporter genes. Estrogenic activity close to regulatory safety limit concentrations was uncovered by receptor-binding assays. The results highlight the need of precautionary actions on the assessment of chemical mixtures even in cases where individual toxicants are present at seemingly harmless concentrations.