AP-1 is a temporally regulated dual gatekeeper of reprogramming to pluripotency (ChIP-Seq). AP-1 is a temporally regulated dual gatekeeper of reprogramming to pluripotency (ChIP-Seq)

Somatic cell transcription factors are critical to maintaining cellular identity and constitute a barrier to human somatic cell reprogramming, yet a comprehensive understanding of the mechanism of action is lacking. To gain insight, we examined epigenome remodeling at the onset of human nuclear reprogramming by profiling human fibroblasts after fusion with murine embryonic stem cells. By ATAC-seq and ChIP-seq we identified enrichment for the AP-1 transcription factor c Jun at regions of early transient accessibility at fibroblast-specific enhancers. Expression of a dominant negative AP-1 mutant (dnAP 1) reduced accessibility and expression of fibroblast genes, overcoming the barrier to reprogramming. Remarkably, efficient reprogramming of human fibroblasts to iPSC was achieved by transduction with vectors expressing SOX2, KLF4, and inducible dnAP 1 demonstrating that dnAP-1 can substitute for exogenous human OCT4. Mechanistically, we show that the AP-1 component c-Jun has two unexpected temporally distinct functions in human reprogramming: i) to potentiate fibroblast enhancer accessibility and fibroblast-specific gene expression and ii) to bind to and repress OCT4 as a complex with MBD3. Our findings highlight AP-1 as a previously unrecognized potent dual gatekeeper of the somatic cell state. Overall design: Chip-seq by ChIPmentation in cultured human fibroblasts

体细胞转录因子（somatic cell transcription factors）对维持细胞身份至关重要，同时亦是人类体细胞重编程的天然屏障，但目前学界对其作用机制仍缺乏全面认知。为深入解析这一调控机制，我们通过对与小鼠胚胎干细胞融合后的人成纤维细胞开展组学分析，考察了人类细胞核重编程起始阶段的表观基因组重塑过程。借助ATAC-seq与ChIP-seq技术，我们在成纤维细胞特异性增强子的早期瞬时染色质可及区域中，鉴定出AP-1转录因子c-Jun的富集现象。显性负性AP-1突变体（dominant negative AP-1 mutant，dnAP-1）的表达可降低染色质可及性并抑制成纤维细胞特异性基因的转录，进而突破重编程障碍。值得注意的是，通过携带SOX2、KLF4以及可诱导型dnAP-1的载体进行转导，我们成功实现了人成纤维细胞向诱导多能干细胞（induced pluripotent stem cell，iPSC）的高效重编程，这表明dnAP-1可替代外源性人类OCT4发挥核心功能。从分子机制层面来看，我们证实AP-1组分c-Jun在人类重编程过程中具备两种前所未有的时序特异性功能：其一，增强成纤维细胞增强子的染色质可及性与成纤维细胞特异性基因的表达；其二，与MBD3形成复合物结合并抑制OCT4的转录活性。我们的研究发现凸显了AP-1作为此前未被认知的强效体细胞状态双重守门人的关键调控作用。整体实验设计：在体外培养的人成纤维细胞中通过ChIPmentation技术完成ChIP-seq测序。