RESUMEN
Prader-Willi syndrome (PWS) is characterized by neonatal hypotonia, developmental delay and hyperphagia/obesity. This disorder is caused by the absence of paternally expressed gene products from chromosome 15q11-q13. We previously demonstrated that knocking out ZNF274, a Kruppel-associated box-A-domain zinc finger protein capable of recruiting epigenetic machinery to deposit the H3K9me3 repressive histone modification, can activate expression from the normally silent maternal allele of SNORD116 in neurons derived from PWS induced pluripotent stem cells (iPSCs). However, ZNF274 has many other targets in the genome in addition to SNORD116. Depleting ZNF274 will surely affect the expression of other important genes and disrupt other pathways. Here, we used CRISPR/Cas9 to delete ZNF274 binding sites at the SNORD116 locus to determine whether activation of the maternal copy of SNORD116 could be achieved without altering ZNF274 protein levels. We obtained similar activation of gene expression from the normally silenced maternal allele in neurons derived from PWS iPSCs, compared with ZNF274 knockout, demonstrating that ZNF274 is directly involved in the repression of SNORD116. These results suggest that interfering with ZNF274 binding at the maternal SNORD116 locus is a potential therapeutic strategy for PWS.
Asunto(s)
Células Madre Pluripotentes Inducidas/patología , Factores de Transcripción de Tipo Kruppel/metabolismo , Neuronas/patología , Síndrome de Prader-Willi/patología , ARN Mensajero Almacenado/genética , ARN Nucleolar Pequeño/genética , Femenino , Humanos , Células Madre Pluripotentes Inducidas/metabolismo , Factores de Transcripción de Tipo Kruppel/genética , Neuronas/metabolismo , Síndrome de Prader-Willi/genética , Síndrome de Prader-Willi/metabolismoRESUMEN
Prader-Willi syndrome (PWS) is characterized by neonatal hypotonia, developmental delay and hyperphagia/obesity and is caused by the absence of paternal contribution to chromosome 15q11-q13. Using induced pluripotent stem cell (iPSC) models of PWS, we previously discovered an epigenetic complex that is comprised of the zinc-finger protein ZNF274 and the SET domain bifurcated 1 (SETDB1) histone H3 lysine 9 (H3K9) methyltransferase and that silences the maternal alleles at the PWS locus. Here, we have knocked out ZNF274 and rescued the expression of silent maternal alleles in neurons derived from PWS iPSC lines, without affecting DNA methylation at the PWS-Imprinting Center (PWS-IC). This suggests that the ZNF274 complex is a separate imprinting mark that represses maternal PWS gene expression in neurons and is a potential target for future therapeutic applications to rescue the PWS phenotype.
Asunto(s)
Impresión Genómica/genética , Factores de Transcripción de Tipo Kruppel/metabolismo , Síndrome de Prader-Willi/metabolismo , Alelos , Animales , Células Cultivadas , Inmunoprecipitación de Cromatina , Metilación de ADN/genética , Epigénesis Genética/genética , Impresión Genómica/fisiología , N-Metiltransferasa de Histona-Lisina/genética , N-Metiltransferasa de Histona-Lisina/metabolismo , Células Madre Pluripotentes Inducidas/metabolismo , Factores de Transcripción de Tipo Kruppel/genética , Ratones , Ratones Noqueados , Síndrome de Prader-Willi/genéticaRESUMEN
Quantitative analyses and models are required to connect a plant's cellular organisation with its metabolism. However, quantitative data are often scattered over multiple studies, and finding such data and converting them into useful information is time-consuming. Consequently, there is a need to centralise the available data and to highlight the remaining knowledge gaps. Here, we present a step-by-step approach to manually extract quantitative data from various information sources, and to unify the data format. First, data from Arabidopsis leaf were collated, checked for consistency and correctness and curated by cross-checking sources. Second, quantitative data were combined by applying calculation rules. They were then integrated into a unique comprehensive, referenced, modifiable and reusable data compendium representing an Arabidopsis reference leaf. This atlas contains the metrics of the 15 cell types found in leaves at the cellular and subcellular levels.