Single-cell epigenome analysis identifies transcriptional events controlling direct conversion of human fibroblasts to pancreatic ductal-like cells [ChIP-seq]. Single-cell epigenome analysis identifies transcriptional events controlling direct conversion of human fibroblasts to pancreatic ductal-like cells [ChIP-seq]

Cell fate can be reprogrammed by ectopic expression of lineage-specific transcription factors (TF). However, the exact cell state transitions during transdifferentiation are still poorly understood. Here, we have generated pancreatic exocrine cells of ductal epithelial identity from human fibroblasts using a set of six TFs. We mapped the molecular determinants of lineage dynamics using a factor-indexing method based on single-nuclei multiome sequencing (FI-snMultiome-seq) that enables dissecting the role of each individual TF and pool of TFs in cell fate conversion. We show that transition from mesenchymal fibroblast identity to epithelial pancreatic exocrine fate involves two deterministic steps: an endodermal progenitor state defined by activation of HHEX with FOXA2 and SOX17, and a temporal GATA4 activation essential for maintenance of pancreatic cell fate program. Collectively, our data suggest that transdifferentiation – although being considered a direct cell fate conversion method – occurs through transient progenitor states orchestrated by stepwise activation of distinct TFs. Overall design: Enrichment of histone marker H3K27ac in control human foreskin fibroblasts (HFFs) and reprogramming cells was analyzed by chromatin immunoprecipitation followed by deep sequencing (ChIP-seq).

细胞命运可通过谱系特异性转录因子（transcription factor, TF）的异位表达实现重编程。然而，转分化过程中的确切细胞状态转变机制仍未得到充分阐释。本研究通过一组共6种转录因子，从人成纤维细胞中诱导获得具有导管上皮表型的胰腺外分泌细胞。为系统解析谱系动态变化的分子决定因素，我们采用基于单细胞核多组学测序（single-nuclei multiome sequencing）的因子索引法（FI-snMultiome-seq），该方法可拆解单一转录因子及转录因子组合在细胞命运转化中的功能角色。研究表明，从间充质成纤维细胞表型向胰腺外分泌上皮表型的转化包含两个确定性步骤：一是由HHEX与FOXA2、SOX17共同激活所定义的内胚层祖细胞状态，二是对维持胰腺细胞命运程序至关重要的时序性GATA4激活。综上，本研究数据显示，尽管转分化被认为是一种直接的细胞命运转化手段，但其实则通过由不同转录因子逐步激活所调控的瞬时祖细胞状态完成。
实验设计：通过染色质免疫共沉淀联合深度测序（chromatin immunoprecipitation followed by deep sequencing, ChIP-seq），分析对照人包皮成纤维细胞（human foreskin fibroblasts, HFFs）及重编程细胞中组蛋白标记物H3K27ac的富集水平。