RESUMEN
This corrects the article DOI: 10.1038/ncb3554.
RESUMEN
Naive pluripotency is established in preimplantation epiblast. Embryonic stem cells (ESCs) represent the immortalization of naive pluripotency. 2i culture has optimized this state, leading to a gene signature and DNA hypomethylation closely comparable to preimplantation epiblast, the developmental ground state. Here we show that Pramel7 (PRAME-like 7), a protein highly expressed in the inner cell mass (ICM) but expressed at low levels in ESCs, targets for proteasomal degradation UHRF1, a key factor for DNA methylation maintenance. Increasing Pramel7 expression in serum-cultured ESCs promotes a preimplantation epiblast-like gene signature, reduces UHRF1 levels and causes global DNA hypomethylation. Pramel7 is required for blastocyst formation and its forced expression locks ESCs in pluripotency. Pramel7/UHRF1 expression is mutually exclusive in ICMs whereas Pramel7-knockout embryos express high levels of UHRF1. Our data reveal an as-yet-unappreciated dynamic nature of DNA methylation through proteasome pathways and offer insights that might help to improve ESC culture to reproduce in vitro the in vivo ground-state pluripotency.
Asunto(s)
Antígenos de Neoplasias/metabolismo , Blastocisto/enzimología , Células Madre Embrionarias/enzimología , Epigénesis Genética , Proteínas de Neoplasias/metabolismo , Proteínas Nucleares/metabolismo , Células Madre Pluripotentes/enzimología , Complejo de la Endopetidasa Proteasomal/metabolismo , Animales , Antígenos de Neoplasias/genética , Blastocisto/citología , Proteínas Potenciadoras de Unión a CCAAT , Proteínas Cullin/metabolismo , Metilación de ADN , Regulación del Desarrollo de la Expresión Génica , Células HEK293 , Humanos , Ratones Endogámicos C57BL , Proteínas de Neoplasias/genética , Proteínas Nucleares/genética , Fenotipo , Dominios y Motivos de Interacción de Proteínas , Estabilidad Proteica , Proteolisis , Interferencia de ARN , Factores de Tiempo , Transcriptoma , Transfección , Ubiquitina-Proteína LigasasRESUMEN
A unique and complex signaling network allows ESCs to undergo extended proliferation in vitro, while maintaining their capacity for multilineage differentiation. Genuine ESC identity can only be maintained when both self-renewal and suppression of differentiation are active and balanced. Here, we identify Pramel7 (preferentially expressed antigen in melanoma-like 7) as a novel factor crucial for maintenance of pluripotency and leukemia inhibitory factor (LIF)-mediated self-renewal in ESCs. In vivo, Pramel7 expression was exclusively found in the pluripotent pools of cells, namely, the central part of the morula and the inner cell mass of the blastocyst. Ablation of Pramel7 induced ESC differentiation, whereas its overexpression was sufficient to support long-term self-renewal in the absence of exogenous LIF. Furthermore, Pramel7 overexpression suppressed differentiation in ESCs in vitro and in vivo. This process was reversible, as on transgene excision cells reverted to a LIF-dependent state and regained their capacity to participate in the formation of chimeric mice. Molecularly, LIF directly controls Pramel7 expression, involving both STAT3-dependent transcriptional regulation and PI3K-dependent phosphorylation of glycogen synthase kinase 3ß. Pramel7 expression in turn confers constitutive self-renewal and prevents differentiation through inactivation of extracellular signal-regulated kinase phosphorylation. Accordingly, knockdown of Pramel7 promotes ESC differentiation in presence of LIF and even on forced STAT3-activation. Thus, Pramel7 represents a central and essential factor in the signaling network regulating pluripotency and self-renewal in ESCs.
