COUP-TF1调控甲状腺激素核受体α1介导的基因转录及其对大脑皮层发育影响的机制研究

Thyroid hormone plays an important role in the brain development, which is mainly mediated by thyroid hormone receptor α1（TRα1）on its target gene transcription. Thyroid hormone controls brain development in a time and space specific mechanism, which is undetermined. Chicken ovalbumin upstream promoter transcription factor 1 (COUP-TF1) belongs to orphan nuclear receptor family, Which can inhibit the gene transcription mediated by nuclear receptors. COUP-TF1 is expressed in the early phase of cerebral cortex development. The window period of thyroid hormone initiates cerebral cortex development is just after the declining expression of COUP-TF1. We speculate during the early time, COUP-TF1 inhibit the gene transcription mediated by TRα1. When the expression of COUP-TF1 declines, this inhibition is removed, followed by the initiation of thyroid hormone induced brain development. In order to confirm this assumption, our previous experiments utilized shRNA technology to inhibit the expression of COUP-TF1 in embryonic stem cells. We found that after COUP-TF1 knockdown, in the presence of T3, the expression of TRα1 mediated target genes were induced earlier. Based on these preliminary studies, we aim to study the molecular mechanism of COUP-TF1 regulates gene transcription mediated by TRα1. We utilized cerebral cortex COUP-TF1 conditional knockout mice to investigate whether COUP-TF1 involved in the mechanism mediated by thyroid hormone on the development of cerebral cortex in a way of specific time and space regulatory mechanism. This study will play an experimental research basis for clarifying the time and space specific regulatory mechanism mediated by thyroid hormone on the brain development.

甲状腺激素(TH)主要是通过甲状腺激素核受体（TR）介导的靶基因转录实现其对脑发育的调控。TH对脑发育的调控存在时空特异性，但其机制不明。COUP-TF1属孤儿核受体家族, 可以抑制核受体介导的基因转录，其在大脑皮层发育早期表达，TH启动大脑皮层发育恰好是COUP-TF1在大脑皮层表达下降后。我们推测在TH启动大脑皮层发育之前，COUP-TF1抑制了TRα1靶基因的转录，当COUP-TF1的表达下降，这种抑制作用得到了解除，从而启动了TH对大脑皮层发育的调控。为了证实这种假设，本课题组前期对小鼠胚胎干细胞的研究中发现:COUP-TF1抑制后促进了TRα1介导的大脑皮层发育相关的靶基因的表达。因此本课题将在前期研究结果基础上,进一步探讨COUP-TF1调控TRα1靶基因转录的分子机制，并利用大脑皮层COUP-TF1条件性敲除鼠，探讨COUP-TF1是否参与TH对大脑皮层的时空特异性调控。

甲状腺激素(TH)在脑发育中发挥重要作用。甲状腺激素作为核受体配体，主要是通过TRα1介导的靶基因转录实现其对脑发育的调控。TH对大脑皮层发育的调控存在时空特异性，但其机制不明。COUP-TF1属孤儿核受体家族, 可以抑制核受体介导的基因转录，其在大脑皮层发育早期表达，TH启动大脑皮层发育恰好是COUP-TF1在大脑皮层表达下降后。我们推测在TH启动大脑皮层发育之前，COUP-TF1抑制了TRα1靶基因的转录，当COUP-TF1的表达下降，这种抑制作用得到了解除，从而启动了TH对大脑皮层发育的调控。为了证实这种假设，本课题组前期对小鼠胚胎干细胞的研究中发现:COUP-TF1抑制后促进了TRα1介导的大脑皮层发育相关的靶基因的表达。为了探讨COUP-TF1调控TRα1靶基因转录的分子机制，我们首先应用ChIP方法证实COUP-TF1和TRα1可能竞争结合了RC3第一内含子区TRE，导致TRα1无法与RC3的TRE结合，抑制了RC3的转录。之后我们应用荧光素酶双报告证实COUP-TF1能够抑制TRα1介导的RC3基因转录。Co-IP证实在体内TRα1和COUP-TF1之间存在相互作用。进一步地，我们在COUP-TF1大脑皮层条件性敲除鼠模型中进行了验证，在脑发育早期，甲状腺激素发挥作用之前，正常对照组即使给与甲状腺激素，甲状腺激素的靶基因表达并无变化，而在COUP-TF1大脑皮层条件性敲除鼠中甲状腺激素的靶基因表达显著增加，而且COUP-TF1敲除还促进了神经前体细胞分化为神经细胞。本研究提示COUP-TF1通过抑制TRα1的基因转录，参与了甲状腺激素对大脑皮层的时空特异性调控。