Mi-2 and E93 coordinate robust cell cycle exit with terminal differentiation through enhancer decommissioning [RNA-seq]

Many postmitotic tissues coordinate robust cell cycle exit with the progression of terminal differentiation. While the signals involved in initiating cell cycle exit during terminal differentiation for some tissues have been described, less is known about the mechanisms that maintain a stable, non-cycling state. We previously found that chromatin accessibility changes at a subset of rate-limiting cell cycle genes occurs during maintenance of cell cycle exit, suggesting chromatin remodelers may play a key role in maintaining a robust postmitotic state during terminal differentiation. Here we show that the chromatin remodeler Mi-2 is required to ensure a stable, postmitotic state in Drosophila eyes and wings. Mi-2 alters chromatin accessibility and gene expression during cell cycle exit and terminal differentiation. Mi-2 and a transcription factor involved in tissue maturation, E93, close chromatin accessibility at an overlapping subset of potential enhancers that include the rate-limiting mitotic regulator string (cdc25c) and genes involved in the progression of terminal differentiation. E93 is also required for a stable, postmitotic state and we verified in vivo that Mi-2 and E93 cooperate to decommission an enhancer for the essential mitotic regulator string, to coordinate the transition to a postmitotic state with terminal differentiation. Enhancer decommissioning at the string locus provides a molecular explanation for the long-term, nearly irreversible postmitotic state observed in many tissues as terminal differentiation progresses. RNA-seq for Mi-2, E93 manipulations with controls in 24h and 44h APF Drosophila eyes

许多有丝分裂后组织（postmitotic tissues）会在终末分化（terminal differentiation）进程中，同步实现稳健的细胞周期退出（cell cycle exit）。尽管已有研究阐明了部分组织在终末分化过程中启动细胞周期退出的相关信号通路，但对于维持稳定非增殖状态的分子机制，目前仍知之甚少。我们此前的研究发现，在维持细胞周期退出的过程中，部分限速细胞周期基因（rate-limiting cell cycle genes）的染色质可及性（chromatin accessibility）会发生改变，这提示染色质重塑因子（chromatin remodeler）可能在终末分化阶段维持稳健的有丝分裂后状态中发挥关键作用。本研究证实，染色质重塑因子Mi-2是果蝇（Drosophila）眼和翅组织维持稳定有丝分裂后状态所必需的。Mi-2可在细胞周期退出与终末分化过程中，调控染色质可及性与基因表达。Mi-2与参与组织成熟的转录因子（transcription factor）E93，可在包含限速有丝分裂调控因子string（cdc25c）以及参与终末分化进程的基因在内的潜在增强子（enhancer）重叠子集区域，关闭染色质可及性。E93同样是维持稳定有丝分裂后状态所必需的；我们通过体内实验（in vivo）证实，Mi-2与E93可协同作用，对核心有丝分裂调控因子string的增强子进行失活退役，从而协调细胞向有丝分裂后状态的转变，并同步推进终末分化进程。string基因座（locus）处的增强子失活退役，为众多组织在终末分化进程中出现的长期、近乎不可逆的有丝分裂后状态提供了分子层面的解释。本研究对蛹化后24小时与44小时（APF, After Puparium Formation）的果蝇眼组织中，Mi-2与E93的基因操纵实验及对应的对照组样本进行了RNA测序（RNA-seq）。