现今所了解的皮层基因表达多为来自神经元本身的自主调控， 我们最新研究发现RORB的表达主要依赖来自丘脑上行轴突的直接突触联系，这种非自主转录调控不是通过突触释放神经递质产生的电活动引起的， 也不由丘脑神经元释放的神经营养因子介导引起。为揭示RORB的非自主跨突触转录调控机制， 我们将采用yeast one-hybrid system 的方法， 以RORB基因的转录调控序列作为bait, 重点扫描丘脑cDNA文库。 扫描得到的阳性转录因子将进行Luciferase Assay 及ChIP sequencing 以确定他们具体DNA结合位点。为证实非自主调控因子的跨突触功能，我们将运用新型细胞隔层培养的研究方法及产生荧光或抗原标记丘脑特异性转基因小鼠模型进行验证。该项目将揭示及分离前所未有的丘脑及皮层之间的跨突触转录调控因子及可塑性的新分子机制，并为开发研究神经环路提供潜在的分子工具。

To date, regulatory mechanisms of gene expression in neocortex are largely known to be cell autonomous. However, we found expression of RORB in layer IV neurons of neocortex depends on thalamocortical axon innervation. Our preliminary experiments showed that the non-autonomous regulation of RORB gene expression is not mediated by neurotransmitter release at thalamocortical synapses nor neurotrophic factors released by thalamocortical axon terminals. In this project, we are going to identify the non-autonomous transsynaptic regulatory mechanism of RORB expression in neocortex. We will employ yeast one-hybrid system to identify thalamus regulatory factors binding to cis regulatory element of RORB gene. The cis regulatory elements of RORB gene will be used as baits to screen cDNA library from thalamus neurons. Positive binding proteins will be subjected for luciferase assays and ChIP sequencing to verify their DNA binding sites. To validate transsynaptic function of the identified trascription factors, we will employ neuronal double culture method and generate thalamus specific transgenic mice of fluorescence or antigen labeled the factors. This project will identify potential transsynaptic molecules that regulate RORB gene expression in neocortex and reveal a novel molecular mechanism of cortical plasticity. Moreover, our study will provide a new molecular tool to study complex neuronal circuits in the brain.