Jin Kang-Xuan, Zuo Rujuan, Anastassiadis Konstantinos, Klungland Arne, Marr Carsten, Filipczyk Adam
Laboratory for Stem Cell Dynamics, Department of Microbiology, Division of Laboratory Medicine, Oslo University Hospital, Rikshospitalet, Oslo 4950, Norway.
Department of Molecular Medicine, Institute of Basic Medical Sciences, University of Oslo, Oslo 1072, Norway.
Proc Natl Acad Sci U S A. 2021 Dec 21;118(51). doi: 10.1073/pnas.2105192118.
N6-methyladenosine (mA) deposition on messenger RNA (mRNA) controls embryonic stem cell (ESC) fate by regulating the mRNA stabilities of pluripotency and lineage transcription factors (TFs) [P. J. Batista et al., 15, 707-719 (2014); Y. Wang et al., 16, 191-198 (2014); and S. Geula et al., 347, 1002-1006 (2015)]. If the mRNAs of these two TF groups become stabilized, it remains unclear how the pluripotency or lineage commitment decision is implemented. We performed noninvasive quantification of Nanog and Oct4 TF protein levels in reporter ESCs to define cell-state dynamics at single-cell resolution. Long-term single-cell tracking shows that immediate mA depletion by Mettl3 knock-down in serum/leukemia inhibitory factor supports both pluripotency maintenance and its departure. This is mediated by differential and opposing signaling pathways. Increased FGF5 mRNA stability activates pErk, leading to Nanog down-regulation. FGF5-mediated coactivation of pAkt reenforces Nanog expression. In formative stem cells poised toward differentiation, mA depletion activates both pErk and pAkt, increasing the propensity for mesendodermal lineage induction. Stable mA depletion by Mettl3 knock-out also promotes pErk activation. Higher pErk counteracts the pluripotency exit delay exhibited by stably mA-depleted cells upon differentiation. At single-cell resolution, we illustrate that decreasing mA abundances activates pErk and pAkt-signaling, regulating pluripotency departure.
信使核糖核酸（mRNA）上的N6-甲基腺苷（mA）沉积通过调节多能性和谱系转录因子（TFs）的mRNA稳定性来控制胚胎干细胞（ESC）的命运[P. J. 巴蒂斯塔等人，《细胞》15，707 - 719（2014年）；王Y. 等人，《细胞研究》16，191 - 198（2014年）；以及S. 盖拉等人，《科学》347，1002 - 1006（2015年）]。如果这两类TF的mRNA变得稳定，那么尚不清楚多能性或谱系定向决定是如何实现的。我们对报告基因胚胎干细胞中的Nanog和Oct4 TF蛋白水平进行了无创定量，以单细胞分辨率定义细胞状态动态。长期单细胞追踪表明，在血清/白血病抑制因子中通过敲低Mettl3使mA立即耗竭，既支持多能性维持，也支持其偏离。这是由不同且相反的信号通路介导的。FGF5 mRNA稳定性增加激活pErk，导致Nanog下调。FGF5介导的pAkt共激活增强Nanog表达。在倾向于分化的形成性干细胞中，mA耗竭激活pErk和pAkt，增加中内胚层谱系诱导的倾向。通过敲除Mettl3实现的稳定mA耗竭也促进pErk激活。更高水平的pErk抵消了稳定mA耗竭的细胞在分化时表现出的多能性退出延迟。在单细胞分辨率下，我们表明降低mA丰度会激活pErk和pAkt信号传导，调节多能性偏离。
