DNA dioxygenases Tet2/3 control epithelial differentiation [RNA-seq]

Execution of lineage-specific differentiation programs requires tight coordination between many regulators including Ten-eleven translocation (TET) family enzymes catalyzing 5-methylcytosine oxidation in DNA. Here, by using Keratin 14-Cre-driven ablation of Tet genes in skin epithelial cells, we demonstrate that ablation of Tet2/Tet3 result in marked alterations of hair shape and length followed by hair loss. We show that through DNA demethylation, Tet2/Tet3 control chromatin accessibility, Dlx3 binding and promoter activity of the Krt25 and Krt28 genes regulating hair shape, as well as regulate interactions between the Krt28 gene promoter and distal enhancer. Moreover, Tet2/Tet3 also control three-dimensional chromatin topology in Keratin type I/II gene loci via DNA methylation-independent mechanisms. These data demonstrate the essential roles for Tet2/3 in establishment of lineage-specific gene expression program and control of Dlx3/Krt25/Krt28 axis in hair follicle epithelial cells and implicate modulation of DNA methylation as a novel approach for hair growth control. We performed RNA-seq from sorted hair matrix keratinocytes of wild type and TetDKO (Krt14-Cre, Tet2 flox/flox, Tet3 flox/flox) mice at P4.5 which are treated or untreated with 5-aza.谱系特异性分化程序的执行，需诸多调控因子间的精密协同，其中包含催化DNA中5-甲基胞嘧啶氧化的十-十一易位（Ten-eleven translocation, TET）家族酶。本研究通过在皮肤上皮细胞中利用角蛋白14-Cre（Keratin 14-Cre）介导的Tet基因敲除，证实Tet2与Tet3的敲除会导致毛发形态、长度发生显著改变，并伴随脱毛现象。研究表明，Tet2与Tet3可通过DNA去甲基化调控染色质开放性、Dlx3的结合以及调控毛发形态的Krt25与Krt28基因的启动子活性，同时还可调控Krt28基因启动子与远端增强子之间的相互作用。此外，Tet2与Tet3还可通过不依赖DNA甲基化的机制，调控I型/II型角蛋白基因座的三维染色质拓扑结构。上述数据证实，Tet2/3在毛囊上皮细胞的谱系特异性基因表达程序建立以及Dlx3/Krt25/Krt28信号轴的调控中发挥关键作用，同时提示调控DNA甲基化可作为毛发生长调控的全新策略。我们对出生后第4.5天（P4.5）的野生型及TetDKO（Krt14-Cre, Tet2 flox/flox, Tet3 flox/flox）小鼠的分选毛发基质角蛋白细胞开展了RNA测序（RNA-seq），部分样本经5-氮杂胞苷（5-aza）处理，其余样本未作处理。