Rasd1 Modulates the Coactivator Function of NonO in the Cyclic AMP Pathway

All living organisms exhibit autonomous daily physiological and behavioural rhythms to help them synchronize with the environment. Entrainment of circadian rhythm is achieved via activation of cyclic AMP (cAMP) and mitogen-activated protein kinase signaling pathways. NonO (p54nrb) is a multifunctional protein involved in transcriptional activation of the cAMP pathway and is involved in circadian rhythm control. Rasd1 is a monomeric G protein implicated to play a pivotal role in potentiating both photic and nonphotic responses of the circadian rhythm. In this study, we have identified and validated NonO as an interacting partner of Rasd1 via affinity pulldown, co-immunoprecipitation and indirect immunofluorescence studies. The GTP-hydrolysis activity of Rasd1 is required for the functional interaction. Functional interaction of Rasd1-NonO in the cAMP pathway was investigated via reporter gene assays, chromatin immunoprecipitation and gene knockdown. We showed that Rasd1 and NonO interact at the CRE-site of specific target genes. These findings reveal a novel mechanism by which the coregulator activity of NonO can be modulated.所有活的生物体均具备自主的每日生理与行为节律，以此帮助自身与周遭环境实现同步。昼夜节律的同步化通过环磷酸腺苷（cyclic AMP, cAMP）与丝裂原活化蛋白激酶信号通路的激活得以实现。NonO（p54nrb）是一种多功能蛋白质，参与cAMP通路的转录激活过程，并参与昼夜节律的调控。Rasd1是一种单体G蛋白，被认为在增强昼夜节律的光诱导与非光诱导响应方面发挥关键作用。本研究通过亲和沉淀、免疫共沉淀与间接免疫荧光实验，鉴定并验证了NonO为Rasd1的相互作用伴侣蛋白。Rasd1的GTP水解活性是二者发生功能性相互作用的必要条件。本研究通过报告基因检测、染色质免疫共沉淀与基因敲低技术，探究了Rasd1与NonO在cAMP通路中的功能性相互作用。我们证实，Rasd1与NonO可在特定靶基因的cAMP应答元件（CRE）位点处发生相互作用。上述研究发现揭示了一种可调控NonO共调节因子活性的全新分子机制。