Department of Molecular, Cellular and Developmental Biology, University of Michigan, Ann Arbor, Michigan.
Neuroscience Graduate Program, University of Michigan, Ann Arbor, Michigan.
Endocrinology. 2020 Nov 1;161(11). doi: 10.1210/endocr/bqaa155.
Thyroid hormone (T3) plays pivotal roles in vertebrate development, acting via nuclear T3 receptors (TRs) that regulate gene transcription by promoting post-translational modifications to histones. Methylation of cytosine residues in deoxyribonucleic acid (DNA) also modulates gene transcription, and our recent finding of predominant DNA demethylation in the brain of Xenopus tadpoles at metamorphosis, a T3-dependent developmental process, caused us to hypothesize that T3 induces these changes in vivo. Treatment of premetamorphic tadpoles with T3 for 24 or 48 hours increased immunoreactivity in several brain regions for the DNA demethylation intermediates 5-hydroxymethylcytosine (5-hmC) and 5-carboxylcytosine, and the methylcytosine dioxygenase ten-eleven translocation 3 (TET3). Thyroid hormone treatment induced locus-specific DNA demethylation in proximity to known T3 response elements within the DNA methyltransferase 3a and Krüppel-like factor 9 genes, analyzed by 5-hmC immunoprecipitation and methylation sensitive restriction enzyme digest. Chromatin-immunoprecipitation (ChIP) assay showed that T3 induced TET3 recruitment to these loci. Furthermore, the messenger ribonucleic acid for several genes encoding DNA demethylation enzymes were induced by T3 in a time-dependent manner in tadpole brain. A TR ChIP-sequencing experiment identified putative TR binding sites at several of these genes, and we provide multiple lines of evidence to support that tet2 contains a bona fide T3 response element. Our findings show that T3 can promote DNA demethylation in developing tadpole brain, in part by promoting TET3 recruitment to discrete genomic regions, and by inducing genes that encode DNA demethylation enzymes.
甲状腺激素（T3）在脊椎动物发育中发挥关键作用，通过核 T3 受体（TR）发挥作用，通过促进组蛋白的翻译后修饰来调节基因转录。脱氧核糖核酸（DNA）中胞嘧啶残基的甲基化也调节基因转录，我们最近发现，在变态过程中，即依赖 T3 的发育过程中，非洲爪蟾蝌蚪大脑中的 DNA 去甲基化占主导地位，这促使我们假设 T3 在体内诱导这些变化。用 T3 处理前期蝌蚪 24 或 48 小时，增加了几个脑区 5-羟甲基胞嘧啶（5-hmC）和 5-羧基胞嘧啶以及甲基胞嘧啶双加氧酶 ten-eleven 易位 3（TET3）的免疫反应性。通过 5-hmC 免疫沉淀和甲基化敏感限制酶消化分析，甲状腺激素处理诱导了靠近 DNA 甲基转移酶 3a 和 Krüppel 样因子 9 基因中已知 T3 反应元件的位置特异性 DNA 去甲基化。染色质免疫沉淀（ChIP）试验表明，T3 诱导 TET3 募集到这些基因座。此外，T3 以时间依赖性方式诱导蝌蚪脑中几种编码 DNA 去甲基化酶的信使核糖核酸的诱导。TR ChIP-seq 实验鉴定了这些基因中的几个基因的假定 TR 结合位点，我们提供了多条证据支持 tet2 包含一个真正的 T3 反应元件。我们的研究结果表明，T3 可以促进发育中的蝌蚪脑的 DNA 去甲基化，部分是通过促进 TET3 募集到离散的基因组区域，并通过诱导编码 DNA 去甲基化酶的基因来实现。
