RESUMO
Cell type specification relies on the capacity of undifferentiated cells to properly respond to specific differentiation-inducing signals. Using genomic approaches along with loss- and gain-of-function genetic models, we identified OCT4-dependent mechanisms that provide embryonic stem cells with the means to customize their response to external cues. OCT4 binds a large set of low-accessible genomic regions. At these sites, OCT4 is required for proper enhancer and gene activation by recruiting co-regulators and RAR:RXR or ß-catenin, suggesting an unexpected collaboration between the lineage-determining transcription factor and these differentiation-initiating, signal-dependent transcription factors. As a proof of concept, we demonstrate that overexpression of OCT4 in a kidney cell line is sufficient for signal-dependent activation of otherwise unresponsive genes in these cells. Our results uncover OCT4 as an integral and necessary component of signal-regulated transcriptional processes required for tissue-specific responses.
Assuntos
Diferenciação Celular , Linhagem da Célula , Células-Tronco Embrionárias/metabolismo , Fator 3 de Transcrição de Octâmero/metabolismo , Células-Tronco Pluripotentes/metabolismo , Via de Sinalização Wnt , Animais , Sítios de Ligação , Diferenciação Celular/efeitos dos fármacos , Reprogramação Celular , Células-Tronco Embrionárias/efeitos dos fármacos , Regulação da Expressão Gênica , Células HEK293 , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Humanos , Camundongos , Fator 3 de Transcrição de Octâmero/genética , Células-Tronco Pluripotentes/efeitos dos fármacos , Regiões Promotoras Genéticas , Interferência de RNA , Receptor alfa de Ácido Retinoico/genética , Receptor alfa de Ácido Retinoico/metabolismo , Receptores X de Retinoides/genética , Receptores X de Retinoides/metabolismo , Transcrição Gênica , Transfecção , Tretinoína/farmacologia , Via de Sinalização Wnt/efeitos dos fármacosRESUMO
Histone deacetylases (HDACs) 1, 2 and 3 form the catalytic subunit of several large transcriptional repression complexes. Unexpectedly, the enzymatic activity of HDACs in these complexes has been shown to be regulated by inositol phosphates, which bind in a pocket sandwiched between the HDAC and co-repressor proteins. However, the actual mechanism of activation remains poorly understood. Here we have elucidated the stereochemical requirements for binding and activation by inositol phosphates, demonstrating that activation requires three adjacent phosphate groups and that other positions on the inositol ring can tolerate bulky substituents. We also demonstrate that there is allosteric communication between the inositol-binding site and the active site. The crystal structure of the HDAC1:MTA1 complex bound to a novel peptide-based inhibitor and to inositol hexaphosphate suggests a molecular basis of substrate recognition, and an entropically driven allosteric mechanism of activation.