Guo Ge, von Meyenn Ferdinand, Rostovskaya Maria, Clarke James, Dietmann Sabine, Baker Duncan, Sahakyan Anna, Myers Samuel, Bertone Paul, Reik Wolf, Plath Kathrin, Smith Austin
Wellcome Trust - Medical Research Council Cambridge Stem Cell Institute, University of Cambridge, Cambridge CB2 1QR, UK
Epigenetics Programme, Babraham Institute, Cambridge CB22 3AT, UK.
Development. 2017 Aug 1;144(15):2748-2763. doi: 10.1242/dev.146811.
Much attention has focussed on the conversion of human pluripotent stem cells (PSCs) to a more naïve developmental status. Here we provide a method for resetting via transient histone deacetylase inhibition. The protocol is effective across multiple PSC lines and can proceed without karyotype change. Reset cells can be expanded without feeders with a doubling time of around 24 h. WNT inhibition stabilises the resetting process. The transcriptome of reset cells diverges markedly from that of primed PSCs and shares features with human inner cell mass (ICM). Reset cells activate expression of primate-specific transposable elements. DNA methylation is globally reduced to a level equivalent to that in the ICM and is non-random, with gain of methylation at specific loci. Methylation imprints are mostly lost, however. Reset cells can be re-primed to undergo tri-lineage differentiation and germline specification. In female reset cells, appearance of biallelic X-linked gene transcription indicates reactivation of the silenced X chromosome. On reconversion to primed status, -induced silencing restores monoallelic gene expression. The facile and robust conversion routine with accompanying data resources will enable widespread utilisation, interrogation, and refinement of candidate naïve cells.
人们将大量注意力集中在将人类多能干细胞(PSC)转变为更原始的发育状态上。在此,我们提供一种通过短暂抑制组蛋白脱乙酰酶进行重编程的方法。该方案对多种PSC系均有效,且可在不改变核型的情况下进行。重编程后的细胞无需饲养层即可扩增,倍增时间约为24小时。抑制WNT可稳定重编程过程。重编程细胞的转录组与已分化的PSC有显著差异,并具有人类内细胞团(ICM)的特征。重编程细胞激活了灵长类特异性转座元件的表达。DNA甲基化整体降低至与ICM相当的水平,且是非随机的,在特定基因座处甲基化增加。然而,甲基化印记大多丢失。重编程细胞可再次分化为三胚层并进行种系定向分化。在雌性重编程细胞中,双等位基因X连锁基因转录的出现表明沉默的X染色体重新激活。在重新转变为已分化状态时,诱导沉默恢复单等位基因表达。这种简便且稳健的转变程序以及相关的数据资源将有助于广泛应用、研究和优化候选原始细胞。